Department of Physiology Tirunelveli Medical College Tirunelveli – 627 011

A CROSS SECTIONAL STUDY TO CORRELATE THE DURATION OF DIABETES, GLYCEMIC CONTROL, AUTONOMIC NEUROPATHY AND DERMATOLOGICAL MANIFESTATIONS IN TYPE 2 DIABETES MELLITUS

Dissertation submitted to

THE TAMILNADU DR. M.G.R. MEDICAL UNIVERSITY,

CHENNAI – 600 032 In partial fulfillment of the requirement for the degree of Doctor of Medicine in Physiology (Branch V)

M.D. (PHYSIOLOGY) APRIL – 2017

DEPARTMENT OF PHYSIOLOGY TIRUNELVELI MEDICAL COLLEGE TIRUNELVELI – 627 011.

CERTIFICATE

This is to certify that the dissertation entitled, “A CROSS

SECTIONAL STUDY TO CORRELATE THE DURATION OF

DIABETES, GLYCEMIC CONTROL, AUTONOMIC NEUROPATHY

AND DERMATOLOGICAL MANIFESTATIONS IN TYPE 2

DIABETES MELLITUS” by Dr. SUJATHA ANDREW, postgraduate in PHYSIOLOGY (2014-2017), is a bonafide research work carried out under our direct supervision and guidance and is submitted to The

Tamilnadu Dr.M.G.R. Medical University, Chennai, for M.D., Degree

Examination in Physiology (Branch V), to be held in April 2017.

Dr. Ratna Manjushree Jayaraman M.D.,D.C.H., Dr.K. Sithy Athiya Munavarah, M.D., Associate Professor and Head , Dean, Department of Physiology, Tirunelveli Medical College, Tirunelveli Medical College, Tirunelveli – 11. Tirunelveli - 11.

ENDORSEMENT BY THE GUIDE

This is to certify that the dissertation entitled, “A CROSS SECTIONAL

STUDY TO CORRELATE THE DURATION OF DIABETES,

GLYCEMIC CONTROL, AUTONOMIC NEUROPATHY AND

DERMATOLOGICAL MANIFESTATIONS IN TYPE 2 DIABETES

MELLITUS’’ is a bonafide research work carried out by Dr. SUJATHA

ANDREW in the Department of Physiology, Tirunelveli Medical College &

Hospital, Tirunelveli – 11 under my direct guidance and supervision in partial fulfillment of the requirement for the award of the degree of M.D., in

PHYSIOLOGY (Branch – V) in April 2017.

GUIDE:

Dr. R.THENMOZHI M.D., D.C.P., Associate Professor, Department of Physiology, Tirunelveli Medical College, Tirunelveli – 11.

DECLARATION

I solemnly declare that the dissertation entitled, “A CROSS SECTIONAL STUDY TO CORRELATE THE DURATION OF DIABETES, GLYCEMIC CONTROL, AUTONOMIC NEUROPATHY AND DERMATOLOGICAL MANIFESTATIONS IN TYPE 2 DIABETES MELLITUS’’ is done by myself at Tirunelveli Medical College Hospital, Tirunelveli.

The dissertation is submitted to The Tamilnadu Dr. M.G.R. Medical University towards the partial fulfillment of the requirement for the award of M.D., Degree (Branch V) in Physiology.

Place :Tirunelveli-11 Dr. SUJATHA ANDREW,

Date : Postgraduate Student,

M.D. (Physiology),

Department of Physiology,

Tirunelveli Medical College,

Tirunelveli.

TIRUNELVELI MEDICALCDLLEGE

INSTITUTIONAL RESEARCIIETHICSCOMNI1TTF:F:

CERYIFICATE OF REGISTRATION &APPROVAL OFTHETIREC REF NO‹ 750/PHY/ 20 \5

Principallnx•estigator

ACKNOWLEDGEMENT

I primarily would like to thank the Lord Almighty for enabling me to complete this thesis.

I am grateful to Prof. Dr. K. Sithy Athiya Munavarah, M.D., Dean,

Tirunelveli Medical College, Tirunelveli for permitting me to carry out this study and for the constant encouragement towards the completion of this thesis.

I am immensely thankful to Dr. S. M. Kannan, M.S., Mch., Vice

Principal, Tirunelveli Medical College, Tirunelveli, for the support and encouragement.

I profoundly thank Dr. Ratna Manjushree Jayaraman, M.D.,

D.C.H., Associate Professor and Head, Department of Physiology,

Tirunelveli Medical College for the indispensible advice, expert guidance, information and support on different aspects of this work.

I take this opportunity to thank Dr. R. Thenmozhi, M.D., D.C.P.,

Associate Professor in Physiology, Tirunelveli Medical College to whom I owe a debt of gratitude for being my guide, who motivated, encouraged, helped and guided me through the completion of this study, which could not have been possible without her expertise.

I would like to thank Dr.B.Sujatha M.D., D.A., Associate Professor in Physiology for her constructive guidance and help in every aspect of my research and was imperative for the completion of this degree. I thank Dr. P. Nirmala Devi, M.D., (Derm) Professor and Head,

Department of Dermatology, Assistant Professors and Postgraduates of

Department of Dermatology, Tirunelveli Medical College Hospital for sparing their valuable time, to help with suggestions for the study.

I thank Dr.S.Saravanan ,M.D, D.M., (Neurology) Professor and

Head, Department of Neurology, Tirunelveli Medical College Hospital for his support and guidance. I wish to thank the Heads of Department of

Medicine and Department of Biochemistry for their help and encouragement.

I express my sincere thanks to Assistant professor Dr. S. Sudha,

M.D., D.D.V.L., other Assistant professors and Tutors of Department of

Physiology for their moral support, advice and help to this study.

I thank my Post Graduate colleagues as their assistance, cooperation and experience were essential for the completion of this work.

My special thanks to the statistician Mr.R.Jeromia Muthuraj,

M.Phil., PhD for his untiring work to accomplish the study in the best way.

I certainly thank the Librarian and staff of library, technical and non- technical staff of Central Laboratory, Department of Neurology, Department of Dermatology and Department of Physiology, Tirunelveli Medical College for their kind cooperation and assistance. I thank the participants of this study without them this study would not have been possible.

Finally, I thank my family members for their help and support during the entire course of this study.

CONTENTS

Sl. No. TITLE PAGE No.

1 INTRODUCTION 1

2 AIM AND OBJECTIVES 5

3 REVIEW OF LITERATURE 6

4 MATERIALS AND METHODS 38

5 RESULT ANALYSIS 43

6 DISCUSSION 70

7 SUMMARY AND CONCLUSION 81

8 LIMITATIONS 84 9 FUTURE SCOPE OF STUDY

10 BIBLIOGRAPHY

11 ANNEXURES

LIST OF ABBREVIATIONS

ANS Autonomic Nervous System

ADA American Diabetes Association

DM Diabetes Mellitus

DPP-4 Di Peptidyl Peptidase 4

FBS Fasting Blood Sugar

FDA Food and Drug Administration

FFA Free Fatty Acid

GLP Glucagon Like Polypeptide

GLUT Glucose Transporters

HbA1c Glycosylated Hemoglobin

HDL High Density Lipoprotein

IAPP Islet Amyloid Polypeptide

IDF International Diabetic Federation

IGT Impaired Glucose Tolerance

IGF Impaired Fasting Glucose

IGF - I Insulin like Growth Factor I

IGF - II Insulin like Growth Factor II

IRS Insulin Receptor Substrates LDL Low Density Lipoprotein

NCD Non-Communicable Disease

PBS Post prandial Blood Sugar

PNS Parasympathetic Nervous System

SEA South East Asia

SGLT Sodium Dependent Glucose Transporters

SSR Sympathetic Skin Response

TGL Triglycerides

WHO World Health Organization

A CROSS SECTIONAL STUDY TO CORRELATE THE DURATION OF DIABETES, GLYCEMIC CONTROL, AUTONOMIC NEUROPATHY AND DERMATOLOGICAL MANIFESTATIONS IN TYPE 2 DIABETES MELLITUS

INTRODUCTION

Diabetes mellitus is a non-communicable disease (NCD). NCD contribute to a huge burden on the healthcare system in India as compared to past when it was due to infectious diseases1,2. More than a half of the deaths in India are attributed to NCD of which diabetes mellitus (DM) is on the forefront and has emerged as an epidemic3. The severity of the present situation in the Indian context can be judged from the alarming figures that during 2004, diabetes has been directly responsible for 109,000 deaths, 1157 years of life lost and 2263 disability adjusted life years4. According to Global Report on Diabetes 2016 by

World Health Organization5DM mortality rate in South East Asia region is 115.3 per 100,000 in both sexes.

Diabetes mellitus is defined by WHO as, ―A metabolic disorder of multiple etiology characterized by chronic hyperglycemia with disturbances of carbohydrate, fat and protein metabolism resulting from defects in insulin secretion, insulin action, or both6. It can lead to long- term complications involving the nervous, cardiovascular, renal, and sensory organ systems. In the sensory organ systems, skin and eyes are affected mainly7.The major cause of morbidity and mortality in DM is mainly because of vascular complications and DM can be considered as a serious disease because of its complications8.

Type 2 diabetes mellitus is the commonest form of diabetes worldwide, attributing 90% of cases globally9, 10.Type 2 DM has now become a major public health problem in our country. Our country has the largest number of diabetics in the world second to China11.Though the fact that large numbers of individuals are at risk, 30-80% of the individuals in India are undiagnosed and are left untreated. This is to a large extent due to limited opportunities for diagnosis and lack of public awareness7. From epidemiological studies it is evident that without effective prevention and control programmes, the burden of diabetes is probable to increase worldwide. Because diabetes is affecting workforce people, it has a deleterious impact on both individual and national productivity both in developed and undeveloped countries. The World

Health Organization predicts the economic losses in national income for

India for about 237 billion7US Dollars.(www.cadiresearch.org).To stress the importance of this global epidemic, on 20th December 2006, the UN

General Assembly declared diabetes an international public health issue and declared World Diabetes Day as United Nations Day. To reduce the economic burden of this disease, it is necessary to diagnose early and to manage the observed risks.

Skin is affected by both acute metabolic default and the chronic degenerative complication of diabetes12. It is observed from studies that more than 33% of diabetic patients have some dermatologic

2 manifestations on a chronic run13. Atherosclerosis, microangiopathy, abnormal carbohydrate metabolism, neuron degeneration and impaired host defense mechanism all attribute to the pathogenesis of cutaneous complications14. UrbachUrbach E. demonstrated that skin sugar levels run parallel to the blood sugar levels. Cutaneous signs and symptoms of diabetes mellitus are extremely valuable to the physician as some may reflect the status of lipid metabolism and glycemic control15. It has been observed that the response to treatment is poor in cutaneous complications without control of diabetes.

Autonomic dysfunction is a major microvascular complication of

DM since it involves cardiovascular system, gastrointestinal system, genitourinary and sudomotor system. Autonomic involvement of CVS had been studied extensively but sudomotor function specifically the peripheral sympathetic nerve fibers which supply the eccrine sweat glands of skin is an unearthed treasure. Karamitsos and colleagues16 have reported that the progression of diabetic autonomic neuropathy is significant during the 2 years following its diagnosis.

Skin being the largest organ and the mirror of internal diseases, which can be easily examined, can act as a valuable tool in diagnosing

DM autonomic neuropathy. Autonomic neuropathy pertaining to skin is tested by Sympathetic skin response (SSR), one among the ANS function

3 tests is performed in this study to correlate autonomic neuropathy with cutaneous manifestations.

