In silico anti-viral assessment of phytoconstituents in a traditional (Siddha Medicine) polyherbal formulation - Targeting Mpro and pan-coronavirus post-fusion Spike protein.

Background and aim: Novel nature of the viral pathogen SARS-CoV-2 and the absence of standard drugs for treatment, have been a major challenge to combat this deadly infection. Natural products offer safe and effective remedy, for which traditional ethnic medicine can provide leads. An indigenous poly-herbal formulation, Kabasura Kudineer from Siddha system of medicine was evaluated here using a combination of computational approaches, to identify potential inhibitors against two anti-SARS-CoV-2 targets - post-fusion Spike protein (structural protein) and main protease (Mpro, non-structural protein). Experimental procedure: We docked 32 phytochemicals from the poly-herbal formulation against viral post-fusion Spike glycoprotein and Mpro followed by molecular dynamics using Schrodinger software. Drug-likeness analysis was performed using machine learning (ML) approach and pkCSM. Results: The binding affinity of the phytochemicals in Kabasura Kudineer revealed the following top-five bioactives: Quercetin > Luteolin > Chrysoeriol > 5-Hydroxy-7,8-Dimethoxyflavone > Scutellarein against Mpro target, and Gallic acid > Piperlonguminine > Chrysoeriol > Elemol > Piperine against post-fusion Spike protein target. Quercetin and Gallic acid exhibited binding stability in complexation with their respective viral-targets and favourable free energy change as revealed by the molecular dynamics simulations and MM-PBSA analysis. In silico predicted pharmacokinetic profiling of these ligands revealed appropriate drug-likeness properties. Conclusion: These outcomes provide: (a) potential mechanism for the anti-viral efficacy of the indigenous Siddha formulation, targeting Mpro and post-fusion Spike protein (b) top bioactive lead-molecules that may be developed as natural product-based anti-viral pharmacotherapy and their pleiotropic protective effects may be leveraged to manage co-morbidities associated with COVID-19.

Nature-inspired Enzyme engineering and sustainable catalysis: biochemical clues from the world of plants and extremophiles.

The use of enzymes to accelerate chemical reactions for the synthesis of industrially important products is rapidly gaining popularity. Biocatalysis is an environment-friendly approach as it not only uses non-toxic, biodegradable, and renewable raw materials but also helps to reduce waste generation. In this context, enzymes from organisms living in extreme conditions (extremozymes) have been studied extensively and used in industries (food and pharmaceutical), agriculture, and molecular biology, as they are adapted to catalyze reactions withstanding harsh environmental conditions. Enzyme engineering plays a key role in integrating the structure-function insights from reference enzymes and their utilization for developing improvised catalysts. It helps to transform the enzymes to enhance their activity, stability, substrates-specificity, and substrate-versatility by suitably modifying enzyme structure, thereby creating new variants of the enzyme with improved physical and chemical properties. Here, we have illustrated the relatively less-tapped potentials of plant enzymes in general and their sub-class of extremozymes for industrial applications. Plants are exposed to a wide range of abiotic and biotic stresses due to their sessile nature, for which they have developed various mechanisms, including the production of stress-response enzymes. While extremozymes from microorganisms have been extensively studied, there are clear indications that plants and algae also produce extremophilic enzymes as their survival strategy, which may find industrial applications. Typical plant enzymes, such as ascorbate peroxidase, papain, carbonic anhydrase, glycoside hydrolases and others have been examined in this review with respect to their stress-tolerant features and further improvement via enzyme engineering. Some rare instances of plant-derived enzymes that point to greater exploration for industrial use have also been presented here. The overall implication is to utilize biochemical clues from the plant-based enzymes for robust, efficient, and substrate/reaction conditions-versatile scaffolds or reference leads for enzyme engineering.

Targeting lipid-sensing nuclear receptors PPAR (α, γ, ß/δ): HTVS and molecular docking/dynamics analysis of pharmacological ligands as potential pan-PPAR agonists.

