Effects of Justicia Carnea Leave on Hematological Parameters in Albino Mice Carried out in Mbingo Annex Hospital Laboratory in Bamenda, North West Region, Cameroon

Journal of Advances in Microbiology

20(10): 43-50, 2020; Article no.JAMB.62608 ISSN: 2456-7116

Effects of Justicia carnea Leave on Hematological Parameters in Albino Mice Carried Out in Mbingo Annex Hospital Laboratory in Bamenda, North West Region, Cameroon

Asakizi Augustine Nji1*, Nchang Chrysanthus2 and Ngombe David Monyongo3

1Department of Biological Sciences, Ministry of Higher Education, University of Bamenda, North West of Cameroon. 2Department of Medical Laboratory Sciences, School of Health Sciences, Ministry of Water Resources and Energy, Jude`s University institute, Regional Delegation for the North West, Divisional Delegation for Boyo, Fundong, St of Cameroon. 3Department of Medical Laboratory Sciences, School of Medical Biological Science, National Polytechnic University Institute Bamenda, Cameroon.

Authors’ contributions

This work was carried out in collaboration among all authors. Author ANA designed the study, performed the statistical analysis, wrote the protocol and wrote the first draft of the manuscript. Author NC managed the analyses of the study. Author NDM managed the literature searches. All authors read and approved the final manuscript.

Article Information

DOI: 10.9734/JAMB/2020/v20i1030289 Editor(s): (1) Dr. C. Ike, Anthony, University of Nigeria, Nsukka, Nigeria. Reviewers: (1) Najma Ismail, Aliah University, India. (2) Ashok Thulluru, Shri Vishnu College of Pharmacy (Autonomous), India. Complete Peer review History: http://www.sdiarticle4.com/review-history/62608

Received 27 August 2020 Original Research Article Accepted 01 November 2020 Published 23 November 2020

ABSTRACT

This study is design to ascertain the hematological status in albino mice treated with Justicia carnea leave, which could aid in the treatment of anemia. The hematological parameters investigated include Mean Cell Volume (MCV), Mean Cell Hemoglobin (MCH), Mean Cell Hemoglobin Concentration (MCHC), Platelets (PLTS), Hemoglobin (Hb), Red Blood Cells (RBCs), Pack Cell Volume (PCV) and White Blood Cells (WBCs). Nine (9) male albino mice of approximately the same weight 300 g were grouped into 3 groups that’s Negative Control (NC) Positive Control (PC) and

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*Corresponding author: E-mail: [email protected];

Nji et al.; JAMB, 20(10): 43-50, 2020; Article no.JAMB.62608

Test Control (TC), each made up of 3 mice. Negative control mice were given normal feed moisten with water, Positive control received diluted iron folate with normal feed, finally Test control mice were administered powdered leaves of Justicia carnea with normal feed for 7 days, after which blood samples were collected using EDTA tubes by exsanguination and run for hematological parameters using auto-hematological analyzer. The results of various groups were found to be; as there was a significant increase in RBCs, Hb, & PCV (P=0.05), between the groups. No significant change was observed on the MCV, MCH, and MCHC. Lymphocytosis was observed in all groups with mark difference in granulocytosis. TC had granulocytosis, NC showed normal granulocyte scores and the PC show granulocytopenia with a statistical difference of P=0.0017. A significant difference was seen in platelets between the groups P=0.02. This study shows that J. carnea leaves possess anti-anemic potential, lending credence to the use of these plant leaves in folk medicine for the management of hemolytic anemia would be helpful. Further research on the various phytochemicals of the plant should be done and also its toxicological aspects.

Keywords: Hematological parameters; albino mice; and justicia caenea.