From many studies it is evident that the duration of diabetes mellitus and glycemic control are directly related to the complications of

DM and generally glycemic control is assessed by HbA1c17-19.Some studies have reported that there is a continuous, but not linear, relationship between the level of glycemia and the risk of development and progression of these complications.20,21

Since the incidence and prevalence of type 2 DM is more, type 2

DM subjects were selected for this study. As ANS function tests pertaining to skin manifestations in DM were not much studied, our study had been designed to correlate the duration of diabetes mellitus, glycemic control and autonomic neuropathy and skin manifestations in type 2 DM.

AIM AND OBJECTIVES

 To study the pattern of dermatological manifestations in

subjects with type 2 diabetes mellitus.

 To correlate duration of type 2 diabetes mellitus with

dermatological manifestations.

 To correlate the glycemic control (HbA1c) with dermatological

manifestations.

 To correlate autonomic neuropathy with dermatological

manifestations.

 To correlate HbA1c with lipid profile.

 To analyze the observed demographic profile of the study

subjects.

REVIEW OF LITERATURE

ANNALS OF DIABETES

Imhotep described polyuria in 3000BC22 and evidence of description of diabetes symptoms was recorded in Ebers papyrus in 1550

BC23. In ancient Egyptian‘s manuscripts descriptions of emptying large quantities of urine was found24. Indian physicians Sushruta and Charaka described madhumeha or honey urine in 400-500 BC by noticing ants attracted towards urine, they also described the association of type-2 with obesity and type-1 with young age24. The term diabetes was coined by

Arateus of Cappadocia in 150 AD meaning ‗siphon‘ to denote liquefaction of flesh and bone into urine25.Thomas Willis used the term mellitus in the 17 thcentury to separate the condition from diabetes insipidus25.

Mathew Dobson demonstrated that the residue after boiling urine to dryness tasted like brown sugar26. Paul Langerhans identified in 1869 the pancreatic cells which produce a substance that controls glucose levels and the islets of Langerhans was named after him27.In 1889 Joseph von Mering and Oscar Minkowski observed that pancreatectomized dogs developed signs and symptoms of diabetes and died28.

In 1910, Insulin was named by Sir Edward Albert Sharpey-Schafer, after the Latin word insula, meaning ―island‖, referring to the B cells of

Langerhans in the pancreas. In1921,Sir Frederick Grant Banting

6 and Charles Herbert Best identified insulin. Banting, Best and Collip purified the hormone insulin from bovine pancreas. Insulin injection was used first in a 14 year old boy on January 23 in 1922. In 1923 Banting and

John MacLeod were awarded Nobel Prize in Physiology or Medicine29. The

Eli Lilly Company produced insulin30 by 1923. In order to honor Banting, since 1991, his birthday is celebrated as World Diabetes Day on November

14. In 1949 Rachmiel Levine, discovered that insulin transports glucose into cells. Nobel Prize was awarded to Sir Fredrick Sanger for describing amino acid sequence of Insulin30.Yalow for discovering radioimmunoassay for insulin,received the 1977 Nobel Prize in Physiology or Medicine31. In1977

Boston researchers described the method of HbA1c testing. In 1978

Researchers at the City of Hope National Medical Center identified that E. coli bacteria can be used to produce insulin identical to human insulin.

In Mid-1990s incretin hormone GLP-1 was discovered. In 1996

Acarbose an alpha-glucosidase inhibitor was introduced. In 1997

Troglitazone was approved by the FDA. Repaglinide was developed in

1998. In 2002 the anti – CD 3 monoclonal antibody, hOKT3gamma1 (Ala-

Ala) was introduced.In2005 Exenatide, an incretin mimetic(GLP-1) drug was approved. In 2006 FDA approved sitagliptin phosphate, aDPP-4 inhibitor. In 2013 – Invokana was approved by FDA, a class of SGLT-2 inhibitors.

EPIDEMIOLOGY

DIABETES MELLITUS - A GLOBAL EMERGENCY32

Diabetes can be considered as a global health emergency of the 21st century. It is estimated that 415 million adults have diabetes currently and

612 million adults will be diabetic in 2040. There are currently 318 million adults with impaired glucose tolerance32.

GLOBAL SCENARIO

2015 estimates indicate that 8.5% (6.8-10.8%) of the adult population has diabetes. This is equivalent to 62.9 to 100.4 million people living with diabetes. Over half (52.1%) of these are undiagnosed. Nearly

47.5% of all adults with diabetes can be found urban area32. Currently 1 in 11 adults has DM. Both in developed countries and in developing countries type 2 DM is the commonest one33-36. Men are affected more than women. Prevalence of DM is more in productive age group.

Globally urban population has more incidence of DM than rural.

IDF estimation says that there are 193 million undiagnosed diabetic people in the world, who are more at risk of complications and currently one in two adults with DM is undiagnosed32. One in fifteen adults is found to have impaired glucose tolerance (IGT), and one in seven births have gestational diabetes32, both are at risk of developing diabetes later.

Worldwide diabetes and its complications are major causes of death and 5

World Diabetic Adults Population

415 million in 2015, 642 million in 2040

Top Ten Countries for Prevalence of Diabetes

Adults who Died from Diabetes, HIV/AIDS, Tuberculosis and Malaria

Prevalence of Diabetes Mellitus in World

2015

One in 11 adults has diabetes

2040

One in ten adults will have diabetes

million deaths are due to DM in 2015. Currently 1 death in every 6 sec is due to DM32. This is more than the combined number of deaths due to infections37namely Human Immuno Deficiency Virus infection,

Tuberculosis and Malaria. China, India, USA and the Russian Federation showed the highest number of deaths due to diabetes. On an average 12% of global health expenditure is spent on DM32.

REGIONAL PERSPECTIVE

In South East Asia, the prevalence rate in age the group 20-79 years is 8.5%. India has 69.2 million (56.2-84.8) people with diabetes, ranking second in the world32. India is expected to have 123.5 million

(99.1-150.3) diabetics by 2040. In South-East Asia the proportion of undiagnosed diabetics are 52.1% and it is 40.8 million in 2015 whereas in world it is 46.5% and 192.8 million32. In SEA expenditure spent on DM in 2015 is 7.3 billion USDollar32. India spent less than 3% of the global total (ID23 billion) expenditure on diabetes. India, Bangladesh and Sri

Lanka make up 99.0% of the SEA‘s total adult diabetes population. India is the largest contributor to regional mortality, with one million deaths attributable to diabetes32. In India 30-80% of the people are undiagnosed and are untreated. This is mainly due to lack of public awareness38.

According to Park‘s text book, 23 rd edition in 2004 there were 37.7 million diabetics in India, 21.4million were living in urban and 16.3 million in rural areas. DALY lost were 2.2 million39.

Population based studies showing the prevalence of type 2

diabetes in different parts of India40-46

2015

Chennai DM prevalence in

2001 - 13.5%

2006 - 14.3%

INDIA 69.2 million32

Estimated number of diabetics in India47,48. CARBOHYDRATE THE MAJOR FUEL

Carbohydrates are the bountiful natural organic molecules which have many functions. Carbohydrate means ―hydrate of carbon‖

(CH2 O)n. Glucose the important monosaccharide is the major metabolic fuel of our body. Constant source of glucose is an absolute requirement for human life49. The external source of glucose is carbohydrate rich foods. Ingested carbohydrate is first acted upon by salivary amylase then pancreatic amylase and in the intestinal lumen by glycosidases. The final products are the monosaccharides namely glucose, galactose and fructose.

Glucose and galactose are absorbed via SGLT-1 and fructose through

GLUT -5. From basolateral side of the enterocytes they enter the portal circulation through GLUT -2.Glucose enters the cells via SGLT or GLUT and utilized by all tissues for energy which is provided by glycolysis of glucose.

Normal Glucose Homeostasis

Normal glucose homeostasis is regulated by following processes:

(1) glucose production in the liver, (2) glucose uptake and utilization by peripheral tissues, chiefly skeletal muscle, and (3) actions of insulin and counter regulatory hormones glucagon, epinephrine, growth hormone and cortisol50.

Insulin reduces the production of glucose from the liver. Fasting plasma glucose levels is determined by hepatic glucose output. Hepatic

10 gluconeogenesis and glycogenolysis are facilitated during fasting by low insulin and high glucagon level. After a meal, insulin levels rise and glucagon levels fall in response to the glucose load. The most important stimulus that triggers insulin release is glucose. In peripheral tissues skeletal muscle and adipose tissue, secreted insulin binds to the insulin receptor, triggering intracellular responses that promote glucose uptake and postprandial glucose utilization, thereby maintaining glucose homeostasis. Abnormalities at various points, from synthesis and release of insulin by beta cells to insulin receptor interactions in peripheral tissues, can result in the diabetic phenotype50.

Glycolysis

In well fed state, glucose enters the cells by the action of insulin. In cells with mitochondria, by aerobic glycolysis pyruvate is formed from glucose and 2 ATPs are generated and lactate is formed in tissues which lack mitochondria or deprived of sufficient oxygen which is known as anaerobic glycolysis.

Glycogenesis

Dietary supply of carbohydrates is sporadic and body has certain mechanisms to store. The storage form is glycogen from which glucose will be supplied in needy time. The glycogen synthesis is named as glycogenesis which occurs during well fed state.

Metabolic Actions of Insulin

Glycogenolysis

In starvation and prolonged inter digestive period insulin secretion is low and glucagon secretion increases in order to supply glucose by glycogen breakdown. Glycogen breakdown is called as glycogenolysis.

Gluconeogenesis

Production of glucose from non carbohydrate sources is called as gluconeogenesis. During prolonged fasting, hepatic glycogen stores are depleted and glucose is synthesized from lactate, pyruvate and glycerol to provide glucose.

Glucose homeostasis by hormones

Glucose homeostasis is maintained by the hypoglycemic hormone insulin and the hyperglycemic hormones namely glucagon, catecholamines, thyroid hormones, growth hormone and adrenal glucocorticoids50 by various mechanisms.

The hormone insulin is secreted by endocrine pancreas. Imbalance in glucose homeostasis leads to hyperglycemia and disordered carbohydrate metabolism. The constellation of symptoms with hyperglycemia is called as Diabetes Mellitus.

MORPHOPHYSIOLOGY OF PANCREAS

In Greek pan means all, kreas means flesh. Pancreas is a soft structure which may be considered two organs in one because it has an exocrine and endocrine portion with entirely different functions.

Embryology

Pancreas has both endocrine tissue consisting of islets and exocrine tissue comprising of acinar cell. It originates from the dorsal and ventral regions of the foregut endoderm as dorsal and ventral buds on days 26 and 32 respectively which at 36 days fuse to form pancreas51.The posterior part of the head of the pancreas, develops from ventral bud while the rest develops from the dorsal bud. The islets are identified in the human embryo from about the 12th week with their own independent blood supply and begin to function by 16thweek51.The A-cell-rich islets develop from the dorsal pancreatic bud, and the F-cell-rich (pancreatic polypeptide-rich) islets from the ventral pancreatic bud50.

Gross anatomy

Pancreas is a 12.5-15 cm long leaf like structure and weighs about

100 gms. It is irregularly prismatic in shape, it has a head (caput pancreatis), neck, body (corpus pancreatis) and a tail (caudapancreatis). It is situated transversely across the posterior wall of the abdomen retroperitoneally52.

The arterial supply is from the lienal, and the pancreaticoduodenal branches of the hepatic and superior mesenteric arteries and venous

13 drainage into corresponding veins. Nerve supply is from the lienal plexus52,53.