The global prevalence of obesity-related systemic disorders, including non-alcoholic fatty liver disease (NAFLD), and cancers are rapidly rising. Several of these disorders involve peroxisome proliferator-activated receptors (PPARs) as one of the key cell signaling pathways. PPARs are nuclear receptors that play a central role in lipid metabolism and glucose homeostasis. They can activate or suppress the genes responsible for inflammation, adipogenesis, and energy balance, making them promising therapeutic targets for treating metabolic disorders. In this study, an attempt has been made to screen novel PPAR pan-agonists from the ZINC database targeting the three PPAR family of receptors (α, γ, ß/δ), using molecular docking and molecular dynamics (MD) simulations. The top scoring five ligands with strong binding affinity against all the three PPAR isoforms were eprosartan, canagliflozin, pralatrexate, sacubitril, olaparib. The ADMET analysis was performed to assess the pharmacokinetic profile of the top 5 molecules. On the basis of ADMET analysis, the top ligand was subjected to MD simulations, and compared with lanifibranor (reference PPAR pan-agonist). Comparatively, the top-scoring ligand showed better protein-ligand complex (PLC) stability with all the PPARs (α, γ, ß/δ). When experimentally tested in in vitro cell culture model of NAFLD, eprosartan showed dose dependent decrease in lipid accumulation and oxidative damage. These outcomes suggest potential PPAR pan-agonist molecules for further experimental validation and pharmacological development, towards treatment of PPAR-mediated metabolic disorders.

Nutraceutical and flavor profiles in underutilized desert legumes of India: gene editing strategies towards sustainable food development.

Climate change has posed a challenge for food security all over the world in the form of fluctuating crop yields and novel disease outbreaks in plants. Human society's overdependence on a few food crops does not seem a wise precedence. There are numerous underutilized/orphan/neglected legumes growing in the Indian desert regions that can come to the rescue and act as balanced and sustainable sources of nutrients and health-benefitting nutraceuticals. However, challenges such as low plant yield, unidentified metabolic pathways and off-flavor in the food products derived from them prevent the realization of their full potential. Conventional breeding techniques are too slow to achieve the desired modifications and cater to the sharply rising demand for functional foods. The novel gene editing tools like CRISPR-Cas provide more precise tool to manipulate the target genes with or without introduction of foreign DNA and therefore, have better chances to be accepted by governments and societies. The current article reports some of the relevant 'gene editing' success stories with respect to nutraceutical and flavor profiles in the popular legumes. It highlights gaps and future potential, along with areas requiring caution, in underutilized edible legumes of the Indian (semi) arid regions like Prosopis cineraria, Acacia senegal and Cyamopsis tetragonoloba.

Investigating the antioxidant activity enhancer effect of Cyamopsis tetragonoloba seed extract on phenolic phytochemicals.

Introduction: Phenolic phytochemicals are known for antioxidant-mediated pharmacological effects in various diseases (diabetes, cancer, CVDs, obesity, inflammatory and neurodegenerative disorders). However, individual compounds may not exert the same biological potency as in combination with other phytochemicals. Cyamopsis tetragonoloba (Guar), an underutilized semi-arid legume which has been used as a traditional food in Rajasthan (India), is also a source of the important industrial product guar gum. However, studies on its biological activity, like antioxidant, are limited. Methods: We tested the effect of C. tetragonoloba seed extract to enhance the antioxidant activity of well-known dietary flavonoids (quercetin, kaempferol, luteolin, myricetin, and catechin) and non-flavonoid phenolics (caffeic acid, ellagic acid, taxifolin, epigallocatechin gallate (EGCG), and chlorogenic acid) using DPPH radical scavenging assay. The most synergistic combination was further validated for its cytoprotective and anti-lipid peroxidative effects in in vitro cell culture system, at different concentrations of the extract. LC-MS analysis of purified guar extract was also performed. Results and discussion: In most cases, we observed synergy at lower concentrations of the seed extract (0.5-1 mg/ml). The extract concentration of 0.5 mg/ml enhanced the antioxidant activity of Epigallocatechin gallate (20 µg/ml) by 2.07-folds, implicating its potential to act as an antioxidant activity enhancer. This synergistic seed extract-EGCG combination diminished the oxidative stress nearly by double-fold when compared with individual phytochemical treatments in in vitro cell culture. LC-MS analysis of the purified guar extract revealed some previously unreported metabolites, including catechin hydrate, myricetin-3-galactoside, gossypetin-8-glucoside, and puerarin (daidzein-8-C-glucoside) which possibly explains its antioxidant enhancer effect. The outcomes of this study could be used for development of effective nutraceutical/dietary supplements.