1. INTRODUCTION against diseases, example of some documented plants used in the treatment of anemia condition Anemia can be define as a decrease in the ability include Coco nucifera, leaves extracts of Tectona of the blood to carry oxygen due to a decrease in grandis etc. The availability of some synthetic the total number of erythrocytes, a diminish drugs used in the treatment of a specific disease concentration of Hb per erythrocyte or a is common but because of the high cost and side condition of both [1]. Anemia is the most effects associated with their use, attention is common blood disorders affecting about a third currently been focused on the use of medicinal of the global population. Anemia increases the plant products in the management or prevention cost of medical care and lowers a person’s of various diseases. The genus Jesticia, name productivity through a decrease ability to work. after the 18 century by a Scottish botanist James When anemia comes on slowly, the symptoms Justice, belongs to the large family of are often vague and may include feeling tired, Acanthaceae consisting of about 600 species of weakness, shortness of breath or poor ability to herbs and shrubs native to the tropics and exercise. Anemia that comes quickly often has subtropics. Justicia carnea is a flowering plant, greater symptoms which may include confusion, widely distributed in various parts of Africa [1]. In feeling like one is going to pass out, loss of Cameroon the shrubs of Justicia carnea are consciousness, or increase thirst. It’s classified grown around homes which easily grow and based on the size of RBCs and the amount of propagate from stems cutting by pushing the hemoglobin in each cell. If the cells are small, it stems 1-2 inches into the soil. A survey among is microcytic anemia while if large, it is the Bakweri population in Cameroon revealed macrocytic anemia while if they are normal size, that, the plant under study is locally called” it is normocytic anemia [2]. Hematology refers to Ewolamajia” meaning a plant that gives blood. the study of the number and morphology of the The deep colour juice from the leaves of this cellular elements of the blood, the Red blood plant is extracted either by soaking or boiling in cells (erythrocytes), white blood cells water which can be drank as tea. This study will (leucocytes), and platelets (thrombocytes) and aid to establish whether individuals using these the use of these results in the diagnosis and leaves as blood show improved hematological monitoring of disease [3]. It has been reported profile which can aid in the treatment of anemia. that, ingestion of drugs can alter the This study is design to ascertain its hematological parameters [4]. They are good hematological status in albino mice treated with indicators of the physiological status of animals Justicia carnea leave. [5]. Therefore hematological parameters could be an important tool in the assessment of effect of 2. MATERIALS AND METHODS drugs [6]. Medicinal plants have been documented as having beneficial properties used 2.1 Study Area for the management of various ailments because they have been discovered to contain bioactive Samples and specimens obtain were compounds called phytochemicals and analyzed at Bingo Annex Hospital Bamenda secondary metabolites that protect humans Cameroon.

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2.2 Plants Material {Final body weight – initial body weight} / {Final body weight} x 100. Fresh leaves of Justicia carnea was harvested from the Bambui locality (Tubah Sub Division, Similarly after exsanguination, the liver and North West Region Cameroon). The leaves were the heart were closely observed for goose removed from the stems, rinse in clean water pathological changes and then weighted and then dried in shadow for weeks. After drying, by the aid of a balance and the relative the leaves were grind to powder with the aid of a organ weight were calculated as; {Relative blender to obtain approximately 30 g. The organ weight= Organ weight [g] / Body weight powered leaves were stored in airtight container [g]. until use. 3.3 Statistical Analysis 2.3 Animal Handling and Grouping The data collected were analyzed using SPSS A total of nine (9) male albino mice of (Statistical Package of Social Science) version approximately the same weight 300 g were used 21. Differences in means were done using the in this research. All experimented animals were least significant difference (LSD) at P= 0.05. The obtained from the research Biopham Centre results were expressed as mean +- standard Dschang. The animals were house in a plastic error of mean (S.E.M.). bowl rapped with net and were acclimatized. The The mean distribution of RBCs and Hb within the animals were allowed free access to food and 3 study groups is presented in Fig. 1. water. After acclimatization, the animals were grouped into 3, that’s Negative Control, Positive The mean distribution values of PCVs within the control and Test Control, each made up of 3 3 study groups are also presented in Fig. 2. mice, using a random sampling technic. The