Physiological anatomy

The endocrine portion of pancreas constitutes the Islets of

Langerhans measuring 76×175μm. They are found more in the tail region than in the body and head. β-islets constitute 2% of the volume of the gland, while the exocrine portion of the pancreas constitutes 80%, rest by ducts and blood vessels50. There is 1 to 2 million islets in humans.

There are four cell types in the islets namely A, B, D, and F cells depending on the staining properties and morphology. They are also called α, β, and δ cells respectively. The A or α cells secrete glucagon, the B or βcells secrete insulin, the D or δcells secrete somatostatin and the F cells secrete pancreatic polypeptide. The B cells are located in the center of each islet and account for 60–75% of the cells in the islets. A cells, D cells, F cells all surround B cells. A cells constitutes about 20% of the cells50.

Islets of Langerhans

PHYSIOLOGY OF INSULIN

Insulin is a polypeptide hormone with molecular weight 5808, has two chains of amino acids linked by disulfide bridges. When two amino acid chains are split, the biological activity is lost54.The amino acid composition differs slightly from species to species. This small difference is not sufficient to affect the biological activity of insulin, but capable of inducing antigenicity50.

Synthesis and secretion of insulin

In the rough endoplasmic reticulum of B cells insulin is synthesized, then transported to the Golgi apparatus for packaging into membrane-bound granules. These granules move to the plasma membrane and their contents are extruded out by exocytosis. Insulin then crosses the basement membrane of the B cell and the adjacent capillary endothelium to reach the bloodstream.

Gene for insulin synthesis is situated on the short arm of chromosome 11. Insulin is initially synthesized as preproinsulin50. From preproinsulin, proinsulin is formed by cleaving54. Proinsulin consists of three chains of peptides namely A, B, and C which are connected by disulphide bonds. C peptide connects the A and B chains, and helps in folding at Golgi apparatus and then is detached in the granules and insulin is formed55.The composition of the released products of B cells constitutes 90-97% of insulin along with equimolar amounts of C peptide

15 and the rest is proinsulin. C peptide can be measured by radioimmunoassay, and it is a measure of endogenous insulin production especially those on exogenous insulin therapy 50. There are 2 phases of insulin release early phase or rapid release56 which falls in 10 minutes and the slow or late phase which occurs over one hr50.

Fate of insulin

Insulin circulates in an unbound form in the blood, with half life of

6 minutes, and within 10 to 15 minutes it is cleared from the circulation.

Insulinase enzyme degrades insulin in the liver, in the kidneys and muscles, and slightly in other tissues54.Insulin like activity which is not suppressed by anti insulin antibodies is known as NSILA (‗non suppressible insulin like activity‘)50that is because of IGF I and IGF II.

Action of Insulin

Insulin is an anabolic hormone and can be called as ―hormone of abundance‖50. Its action can be classified into rapid, intermediate and delayed actions depending on the effects of insulin. Immediate action is increased transport of glucose, amino acids, and K + into insulin-sensitive cells. It promotes protein synthesis, inhibits protein degradation and enhances glycolysis, glycogen synthesis and lipid synthesis. The net effect of the hormone is storage of carbohydrate, protein, and fat50.

Mechanism of action of insulin

First insulin binds with and activates insulin receptor protein the activated receptor causes the ensuing effects. Once insulin binds with alpha unit subsequently beta subunit is autophosphorylated which is an example of an enzyme-linked receptor, which successively activates tyrosine kinase, which in turn causes phosphorylation of multiple intracellular enzymes including insulin-receptor substrates54 (IRS).

Effects of insulin on various tissues

Adipose tissue:Increased glucose entry, fatty acid synthesis, glycerol phosphate synthesis, triglyceride deposition and K + uptake, activation of lipoprotein lipase and inhibition of hormone-sensitive lipase.

Muscle: Increased glucose entry, glycogen synthesis, aminoacid uptake, protein synthesis in ribosome, ketone uptake, and K + uptake, decreased protein catabolism and decreased release of gluconeogenic amino acids.Liver: Decreased ketogenesis and glucose output, increased protein synthesis, lipid synthesis, glycogen synthesis, and glycolysis. General:

Increased cell growth50.

Glucose transporters

Glucose is transported through Glucose Transporters (GLUT) and

Sodium Dependent Glucose Transporters (SGLT). Glucose move into cells by facilitated diffusion through GLUT, by secondary active transport with Na2⁺ in intestine and kidney through SGLT. Seven GLUTs

Early and Late phases of insulin secretion

Mechanism of Action of Insulin

have been characterized50. In insulin responsive tissues GLUT 4 is the major glucose transporter.

Insulin Glucagon Molar Ratio

In balanced diet it is 2.3, in starvation I: G low, in well fed state

I:G high.

PHYSIOLOGICAL ANATOMY OF AUTONOMIC NERVOUS SYSTEM ANS is divided into two divisions: 1. Sympathetic division 2. Parasympathetic division Sympathetic system has preganglionic and postganglionic neurons. Cell body of preganglionic neuron is present in intermediolateral horn of spinal cord from T1 to L2. The preganglionic fibers synapse in the sympathetic ganglia and post ganglionic fibers pass to the effector organs. Sympathetic post ganglionic fibers pass to spinal nerves which are small type C fibers and supplies blood vessel, sweat glands and piloerector muscles of hair. In parasympathetic system preganglionic fibers synapse with the neuron present in the walls of effector organ and very short postganglionic fibers supply the organ. The main parasympathetic nerve is vague nerve which supplies the heart, lungs, esophagus, stomach, entire small intestine, proximal half of the colon, liver, gallbladder, pancreas, kidneys, and upper portions of the ureters. All preganglionic neurons are cholinergic in both the sympathetic and the parasympathetic nervous (PNS) systems and almost all of the postganglionic neurons of the PNS are also cholinergic. Most of the

Autonomic Nervous System

postganglionic sympathetic neurons are adrenergicbut the postganglionic sympathetic nerve fibers to the sweat glands are cholinergic since the neurotransmitter is acetylcholine54. When the sympathetic nerves are stimulated, the sweat glands secrete large volume of sweat.Eccrine sweating hydrates the skin and the bioelectrical property of skin is related to sweating. This skin biopotential can be recorded by using SSR.

SKIN FUNCTIONAL ANATOMY

Skin has superficial epidermis, mid dermis and deep sub cutis. The eccrine sweat gland is situated in reticular dermis and the duct passes through the papillary dermis epidermis and opens on the surface of epidermis through eccrine pore. The main function of eccrine gland is secretion of sweat. Eccrine gland is innervated by sympathetic postganglionic nerve fibers with acetylcholine as neurotransmitter. Sweat glands are distributed all over the body except lip, ear canal, glans penis, prepuce, labia minora and clitoris. In palms and soles the number of eccrine glands are more. Sweat has many functions. They play important role in water regulation, thermoregulation, excretory function and it hydrates the superficial layers of skin. Skin hydration is affected in autonomic neuropathy because of reduced secretion of sweat which may lead on to dry skin. The skin potentials are altered in dry skin. This is the basic principle behind SSR.

PRINCIPLE OF SSR

SSR is a change in potential recorded from surface of the skin and it represents sudomotor activity. It is a poly synaptic reflex. The effector as well as the generator of potential change are activated eccrine sweat glands.

Efferent of SSR reflex arc are sympathetic fibers from intermediolateral nucleus in T1-L2 part of spinal card. Post ganglionic fibers are unmyelinated type C fibers that innervate eccrine glands.

Central connection is presumably with hypothalamus, ventrolateral part of brainstem, medial and basal part of frontal lobe and medial part of temporal lobe.

According to Kucera P et al., SSR can be evoked by different stimuli namely electrical stimuli, deep inspiration, clicking sound via binaural earphones, magnetic stimulation, startle stimuli, LASER stimulation of skin, reflex hammer percussion on sternum, blinking, light stimuli, deglutition and skeletal movements.

Principle of Sympathetic Skin Response

PATHOPHYSIOLOGY OF DIABETES MELLITUS

Diabetes mellitus can be best described as ―Starvation in the midst of plenty‖50 because of reduced entry of glucose into various peripheral tissues and increased liberation of glucose into the circulation from the liver. Extracellular glucose excess and an intracellular glucose deficiency occur in many cells. Diabetes mellitus is classified mainly into Type1

DM and Type 2 DM, Gestational DM, Secondary DM due to other causes50.The disordered glucose homeostasis lead on to hyperglycemia.

Type 1 Diabetes Mellitus

Type1 DM results from autoimmune beta cell destruction and rarely by non immune mechanism. Type1 DM is the result of interactions of genetic, environmental, and immunologic factors57 that lead on to the destruction of the pancreatic beta cells and insulin deficiency. Type 1 DM can develop at any age but develops most commonly before 20 years of age11.

Type 2 Diabetes Mellitus

The key factors to the development of type 2 DM are insulin resistance and or abnormal insulin secretion. Insulin secretary defect is preceded by insulin resistance but diabetes develops only when insulin secretion becomes inadequate.

Disordered Plasma Glucose Homeostasis in Insulin Deficiency

Genetic considerations of type 2 diabetes mellitus

There is a strong genetic component for type 2 DM. The concordance of identical twins is between 70 and 90% in type 2 DM.

Individuals are at risk of developing DM if a parent is having type 2 DM.

If both parents have type 2 DM the risk is 40%. The etiology of type 2

DM is multifactorial. Both increased or reduced birth weight increases the risk of type 2 DM. Eventhough the genes that make individuals susceptible to type 2 DM are not well identified, it have been discovered that >70 genes has been associated with type 2 DM11. Currently genes encoding the peroxisome proliferator–activated receptor γ, inward rectifying potassium channel, zinc transporter, IRS and calpain 10 are identified.

Pathophysiology

In the early stages of the type 2 DM glucose tolerance tends to remain near-normal by beta cell secretion of insulin, but in some is unable to continue the hyperinsulinemic state as insulin resistance and compensatory hyperinsulinemia progress11 then impaired glucose tolerance develops. Still further decline in insulin secretion and an increase in hepatic glucose production give rise to overt diabetes. The

22 relative contribution of insulin resistance and impaired secretion of insulin to DM varies from individual to individual11.

I) Insulin resistance

Insulin resistance is defined as the failure of target tissues to respond normally to insulin57. It gives rise to decreased uptake of glucose in muscle, reduced glycolysis and fatty acid oxidation in the liver, and an

57 inability to control hepatic gluconeogenesis .

a) Obesity and Insulin Resistance:

Visceral obesity has been associated with type 2 DM patients.

