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Post-injury nerve regeneration in diabetic mice
Judyta Juranek firstname.lastname@example.org
Neurological complications of diabetes
‘A descriptive term meaning a demonstrable disorder, either clinically evident or subclinical, that occurs in setting of diabetes mellitus without other causes for peripheral neuropathy. The neuropathic disorder includes manifestations in the somatic and/or autonomic parts of the peripheral nervous system’.
American Diabetes Association, 1988 adapted from The Epidemiology of Diabetes Mellitus, the International Perspective, 2001
Diabetologia.distal symetric sensorimotor and autonomic syndroms C) Focal syndroms According to American Diabetes Association .determined by abnormalities in elecrodiagnostic and quantitative sensory testing B) Diffuse clinical neuropathy .Types of Diabetic Neuropathy A) Subclinical neuropathy . 2000. 43: 957-973 .
43: 957-973 .Symptoms of Diabetic Neuropathy Sensorimotor neuropathy: Small fibre: manifested by pain and paresthesia early in the course of diabetes Associated with the onset of insulin therapy. 2000. Large fibre: impaired motor functions.symptoms vary depending on which organ/system is affected Diabetologia. position sense And cold thermal perception Autonomic neuropathy : / 0 9$8 '"!'"$ -. vibration perception.
Pathogenesis of Diabetic Neuropathy HYPERGLYCEMIA ANGIOPATHY DECREASED LEVEL OF NGF AND IGF .
Advanced End-Glycation Products Oxidative stress. inflammation AGE Receptor (RAGE) Lukic et al. 2008 .Pathogenesis of Diabetic Neuropathy HYPERGLYCEMIA Polyol pathway Level of frucose and sorbitol Non-enzymatic protein glycation AGEs.
HYPOTHESIS RAGE deletion facilitates nerve regeneration and protects nerves from adverse consequences of hyperglycemia MAP kinase AGEs – RAGE binding NFkappaB P3I Inflammation Degeneration .
C57 B6 .org Induction of hyperglycemia: Intraperitoneal injections of Streptozotocin (STZ) (60 mg/kg dissolved in cytrinian buffer.jax. daily for 5 consecutive days) Permanent destruction of pancreatic beta-cells .Materials and Methods Animals: Control and hyperglycemic groups .RKO B6 www.
8 wk old mice injected with Strepotozotocin (STZ) (induced damage of pancreatic beta-cells resulting in hyperglycemia) . spinal cord.10 days after 1st STZ injection . dorsal root ganglia) .3 wks after the crush .euthanasia .2 mths after hyperglycemia was confirmed .glucose level check .sciatic nerve crush .specimen collection (sciatic nerve.EEG (electrography) – nerve condution velocity tests .Methodology (1) Data points .
com en.net biology.ru cytochemistry.edu .Quantification of the number of apoptotic cells in the dorsal root ganglion drpinna.uc.Assessement of myelinated fibre number in the crushed nerve .Immunohistochemical and biochemical assessement of RAGE expression in the nerve .clc.Comparative analysis of macrophage influx in the nerve between all groups of animals .Methodology (2) Techniques .academic.
Nerve condution velocity (NCV) Electromiograph Checking temperature Inserting electrodes .
Diabetes 57:1002–1017.NCV – study background (1) Control and hyperglycemic C57 mice Toth et al. 2008 .
Diabetes 57:1002–1017.db Toth et al.ct RKO .ct WT -db * * RKO .NCV – study background (2) C57 and RAGE KO mice – control and hyperglycemic Motor nerve * CV (m/s) WT .ct RKO . 2008 .db Sensory nerve * CV (m/s) WT .ct * * WT -db RKO .
NCV – study results Motor nerve conduction velocity Control Sham Crush Przewodzenie sygnału w nerwie czuciowym Control Sham Crush .
db RKO .ctr WT .ctr RKO .db .Myelined fibre density (MFD) CONTROL WT .
db RKO .ctr RKO .db .Myelined fibre density (MFD) SHAM WT .ctr WT .
Myelined fibre density (MFD) CRUSH WT .db .db * * * RKO .ctr WT .ctr RKO .
ct * * RKO .ct WT -db RKO .db RKO .db .ctr RKO .db * CV (m/s) WT .CONCLUSION WT .db RKO diabetic (hyperglycemic) mice have better nerve regeneration index compared to their wild type C57 counterparts * * * * * CV (m/s) WT .ct * RKO .ctr WT .ct WT -db RKO .
RAGE and Nerve Regeneration TWO PATHWAYS GLOBAL CHANGES LOCAL CHANGES .
in sciatic nerve of control mice WT RKO Increased level of AGEs CML BASELINE levels No. of fibres per ROI 12 100 90 80 IF signal (AIU) 10 8 6 4 * 70 60 50 40 30 * 2 0 20 10 0 WT RKO WT RKO .Expression of CML – AGE precurosor.
Fast axonal transport of mDia – immunofluorescent study WT RKO WT RKO WT RKO Proximal 1h Axonal transport impairment 3h 8h Distal .
WT Control RKO Lower Macrophage activity Diabetic 25 20 15 Macrophage no. DAPI and30 F4/80 distribution in sciatic nerve 30 20 10 0 * Diabetic Baseline levels 10 5 0 WT-ctr RKO-ctr WT-db RKO-db WT RKO .
of stained neurons 200 150 100 50 - * Diabetic Baseline levels WT RKO .WT Control RKO Diabetic Control Increased Apoptosis Diabetic Cleaved Caspase expression in DRG No.
Further Studies RAGE/HMB DIABETES RAGE/mDIA Pharmacological blockage of RAGE in mice Comparative distribution of RAGE in sciatic nerve of human diabetic patients RAGE HMB RAGE mDIA .
All Lab Members Special thanks to Ann Marie SCHMIDT Matthew GEDDIS Ravichandran Ramasamy Marta BANACH Jose GARCIA Thomas BRANNAGAN Rosa ROSARIO Fei SONG .