Shafeeque Ahmad

Hypolipidemic and antioxidant activities of thymoquinone and limonene in atherogenic suspension fed rats

The hypolipidemic and antioxidant actions of thymoquinone (TQ) and limonene (LMN) were investigated by giving 1 ml of 10mg TQ or 200mg LMN suspension, by gavage in two equal doses (morning and evening) of 0.5 ml each for 30 days, in rats, fed an atherogenic suspension. These compounds effectively ameliorated all the altered cardiovascular risk parameters via a reduction in HMG-CoA reductase activity, along with an increase in arylesterase activity. The compounds significantly blocked the shift in buoyancy from less atherogenic lb-LDL to highly atherogenic sd-LDL, restoring the percent distribution of LDL-C and apoB into sd-LDL and lb-LDL to near normal levels. These compounds also blocked basal and maximal formation of CD and malondialdehyde, and lengthened the lag times of LDL, sd-LDL and lb-LDL in the order TQ>LMN. Our results strongly suggest an important therapeutic use of test compounds, especially TQ, in the prevention of cardiovascular disease risks parameters.

Elucidation of mechanisms of actions of thymoquinone-enriched methanolic and volatile oil extracts from Nigella sativa against cardiovascular risk parameters in experimental hyperlipidemia

BackgroundNigella sativa belonging to the Ranunculaceae family has been reported to use for thousands of years as protective and curative traditional medicine against a number of diseases. GC-MS analysis of methanolic extract (ME) and volatile oil (VO) extracted from Nigella sativa seed oil was performed by two different mass spectrometry libraries, WIlEY8 and NIST05s. The cholesterol lowering and antioxidant actions of VO and ME fractions were investigated in atherogenic suspension fed rats.MethodsIn this study, four groups of male Wistar rats were used: normolipidemic control (NLP-C), hyperlipidemic control (HLP-C), methanolic extract (HLP-ME) and volatile oil treated (HLP-VO) groups for 30 days of duration. P value < 0.05 was assumed as significant data in groups.ResultsAdministration of atherogenic suspension to male Wistar rats for 30 days resulted in a marked increase of plasma triglycerides and total cholesterol, and significant change in plasma lipoprotein levels along with a decrease in antioxidant arylesterase activity in hyperlipidemic control (HLP-C) group. The oral feeding of 100 mg ME or 20 mg VO per rat/day effectively reduced the plasma triglycerides to near normal level, while high density lipoprotein cholesterol and its subfraction along with arylesterase activity levels were significantly increased. The test fractions elicited a significant decrease in hepatic HMG-CoA reductase activity. The fractions significantly blocked the ex vivo basal and in vitro maximal formation of conjugated diene and malondialdehyde, and lengthened the lag times of low density lipoprotein, small dense low density lipoprotein and large buoyant low density lipoprotein. ME possessing ω-6 linoleic acid along with palmitic acid active compounds was more effective than VO extract containing thymol and isothymol phenolic antioxidant compounds, thymoquinone phenolic compound common to the both extracts, via reduction in hepatic HMG-CoA reductase activity as well as antioxidant mechanisms.ConclusionThe both extracts especially, ME significantly improve cardiovascular risk parameters in treated rats, and can be used in reactive oxygen species disorders such as cardiovascular diseases.

Hyperglycemia induced reactive species trigger structural changes in human serum albumin of type 1 diabetic subjects

Chronic oxidative stress fuels pathogenesis of a large set of diseases. Oxidative stress is the cause and consequence of numerous diseases including type 1 diabetes mellitus (T1DM), in which there is selective destruction of insulin producing pancreatic β-cells. Studies have documented that hyperglycemia produces profound stress. In vivo production of numerous reactive oxygen, nitrogen, chlorine species and lipid/sugar oxidation products in T1DM patients may be the result of persistent hyperglycemia. Post-translational modifications by reactive species may create new antigenic epitopes and play a role in the development of autoimmune response. In this paper our main focus was to establish the effect of existing hyperglycemia induced oxido-nitrosative stress in T1DM patients on the integrity of human serum albumin. Raised nitric oxide, carbonyl, RBC hemolysis, lowered ferric reducing antioxidant power (FRAP), thiol and deformed RBC in T1DM are all highly suggestive of persistent oxido-nitrosative stress. Hyperglycemia induced generation of advanced glycation end products (AGEs) was established by LCMS. Chronic oxido-nitrosative stress can modify HSA in T1DM patients, producing immunologically active albumin. Therefore, it is speculated that the aberrant HSA may play a role in the initiation/progression of T1DM.

Effect of peroxynitrite on human serum albumin: a multi technique approach.

In this study, human serum albumin (HSA), the most abundant protein of blood plasma, was modified with varying concentrations of peroxynitrite. The peroxynitrite-induced changes in HSA was monitored by spectroscopy, SDS-PAGE, 1-anilinonaphthalene-8-sulfonic acid (ANS), thermal denaturation studies, and matrix-assisted laser desorption/inonization-time of flight mass spectrometry (MALDI-TOF MS). Aggregate formation was studied by thioflavin T binding and scanning electron microscopy (SEM). The results indicated formation of 3-nitrotyrosine, 6-nitrotryptophan, dityrosine, and carbonyls in modified samples and showed retarded mobility in SDS–polyacrylamide gel. Reduction in α-helicity and surface protein hydrophobicity confirmed the secondary and tertiary structure alterations in peroxynitrite-modified-HSA. Also, attachment of nitro group and increase in melting temperature was observed in modified sample. Furthermore, significant enhancement in the fluorescence intensity of ThT upon binding with peroxynitrite-modified-HSA and images under scanning electron microscope are suggestive of protein aggregation. It is, therefore, speculated that HSA modified by endogenously formed peroxynitrite might act as a trigger for nitration/aggregation and suggested the role of peroxynitrite-modified-HSA in SLE.

SLE autoantibodies are well recognized by peroxynitrite-modified-HSA: Its implications in the pathogenesis of SLE

Systemic lupus erythematosus (SLE) is an autoimmune disorder where the role of inflammatory processes in the etiopathogenesis is well documented. Despite extensive research, the trigger for initiation of the disease has not been identified. Peroxynitrite, a strong nitrating/oxidizing agent has been reported in SLE and other autoimmune diseases. In this study, human serum albumin (HSA) was exposed to peroxynitrite for 30min at 37°C. The structure of HSA was grossly perturbed when examined by various physico-chemical techniques. Peroxynitrite mediated nitration of HSA was confirmed by LCMS/MS. Furthermore, increase in hydrodynamic radius of peroxynitrite-modified-HSA suggests the attachment of nitro group(s). Aggregation in peroxynitrite-modified-HSA was evident in a TEM scan. Nitration, oxidation, cross linking, aggregation etc conferred immunogenicity on peroxynitrite-modified-HSA. High titre antibodies were elicited in rabbits immunized with peroxynitrite-modified-HSA. Induced antibodies were highly specific for peroxynitrite-modified-HSA but showed considerable binding with other nitrated molecules. Direct binding/inhibition ELISA carried out with autoantibodies in SLE sera showed preferential binding with peroxynitrite-modified-HSA. Anti-nDNA positive IgG from SLE sera showed preference for peroxynitrite-modified-HSA when subjected to immunoassay (direct binding and inhibition) and mobility shift assay. Our results reinforce the role of augmented inflammation in SLE progression.