Insulin resistance is present even with simple obesity without hyperglycemia. It is because of abnormal insulin signaling in circumstances of excess fat. The Adipo-insulin axis connection between adipose tissue and insulin is yet to be explained57. Metabolic syndrome has also been attributed to visceral obesity, which has components of insulin resistance, glucose intolerance, and cardiovascular risk factors such as hypertension and abnormal lipid profiles. The risk of diabetes increases as the body mass index increases. East and South Asians seem to develop type 2 DM at younger age and a lower BMI11.

b) Role of excess Free Fatty Acids:

An inverse correlation exists between fasting plasma FFAs and insulin sensitivity. The increased adipocyte mass lead on to more FFA

Pathogenesis of Type 2 Diabetes Mellitus

Mechanism of Insulin resistance and Beta cell dysfunction

formation and fat cell products in the circulation. Excess circulating FFAs are deposited in muscle and liver tissues in obese persons and causes more intracellular triglycerides level. Intracellular triglycerides and fatty acid metabolism products are potent inhibitors of insulin signaling and result in an acquired insulin resistance state. Through decrease in activity of key insulin-signaling proteins these lipotoxic effects of FFAs are mediated57.

c) Role of inflammation:

An inflammatory milieu by proinflammatory cytokines secreted in response to excess nutrients such as FFAs results in both peripheral insulin resistance and beta cell dysfunction. FFAs within macrophages and beta cells attract inflammasome, a multiprotein cytoplasmic complex that leads to secretion of the cytokine interleukin IL-1β. IL-1β, in turn, induces secretion of other pro-inflammatory cytokines from macrophages, islets, and other cells that are released into the circulation and act on the sites of insulin action to promote insulin resistance. Thus excess FFAs can impede insulin signaling directly within peripheral tissues, and indirectly through cytokines57.

d) Role of Adipokines:

Adipokines are released by adipose tissue in response to extracellular stimuli or changes in metabolic status. In response to excess

FFAs, adipocytes also release IL-1β and other proinflammatory cytokines

24 which promote peripheral insulin resistance. But adiponectin, an adipokine with insulin sensitizing activity decreases the inflammatory response.

e) Peroxisome proliferator-activated receptor-γ (PPARγ):

PPARγ58 is important for adipocyte differentiation. Activation of

PPARγ encourages secretion of anti hyperglycemic adipokines like as adiponectin, and relocates the deposition of FFAs toward adipose tissue and away from liver and skeletal muscle57.Gene mutation related to

PPARγ is currently identified to be one of the genetic factor responsible for insulin resistance.

II) Beta cell dysfunction

Beta cell dysfunction is due to the inability of beta cells to adjust themselves to the long-term demands of peripheral insulin resistance and increased insulin secretion. In insulin resistance insulin secretion is initially higher for each level of glucose than in controls. Insulin secretion initially increases in response to insulin resistance to preserve normal glucose tolerance. To begin with, the insulin secretory defect is mild57.

The response to other non glucose secretagogues, such as arginine, is preserved, but overall function is decreased by 50% at the onset of type 2

DM11. Increased secretion of proinsulin in type 2 DM reflects abnormalities in proinsulin processing. This is assumed to be due to a

Mean rates of insulin secretion in controls and in type 2 diabetes mellitus

second genetic defect superimposed upon insulin resistance leads to beta cell failure. Ultimately insulin secretary defect is progressive.

FFAs and glucose promote pro-inflammatory cytokines secretion from beta cells which leads to recruitment of macrophages and T cells into the islets resulting in local cytokine production and finally beta cell dysfunction and beta cell death. Replacement of islets by amyloid is seen in persons with long duration of type 2 DM. The abnormal aggregation of amylin, the islet amyloid polypetide (IAPP) secreted by the beta cells in conjunction with insulin is deposited as amyloid. IAPP also engages the inflammasome and encourages IL-1β secretion and sustains the inflammatory onslaught on surviving beta cells even late in the disease57.

METABOLIC ABNORMALITIES

I) Abnormal muscle fat metabolism:

Insulin resistance impedes glucose utilization by insulin-sensitive tissues and increases hepatic glucose output. Increased hepatic glucose output is predominantly responsible for increased FPG levels where as decreased peripheral glucose usage results in postprandial hyperglycemia.

But in red blood cells and brain glucose metabolism is not altered because they are insulin independent structures for glucose uptake.

Secondary to hyperinsulinemia insulin receptor levels and tyrosine kinase activity in skeletal muscle are reduced. Postreceptor defect play principal role in insulin resistance. Fat deposition in myocytes decreases oxidative phosphorylation and decrease insulin-stimulated ATP production. Because of diminished fatty acid oxidation the reactive oxygen species namely lipid peroxides are generated. Cell growth and differentiation pathways using protein kinase pathway are not resistant to insulin action as a result, hyperinsulinemia accelerates atherosclerosis11.

Diabetes mellitus has been labeled as ―more a disease of lipid than of carbohydrate metabolism‖50.FFA level is inversely proportional to insulin sensitivity. Insulin secretion and sensitivity are interrelated.

II) Glucose Toxicity and Lipotoxicity:

Chronic hyperglycemia diminishes islet function (―glucose toxicity‖).

Improvement in glycemic control is usually associated with improved beta cell function. In addition, elevation of free fatty acid levels (―lipotoxicity‖) and dietary fat may also affect islet function.

III) Increased hepatic glucose and lipid production:

In type 2 DM there is failure of insulin to suppress gluconeogenesis which gives rise to fasting hyperglycemia. As a result of insulin resistance, in adipose tissue lipolysis and free fatty acid flux from adipocytes are increased leading to increased lipid synthesis in hepatocytes. This is also responsible for the dyslipidemia in type 2 DM11.

According to Rolo and Palmeira four major59 molecular mechanisms have been implicated in hyperglycemia-induced tissue damage: activation of protein kinase C isoforms, increased hexosamine pathway, increased

Advance Glycation End product formation, and increased polyol pathway.

Overproduction of superoxide due to hyperglycemia appears to be the link between high glucose and tissue injury. Mutations in mitochondrial DNA

(mt DNA) and decreases in the copy number have been linked to the pathogenesis of type 2 DM. Lee and associate have shown an independent association between peripheral blood (leukocytes) mtDNA content and visceral adiposity. They conclude that the mtDNA copy number is a potential predictive marker for metabolic disturbances.

COMPLICATIONS OF DIABETES MELLITUS

Various systems in our body are affected in DM related complications and are responsible for the morbidity and mortality associated with the disease. Many individuals with type 2 DM have complications at the time of diagnosis because they usually have a long term of asymptomatic hyperglycemia before type 2 DM diagnosis. Early diagnosis will prevent complications of DM.

Complications related to DM could be either vascular and or non vascular complications. The vascular complications of DM are microvascular (retinopathy, neuropathy, and nephropathy) and macrovascular (coronary arterial disease, peripheral arterial disease, cerebrovascular disease). The diabetes specific complications are mainly related to microvascular pathology. Gastroparesis, infections, skin changes, and hearing loss3 are the non vascular complications.

Chronic hyperglycemia is the major contributor to the microvascular complications58 of both type 1 and type 2 DM. In the vasanervorum, degeneration of endothelial cells and pericytes occur60and these microvascular changes may precede the development of diabetic peripheral neuropathy61. Coronary heart disease correlate with fasting and postprandial plasma glucose levels as well the HbA1c. In macrovascular complications other factors such as hypertension and dyslipidemia also play important roles62. The Diabetes Control and Complications Trial

Association between HbA1c and Diabetes Mellitus

Complications

(DCCT) provided corroborative evidence that reduction in chronic hyperglycemia can prevent many complications63.

The DCCT study exhibited that improvement of glycemic control reduced neuropathy (60% reduction), nephropathy (54% reduction), microalbuminuria (39% reduction) and nonproliferative and proliferative retinopathy (47% reduction). Improved glycemic control also slowed the progression of early diabetic complications63.The results of the DCCT also speculated that persons in the intensive diabetes management group would gain 5.6 years free from lower extremity amputations, 5.8extra years free from end-stage renal disease (ESRD), and 7.7 more years of vision63. When all the DM complications were combined the intensive diabetes management group individuals would encounter 15.3 additional years of life without significant microvascular complications of DM compared to persons who received the standard treatment. The 30-year follow up data in the intensively treated group showed a sustained reduction in complications11.

The United Kingdom Prospective Diabetes Study (UKPDS) used many treatment regimens and monitored intensive glycemic control effect on the development of diabetic complications. UKPDS study showed that each decrease in percentage in HbA1c was associated with a 35% reduction in microvascular complications. Like DCCT, it showed sustained relationship between glycemic control and development of complications. Improved glycemic control also reduced the cardiovascular disease occurrence rate in the follow up period of >10 years63.

UKPDS‘s another major finding was that stringent blood pressure control remarkably decreased both macro and microvascular complications. Lowering blood pressure reduced the probability of DM- induced death, stroke, microvascular end points, retinopathy, and heart failure risk reduced between 32% and 56%. Kumamoto study showed reductions in the risks of retinopathy and nephropathy in lean type 2 DM

Japanese individuals with good Glycemic control. All these study results demonstrate the effectiveness of improved glycemic control in individuals of different nationality.

The Action to Control Cardiovascular Risk in Diabetes58,63

(ACCORD) and Action in Diabetes and Vascular Disease: Preterax and

Diamicron MR Controlled Evaluation (ADVANCE) trials also reported that improving the glycemic control lowered microvascular complications. Many studies showed development of neuropathy and other microvascular complications correlate well with the duration of diabetes and glycemic control62. However in the ACCORD study, there was an observation that significant increase in mortality in the intensively treated arm of glycemic control58. So the Glycemic control must be individualized.

The salient features of diabetes-related complications include

(1) Duration and degree of hyperglycemia correlate with complications

(2) Intensive glycemic control is rewarding in all types of DM (3) Blood

Complications of Diabetes Mellitus

pressure control is crucial chiefly in type 2 DM. (4) Survival in patients with type 1 DM is improving, and DM related complications are decreasing. (5) Not all individuals with DM develop DM associated complications. Certain individuals never develop nephropathy or retinopathy eventhough they have long-standing DM suggesting genetic susceptibility.

MECHANISM OF COMPLICATIONS

An important etiologic factor leading to complications of DM is chronic hyperglycemia. A hypothesis is that hyperglycemia leads to epigenetic changes that influence gene expression in affected cells.

The theories, how hyperglycemia direct to the complications of DM are (1) Formation of advanced glycosylation end products causes nonenzymatic glycosylation of intra- and extracellular proteins. (2)

Glucose metabolism is increased by means of sorbitol pathway by enzyme aldose reductase. (3) Increased formation of diacylglycerol results in activation of protein kinase C. (4) Increased flux through the hexosamine pathway, which generates fructose-6-phosphate. Vascular endothelial growth factor A is associated with diabetic proliferative retinopathy, in diabetic nephropathy TGF-β is increased which encourages basement membrane synthesis. There is increased production of reactive oxygen species in the mitochondria because of hyperglycemia and reactive oxygen species may activate all the four pathways.

DIABETIC NEUROPATHY

Diabetic neuropathy affects 50% -90% of individuals8 with type 2 DM.

It may present as polyneuropathy, mononeuropathy, and/or autonomic neuropathy. The duration of DM, glycemic control, increased BMI all increase the risk of neuropathy. Smoking, coronary artery disease, elevated triglycerides, and hypertension are also related with development of diabetic peripheral neuropathy63.

Natural History of Neuropathy

Neuropathies have two distinct entities: 1) those that progress gradually with increasing duration of diabetes 2) those that remit,usually completely. Sensory and autonomic neuropathies typically progress although the symptoms of mononeuropathies, radiculopathies, tend to recover64. Progression of neuropathy is related to glycemic control in both type 1DM and type 2 DM65,66.

DM neuropathy is broadly classified into somatic and visceral62.

Polyneuropathy/Mononeuropathy

Distal symmetric polyneuropathy is the commonest form of diabetic neuropathy which manifests as distal sensory loss and pain. The pain may be acute or chronic, but in due course, the pain subsides and disappears, but sensory deficit persists. Mononeuropathy presents as pain and motor weakness in a single nerve, the third cranial nerve is most commonly affected.