Negative control mice were given 25 g of normal The mean distribution values of PLTs within feed (that’s grind corn mixed with soya bean, fish the 3 study groups are also presented in & vitamin), moisten with water. Positive control Fig. 4. mice received 4ml of diluted iron folate mixed with 25 g normal feed. Lastly, Test control mice 4. RESULTS were been administered 1-2 preparation of powered leaves of Justicia carnea and normal In this study Justicia carnea leaves induced feed, of which 25 g of it were administered orally changes in erythrocytes and related parameter for 7 days. profiles such as Hb in the study albino mice (Table 1 or fig. 1). Seven days after 3. HEMATOLOGICAL ANALYSIS administration of the leave at the dose levels of approximately 25g, there were significant Hemoglobin (Hb), Red Blood Cells (RBCs), Pack increase in erythrocytes and Hb levels. When Cell Volume (PCV), Platelets (PLTs), Mean Cell comparing the test (T) to the negative control Volume (MCV), Mean Cell Hemoglobin (MCH), (NC) group, there was no statistical significance Mean Cell Hemoglobin Concentration (MCHC) between their RBC values, despite the fact that and White Blood Cell (WBC) were analyzer and their Hb levels significantly differed. When results printed by the aid of auto-hematological compared with the positive control (PC), the RBC analyzer. and Hb values between the T and PC group were significantly different. Furthermore, With 3.1 Collection of Blood Sample regards to PCV, there were statistical significant (P < 0.05) differences between the T group, After 7 days of administration, blood negative and positive control groups (Table 2 or samples were collected by exsanguination Fig. 2). In addition, Despite the randomization in and placed in EDTA tubes for Hematological occurrence of the MCV, MCH and MCHC values studies. (as seen in Fig. 3 above), there was no statistical significant difference between the Mean 3.2 Body Weight and Relative Organ ±standard deviation values within and between Weight Determination the three groups in this study (P ˃ 0.05). The study also revealed that although there were The percentage change in the body weight of the significant increase in the WBC count above experimental animals was calculated as: normal in the Test group (leucocytosis)

Nji et al.; JAMB, 20(10): 43-50, 2020; Article no.JAMB.62608

compared to the NC and PC groups; there was granulocytosis (Table 3). The differences no statistically significant difference between the between the mean ±standard deviation scores in mean ±standard deviations between the three the three groups was statistically significant at groups. The same was also observed in the 0.05 significance level (P = 0.0017).To conclude, lymphocyte percentage scores within the study no group was observed with thrombocytopenia or groups. All the three groups had lymphocytosis. thrombocytosis but test group had higher The ANOVA output found no statistical difference thrombocyte counts (which were still within between the groups (P = 0.062). But with respect normal limits) compared to the negative and to the granulocyte percentage, the negative positive control groups. There was a statistically control group had normal granulocyte percentage significant difference between the three study scores. The positive control rather had groups (P = 0.002). This is presented in Table 4 granulocytopenia while the test group had above.

Table 1. RBC and Hb distribution between the study groups

RBCs and Hb RBC (cells x 1012/L) Hb (g/dl) Negative control (NC) 4.21±0.32 12.4±0.60a Positive control (PC) 3.96±0.95b 11.73±0.94b Test (T) 5.13±0.46 15.57±0.61 F – VALUE 68.21 89.74 P - VALUE 0.0001 0.0001 a = P < 0.05 when NC is compared with T, b = P < 0.05 when PC is compared with T

e u l a v

n a e M

Negative control Positive control Test (T) (NC) (PC) RBC 4.21 3.96 5.13 Hb 12.4 11.73 15.57

Fig. 1. Mean RBC and Hb distribution between the study groups

Table 2. PCV distribution between the study groups

Pack cell volume (%) PCV (%) Negative control (NC) 36.80±0.71a Positive control (PC) 35.13±0.52b Test (T) 46.70±0.46 F – VALUE 42.61 P - VALUE 0.0001 a = P < 0.05 when NC is compared with T, b = P < 0.05 when PC is compared with T

