Although multiple toxins have been implicated in the development of uraemic neuropathy, no causative agent has been identified. In the present study, the excitability properties of lower limb motor nerves in patients with end-stage kidney disease treated with haemodialysis were measured before, during and after a standard 5 h haemodialysis session, in an attempt to explore the pathophysiology of uraemic neuropathy. Compound muscle action potentials were recorded from tibialis anterior and extensor digitorum brevis, following stimulation of the common peroneal nerve in 14 patients. Measures of excitability were assessed in relation to changes in serum levels of potential neurotoxins, including potassium, calcium, urea, uric acid, parathyroid hormone and beta-2-microglobulin. Before dialysis, measures of nerve excitability were significantly abnormal in the patient group for axons innervating tibialis anterior and extensor digitorum brevis, consistent with axonal depolarization: refractoriness was increased and superexcitability and depolarizing threshold electrotonus were reduced. Pre-dialysis excitability abnormalities were strongly correlated with serum K+. Correlation was also noted between the severity of symptoms and excitability abnormalities. Haemodialysis normalized the majority of nerve excitability parameters. In conclusion, lower limb motor axons in uraemic patients are depolarized before dialysis. The correlation between serum K+ and excitability measures indicates that hyperkalaemia is primarily responsible for uraemic depolarization, and a likely contributing factor to the development of neuropathy.

Objective: Little is known of the metabolism of different isoforms of adiponectin. We therefore (a) characterised the size distribution of human adiponectin in relation to gender, body composition and following a challenge with a fat meal or oral glucose in humans, and (b) studied the metabolism of isoforms of human adiponectin in rabbits. Method: Electrophoresis, blotting and chromatography were used to characterise human adiponectin in 36 healthy subjects, including 15 with at least two first-degree relatives with type 2 diabetes, before and after consumption of a fatty meal or glucose. The metabolism of column-fractionated human adiponectin was studied in rabbits, some of which were coinjected with insulin. Results: Females had a higher proportion of high molecular weight (HMW) and hexameric adiponectin (P ¼ 0.002 and 0.004 respectively), and a lower proportion of trimers (P , 0.0001) than males. Females also showed a strong negative relationship between body fat measures and the proportion of HMW adiponectin. There were no differences in isoforms between insulin-resistant and -sensitive subjects, or following oral glucose or a fat meal. Adiponectin in rabbits had an extravascular/intravascular ratio of 0.71, and a half-life (T1/2) of 14.3 h. Metabolism was not influenced by insulin or reduction of sulphydryl bonds. HMW and trimeric isoforms had a significantly different T1/2 of 13.0 and 17.5 h respectively (P , 0.05), and these isoforms did not interconvert in vivo. Conclusions: Human adiponectin is present as trimers, hexamers and HMW forms. Females had a higher proportion and absolute amount of HMW species compared with males, and female, but not male, subjects showed a strong negative relationship between measures of body fat, and the proportion of HMW species. These isoforms did not respond to challenge in man with a fatty meal or oral glucose, and in the rabbit, to injected insulin. HMW adiponectin was more rapidly metabolised than the trimeric form, but both were stable in vivo, and did not interconvert. We conclude that human adiponectin is much longer-lived than is the case with other hormones, a finding with positive implications for the potential to supplement levels of adiponectin in man.European Journal of Endocrinology 153 409-417

OBJECTIVE -Inflammatory markers such as C-reactive protein (CRP) are associated with insulin resistance, adiposity, and type 2 diabetes. Whether inflammation causes insulin resistance or is an epiphenomenon of obesity remains unresolved. We aimed to determine whether first-degree relatives of type 2 diabetic subjects differ in insulin sensitivity from control subjects without a family history of diabetes, whether first-degree relatives of type 2 diabetic subjects and control subjects differ in CRP, adiponectin, and complement levels, and whether CRP is related to insulin sensitivity independently of adiposity.RESEARCH DESIGN AND METHODS -We studied 19 young normoglycemic nonobese first-degree relatives of type 2 diabetic subjects and 22 control subjects who were similar for age, sex, and BMI. Insulin sensitivity (glucose infusion rate [GIR]) was measured by the euglycemic-hyperinsulinemic clamp. Dual-energy X-ray absorptiometry determined total and abdominal adiposity. Magnetic resonance imaging measured abdominal adipose tissue volumes.RESULTS -First-degree relatives of type 2 diabetic subjects had a 20% lower GIR than the control group (51.8 Ϯ 3.9 vs. 64.9 Ϯ 4.6 mol ⅐ min Ϫ1 ⅐ kg fat-free mass Ϫ1 , P ϭ 0.04). However, first-degree relatives of subjects with type 2 diabetes and those without a family history of diabetes had normal and comparable levels of CRP, adiponectin, and complement proteins. When the cohort was examined as a whole, CRP was inversely related to GIR (r ϭ Ϫ0.33, P ϭ 0.04) and adiponectin (r ϭ Ϫ0.34, P ϭ 0.03) and positively related to adiposity (P Ͻ 0.04). However, CRP was not related to GIR independently of fat mass. In contrast to C3 (r ϭ 0.41, P ϭ 0.009) and factor B (r ϭ 0.43, P ϭ 0.005), CRP was unrelated to factor D.CONCLUSIONS -The insulin-resistant state is not associated with changes in inflammatory markers or complement proteins in subjects at high risk of type 2 diabetes. Our study confirms a strong relationship between CRP and fat mass. Increasing adiposity and insulin resistance may interact to raise CRP levels. Diabetes Care 27:2033-2040, 2004I nsulin resistance is fundamental to the pathogenesis of the metabolic syndrome and type 2 diabetes (1) and is a heritable trait (2). Type 2 diabetes is also strongly genetically determined (3). Consequently, first-degree relatives of subjects with type 2 diabetes demonstrate the metabolic accompaniments of insulin resistance before they develop overt diabetes (4). Because hyperglycemia further impairs both insulin action and insulin secretion (5), the study of primary metabolic defects leading to insulin resistance is best undertaken before insulin secretion begins to fail and blood glucose rises (6). In addition to genetic factors, excess body fat, particularly visceral fat, has been linked to the pathogenesis of insulin resistance and type 2 diabetes. We have reported that increased central adiposity is strongly related to reduced insulin sensitivity in subjects with and without firstdegree relatives with type 2 diabetes (7). Furthermo...