Autonomic Neuropathy

The ANS supplies all organs in the body and consists of an afferent and an efferent system and a third component the neuropeptidergic system and its neurotransmitters substance P, vasoactive intestinal polypeptide , and CGRP58.The organ systems affected in DM autonomic neuropathy includes the sweat glands, pupil, genitourinary system, adrenal medullary system, gastrointestinal system and cardiovascular system58, 67, 68. Parasympathetic or sympathetic nerves are affected in one or more visceral systems. 30–50% of patients are dead within 10 years on developing symptoms of autonomic neuropathy from sudden cardio- respiratory arrest62. Patients with postural hypotension (a drop in systolic pressure of 30 mm Hg or more on standing from the supine position) have the highest subsequent mortality62. Silent MI, respiratory failure, amputations, and sudden death are hazards for diabetic patients with cardiac autonomic neuropathy69-72 and the increase in sudden death in the

Simplified view of Peripheral Nervous System

ACCORD study is likely due to the presence of autonomic dysfunction60.

Because of autonomic neuropathy and failure of glucose counterregulation there may be absence of warning signs of impending neuroglycopenia known as ―hypoglycemic unawareness‖ which may be detrimental to the individuals. Karamitsos and colleagues73 have reported that progression of diabetic autonomic neuropathy occurs during progression of duration.

Autonomic neuropathy and skin

In hairy skin a functional defect is noticed before neuropathy develops74, and it is correctable with antioxidants75. The clinical counterpart of autonomic neuropathy affecting skin manifest as skin that is dry and cold(xerosis), loses the ability to sweat, and develops fissures and cracks that are portals of entry for organisms leading to infectious ulcers and gangrene58,63.

Sweat disturbances

Hyperhidrosis of the upper bodymainly gustatory sweating63, is a characteristic feature of autonomic neuropathy related to sudomotor function. Anhidrosis of the lower body is also common in autonomicneuropathy63. Loss of lower body sweating can cause dry, brittle skin that cracks easily, predisposing the patient to ulcer formation58 that can lead to loss of a limb11,58,63.

Diabetic foot

Foot problems are the most preventable among the late complications. Joslin wrote in 1934 that ―diabetic gangrene is not heaven- sent, but earthborn‖. Autonomic neuropathy encourages drying of the skin and fissure formation allowing them to become large and infected11, and a significant foot ulcers will finally go for amputation. Risk factors for ulcer foot or amputation include smale gender, diabetes for >10 years, smoking, visual impairment, peripheral neuropathy, and poor glycemic control11.

All three components of neuropathy—sensory, motor, and autonomic can contribute to ulceration in the foot58. Sympathetic autonomic neuropathy affecting the lower limbs results in reduced sweating, dry skin, and development of cracks and fissures58. Neuropathy as a contributory cause to foot ulceration and ultimately to amputation had been confirmed. Hence autonomic neuropathy can be considered serious in DM complications76. One study observed that there is seven fold higher risk for foot ulcer in individuals with neuropathy than in those without neuropathy77.

ADA CRITERIA FOR DIABETES DIAGNOSIS OF DM78

FPG ≥126 mg/dL (7.0 mmol/L)

Fasting is defined as no caloric intake for ≥8 hours

2-hr PG ≥200 mg/dL(11.1 mmol/L) during OGTT (75-g)

A1C ≥6.5% (48 mmol/mol)

Random PG ≥200 mg/dL(11.1 mmol/L)

Type 2 Diabetes Risk Factors  Physical inactivity  First-degree relative with diabetes

 High-risk race or ethnicity  Women who delivered a baby weighing >9 lb or who have previously been diagnosed with GDM

 HDL-C <35 mg/dL and/or TG >250 A1C ≥5.7%, IGT, or IFG mg/dL

 Hypertension (≥140/90 or on treatment)  History of cardiovascular disease

 Conditions associated with Insulin Resistance

MATERIALS AND METHODS

METHODOLOGY

This study was conducted at Tirunelveli Medical College Hospital.

The subjects were randomly selected from Diabetology outpatient department and Dermatology outpatient department. Institutional ethical committee clearance was obtained. Study subjects were selected randomly by using following inclusion and exclusion criteria.

Study design

Cross sectional descriptive observational study.

Duration of study

6 months.

Inclusion criteria

1. Type 2 diabetes mellitus subjects.

2. Age group 30 to 70 years.

3. Both genders.

Exclusion criteria

1. Type 1 diabetes mellitus.

2. Drug induced diabetes.

3. Gestational diabetes.

4. Co morbidities – Thyroid disorders, Epilepsy, Bronchial

Asthma and Tuberculosis.

5. Subjects on medications which can induce neuropathy.

6. Alcoholics.

7. Hansen‘s disease.

Materials used for the study

 Consent form.

 Proforma - to register the demographic details, to record the

clinical details and investigation results.

 Stadiometer to measure height in cm.

 Weighing machine to record body weight in kg.

 Mercury sphygmomanometer ( Diamond) to record blood pressure.

 Appropriate test tubes for blood sample collection.

 Fully automated Biochemical Analyzer EM360, ERBA, for

estimation of blood sugar, urea, creatinine and lipid profile.

 ERBA Chem 5x ,Trans Asia semi auto analyzer for estimation of

HbA1c.

 MEDICAID EMG-2000.Digital polygraph – 4 channel, Neurostim

machine to elicit sympathetic skin response.

The study was carried out after explaining the procedure to the subjects and after obtaining the informed written consent.

Study Protocol:

The demographic details of the subjects namely age, sex, education, occupation, address were registered. History related to personal habits like smoking, alcoholism and family history regarding DM in parents and siblings were enquired. History related to duration of DM, medications for DM whether on oral hypoglycemic agents or on insulin or on both or

39 on any alternative medicine was enquired. Pre and present history of skin diseases was elicited. History of thyroid disorder, bronchial asthma, tuberculosis and epilepsy was enquired. History of any other drug intake, history of neuropathy due to other diseases were enquired.

Weight was measured by using standard digital weighing machine.

Height was measured by using stadiometer to the nearest 0.5cm. Body mass index was calculated by using Quetelet‘s formula. (Weight in kg/Height in m2).Pulse rate was counted manually and blood pressure was recorded with Diamond sphygmomanometer after making the subject to rest for 30 minutes. (Pickering et al 2005) Blood pressure

>140/90mmHg was considered as high.

Subjects were subjected to detailed dermatological examination by dermatologist. Under strict aseptic precautions venous blood sample was collected in appropriate containers both in fasting state and in post prandial state. The blood samples were analyzed in Central Laboratory

Tirunelveli Medical College Hospital, Tirunelveli for HbA1c, sugar, urea, creatinine and lipid profile. HbA1c was tested in ErbaChem 5x semi automated analyzer by Particle enhanced Immuno-Turbidimetric method.

This is an IFCC approved reference method. Measuring range in this method is 3.98% - 15.42%. HbA1C reference values are as follows: for non diabetic 4- 6 % is normal, for DM subjects< 7% good control, 7% -

8%fair control, > 8% poor control.

MEASUREMENT OF HEIGHT WEIGHT

RECORDING OF BLOOD PRESSURE

COLLECTION OF BLOOD SAMPLE FOR INVESTIGATIONS

ERBA CHEM 5X MACHINE FOR HbA1c ANALYSIS

HbA1c ANALYSIS

FULLY AUTOMATED BIOCHEMICAL ANALYZER - EM 360 - BLOOD ANALYSIS

Blood sugar, urea, creatinine and lipid profile were tested in EM

360 Fully automated Biochemical analyzer. Blood sugar was tested by using Trinder‘s method, End point / Fixed Time method. Blood sugar normal range for Fasting is 70-110mgm/dL, for Postprandial is 110-140 mgm/dL.

Normal range for blood urea is 15-40 mgm%. Blood urea was analyzed by Kinetic UV test – F.T9 (UV- SLR). Serum creatinine was tested by Jaffe‘s Method, Initial Rate. The normal range is 0.2 -1.2 mgm%. Lipid profile normal range for TGL is 70-200 mg/dL, for LDL is

70-130mg/dL, for HDL is 35-60 mg/dL.TGL was tested by GPO – PAP

Method, End Point, Total cholesterol by Enzymatic calorimetric test method, HDL by precipitation method.LDL was calculated by following

TGL formula.LDL= Total Cholestrol - (HDL + VLDL). VLDL = 5

SSR Test

SSR was done in Department of Neurology by using Medicaid EMG-

200 Digital polygraph 4-channel Neurostim machine. We followed the standard method which was described by Shahani et al.,(1984) and Kucera et al., in his study, Sympathetic Skin response: review of the method and its clinical use (2004). Test was performed with subject supine, relaxed in a semi-darkened quiet room with ambient temperature 22⁰C to 24⁰C. Body temperature was checked and found normal during the test. The surface silver disc electrodes were used in this study.

MEDICAID EMG-2000 DIGITAL POLYGRAPH

SYMPATHETIC SKIN RESPONSE

SYMPATHETIC SKIN RESPONSE – PROCEDURE

After cleaning the area thoroughly the active electrode was placed with electrode jelly over palm and sole of both side extremities. Reference electrode was placed over dorsal aspects of palm and sole of both side extremities. Single electrical stimulus was given for 0.1msec duration with

10mA intensity to contra lateral median nerve, contra lateral posterior tibial nerve as well as to ipsilateral median nerve and ipsilateral posterior tibial nerve. Recording from all the four extremities was done. The absence of

SSR from at least two extremities after electrical stimulation followed by deep inspiration was considered abnormal. To avoid habituation one stimuli per minute was given, and after 2 minutes next stimuli was given. Likewise

5 stimuli were given. Presence or absence of response was observed.

All the results were tabulated in MS office Excel sheets and descriptive statistical analysis was done by using IBM SPSS statistical software version 20.Demographic profile was analyzed .Pearson correlation analysis was done. Skin manifestations were cross tabulated with duration of

DM and analyzed. Glycemic control was analyzed with skin manifestations.

SSR test was cross tabulated and analyzed with skin manifestations. HbA1c and lipid profile was analyzed by using Pearson correlation analysis. ‗p‘ value < 0.05 was considered significant.

RESULT ANALYSIS

TABLE – 1: DESCRIPTIVE ANALYSIS

Descriptive Statistics

Range Minimum Maximum Mean Standard Variance Deviation

AGE in years 39.00 31.00 70.00 56.0200 10.30542 106.202

HEIGHT (cm) 42 138 180 160.36 9.430 88.920

WEIGHT (kg) 54 44 98 65.63 12.069 145.670

BMI 17.00 18.49 35.49 25.4883 4.00730 16.058

HbA1c % 6.40 4.40 10.80 7.7990 1.39070 1.934

BLOOD SUGAR 214.00 82.00 296.00 146.1100 41.72151 1740.685 FASTING (mgm/dL)

BLOOD SUGAR 276.00 120.00 396.00 253.5600 70.81322 5014.512 POSTPRANDIAL(mg/dL)

BLOOD UREA (mgm %) 166.00 15.00 181.00 27.4900 18.00421 324.151

SERUM CREATININE 8.70 0.50 9.20 0.9558 0.86960 0.756 (mgm %)

TRIGLYCERIDES 179 68 247 158.40 46.797 2189.980 (mgm/dL)

LOW DENSITY 123 65 188 116.72 28.789 828.789 LIPOPROTEIN(mgm/dL)

HIGH DENSITY 27 23 50 37.98 6.184 38.242 LIPOPROTEIN(mgm/dL)

DEMOGRAPHIC PROFILE

TABLE – 2: AGE

Age in years Frequency Percentage

30 - 40 7 7.0%

40 - 50 26 26.0%

50 - 60 31 31.0%

60 - 70 36 36.0%

TOTAL 100 100.0%

AGE IN YEARS

40 35 30 25 20 31% 36% 15 26% 10 5 7%

0 NUMBER OF TYPE2 DIABETICS 30 - 40 40 - 50 50 - 60 60 - 70 AGE IN YEARS

Age: Mean age being 56.02 years, minimum age was 31 years and maximum was 70 years. Distribution of age group was 60- 70 years 36%,

50-60 years 31%, 40-50 years 26%, 30-40 years 7%. Number of subjects in< 60 years was 64 %,> 60 years was 36%.