Nji et al.; JAMB, 20(10): 43-50, 2020; Article no.JAMB.62608

PCV (%), Positive control (PC), 35.13, 30% PCV (%), Test (T), 46.7, 39%

PCV (%), Negative control (NC), 36.8, 31%

Fig. 2. Mean PCV distribution between the study groups

Table 3. Red cell indices distribution between the study groups

Red cell indices MCV (fl) MCH (pg) MCHC (g/dL) Negative control (NC) 88.45±1.01 29.75±0.21 33.70±0.40 Positive control (PC) 88.80±1.41 29.67±0.94 33.40±0.23 Test (T) 91.00±2.47 30.33±1.01 33.33±0.26 F – VALUE 54.21 63.82 59.67 P - VALUE 0.129 0.184 0.188

n o i t u b i r t s i d

n a e M

MCV (fl) MCH (pg) MCHC (g/dL) Negative control (NC) 88.45 29.75 33.7 Positive control (PC) 88.8 29.67 33.4 Test (T) 91 30.33 33.33

Fig. 3. Mean red cell indices distribution between the study groups

Table 4. PLT distribution between the study groups

Platelets PLT (x109/L) Negative control (NC) 501.00±3.94 Positive control (PC) 484.33±2.61 Test (T) 632.37±3.28 F – VALUE 532.04 P - VALUE 0.002

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PLT (x109/L) n o i t u b i r t s i d

n a e M

Negative Positive Test (T) control control (NC) (PC) PLT (x109/L) 501 484.33 632.37

Fig. 4. Mean PLT distribution between the study groups

5. DISCUSSION Owoyele et al in Nigeria in 2011 [13], which show no significant increase in RBCs, Hb and PCV. The actual physiological status of organisms can Furthermore, the present study reveals no be diagnosed through use of blood parameters. significant change in the Mean Cell Volume For normal functioning of the body of an (MCV), Mean Cell Hemoglobin (MCH) and Mean organism, it must keep its blood composition and Cell Hemoglobin concentration (MCHC). This constituent under natural conditions. The present could be attributed to the fact that the mice were study showed that leaves of Justicia carnea not induce by phenyl hydrazine to become demonstrated varying degrees of hematological anemic. The study is in line with that carryout by changes at the dose levels of 25 g. The Mishra and Tandon in India on extracts of significant (p<0.05) increase in erythrocytes and ornamental plants in Swiss albino mice in 2011 hematocrit counts (i.e RBCs, Hb and PCV) after [14], and disagrees with that done by Obel et al. oral administration of Justicia carnea leaves in Kenya on Carissa edulis extract in 2016 [15] (TC), iron folate (PC) and normal feed (NC) as which revealed a significant increase in MCV, shown in Table 1 and Table 2, suggests that the MCH and MCHC. In addition, the present study leaf may contain phytochemicals and compounds show a significant increase in WBCs most that stimulate the secretion or formation of especially in the Test control with all groups erythropoietin in the stem cells of normal mice. having lymphocytosis but a mark difference in Stimulation of stem cells in the bone marrow to granulocyte showing normal granulocyte score in produce red blood cells occurs due to the action the Negative control, granulocytopenia in the of erythropoietin. Phytochemicals such as positive control and finally granulocytosis in the saponin, alkaloids, tamin, and flavonoid have Test control(P = 0.0017) Table 3. Increase WBC been documented to have erythropoietic have been report to boost the immune system properties [7], which if present in the leaf [16], the significant increase in WBC count, explains the significant increase in RBCs, Hb and lymphocyte count and granulocyte count cause PCV. This may also suggest that the plant may by the plant leaves reflect leucopoietic and cause polycythemia [8,9,10,11]. This is similar to possible immunomodulatory effects of the extract the research carry out by Chimaraoke et al., [1] which augmented the production of more WBC in Nigeria on the same plant in 2017 and and lymphocyte [17]. This agrees with research disagrees with a study carry by Mishra and performed by Chimaraoke et al., on Justicia Tandon in India on Bougainvillea spectabilis carnea extracts in Nigeria in 2017 [1], and leaves in Swiss albino mice in 2011 [12], which disagrees with that performed by Olusegum et show a significant reduction in RBCs and Hb, al., on Ocinum gratissimum in Wister rats in and also disagrees with that conducted by Nigeria in 2008. [18]. The study also revealed a