COVID-19 infection and metabolic comorbidities: Mitigating role of nutritional sufficiency and drug - nutraceutical combinations of vitamin D.

The vulnerability of human health is amplified in recent times with global increase in non-communicable diseases (due to lifestyle changes and environmental insults) and infectious diseases (caused by newer pathogens and drug-resistance strains). Clinical management of diseases is further complicated by disease severity caused by other comorbid factors. Drug-based therapy may not be the sole approach, particularly in scenarios like the COVID-19 pandemic, where there is no specific drug against SARS-CoV-2. Nutritional interventions are significant in armouring human populations in disease prevention, and as adjunctive therapy for disease alleviation. Amidst ongoing clinical trials to determine the efficacy of Vit. D against infections and associated complications, this review examines the pleiotropic benefits of nutritional adequacy of vitamin D (Vit. D) in combating viral infections (COVID-19), its severity and complications due to co-morbidities (obesity, diabetes, stroke and Kawasaki disease), based on research findings and clinical studies. Supplements of Vit. D in combination with other nutrients, and drugs, are suggested as promising preventive-health and adjunct-treatment strategies in the clinical management of viral infections with metabolic comorbidities.

Effect of Different Proportions of Phenolics on Antioxidant Potential: Pointers for Bioactive Synergy/Antagonism in Foods and Nutraceuticals.

Phenolic compounds include a broad variety of antioxidant plant substances such as flavonoids that have in common an aromatic ring with one or more hydroxyl groups. Nutraceuticals and health food supplements are designed from flavonoids as well as pure phytochemicals, often in isolation. However, studies on synergistic and antagonistic effects of such compounds are relatively few. In the current study, dual combinations prepared from five phenolic compounds (flavonoid and non-flavonoid) including rutin hydrate, quercetin dihydrate, hydroquinone, kaempferol, and resveratrol were tested for their antioxidant activities using DPPH · radical scavenging assay. The synergistic antioxidant interactions among these phenolics were evaluated by comparing their individual antioxidant effect with that obtained by a mixture of two compounds in various ratios. Quercetin dihydrate showed the highest antioxidant activity. Many combinations were found statistically synergistic in particular ratios. Rutin hydrate and resveratrol showed maximum synergy (1:1, 2:1, and 3:1 ratio). Antagonistic interactions were also identified. The results of this study could be used by industries to develop more potent nutraceutical supplements or guide the researchers for further bioactivity validation using in vivo assays.

In silico and in vitro analysis of PPAR - α / γ dual agonists: Comparative evaluation of potential phytochemicals with anti-obesity drug orlistat.

Obesity is an abnormal fat accumulation disorder in the metabolic syndrome constellation, and a risk factor for diabetes, cardiovascular disorders, non-alcoholic fatty liver disease (NAFLD), and cancer. Nuclear receptors (Peroxisome proliferator-activated receptor, PPAR) are implicated in metabolic syndrome and NAFLD, and have potential for therapeutic targeting. Nuclear receptors are ligand-dependent transcription factors that have diverse roles in metabolism, including regulating genes involved in lipid and glucose metabolism, modulating inflammatory genes, and are crucial for maintaining metabolic flexibility. PPAR activates adipose triglyceride lipase, which then releases fatty acids as ligands for PPAR, indicating the interdependency of nuclear receptors and lipases. Here, molecular docking was performed with selected phytochemical ligands that can bind with PPAR-α/γ (PDB ID: 2ZNN and 2ATH, respectively) using Glide module of Schrodinger software followed by molecular dynamics simulation study using Desmond module, and ADMET analysis. Interestingly, orlistat which is a well-known lipase and fatty acid synthase inhibitor also demonstrated favorable binding affinity with both PPAR-α/γ (-10.96 kcal/mol against PPARα and -10.26 kcal/mol against PPARγ). The highest docking scores were however shown by the flavonoids - rutin (-14.88 kcal/mol against PPARα and -13.64 kcal/mol against PPARγ), and its aglycone, quercetin (-10.08 kcal/mol in PPARα and -9.89 kcal/mol in PPARγ). The other phytochemicals (genistein, esculin, daidzin, naringenin, daidzein, dihydroxy coumarin, hydroquinone) showed lower binding affinity as dual agonists. The anti-obesity effects were experimentally validated in cultured adipocytes, which revealed better lipid inhibition by rutin and quercetin than orlistat (quercetin > rutin > orlistat) pointing to their strong potential in anti-obesity treatment.

