Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine 351 Kidney Hyaluronan Regulatory Aspects During Different States of Body Hydration, Nephrogenesis & Diabetes LOUISE RÜGHEIMER ACTA UNIVERSITATIS UPSALIENSIS ISSN 1651-6206 UPPSALA ISBN 978-91-554-7199-6 2008 urn:nbn:se:uu:diva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ist of Papers This thesis is based on the following papers, which are referred to in the text by their Roman numerals: I. Hormonal Regulation of Renomedullary Hyaluronan. Rügheimer L, Johnsson C, Maric C, and Hansell P Acta Physiol (Oxf.) 2008, In press. II. Hyaluronan Synthases and Hyaluronidases in the Kidney During Changes in Hydration Status. Rügheimer L, Olerud J, Takahashi T, Shimizu K and Hansell P In manuscript 2008. III. ACE Inhibition During Completion of Nephrogenesis Obstructs Neonatal Dissipation of Interstitial Hyaluronan, not Correlated to Lymph Vessel Expression. Hansell P, Rügheimer L, Åstrand ABM, Johnsson C, Friberg P and Kerjaschki D In manuscript 2008. IV. Renal Hyaluronan Content During Experimental Uncontrolled Diabetes in Rats. Rügheimer L, Carlsson C, Johnsson C and Hansell P J Physiol Pharmacol 2008; 59(1): 115-128. Reprints of papers and figures are with permission from the publishers, Paper I from Blackwell Publishing and Paper IV from Polish Physiological Society. 3 Vita non est vivere sed valere vita est 4 Contents Abbreviations .................................................................................................. 7 Introduction ..................................................................................................... 9 The Kidney ................................................................................................. 9 Vasopressin/ADH/AVP ......................................................................... 9 The renin-angiotensin system (RAS) ..................................................... 9 Nitric oxide (NO) ................................................................................. 10 Prostaglandins ...................................................................................... 10 Hyaluronan ............................................................................................... 11 Hyaluronan receptors ............................................................................... 12 Turnover of hyaluronan - Synthesis ......................................................... 12 Turnover of hyaluronan - Catabolism ...................................................... 13 The role of hyaluronan in the kidney ....................................................... 14 Renomedullary interstitial cells (RMIC) ............................................. 16 Nephrogenesis – postnatal development .................................................. 16 Diabetes mellitus ...................................................................................... 17 Animal model of diabetes ........................................................................ 18 Aims .............................................................................................................. 19 Materials and Methods .................................................................................. 20 Animals .................................................................................................... 20 Treatment with ACE-inhibitor enalapril (Paper III) ................................. 20 Induction of diabetes (Paper IV) .............................................................. 20 Anesthesia ................................................................................................ 21 Surgical procedures .................................................................................. 21 Experimental protocols in vivo ................................................................. 21 Experimental protocols in vivo (Paper I) ............................................. 22 Experimental protocols in vivo (Paper II) ............................................ 22 Experimental protocols in vivo (Paper IV) .......................................... 22 Hyaluronan analysis and determination of water content ........................ 23 Histochemistry ......................................................................................... 23 Staining for hyaluronan ....................................................................... 23 Staining for CD44 ................................................................................ 24 Staining for podoplanin ....................................................................... 24 Staining for LYVE-1 ........................................................................... 25 5 Morphology .............................................................................................. 25 Urine analysis ........................................................................................... 25 Hyaluronidase activity measurements ................................................. 25 HAS 2 mRNA isolation and preparation of cDNA .................................. 26 Gene expression analyses ......................................................................... 26 Analysis of HAS 1-3 mRNA expression (Paper II) ............................. 26 Analysis of HAS 2 mRNA expression (Paper IV) .............................. 27 RMICs in culture (Paper I) ....................................................................... 28 Experimental protocol in vitro (Paper I) .................................................. 28 Statistical analysis .................................................................................... 29 Results ........................................................................................................... 30 Hormonal regulation of renomedullary hyaluronan (Paper I) .................. 30 Hyaluronan synthases and hyaluronidases in the kidney during changes in hydration status (Paper II) ........................................................................ 32 ACE inhibition during completion of nephrogenesis obstructs neonatal dissipation of interstitial hyaluronan, not correlated to lymph vessel expression (Paper III) ............................................................................... 35 Renal hyaluronan content during experimental uncontrolled diabetes in rats (Paper IV) .......................................................................................... 38 Discussion ..................................................................................................... 41 The role of hyaluronan in homeostasis ..................................................... 41 Hyaluronan and different states of body hydration .................................. 44 Heterogeneous distribution of hyaluronan in nephrogenesis ............... 47 Water handling in the diabetic kidney ................................................. 50 What regulates renal hyaluronan? ............................................................ 52 Conclusions ................................................................................................... 53 Svensk sammanfattning / Summary in Swedish ........................................... 54 Acknowledgement / Tack! ............................................................................ 56 References ..................................................................................................... 59 6 Abbreviations ABC Avidin biotin complex ACE Angiotensin converting enzyme Ang II Angiotensin II ANOVA Analysis of variance AQP Aquaporines AT1-receptor Angiotensin II receptor type 1 AT2-receptor Angiotensin II receptor type 2 AVP, ADH Vasopressin, Antidiuretic hormone BG Blood glucose bw Body weight CD44 A cell surface receptor for hyaluronan cDNA complementary deoxyribonucleic acid cGMP Cyclic guanosine monophosphate COX Cyclooxygenase Da Dalton dw Dry weight ELISA Enzyme-Linked Immunosorbent Assay eNOS endothelial eNOS FITC Fluorescein isothiocyanate. a derivative of fluorescein GlcNAC N-acetyl-D-glucosamine GlcUA D-glucuronic acid GLUT-2 Insulin-independent glucose transporter HA Hyaluronan (hyaluronic acid) HABP Hyaluronan binding protein HAS Hyaluronan synthase HYAL Hyaluronidase IL-1 Interleukin 1 beta INDO Indomethacin iNOS Inducible nitric oxide synthase L-NAME N-nitro-L-arginine methyle ester LYVE-1 A receptor for hyaluronan in the lymphatic vessel endothelium MAPK Mitogen activated protein kinase MP Methylprednisolone mRNA Messenger ribonucleic acid nNOS neuronal nNOS 7 NO Nitric oxide NOS