OBJECTIVE:Adiponectin is an adipose-specific protein with short-term effects in vivo on glucose and fatty acid levels. We studied the plasma concentration and the proteolytic activation status of adiponectin following the consumption of a high-fat, lowcarbohydrate meal. DESIGN: Analysis of adiponectin concentration and polypeptide structure after consumption of a fat meal. SUBJECTS: Normal subjects (n ¼ 24) and first-degree relatives of patients with type II diabetes (n ¼ 20). MEASUREMENTS: All subjects had a normal fasting plasma glucose and glucose tolerance. Blood was collected for the determination of plasma insulin, adiponectin, triglyceride, and free fatty acids. Body composition was assessed with dual-energy X-ray absorptiometry and whole-body insulin sensitivity with a euglycaemic, hyperinsulinaemic clamp. Postprandial response over 6 h was determined for plasma adiponectin, glucose, insulin, triglyceride, and free fatty acids. Adiponectin was measured by commercial RIA and its polypeptide structure examined by Western blotting. RESULTS: The relatives were more insulin resistant and had increased adiposity compared with control subjects. There was no significant difference in postprandial response in fatty acids, triglyceride, or insulin between the groups. Postprandial levels of adiponectin measured by radioimmunoassay were not significantly different from fasting levels, and no breakdown products of adiponectin were detectable in postprandial samples by Western blotting. CONCLUSIONS: Levels of circulating adiponectin do not alter in response to a fat meal, despite evidence in mice that acute changes in adiponectin significantly affect postprandial fatty acid flux. Moreover, a fat meal challenge did not lead to significant activation of adiponectin by proteolytic conversion.

Ischemia/reperfusion (IR) injury is a leading cause of acute renal failure and an important contributor to allograft damage. Tissue factor (TF) is up-regulated during IR, and TF inhibition reduces renal injury. However, the underlying mechanisms by which TF contributes to injury have not been elucidated. We postulated that TF contributes to IR injury by production of coagulation proteases and subsequent signaling by protease activated receptor (PARs). We compared renal injury after 25 minutes of bilateral renal ischemia and varying periods of reperfusion in C57BL/6 mice, those expressing low levels of TF (low-TF), hirudin-treated C57BL/6, and mice lacking either PAR-1 or PAR-2. C57BL/6 mice developed severe renal failure and died within 48 hours of reperfusion. In contrast, low-TF, hirudin-treated C57BL/6, and PAR-1-/- mice were protected from renal failure and had reduced mortality, tubular injury, neutrophil accumulation, and lower levels of the chemokines KC and MIP-2. Importantly, PAR-1-/- mice had lower chemokine levels despite up-regulation of TF and fibrin deposition. In addition, treating PAR-1-/- mice with hirudin conferred no additional benefit. Somewhat surprisingly, PAR-2 deficiency did not protect from renal failure. These experiments indicate that increased TF activity after renal IR leads to increased CXC chemokine expression and subsequent neutrophil-mediated injury predominantly by thrombin-dependent PAR-1 signaling.

SUMMARYIn liie activated complement system, vitronectin (complement S-protcin) occupies ihe metastabie membrane binding site of the nascent precursor compiex C5b-7, so that the newly formed SC5b-7 is unable to insert into celi membranes. Some evidence also indicates that vitronectin limits on-going membrane-associated pore ibrmation by inhibiting C9 poiynierization. It has been assumed that these two stages of terminal complement complex (TCC) inhibition taice piace through charge interactions between the heparin-binding region of vitronectin and homoiogous cysteine-rich sequences of the late complement proteins C6, C7, C8 and C9. We examined SC5b-7 formation and inhibition of C9 binding in the TCC using separate haemolylie assays. The mode of action of vitronectin in these assays was compared with iwo 15mer peptides which span residues .148-379 of ihe heparin-binding region, and a heparin-atlinity polypeptide. protamine sulphate. The results showed that vitronectin acts predominantly through SC5b-7 production with a lesser effect on the inhibition of C9 lytic pore formation, in contrast, protamine suiphate did not prevent C5b-7 membrane attachment, but was a potent inhiiiitor of C9-mediated lysis. The peptides did not inhibit C5b-7 membrane insertion and only one aiTeeted C9 binding. These data suggest that the two stages of TCC inhibition involve separate binding sites on the vitronectin molecule. The site for association with nascent C5b-7 is unknown, whereas inhibition of C9 binding and pore formation taices place through the heparin-binding region.

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