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significant increase in platelet count, most COMPETING INTERESTS especially in the test group when compared to the Negative and Positive control. Platelets are Authors have declared that no competing the blood cells involved in Coagulation. interests exist. Coagulation of blood requires that the platelets should be in sufficient size, number and function. REFERENCES The increase in the platelet levels observed in this study may be explained by stimulatory effect 1. Chimaraoke O, Ngozi KA., Ekeleme EAC, on thrombopoietin. The significant increase in Chidinma UE, Chidinma KO. platelets and related parameter profiles after oral Haematological and Biochemical studies administration of Justicia carnea leaves suggests on Justicia carnea leave extracts in that the leaves contain compounds and phenylhydrazine induce anemia in albino phytochemicals that may have stimulated mice, Abia State Nigeria; 2017. thrombopoietic process in the albino mice. This 2. Janz TG, Johnson RC, Rubenstein SD. agrees with that conducted by Obel et al in Anemia in the emergency department Kenya on Carissa edulis extract in 2016 [15] and evaluation and treatment; 2013. disagrees with that carried out by Olusegum et 3. Merck Manual. Haematologic reference al., [18] on Ocinum gratissimum in Wister rats in ranges. Mareck Veterinary Manual; 2012. Nigeria in 2008, which showed a decrease in Retrieved from Platelet count. http://www.merckmanuals.com/. 4. Ajagbonna CP, Onifade KI, Suleiman U. 6. CONCLUSION Hematological and Biochemical Changes in rats given extracts of Catotropics The approximated values in this leaves base on procera. Sokoto. J vet Sci. 1999;1:36-42. the aid of automated analysis and statistical 5. Etim N, Joseph E, Glory E, Metiabasi U, analysis were very reasonable, which suggests Edem OM, AnthonyO. Hematological that the leaves of this plant can make valuable parameters and factors affecting their contributions to improved nutrition, blood values, Akwa Ibom state university;2014. parameters and well-being. This study shows 6. Yakubu M, Musbau AA, Oladiji AJ. that J. carnea leaves possess anti- H e m atological evaluation in male mice. anemic potential, lending credence to the use of Rats following chronic administration of these plant leaves in folk medicine for the aqueous extracts of Fadogia agrestis management of hemolytic anemia is helpful. stem. Article from Pharmacognosy Further studies are warranted to determine the magazine. 2007;3(9):2007. bioactive component present in J. carnea leaves 7. Shravan KD, Rajeshwar V, Anusha V, that could be responsible for its anti-anemic Tyothipriya J, Ravali T, Mounika M. Anti- effects. anaemic activities of ethanolic extract of Punica granatum seeds on COVER STATEMENTS phenylhydrazin-induce anemic rats, India; 2016. This is to indicate that, this article is the fruits of 8. Iranloye BO. Effect of chronic garlic our original Research and that this article has feeding on some hematological not been submitted elsewhere for publication. parameters. African Journal of biomedical Where ever the information has been adapted, Research. 2002;5:81-82. the author has provided adequate citation. 9. Mansi K, Lahham J. Effects of pride of Artemisia sieberi Besser (A/ herba-alba) ACKNOWLEDGEMENT on heart rate and some hematological values in. normal and alloxan induced The authoracknowledge the efforts of Mr. Asakizi diabetic rats. Journal of Basic and Nji Augustine, for his kind attention in supervising Appliced Sciences. 2008;4:57-62. this Research. Joyful thanks to the Dean of 10. Kuppast IJ, Vasudeva NP, Ravi MC, Medical and Biomedical Sciences and the entire Biradar SS. Studies on the hematological school staff for making this Research e f fect of the extracts of Cordia dichotoma Project possible. Lastly, thanks to Mr lukong Forst. F. Fruits. Research Journal of Hubert for all his efforts made in this research Agriculture Science and Forestr. 2009;1: project. 195-204.

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