Desert legume Prosopis cineraria as a novel source of antioxidant flavonoids / isoflavonoids: Biochemical characterization of edible pods for potential functional food development.

Flavonoids and isoflavonoids in foods are attracting attention as they are significant antioxidant and phytoestrogenic compounds that are beneficial for human health. In this study, the edible pods of the underutilized desert legume Prosopis cineraria from Rajasthan, India were used to extract flavonoids. The pods from semi-arid zone showed the highest flavonoid content (432 mg Rutin hydrate/gm). UV spectrophotometric analysis was also done to characterize flavonoids. The flavonoids and isoflavonoids were further purified from semi-arid zone plants using column chromatography with Amberlite XAD7HP and Sephadex LH-20. LC-MS analysis revealed the presence of medicinally valuable antioxidant flavonoids and isoflavonoids in the pods, viz. vitexin, puerarin, phloridzin, and daidzein. It was seen that the flavonoids/isoflavonoids are present in the selected legume in different forms i.e. pure aglycone, C-glycoside as well as O-glycoside. This finding makes P. cineraria an attractive source candidate for extraction of these nutraceuticals with a potential for development into functional food.

Towards safer non-volatile tissue fixatives: Evaluation of choline-based ionic liquids for fixing ocular tissues.

Volatile organic chemicals (VOCs) are routinely used for processing biological tissue samples in clinical laboratories. Recognizing their serious health and environmental impacts, a few non-volatile green solvents (choline based ionic liquids, ILs) were evaluated as tissue fixatives here. Microscopic evaluation of histo-morphology, fixation and staining quality, and macromolecular integrity (DNA and proteins) were assessed in human eye tissues (sclera, choroid, retinal layers and retinal pigmented epithelium, eyelid and orbit) after IL-fixation. Formalin-fixed tissues were used as standard reference. Microscopic examination revealed favorable histomorphology, tissue fixation and staining characteristics in most tissues immersed in ILs. Time taken to fix, and stability over a period of time (24 h, 48 h, 1 week, 1 month) was also recorded. Electrophoretic analysis revealed stability of cellular proteins and nucleic acids in IL-fixed scleral tissues. Heterogeneity in tissue fixation property relative to the type of ocular tissue, duration of fixation and storage, warrant further design and optimization of ILs to fix biological tissues. The simple cholinium salts based ILs tested here show favorable potential for tissue fixation application, and as an alternative approach to the use of VOCs, towards sustainable biomedical practice.

Clinico-pathological correlations of fatty acid synthase expression in retinoblastoma: an Indian cohort study.

