Molecular mechanisms underlying the effects of cyclosporin A and sirolimus on glucose and lipid metabolism in liver,skeletal muscle and adipose tissue in an in vivo rat model

Cyclosporin A (CsA) and sirolimus (SRL) are immunosuppressive agents (IAs) associated with dyslipidemia, insulin resistance and new onset diabetes after transplantation (NODAT). However, the molecular mechanisms involved are not fully understood. We investigated the effects of six-week treatment of either CsA or SRL on glucose and lipid metabolism in Wistar rats. The results show that, compared with vehicle-treated rats, SRL-treated rats were significantly lighter starting at week 5. CsA or SRL caused glucose intolerance, increased storage of lipids in the liver and skeletal muscle, and decreased the insulin-stimulated glucose uptake in isolated adipocytes. Furthermore, these agents significantly decreased genes involved in insulin action and glucose uptake, such as, IRS-1, Glut4 and Glut1, and increased genes and/or proteins involved in hepatic lipogenesis and gluconeogenesis, while decreasing them in adipose tissue. After either treatment PGC1α gene expression was down regulated in skeletal muscle, an important player in fatty acid oxidation. Moreover, there was an increase in IL-6 gene expression in adipose tissue in the SRL-treated rats, suggesting stimulation of lipolysis. The results of the present study suggest that CsA and SRL lead to metabolic alterations in liver, muscle and adipose tissue, which may contribute to the development of dyslipidemia and insulin resistance associated with immunosuppressive therapy.     

Quantitation of activated factor VII levels in plasma using a tissue factor mutant selectively deficient in promoting factor VII activation

Although the majority of factor VII (FVII) circulates in the zymogen form, low levels of activated factor VII (FVIIa) have been postulated to exist in plasma and to serve a priming function for triggering of the clotting cascade. However, direct measurement of plasma FVIIa has not previously been possible. We have quantified plasma FVIIa levels using a novel, highly sensitive assay that is free from interference by FVII. Specificity of this clot-based assay results from the use of a mutant tissue factor that is selectively deficient in promoting FVII activation, but retains FVIIa cofactor function. In normal adults, FVIIa was found to be present in plasma (mean: 3.6 ng/mL) with considerable variation between individuals (range: 0.5 to 8.4 ng/mL). FVIIa levels were only loosely correlated with FVII coagulant activity, but were elevated in pregnancy and reduced with oral anticoagulant therapy. Incubation of plasma on ice in glass containers (cold activation) resulted in substantial FVIIa generation. Measurement of plasma forms of factor VII is of potential clinical importance because elevated FVII coagulant activity has been implicated as a significant risk predictor for ischemic heart disease. Clinically, this new assay will now permit direct assessment of the role of plasma FVIIa in thrombotic disorders.     

Comparison of the hemagglutination inhibition assay kit for erythropoietin (ESF) with the fetal mouse liver cell bioassay in vitro

The commercially available hemagglutination inhibition (HAI) assay kit for erythropoietin (ESF) was compared with the fetal mouse liver cell (FMLC) bioassay. No correlation was obtained ESF levels determined by both methods in a variety of pathologic sera. The HAI kit showed a great batch variability. Significant immunoreactivity was found in those fractions of a normal human serum and a human urinary ESF preparation that were not active in the FMLC bioassay. A very poor recovery of immunoreactivity was found when the international reference preparation for erythropoietin (second IRPE) was added to a normal human serum.     

Expression of p53 in oral mucosal hyperplasia, dysplasia and squamous cell carcinoma

OBJECTIVE: To assess p53 expression in a range of oral mucosal lesions and to relate the results to the clinical outcome in patients with dysplastic oral mucosal lesions and oral squamous cell carcinomas (OSCC).
MATERIALS AND METHODS: Archival tissue was available for eight cases of normal oral mucosa, 50 cases of oral mucosal hyperplasia, 41 cases of oral mucosal dysplasia and 48 cases of OSCC. The monoclonal antibody DO-7, reactive to p53 protein, was applied to paraffin-embedded sections using microwave pretreatment and immu-nohistochemical techniques.
RESULTS: The results showed that normal oral mucosa did not express p53.Positive nuclear staining was found in 18/50 (36%) cases of hyperplasia, 35/41 (85%) cases of dysplasia and 45/48 (94%) cases of OSCC.None of the p53 negative dysplasias progressed, while 19% of p53 positive cases of dysplasia recurred following excision and 11% of the cases underwent neoplastic transformation. Five out of 10 (50%) cases of severe dysplasia which were p53 positive resolved.
CONCLUSION: The proportion of cases with positive p53 expression increased from hyperplasia to dysplasia to OSCC. These results may indicate an involvement of p53 in neoplastic transformation as well as in proliferative events although the presence or absence of p53 staining could not be used to predict the outcome of potentially malignant oral mucosal lesions.     

Inflammation and dyslipidaemia: a possible interplay between established risk factors in North Indian males with coronary artery disease

Objectives

Coronary artery disease (CAD) is a leading cause of morbidity and mortality in the developed world and is rapidly assuming epidemic proportions in developing countries, including India. This has led to extensive research to determine the risk factors and the pathways that may predispose to the elevated risk of this disease. Important among them include lipoproteins, homocysteine, lipoprotein (a), pro-inflammatory cytokines and others. The following study was undertaken to determine a possible inter-relationship between inflammation and dyslipidaemia, which are important risk factors for CAD in the atherosclerosis-prone North Indian male population.

Methods

The study groups comprised 150 clinically assessed North Indian male patients with acute myocardial infarction (AMI), diagnosed on electrocardiographic and biochemical criteria, and 150 healthy controls. Apolipoprotein-AI (Apo-AI), apolipoprotein-B (Apo-B) and C-reactive protein (CRP) levels were estimated using kits based on the immunoturbidimetric assay from Randox, UK. Tumour necrosis factor-α (TNF-α) and lipoprotein (a) were assayed using commercially available ELISA kits from Diaclone Research, Belgium and Innogenetics, Belgium, respectively.

Results

The patients with AMI showed highly significant elevations in the levels of total serum cholesterol, triglycerides, LDL cholesterol, Apo-B and a significant decline in HDL cholesterol, compared with healthy controls. Significantly elevated serum levels of inflammatory markers, TNF-α and CRP were seen in patients with AMI, compared to the control subjects. A significantly positive correlation of TNF-α was observed with lipoprotein (a) in patients with CAD.

Conclusion

The data clearly underlines a possible interplay between inflammation and dyslipidaemia in the pathogenesis of CAD in the Indian context. This insight into the aetiopathogenesis of CAD will prove highly beneficial for devising better preventive measures and pharmacological interventions for CAD.