TABLE – 3: SEX

Sex Frequency Percentage

Male 36 36.0%

Female 64 64.0%

Total 100 100.0%

36 were males and 64 were females. Male: Female ratio 1:1.77.

SEX

70 60 50 64% 40 30 36% 20 10 0 MALE FEMALE

TABLE – 4: EDUCATION

Educational status Frequency Percentage Illiterate 3 3.0% Schooling 64 64.0% Diploma/ITI 8 8.0% UG* 18 18.0%

PG** 3 3.0% Professional 4 4.0%

Total 100 100.0%

UG* undergraduates PG** postgraduates

EDUCATIONAL STATUS

ILLITERATE SCHOOLING DIPLOMA/ITI UG PG PROFESSIONAL

3% 4% 3%

18%

8% 64%

Educational status: 64% had done schooling, 18% undergraduates, 8% diploma/ITI, 4% professionals, 3% post graduates and 3% illiterates.

TABLE – 5: OCCUPATION

Occupation Frequency Percentage

Cooley 19 19.0%

House wife 40 40.0%

Business 8 8.0%

Private 16 16.0%

Government 17 17.0%

Total 100 100.0%

Occupational status: house wives 40%, cooley 19%, government employee17%, private sector 16% and business 8%

OCCUPATION 40% 40 35 30 25 19% 16% 17% 20 15 8% 10 5

0 NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS

OCCUPATION

TABLE – 6: URBAN/RURAL

Place Frequency Percentage

Rural 42 42.0%

Urban 58 58.0%

Total 100 100.0%

58 subjects were from urban area and 42 subjects were from rural area

Urban: Rural ratio 1.08:1

PLACE

60 50 40 58% 30 42% 20 10

0 NUMBER OF TYPE2 OF DIABETICS TYPE2 NUMBER RURAL URBAN PLACE

TABLE –7: SMOKING

Smoking Frequency Percentage

Non smoker 75 75.0%

Smoker 25 25.0%

Total 100 100.0%

Smokers were 25% non smokers 75%

SMOKING

60 75% 40 25% 20

0 NUMBER OF TYPE2 OF DIABETICS TYPE2 NUMBER SMOKER NON SMOKER SMOKING

FAMILY HISTORY

56% 60 50 40 20% 30 12% 12% 20 10 0

NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS MOTHER AND MOTHER FATHER NEITHER FATHER ALONE ALONE PARENT FAMILY HISTORY 1

Family history of DM was positive in 44%, negative in 56%.

Among positive family history, the subjects whose father alone had

DM was 12%, mother alone had DM was 20%, father and mother both had DM were 12%.

DIABETES IN SIBLINGS

59% 60 50 40 30 13% 15% 13% 20 10 0

NUMBER OF TYPE2 DIABETICS BROTHER BROTHER SISTER ALONE NEITHER AND SISTER ALONE SIBILINGS SIBLINGS

History of siblings having DM showed that both brother and sister had DM in 13%, brother alone had DM in 13%, sister alone had in DM

15%, neither in 59%

TABLE -8: BODY MASS INDEX†

Weight Frequency Percentage

Under Weight 1 1.0%

Healthy weight 45 45.0%

Over Weight 37 37.0%

Obesity 1 16 16.0%

Obesity 2 1 1%

Total 100 100.0%

BMI† was calculated by using Quetelet‘s formula.

Weight in kg / Height in m2

BMI Under weight : < 18.5

Normal : 18.5 – 24.9

Over weight : 25.0 -25.9

Obesity 1 : 30.0-34.9

Obesity 2 : 35.0-39.9

Extreme obesity : ≥ 40

Adapted from WHO 1999, WHO 2000, WHO 2004

BMI

35 45% 30 37% 25

15 16% NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS 10

5 1% 1%

0 UNDER HEALTHY OVER WEIGHT OBESITY 1 OBESITY 2 WEIGHT WEIGHT BMI

Among 100 DM subjects observed mean body mass index was

25.48, minimum BMI was 18.49, maximum was 35.49, SD 4.00. under weight 1%, healthy weight (Normal) 45%, over weight 37%, obesity

1- 16%, obesity 2- 1%

TABLE –9: DURATION OF DIABETES MELLITUS

Years Frequency Percentage <1 13 13.0% 1 - 5 38 38.0% 5 - 10 28 28.0% 10 - 20 19 19.0% >20 2 2.0% Total 100 100.0%

The observed duration of diabetes mellitus findings were< 10 years

79% and >10 years 21%.More subjects were in < 10years duration.

Sub grouping showed < 1 year 13%, 1-5 years 38%, 5-10 years

28%,10-20 years 19%, > 20 years 2%.Subjects with duration of DM for

1-5 years outnumbered others.

DURATION

38% 40 35 28% 30 25 19% 20 13% 15 10 2% 5

0 NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS <1 1 to 5 5 to 10 10 to 20 >20 DURATION OF DIABETES IN YEARS

TABLE – 10: SKIN MANIFESTATIONS

Skin Manifestations Frequency Percentage Bacterial Infection 13 13% Fungal Infection 27 27% Viral Infection 4 4% Parasitic infection 1 1% Psoriasis Vulgaris 4 4% Pityriasis Rosea 1 1% Lichen Planus 1 1% Xerosis 26 26% Diabetic Dermopathy 2 2% Diabetic Bullae 1 1% Diabetic Ulcer foot 1 1% Vitiligo 4 4% Acanthosis Nigricans 3 3% Pruritus 2 2% Lipodystrophy 1 1% Lichen Simplex Chronicus 1 1% Telogen Effluvium 1 1% Melasma 1 1% Polymorphic light eruption 1 1% Henoch Schonlein Purpura 1 1% Pompholyx 1 1% Erythrodrema 1 1% Urticaria 1 1% Allergic Contact Dermatitis 1 1% Total 100 100%

Skin manifestations showed: Infections ( bacterial 13%, fungal

27%, viral 4% and parasitic1%) totally 45%, Xerosis 26% , Psoriasis vulgaris 4%, Vitiligo vulgaris 4%, Acanthosis nigricans 3%, Diabetic dermopathy 2% Pruritus 2%, Diabetic bullae, Diabetic ulcer foot,

Lipodystrophy, Pityriasis rosea, Lichen planus, Erythroderma, Henoch

Schonlein Purrpura, Polymorphic light eruption, Pompholyx, LSC,

Telogen effluvium, Urticaria, Melasma, Allergic contact dermatitis all each 1%. For correlation study acanthosis nigricans 3%, Pruritus 2%,

Lipodystrophy 1%, Erythroderma 1%, HSP 1%, Polymorphic light eruption 1%, Pompholyx 1%, LSC 1%, Telogen effluvium1% ,

Urticaria1%, Melasma 1%, Allergic contact dermatitis 1% totally 15% were considered as others.The observation was Infections were the predominant skin lesions 45%, followed by xerosis 26%.

DIABETIC ULCER – FOOT

HERPES ZOSTER

DIABETIC BULLAE

XEROSIS

DIABETIC DERMOPATHY

CELLULITIS LEG

TABLE -11: HbA1c%

HbA1c% Frequency Percentage < 7 28 28.0% 7 - 8 22 22.0% > 8 50 50.0%

Total 100 100.0%

< 7 % Good control

7-8% Fair control

>8% Poor control

22% good control, 28% fair control and 50% showed poor control.

Observed HbA1c values ranged from 4.40% to 10.80%. Mean value was

7.799%. Standard deviation was1.390.

HbA1c%

50 40 50% 30 20 28% 22% 10

0 NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS <7 7 to 8 >8 HbA1c%

TABLE-12: BLOOD SUGAR FASTING

Blood sugar( F)mgm/dL Frequency Percentage

Normal (70 -110) 20 20.0%

Abnormal (>110) 80 80.0%

Total 100 100.0%

Fasting blood sugar was normal in 20% and high in 80%. The average value was 146.11mgm/dL. Lowest value in the range was

82mgm/dL and the highest value in the range was 296mgm/dL.

BLOOD SUGAR FASTING (mg/dl)

40 80% 20 20%

0 NUMBER OF TYPE2 OF DIABETICS TYPE2 NUMBER NORMAL (70 - 110) ABNORMAL (>110) BLOOD SUGAR FASTING (mg/dl)

TABLE –13: BLOOD SUGAR POSTPRANDIAL

Blood sugar (PP)mgm/dL Frequency Percentage

Normal (110-140) 11 11.0%

High (>140) 89 89.0%

Total 100 100.0%

Postprandial blood sugar values ranged between 120-396mgm/dL.

Mean value obtained was 253.56mgm/dL. Subjects with normal values were 11%. and abnormal values were 89%

BLOOD SUGAR POSTPRANDIAL (mg/dl)

100

80 89% 60 40 11% 20

NUMBER OF TYPE2 OF NUMBER TYPE2 DIABETICS 0 NORMAL (110 - 140) ABNORMAL (>140) BLOOD SUGAR POSTPRANDIAL (mg/dl)

TABLE –14: BLOOD UREA

Blood Urea (mgm%) Frequency Percentage

Normal 93 93.0% (15 - 40)

Abnormal 7 7.0% (>40)

Total 10 100.0%

Observed urea values showed 93% having normal result 7% having high value. Mean value was 27.49 mgm%. Range being between 15-181 mgm%. SD 18.004

TABLE-15: SERUM CREATININE

Serum creatinine (mgm%) Frequency Percentage

Normal 92 92.0% (0.2 - 1.2)

Abnormal (>1.2) 8 8.0%

Total 100 100.0%

It was observed in this study that 92% of subjects were having values within normal range 8% of subjects were having high creatinine values. Mean value being 0.9558mgm%. Minimum observed value was

0.5mgm% and maximum observed value was 9.2 mgm%. One subject was having abnormal creatinine value but normal blood urea level.SD0.869

TABLE- 16: SYMPATHETIC SKIN RESPONSE

SSR Frequency Percentage

Absent 28 28.0%

Present 72 72.0%

Total 100 100.0%

SSR was present in 72% and absent in 28%.

SYMPATHETIC SKIN RESPONSE

40 72%

20 28%

0 NUMBER OF TYPE2 OF DIABETICS TYPE2 NUMBER PRESENT ABSENT SYMPATHETIC SKIN RESPONSE

TABLE – 17 : DURATION OF DM AND SKIN

MANIFESTATIONS - CROSS TABULATION

Duration In Years Total no Skin of Manifestations <1 1 - 5 5 - 10 10 -20 >20 subjects Bacterial Infection 0 5 4 4 0 13 Fungal Infection 7 11 7 2 0 27 Viral Infection 0 3 1 0 0 4 Parasitic Infection 0 1 0 0 0 1 Psoriasis Vulgaris 0 3 0 1 0 4 PityriasisRosea 0 1 0 0 0 1 Lichen Planus 0 0 0 1 0 1

Xerosis 1 4 12 7 2 26 Diabetic 0 0 0 2 0 2 Dermopathy Diabetic Bullae 0 0 0 1 0 1 Diabetic Ulcer foot 0 0 0 1 0 1 Vitiligo 0 1 3 0 0 4 Others 5 9 1 0 0 15 Total 13 38 28 19 2 100

In relation to duration subjects were sub grouped into < 1 year , 1-5 years, 5-10 years , 10-20 years ,>20 years and correlated with skin manifestations.Diabeticdermopathy, diabetic bullae, diabetic foot ulcer were observed in subjects with > 10 year duration of DM. The other observed manifestations occurred irrespective of duration of DM.