Retinoblastoma (RB), the most common intra-ocular malignancy in children under 5 years of age, has an estimated incidence of about 2000 a year in India, where most cases are in advanced stage at the time of diagnosis. Newer therapeutic approaches would reduce the morbidity of chemotherapy in children with RB. Fatty Acid Synthase (FASN), a lipogenic multi-enzyme complex, is minimally expressed in normal human tissues and over expressed in many cancers, making it an attractive target for cancer therapy. We analyzed RB tissues for FASN protein expression by immunohistochemistry, western blot, and ELISA, and FASN mRNA expression by RT-PCR. FASN expression was correlated with the clinico-pathological characteristics of the tumors. FASN immunostaining was positive in all the 44 RB tissues analyzed (100%). However, FASN expression was heterogeneous within the tumor samples. Tumors with invasion of choroid, optic nerve, orbit and/or retinal pigment epithelium showed significantly higher FASN immunoreactivity than the tumors without invasion (P<0.05), supported by western analysis (P<0.05). FASN expression was significantly high in poorly differentiated retinoblastomas (P<0.05). FASN protein and FASN mRNA estimated by ELISA and RT-PCR respectively showed multi-fold expression over the non-neoplastic muller glial cells that varied quantitatively between tumor tissues. FASN mRNA over-expression was substantially lower than the corresponding FASN protein expression values. The present study reports (i) markedly high expression of FASN protein in poorly differentiated and in invasive retinoblastomas, and (ii) multi-fold over-expression of FASN mRNA and protein in RB tissues, although at varying levels, indicating FASN to be a potential therapeutic target in retinoblastoma management.

Evaluation of suitable solvents for testing the anti-proliferative activity of triclosan - a hydrophobic drug in cell culture.

Triclosan, a broad spectrum antibiotic is currently being evaluated for its anti-cancer property. Though several solvents are available to dissolve lipophilic (hydrophobic) drugs, solubility and toxicity aspects pose a challenge, when combined with the cell culture medium. In this paper, we present a simple approach based on physico-chemical and biologic criteria to choose a suitable solubilizing agent to study the anti-proliferative property of triclosan in breast cancer cell line MCF-7. Triclosan was dissolved in five different solvents viz. DMSO, absolute ethanol, 1 N NaOH, 55% polyethylene glycol + 45% ethanol mixture (PEM) and acetone and diluted with the culture medium (1 mg/ml). Although triclosan dissolved completely in all five solvents, on dilution with culture medium, turbidity was observed in DMSO, 1 N NaOH and ethanol. Cell viability was 95.23% in 10 microl of acetone, when compared with 49.45% at the same volume of PEM. This non-toxic nature of acetone was supported by DNA fragmentation analysis and phase contrast microscopy. A significant decrease in cancer cell proliferation at 100 microg/ml of acetone-solubilized triclosan, compared with 100 microg/ml of PEM-solubilized triclosan (p<0.05) indicated stronger anti-proliferative effect and greater drug-sensitivity of triclosan when solubilized in acetone. Results showed that acetone-solubilized triclosan was suitable for anti-cancer investigations in cultured MCF-7 cells.

Chemical inhibition of fatty acid synthase: molecular docking analysis and biochemical validation in ocular cancer cells.

Fatty acid biosynthesis is an attractive target for anti-cancer therapeutics. The ocular cancer, retinoblastoma cells were treated with fatty acid synthase (FASN) enzyme inhibitors: cerulenin, triclosan and orlistat. The IC(50) and dose-dependent sensitivity of cancer cells to FASN inhibitors decrease in biologic enzyme activity, and cell morphology alterations were analysed. Molecular interactions of enzyme-inhibitor complexes were studied by molecular modelling and docking simulations. The crystal structures of ketoacyl synthase (PDB ID:3HHD) (cerulenin) and thioesterase (PDB ID:2PX6) (orlistat) domains of human FASN were utilized for docking, while for the non-crystallised human FASN enoyl reductase domain (triclosan), homology model was built and used for docking. All three inhibitors showed significant binding energy indicating stable complex formation with their respective FASN subunits. The predicted Ki value of the FASN inhibitors corroborated well with their corresponding anti-cancer effects.

Oxidative stress in retinoblastoma: correlations with clinicopathologic features and tumor invasiveness.

