Effects of thalidomide on endothelial and metabolic dysfunction in male Wistar rats exposed to chronic mild stress (CMS) and atherogenic diet: molecular and immunohistochemical findings

BAM Ismail1, SA Aboul-Fotouh1, AA Ibrahim2, HH Shehata2, EA Ibrahim3, MI Salman3, OA Hassan1, AM Abdel-tawab1. 1Department of Pharmacology, Facilty of Medicine, Ain Shams University, Cairo, 11566, Egypt, 2Department of Biochemistry, Faculty of Medicine, Ain Shams University, Cairo, 11566, Egypt, 3Department of Pathology, Faculty of Medicine, Ain Shams University, Cairo, 11566, Egypt.

Introduction There is evidence that depression is a risk factor for cardiovascular diseases. The current study aimed at examining the hypothesis that the proinflammatory cytokine, TNF-α, would partially explain the link between depression, insulin resistance and atherosclerotic endothelial changes. Thalidomide ‘THAL’ was used as a pharmacologic probe to annul the expression of TNF-α.

Methods Forty-one male Wistar rats (200-250 g) were divided into 5 groups: naïve; exposed to chronic-mild-stress (CMS); CMS with cholesterol-cholic acid-thiouracil ‘CCT’ diet for 8 weeks; and two groups exposed to CMS-CCT and administered thalidomide ‘THAL’ 100 mg/kg/day i.p. for the last 2 weeks or imipramine ‘IMIP’ 20 mg/kg/day i.p. for the last 3 weeks. Rats were assessed for behavioural changes (sucrose preference, open field and forced- swimming-immobilization), body weight, metabolic parameters (lipid profile, HOMA index) and serum TNF-α were assayed. Animals were killed under anaesthesia (urethane: 1.2 g/kg). Then, isolated aortic rings were mounted in tissue baths for isometric tension measurement. Nitric oxide metabolites in aortae were assessed. Aortic and hepatic TNF-α was assessed by ELISA and immunohistochemistry, TNF-α mRNA was assessed using semi-quantitative reverse-transcription-polymerase-chain-reaction (sQ-RT-PCR). Data were statistically analysed using ANOVA with post hoc tests to compare the means of the different groups. GraphPad software was used in analysis and presentation. Data are presented as means and SD.

Results THAL and IMIP exposed rats showed amelioration of CMS/CCT-related behavioural and body weight changes. Exposure to CMS/CCT protocol significantly decreased heart rate compared to control. Imipramine significantly increased heart rate while thalidomide showed insignificant effect on heart rate. Imipramine and to more extent thalidomide, induced significant reduction in systolic blood pressure, compared to untreated CMS/CCT group. A reduction in serum TNF-α was significant (P < 0.001) with THAL but not with IMIP in comparison to CMS-CCT [33.17±4.50, 58±15.53, 71.95±11.28 pg/mL, respectively]. THAL improved CMS/CCT-induced metabolic and endothelial changes that were worsened by IMIP (except HOMA index). Thalidomide, but not imipramine, improved CMS/CCT induced changes in vascular reactivity of isolated aortic rings. Atherogenic changes were assessed microscopically. RT-PCR showed significant (P < 0.001) percent reduction of aortic TNF-α mRNA expression with THAL and (P < 0.05) with IMIP, in comparison to the CMS-CCT group [32.23±1.6, 56.1±6.1, 67.75±5.49% respectively]. These data were parallel to the ELIZA and immunohistochemical findings in aortic tissues. Similar patterns of findings were noticed with TNF-α immunohistochemical findings in the liver

Conclusion TNF-α provides a target link between depression, metabolic syndrome and endothelial dysfunction. This would open a new therapeutic approach that addresses the co-morbidities of depression.