TABLE -18 : HbA1c% AND SKIN MANIFESTATIONS – CROSS TABULATION

HbA1c Skin manifestations Total <7% 7 – 8% >8%

Bacterial Infection 0 1 12 13

Fungal Infection 3 7 17 27

Viral Infection 1 1 2 4

Parasitic Infection 1 0 0 1

Psoriasis Vulgaris 3 1 0 4

Pityriasis Rosea 1 0 0 1

Lichen Planus 1 0 0 1

Xerosis 5 8 13 26

Diabetic Dermopathy 0 0 2 2

Diabetic Bullae 0 0 1 1

Diabetic Ulcer foot 0 0 1 1

Vitiligo 3 0 1 4

Others 10 4 1 15

Total 28 22 50 100

Glycemic control was assessed by HbA1c% and was sub grouped into <7% good control, 7% - 8% fair control, >8% poor control. These sub groups were correlated with skin manifestations. The observation was major percentage of skin infections, diabetic bulla, diabetic ulcer foot,diabetic dermopathy all occurred with poor glycemic control. Out of 45 subjects with infection 31 subjects had poor(>8%) glycemic control. Totally 26subjects had xerosis, among that 13 had xerosis with poor glycemic control, 8 subjects with fair control and 5 subjects with good control.

TABLE–19: SSR AND SKIN MANIFESTATIONS CROSS

TABULATION

SSR Skin manifestations Total Not recordable Recordable Bacterial Infection 13 4 9 Fungal Infection 27 1 26 Viral Infection 4 1 3 Parasitic Infection 1 0 1 Psoriasis Vulgaris 4 0 4 Pityriasis Rosea 1 0 1 Lichen Planus 1 0 1 Xerosis 26 15 11 Diabetic Dermopathy 2 2 0 Diabetic Bullae 1 1 0 Diabetic Ulcer foot 1 1 0 Vitiligo 4 1 3 Others 15 2 13 Total 100 28 72

Sympathetic skin response was tested and observed whether recordable and not recordable. Our observation was in 72% it was recordable and in 28% SSR was not recordable.15 subjects with xerosis showed no response and 11 showed response. In diabetic bullae, diabetic dermopathy, diabetic ulcer foot SSR was not recordable.

TABLE - 20

Descriptive Statistics Standard Lipid Profile Range Minimum Maximum Mean Variance Deviation TRIGLYCERIDES 179 68 247 158.40 46.797 2189.980 (mgm/dL) LOW DENSITY 123 65 188 116.72 28.789 828.789 LIPOPROTEIN(mgm/dL) HIGH DENSITY 27 23 50 37.98 6.184 38.242 LIPOPROTEIN(mgm/dL)

Lipid profile normal range for TGL is 70-200 mg/dL, for LDL is 70-

130mg/dL, for HDL is 35-60 mg/dL.

The observed laboratory values:

TGL ranged from 68 to 247mgm/dL, mean value was 158.40, SD 46.797.

LDL ranged from 65mgm/dL, to 188mgm/dL, mean value 116.72mgm/ dL, SD28.789 .

HDL ranged from 23mgm/dL to 50mgm/dL, mean value was 37.98, SD

6.184.

TABLE –21 : CORRELATION BETWEEN HbA1c AND TGL

Test Statistical analysis HbA1c TGL Pearson Correlation 1 .541 HbA1c Sig. (2-tailed) .0001* N 100 100 Pearson Correlation .541 1 TGL Sig. (2-tailed) .0001* N 100 100

There was positive correlation of glycemic control and TGL value (.541)

p value <0.05 was considered as significant . Observed p value =

0.0001* . Highly significant correlation between HbA1c and TGL was observed.

TABLE –22 : CORRELATION BETWEEN HbA1c AND LDL

Test Statistical Analysis HbA1c LDL

Pearson Correlation 1 .490

HbA1c Sig. (2-tailed) .0001*

N 100 100

Pearson Correlation .490 1

LDL Sig. (2-tailed) .0001*

N 100 100

Observed positive correlation between HbA1c and LDL (.490) and a highly significant p value was noted p value = 0.0001*

P value <0.05 was considered significant.

Correlation between HbA1c and HDL

It was observed there was negative correlation between HbA1c and HDL

DISCUSSION

This study was attempted to establish the association between the duration of diabetes, individual glycemic control, autonomic neuropathy and manifestations of the skin in type 2 diabetes mellitus individuals. A total of 100 type 2 diabetes subjects with cutaneous manifestations, who visited the Dermatology OPD and Diabetology OPD were selected at random and the above variables were studied.

Demographic Profile

Analysis of demographic profile showed that the mean age in this study was 56.02 years79. Age distribution was < 60 years 64 subjects, and

> 60 years 36 subjects. Our study population age group distribution was consistent with Goyal A et al., study in which the mean age was 57.44 years. According to National Center for Chronic Disease Prevention and

Health Promotion, National Diabetes Statistics Report 2014, the incidence of DM, the prevalence and undiagnosed DM are more in 45-64 year age group. Our study population age distribution also showed more number of subjects in < 60 years of age, which is the economically productive group. The economically active population is considered from

15 to 59 years (ref: censusindia.gov.in).Almost half of diabetes deaths occur in people under the age of 70 years (ref:cadiresearch). Diabetes results in high healthcare costs, loss of labor productivity, and decreased rates of economic growth.

The study subjects of this study showed male: female ratio of

1:1.77. The female predominance was seen in our study as reported by

Mahajan et al80 and Romano et al81.According to IDF Diabetes Atlas seventh edition 2015 report there is slight preponderance of male gender in DM. But death due to DM is seen more in females. According to CADI

82research 55% of diabetes deaths are in women.

Residential status showed urban: rural83 ratio of 1.08:1.IDF statistics showed more incidence and prevalence in urban population when compared to rural population. But nowadays incidence and prevalence of DM among rural population is in increasing trend.

Educational status observed in this study was illiterates 3% (criteria based on census India) and 97% literates. House wives were more in this study.

Family history of DM revealed 44 subjects had positive family history and 56% had negative family history. Among positive family history 12 subjects‘ father had DM and 20 subjects‘ mother had DM, both father and mother had DM in 12% of subjects. Strong genetic component is suggested for the etiology of type 2 DM11.Our study observation also showed significant positive family history. If both parents had DM, then the chance of an individual getting DM increase11 by 40%.If father alone is diabetic the risk of developing DM is 9% and if the mother alone is diabetic the risk11 is 3% .Alcoholics were excluded from the study because alcohol induced neuropathy can induce abnormalities84 in SSR.

Among the study subjects smokers were 25% .Smoking along with hyperglycemia and disordered lipid metabolism make the individual more prone for both microvascular and macrovascular complications of DM

Increase in BMI is one of the risk factors11 of DM. BMI results of this study showed that 45% of subjects had normal BMI. According to studies, Asian Indians develop DM even with normal BMI11. 54% study subjects had overweight and obesity.

Skin Manifestations

In our study skin infections were the predominant skin manifestations and seen in 45% of the total study sample population80,81.

The observed manifestations of infective origin included 27% of fungal,

13% of bacterial, 4% of viral and 1% of parasitic. It was similar with studies done by Mahajanet al80 and Nigametal85. Our observation was among 27% fungal infections 13% had candidal infection, 12% had dermatophyte infection, 1% onychomycosis, 1% Tinea versicolor.

Furunculosis was the most observed bacterial infection and observed in

9%, followed by cellulitis in 4%.It is notable that diabetic patients are susceptible to infections presumably due to hyperglycemia and defects in polymorphonuclear leukocyte function. Viral and parasitic infections

(scabies) in diabetic patients have not been much expressed in earlier studies while it has been observed in this study. 3 subjects had herpes zoster, one subject had viral wart and one had scabies. According to

Mahajan80 dermatophyte infection was the most common (37.16%) followed by bacterial infection (33.78%) candidal infection (22.97%)and viral infection (6.08%) and Al-Mutairi outlined bacterial infection as more common infection than dermatophyte, candida and viral infections.

Cutaneous infections are considered as important markers to diagnose

DM86.Naheed et al.,87from Lahore, Miracle et al88 and Baloch et al89all observed skin infection as commonest skin manifestation. Our study was consistent with Nawaf et al.,90study. Radhu et al91noted fungal infections more common in type 2 diabetics. In our study also fungal infections were the predominant infection. In contrary, one study from Sargodha

Pakistan92 recorded skin infection only in 23% of diabetic patients.

Skin Manifestations

The relative high observation of skin infections of our study can be explained as uncontrolled diabetes mellitus which increase the risk of development of micro-angiopathy and related sequelae. The percentage of skin infection was 31% in poor glycemic control group, 9% in fair control group and 5% in good control group. From this study we observed that cutaneous infections were the common skin problem among the skin manifestations in diabetics.

The second most common cutaneous manifestation observed during this study was xerosis. Previous studies on skin lesions in diabetic patients did not reveal on the frequency of xerosis. But in our study the

73 diabetics were found to have more incidence of xerosis. The clinical observations are supported by objective findings of a reduced hydration state of the stratum corneum and decreased sebaceous gland activity in patients with diabetes, without any impairment of the stratum corneum barrier function93. Autonomic neuropathy has been attributed to the pathogenesis of ichthyosis, xerosis, and pruritus.

Autoimmune dermatoses like vitiligo are known to occur in DM as apart of polyglandular autoimmune syndrome. The incidence of non- specific cutaneous disorders also has pathogenetic, prognostic and therapeutic importance in diabetic individuals. In our observation we found vitiligo in 4% subjects. This prevalence was similar with previous studies reported from India94.In our study psoriasis vulgaris was observed in 4%. Sezai et al.,95noted in their study that 11.2% had psoriasis with type 2 DM. There have been reports of a significant association of DM and psoriasis in a large series of studies96,97. With reference to Avci et al.,96observation that various hues of erythema in psoriatics may alert the physician to possible impaired glucose tolerance in the subjects. There were reports on association between psoriasis and increased cardiovascular risk and metabolic syndrome97.

Diabetic dermopathy, a form of small, brown-scar-like macules on both shins was observed in 2% of study subjects in our study. Diabetic dermopathy and retinopathy are both considered by certain authors as

74 manifestations of diabetic microangiopathy. Various studies showed that retinopathy was found to be more commonin patients with dermangiopathy than in those without it98,99. Diabetic dermopathy may develop from the factors that lead to the development of vascular complications of diabetes and it may serve as a clinical sign of an increased vascular complications in diabetic subjects100. Diabetic dermopathy correlated with neuropathy in a study done by Gerald E et al101.

Acanthosis nigricans was observed in 3 % of patients in this study, though Mashkoor et al102 found acanthosis nigricans in 11% of patients which is almost three times more than our study. Pruritus was found in

2% of our study sample population103. Some similar previous studies reported the prevalence of pruritus as 15.1% and 49% 104,105 which is much higher than our study.