PURPOSE: Retinoblastoma (RB) is the most common primary intraocular malignancy of infancy and childhood, where tumor invasion into the choroid, optic nerve, and/or orbit are risk factors for metastasis. Here we have correlated oxidative stress with the clinicopathologic characteristics of retinoblastoma. METHODS: Tumor samples were processed for histopathologic examination. Malondialdehyde, a biomarker of oxidative stress, was immunohistochemically analyzed in 34 archival retinoblastoma tumor specimens, which included 17 tumors that did not have any invasion of the choroid, optic nerve, and/or orbit, and another 17 tumors that had some form of invasion of the choroid, optic nerve, and/or orbit. Lipid peroxidation levels were biochemically measured in another cohort of retinoblastoma tissue samples (n = 16), and correlated clinicopathologically. RESULTS: Malondialdehyde immunostaining was positive in all 34/34 (100%) tumors, and their corresponding clinicopathologic features were recorded. Malondialdehyde levels were significantly higher in tumors with invasion of the choroid, optic nerve, and orbit, when compared with tumors with no invasion (p < 0.05). No significant correlation was noted between malondialdehyde immunoreactivity and the differentiation/laterality of the tumors. Lipid peroxidation levels increased significantly in tumors with invasion of the choroid, optic nerve, and orbit than in tumors without invasion (p < 0.05). CONCLUSION: RB with invasion of the choroid, optic nerve, and/or orbit strongly correlates with increased oxidative stress.

Favourable modulation of the inflammatory changes in hypercholesterolemic atherogenesis by a low-molecular-weight heparin derivative.

BACKGROUND: In the hypercholesterolemic state, the net result of combined oxidative and nitrosative stress is a pro-inflammatory phenotype that is manifested as increased adhesion molecule expression, enhanced leucocyte trafficking, and increased vascular permeability. The present work explores the inflammatory aspects of hypercholesterolemic atherogenesis, and also evaluates the role of a low-molecular-weight heparin derivative (LMWH), Certoparin, on a biochemical basis. METHODS AND RESULTS: Two groups of male Wistar rats were fed an atherogenic diet (normal rat chow plus 4% cholesterol, 1% cholic acid and 0.5% thiouracil-CCT diet) for 2 weeks. While one was left untreated, the other was administered LMWH (300 microg/day/rat commencing on day 8 and continued for a week). Increased concentrations of plasma C-reactive protein and fibrinogen and cardiac TNF-alpha indicated severe inflammation in the atherogenic diet fed rats. In comparison, these biochemical indices of inflammation diminished significantly in the LMWH treated group (p < 0.001). Significant depletion of thiols, along with accentuated activities of the glutathione metabolising was observed in the cardiac and hepatic tissues of the untreated atherogenic rats, indicating heightened oxidative response. Tissue damage was marked by elevated levels of plasma and tissue hexose, hexosamine, hexuronic acid and sialic acid, which were reversed towards normalcy on LMWH administration. The activities of lysosomal enzymes (N-acetyl glucosaminidase, beta-glucuronidase, beta-galactosaminidase and cathepsin-D) showed a marked increase in the CCT-diet fed rats, while LMWH treated rats showed normal activities (p < 0.001). The osmotic fragility test revealed that the untreated hyperlipidemic rat erythrocytes were significantly fragile at high salt concentrations, while the response was normalized in the LMWH treated group (p < 0.05). Further, hypercholesterolemia induced downregulation of physiological nitric oxide levels was corrected upon treatment with heparin-derivative. CONCLUSION: The results of this work highlight the inflammatory changes in atherogenic conditions and that the low-molecular-weight heparin derivative affords substantial protection.

Influence of a low-molecular-weight heparin derivative on the nitric oxide levels and apoptotic DNA damage in adriamycin-induced cardiac and renal toxicity.

The spectrum of the anti-apoptotic potential of heparin is currently under scrutiny in various tissues and under various pathological situations. In this study, the role of a low-molecular-weight heparin derivative (LMWH), certoparin in adriamycin-induced oxidative DNA damage has been evaluated in the cardiac and renal tissues. Two groups of male albino rats of the Wistar strain (140+/-10 g) received a single intravenous injection of adriamycin (7.5mg/kg), and one of them received low-molecular-weight heparin (Certoparin Sodium, 300 microg/day/rat s.c.) treatment, commencing on day 8, continued for a week. The nitrosative stress in ADR cytotoxicity is indicated by the 1.51-fold cardiac and 2.36-fold renal increase in reactive nitrogen species (RNS), while LMWH treatment restores normalcy (p<0.001). The influence of LMWH on the pro-inflammatory and pro-apoptotic cytokine, TNF-alpha was studied. Renal and cardiac levels of TNF-alpha showed a significant rise (p<0.001) in the ADR cytotoxic group, while the TNF-alpha values departed towards control levels in the LMWH treated group (p<0.001). DNA damage indicated by the fragmentation pattern (agarose gel electrophoresis) and the significantly increased comet tail length (p<0.001) observed after alkaline single cell gel electrophoresis confirmed the toxicity induced by ADR on DNA in the untreated group. In the LMWH-treated group, the observation of intact DNA band after agarose gel electrophoresis, and the finding of comet tail length being comparable with that of the control substantiated the protection rendered by the LMWH, certoparin. In short, the results suggest that the low-molecular-weight heparin derivative, certoparin exerts beneficial effects on the nitrosative status, and on the biological macromolecules as DNA and curtails the rise of the pro-apoptotic and pro-inflammatory cytokine, TNF-alpha in the cardiac and renal tissues.

Biochemical evaluation of the inflammatory changes in cardiac, hepatic and renal tissues of adriamycin-administered rats and the modulatory role of exogenous heparin-derivative treatment.

The aim of the present work is to evaluate the role of a heparin derivative, low molecular weight heparin (LMWH), certoparin on the inflammatory changes in adriamycin (ADR) cytotoxicity on a biochemical basis. Male Wistar rats (140+/-10g) were divided into four groups: untreated control, ADR group (a single dose intravenous injection of 7.5 mg/kg ADR), LMWH control (300 microg/(day rat) s.c. for 1 week) and ADR plus LMWH group (7.5 mg/kg ADR on day 1 of study period followed by LMWH treatment, 300 microg/(day rat) commencing on day 8 and continued for 1 week). At the end of the 2-week experimental period, biochemical assessment of the inflammatory status was carried out in the plasma, cardiac, hepatic and renal tissues. Increased concentrations of plasma C-reactive protein (CRP) and fibrinogen indicated severe inflammation in the ADR cytotoxic rats. These acute-phase inflammatory markers diminished significantly in the LMWH treated group, when compared with the cytotoxic group (p<0.001). Tissue damage was marked by elevated levels of plasma and tissue hexose, hexosamine, hexuronic acid and sialic acid, which were reversed on LMWH administration (p<0.001). The activities of lysosomal enzymes was measured in the experimental groups, and it was observed that the ADR induced rats showed a marked increase in the enzymic activities, while LMWH treated rats revealed normal activities. The present study throws light on the inflammatory changes in the ADR-challenged heart, liver and kidney tissues, and projects the biochemical basis for the anti-inflammatory property of the LMWH, certoparin.

Beneficial cardio-renovascular effects of a low-molecular-weight heparin-derivative on adriamycin-induced glycosaminoglycanuria and tissue lipid abnormalities.

The present work includes a study on the glycosaminoglycanuric condition induced by adriamycin (ADR, a chemotherapeutic agent) and the accompanying secondary hyperlipidemia, wherein the treatment with a low-molecular-weight heparin-derivative (LMWH), certoparin, is evaluated for its protective role (if any) on these parameters. Two groups of male albino rats of the Wistar strain (140+/-10 g) received a single intravenous injection of adriamycin (7.5 mg/kg), and one of these groups was treated with a low-molecular-weight heparin-derivative (Certoparin Sodium, Troparin; 300 microg/day/rat s.c.), commencing on day 8, for a week. Urinary total glycosaminoglycans excretion of the untreated ADR-induced group was found to increase on the 8th and the 15th days of observation, when compared with the controls. The LMWH treatment commencing on day 8 resulted in minimising the glycosaminoglycans (GAGs) excretion by day 15 (p<0.001). Plasma, cardiac, hepatic and renal lipids (cholesterol, triglycerides and phospholipids) showed a sharp increase in the pathologic group, along with a rise in plasma LDL and VLDL cholesterol and drop in HDL cholesterol levels, paralleled by abnormal activities of the enzymes involved in lipid metabolism. LMWH treated group showed a normalised lipid profile and the activities of the lipid-metabolising enzymes was close to that of controls. It is concluded herein that adriamycin administration resulted in severe nephropathy manifested by increased glycosaminoglycanuria and abnormal lipid metabolism, and that LMWH treatment afforded substantial protection by restoring glomerular structure and function, and normalised the plasma and tissue lipid levels, lipoprotein profile and the activities of lipid-metabolising enzymes.