Bullous diabeticorum (diabetic bullae) was seen in 1% of study subjects. Some of the other skin manifestations observed in this study were melasma(2%), pityriasis rosea(1%), lichen planus(1%) while Roshni

Vahora et al32observed 4% lichen planus in their study. We also found telogen effluvium(1%), lipodystrophy (1%), pompholyx (1%) and erythroderma (1%) in the study population. Lipodystrophy to insulin at injection site was seen in 1 subject out of 14 subjects who were on insulin.Adverse drug reaction to oral hypoglycemic was not seen in our study. We observed that out of 100 subjects, 7 were having more than one

75 skin manifestations. Cutaneous manifestations are prevalent in type 2 diabetes mellitus and whenever multiple cutaneous manifestations103are present, evaluation for diabetes mellitus is essential.

Duration of DM

Duration of diabetes mellitus and glycemic control are crucial factors involved in the microvascular and macrovascular complications86.

On considering the duration of DM,13% were diabetic for less than a year,38% were diabetic for 1-5 years, 28% were diabetic for 5-10 years,

19% were diabetic for 10-20 years, and 2% were diabetic for more 20 years. Mean duration of DM was 7.55years. Out of 100 subjects,79 subjects had DM for less than 10years duration, and 21 subjects for more than 10 years duration. Our study observation of more number of subjects in < 10year duration and less number of subjects in > 10 year duration was consistent with Chatterjee N et al., study98.Long-term effects of DM on the microcirculation and on dermal collagen eventually result in skin disorders in almost all the diabetics79.

On considering duration and skin manifestations, diabetic dermopathy, diabetic bullae, diabetic foot ulcer were observed in subjects with > 10 year duration of DM. Diabetic dermopathy was observed in 2% of study subjects and it may serve as a clinical sign of an increased likelihood of vascular complications in diabetic patients106.According to a study from

SaudiArabia, by Shahzad M et al., positive correlation of skin lesions with disease duration was observed107.

From Chatterjee N. et al.,98 study it was observed that diabetic dermopathy, diabetic bullae and diabetic ulcer foot prevalence had increased with long duration of DM. We also observed diabetic dermopathy, diabetic bullae and diabetic ulcer foot in subjects with > 10 years duration.

Xerosis was observed in 26% in the total study population which was the second commonest observation in our study108. Among 26% it was observed in 12% of subjects with 5-10 year duration of DM, 7% with

10-20 year duration, 4% with 1-5 year duration and 2% with > 20years duration and 1% with < 1year duration. Xerosis is due to abnormal, persistent cohesion between corneocytes, thickening and impaired moisturization of the stratum corneum, increased transit time of corneocytes in the SC, and altered skin barrier function.93,109-111.

Xerosis and reduced stratum corneum elasticity induce cracks, followed by bacterial invasion and infection. Recently it was observed that an increased severity in xerosis was linked with the duration of

DM101. Our study did not reveal increased incidence of xerosis with increased duration. Xerosis was observed even in less than 1year duration of DM and more subjects with xerosis were found in <10 year duration of

DM in our study. Skin infections were observed in 45% of subjects

77 among that 39% in < 10 years duration 6% in > 10 year duration. Our observation showed no increasing incidence of infections with increased duration of DM but with poor glycemic control.

Glycemic control

HbA1c reflects average plasma glucose during the last 8 to 12 weeks; it does not require any special preparation such as fasting and can be performed at any time of the day. Because of these properties it is preferred as the test for assessing glycemic control in people with diabetes. Adequate glycemic control decreases the incidence and severity of skin disorders86.Our study showed subjects with good glycemic control

28%, fair control 22%, poor control 50%. In our study subjects, we observed 69% cutaneous infections with poor glycemic control, 20% in fair control and 11% in good control. This is consistent with Chatterjee N et al., 98 study in that the mean HbA1c level was higher in patients with infective lesions. In our study out of 45 subjects with skin infection, 31 subjects had poor glycemic control, 9 subjects had fair control and 5 had good control. In total 27 subjects with fungal infections, 13 had candida infections, 12 had dermatophytosis, onychomycosis in 1 and Tinea versicolor in 1 subject. The observation was poor glycemic control correlated with skin infections. Severe form of bacterial infections were observed with poor glycemic control.

Autonomic neuropathy test - SSR

In our study SSR was recordable in 72% of study subjects and not recordable in 28% of study subjects. In xerosis subjects of our study SSR was absent in 57.69%. Al-Moallem MA et al112., study showed significant association of autonomic neuropathy with SSR ..Jha S, Nag D113 found

SSR was not recordable in 42% of asymptomatic diabetics but in DM subjects with autonomic dysfunction SSR was not recordable in 66%.

From one study114 it was noted that SSR abnormalities were seen in 66–

83% of diabetics. As the disease progress from asymptomatic to symptomatic polyneuropathy and to symptomatic autonomic neuropathy, the frequency of abnormalities increases. SSR is used to assess the sympathetic cholinergic function but correlation with generalized sympathetic dysfunction is poor. Importantly, a normal SSR cannot exclude a defective sympathetic noradrenergic function115.Sweating is reduced on the lower extremities of diabetics but there is gustatory hyperhidrosis and compensatory hyperhidrosis on the upper part of the body116-119. The sweat secretion is important in generation of cutaneous biopotential. Xerosis with absent SSR subjects must be evaluated for other ANS dysfunction mainly cardiovascular ANS dysfunction.

The other co- observations noted in this study were lipid profile, blood urea and serum creatinine. On correlating HbA1c with TGL and

LDL statistically significant correlation was observed, the ‗p ‗value was

0.0001.With poor glycemic control the TGL and LDL value had increased. Insulin resistance along with hypertriglyceridemia makes the individuals prone for Metabolic X syndrome. Out of 100 study subjects 8 had nephropathy with serum creatinine > 1.2 mgm% and all the 8 subjects had poor glycemic control. Out of 8 subjects with increased serum creatinine level 4 had severe bacterial infection (cellulitis), 2 had diabetic dermopathy120,1had diabetic bullae and 1had fungal infection.

Poor glycemic control is associated with cutaneous infections and diabetic complications.

CONCLUSION

SUMMARY

This is a cross sectional observational study where we have randomly selected 100 type 2 DM subjects with skin disease aged between 30-70 years of both genders. The observations made were the average age was 56.02 years which is a economically productive age. The male: female ratio of 1:1.77 with urban : rural 1.08:1 which showed almost equal number of study subjects were from rural area. 45% study subjects had positive family history of DM that shows the significance of genetic predisposition to DM.25% were smokers and BMI was normal in45% of subjects and increased BMI in 54% of subjects. Smoking along with increased BMI are risk factors for DM microvascular complications.

Average duration of DM was 7.55years. More subjects were in< 10 years group.28 subjects had good glycemic control whereas 22 had fair control and 50% had poor control. Commonest skin manifestation observed was skin infections followed by xerosis. Diabetic dermopathy, diabetic bullae, diabetic ulcer foot were observed in >10years duration subjects. Poor glycemic control was observed in skin infections especially in severe bacterial infections and in Diabetic dermopathy, Diabetic bullae and in Diabetic ulcer foot. Sympathetic skin response was absent in 28 subjects predominantly xerosis subjects showed no response due to peripheral sympathetic autonomic neuropathy.

CONCLUSION

It can be concluded from this study that,

As duration of diabetes becomes longer, the complications become more severe. To prevent the morbidity and mortality associated with microvascular complications, it is imperative to diagnose diabetes mellitus early. From IDF statistics, it is learnt that the people with undiagnosed hyperglycemia is more in our country. This also contributes to the complications of DM as the duration of hyperglycemia is a key factor in pathogenesis of microvascular complications of DM. The lack of awareness, lack of knowledge about diabetes and limited opportunities for diagnosis are considered as main reasons for undiagnosed hyperglycemia.

Also DM is in increasing trend among rural population. Proper awareness programmes can bring down the percentage of undiagnosed diabetics by bringing the individuals voluntarily to the healthcare providers. The health education about the life style modification and the knowledge about the risk factors are essential. Non communicable disease (NCD) screening is an effective mode of picking up the hyperglycemic individuals early and with proper treatment we can prevent or delay the complications.

Commonest observed skin manifestations in our DM study subjects were skin infections. The factor responsible for this was

82 considered to be poor glycemic control. Glycemic control is directly correlated with skin infections in our study. Infections must be diagnosed early and adequately controlled with proper treatment to prevent the morbidity and mortality.

Xerosis which was observed in majority of subjects in our study could be considered as earliest indicator of DM cutaneous autonomic neuropathy. Autonomic neuropathy is responsible for silent myocardial infarction and masking of hypoglycemic symptoms. Relation between cutaneous autonomic neuropathy and other autonomic neuropathies is an unearthed treasure. It is better if all the high risk individuals and diagnosed diabetics are subjected to routine skin examination. SSR was not recordable in reasonable number of diabetic subjects. Absent SSR signifies that the subjects are having autonomic neuropathy related to sudomotor function. So skin examination and SSR test can be included in the routine examination of diabetic individuals, because these are rather simple, less time consuming, non- invasive methods.

―Prevention is better than cure‖ this statement holds good in prevention of diabetes induced complications. The early diagnosis of hyperglycemia, adequate glycemic control and simple tests to pick up the complications early will definitely improve the quality of life and reduce the morbidity and mortality of DM individuals.

FUTURE SCOPE

FUTURE SCOPE OF STUDY

LIMITATIONS

1. This study was conducted in a tertiary care hospital and not

included those attending the other health care sectors.

2. Number of study subjects.

3. Other types of DM subjects were not included.

4. SSR was used as a qualitative test.

FUTURE SCOPE

1. Large number of DM subjects with skin disease including those

attending all sectors of health care systems.

2. Including other types of DM subjects.

3. SSR test with quantitative analysis.

4. Incorporating and correlating SSR with other ANS tests.

5. Correlating skin lesions with other microvascular and

macrovascular complications.

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ANNEXURES

CONSENT FORM

PROFORMA

ID NO : ______Date:

NAME : ______O.P no:

AGE/SEX : ______

ADDRESS : ______

OCCUPATION : ______

EDUCATION : ______

HISTORY

1. Duration of diabetes

2. Treatment particulars oral hypoglycemic drugs/ insulin/both/

others

3. Skin disease

Symptoms

Site: skin/mucosa/hair/ nail

Duration of skin disease

Past or family history of similar lesion

Any other associated skin diseases.

4. Other co morbidities

Thyroid disorders

Bronchial asthma

Tuberculosis

Epilepsy

Others

5. Drug history

6. Personal history smoking/ alcoholism/others

7. Family history of DM Parents: Father/Mother/Both

Siblings: Brother/sister/Both

EXAMINATION

Height in cm

Weight in kg

Body mass index

CLINICAL EXAMINATION

Anemia

Icterus

Cyanosis

Pedal edema

Lymphadenopathy

Pulse rate / mt

Blood pressure in mm Hg

System examination

R.S

C.V.S

Abdomen

C.N.S

DERMATOLOGICAL EXAMINATION SKIN MUCOSA HAIR NAIL DIAGNOSIS

NAME ID no

INVESTIGATIONS

1. Blood sugar Fasting (mgm/dL) :

2. Blood sugar Post prandial (mgm/dL) :

3. HbA1c (%) :

4. Lipid profile TGL (mgm/dL) :

LDL (mgm/dL) :

HDL (mgm/dL) :

5. Blood urea (mgm %) :

6. Serum creatinine (mgm %) :

SYMPATHETIC SKIN RESPONSE

Present/Absent

CONSENT: I hearby give my volunteer consent for the above study

Signature.

MASTER CHART