Atheroprotective effect of exogenous heparin-derivative treatment on the aortic disturbances and lipoprotein oxidation in hypercholesterolemic diet fed rats.

BACKGROUND: The present work explores the myriad of biochemical and cellular changes that are featured in the early stages of atherosclerosis; if unchecked these changes lead to the complicated atherosclerotic plaque formation. The influence of a low-molecular-weight heparin derivative on the aortic aberrations and lipoprotein oxidation has been assessed in an experimental model of hypercholesterolemic atherogenesis. METHODS: Two groups of male Wistar rats (140+/-10 g) were fed a hypercholesterolemic atherogenic diet (rat chow supplemented with 4% cholesterol, 1% cholic acid and 0.5% thiouracil; CCT diet) for 2 weeks; one of these groups received LMWH (Certoparin) treatment of 300 microg/day/rat, s.c. for 7 days. An untreated control and a LMWH drug control group were also included. RESULTS: Abnormal increase in the aortic lipids -glycosaminoglycans levels (p<0.001) in CCT-diet fed group was circumvented by the exogenous glycosaminoglycan (LMWH) treatment (p<0.001). The escalation of oxidative stress (markers-lipid peroxidation and thiol levels, superoxide dismutase and catalase activities) in the atherogenic aorta was minimised by LMWH treatment. Further, an increased susceptibility of the apo B-containing lipoproteins (LDL+VLDL) to oxidation in vitro, induced by copper ions and Fenton's reagent, was observed in the untreated CCT diet fed group. This paper reports the favorable modulation of these oxidative changes by LMWH administration. Vascular protection by LMWH is further substantiated by the normal aortic histologic findings as against the appearance of foam cells in the untreated atherogenic group. CONCLUSION: The exogenous heparin-derivative (LMWH) treatment attempted in this experimental model of hypercholesterolemic atherogenesis affords substantial protection against abnormal levels of aortic lipids and glycosaminoglycans, aortic oxidative stress and also stunts the lipoprotein peroxidative process, thereby proving its multi-faceted anti-atherogenic effects.

Salubrious effect of low molecular weight heparin on atherogenic diet-induced cardiac, hepatic and renal lipid peroxidation and collapse of antioxidant defences.

The present work showcases the distressing picture of oxidative stress in the cardiac, hepatic and renal tissues, in an experimental model based on early phase atherogenesis. The protection rendered by LMWH intervention forms part of the same study. Male Wistar rats of 140 +/- 10 g were categorized as four groups. One group served as untreated control and another as LMWH drug control group. Two groups were fed a hypercholesterolemic atherogenic diet (rat chow supplemented with 4% cholesterol, 1% cholic acid and 0.5% thiouracil; CCT diet) for 2 weeks; one of these groups received LMWH treatment of 300 microg/day/rat for 7 days. The biochemical index of tissue lipid peroxidation (LPO) was assessed in terms of MDA formation. Heart, liver and kidney tissues of CCT-diet fed rats showed significantly elevated levels of LPO. In the early phase atherosclerotic group, we observed abnormal changes in the activities/levels of tissue enzymic (superoxide dismutase, catalase and glutathione peroxidase) and non-enzymic (reduced glutathione, ascorbate and alpha-tocopherol) antioxidants. We report normalized LPO levels and antioxidant defences in the atherogenic rats treated with LMWH. Thus the present study highlights the hepatic, cardiac and renal oxidative changes induced by experimental atherogenesis, and the protection rendered by LMWH treatment in atherosclerotic cardiovascular conditions.