Molecular Characterization of Lineage-IV Peste Des Petits Ruminants Virus and the Development of In-House Indirect Enzyme-Linked Immunosorbent Assay (IELISA) for its Rapid Detection".

Peste des petits ruminants (PPRV), a highly contagious viral disease, causes significant economic losses concerning sheep and goats. Recently, PPR viruses (PPRVs), have adopted new hosts and lineage IV of PPRVs represents genetic diversity within the same lineage. 350 samples, including blood, swabs, and tissues from sheep/goats, were collected during the 2020-2021 disease outbreaks in Pakistan. These samples were analysed through RT-PCR and three isolates of PPRV with accession numbers, MW600920, MW600921, and MW600922, were submitted to GenBank, based on the partial N-gene sequencing. This analysis provides a better understanding of genetic characterizations and a targeted RT-PCR approach for rapid PPRV diagnosis. An IELISA test was developed using the semi-purified antigen MW600922 isolate grown in Vero cells. The PPRV isolates currently present high divergence with the Turkish strain; conversely, similarities equivalent to 99.73% were observed for isolates collected from Pakistan. The developed indirect ELISA (IELISA) test demonstrated antibody detection rates at dilutions of 1:200 for antibodies (serum) and 1:32 for antigens. In comparison to cELISA, high specificity (85.23%) and sensitivity (90.60%) rates were observed. In contrast to the virus neutralization test (VNT), IELISA was observed to be 100% specific and 82.14% sensitive in its results. Based on these results, serological surveys conducted for PPR antibodies using IELISA can be a more effective strategy on a larger scale. Furthermore, our results demonstrate a significant breakthrough in the research in terms of cost-effectiveness and storage efficiency, and the developed IELISA test is highly recommended for use in developing countries.

Peste des petits ruminants (PPRV) is a transboundary, highly contagious, and economically significant viral disease affecting small ruminants and wildlife. PPRV, a disease that only targets animals, is the focus of the Global Eradication Programme (PPRV GEP), which aims to eradicate the disease by 2030. Following the completion of the first phase of the GEP (2017­2021), Pakistan has initiated the second phase: PPRV presence and the implementation of a control strategy. Rapid and accurate laboratory diagnosis is vital to the disease's effective control and eradication. In the present study, we have improved diagnosis by reverse transcriptase polymerase chain reaction (RT-PCR), which not only can detect low viral concentrations but also contributes to the genetic analysis of lineage-IV viruses. However, the development of cost-effective indirect ELISA (iELISA) may allow for the analysis of serum samples obtained from larger populations of small ruminants.

Determination of IL-6 Gene Promoter Polymorphism in Patients with Hepatitis C and Its Impact on RNA Secondary Structure.

Background and Objectives: A polymorphism in the promoter region of the IL-6 gene would influence the level of IL-6 expression in patients with HCV, resulting in a pro-inflammatory response. Few studies have shown the association between -174G>C (rs1800795) and -1363G>T (rs2069827) polymorphisms and HCV infection, and their results have been contradictory. There are no data published in our population to study such an IL-6 stimulus against HCV infection and its impact on RNA secondary structure. Therefore, we isolated human subjects from the province of Punjab, Pakistan. The objective was to screen for IL-6 gene promoter polymorphisms -174G/C and -1363G/T and those correlated with serum concentrations of IL-6 in patients with HCV and compared with a control. Materials and Methods: In conventional PCR, measurement of serum IL-6 by CLIA and statistical analysis were performed to observe the genotype association studies. By integrating bioinformatics and computational tools, our study aimed to provide a comprehensive understanding of how variations in the promoter region of IL-6 may have functional implications on gene expression. Results: The -174G>C and -1363G>T genotypes in the promoter region of patients with HCV were in strong allelic association (Δ = 0.97, p < 0.001). Interestingly, the bioinformatics analysis was well aligned with our experimental data. Conclusions: Based on the data, it can be inferred that IL-6 gene promoter polymorphisms are important in the dysregulation of IL-6 levels in patients with HCV.

Body mass index and altered lipid profile as major risk markers for breast cancer progression: a cross-sectional study of postmenopausal women in Pakistan.

BACKGROUND: In Pakistan, the death rate for post-menopausal women with breast cancer is significant due to late detection and delayed referral to proper facilities. There are a few reports on Pakistan's epidemiology and breast cancer risk factors. There are modifiable and non-modifiable risk factors associated with the development of breast carcinoma; of which body mass index (BMI), central obesity, and lipid profile are considered as major risk markers. METHODS: This was a cross-sectional analytical study. A total of 384 women constituted the present study sample. Purposive sampling was used to collect 192 confirmed new breast cancer cases throughout the study. By using basic random sampling, an equal number of controls were chosen. Studied parameters included age, fasting blood sugar, cholesterol, triglyceride, serum high-density lipoprotein, cholesterol, serum low-density lipoprotein cholesterol, weight, height, BMI, waist circumference, and waist-to-hip ratio. The inclusion criteria of this study were post-menopausal women (45-65 years) in Pakistan. The confirmation of breast carcinoma was done through histopathology. Breast cancer occurrence was taken as a dependent variable, whereas BMI, central obesity, and lipid profile were taken as independent variables. RESULTS: Studied risk factors (cholesterol, BMI, and central obesity) significantly correlated with breast cancer. Cholesterol has a significantly high positive correlation (0.646) with breast cancer. BMI has a positive significant correlation (0.491) with breast cancer, and central obesity has a low but positive significant correlation (0.266) with breast cancer. Moreover, the binary logistic regression model also showed a significant association between biochemical factors and breast cancer occurrence. Regression analysis depicted a linear relationship between a dependent variable (breast cancer occurrence) and independent variables (central obesity, cholesterol, BMI). CONCLUSION: Postmenopausal overweight (central obesity), increased BMI and high cholesterol levels are major risk factors for breast cancer. Moreover, high total cholesterol proved to be the most significant risk marker for the occurrence of breast cancer in post-menopausal women of Pakistan.

Exploring the association of ESR1 and ESR2 gene SNPs with polycystic ovary syndrome in human females: a comprehensive association study.

BACKGROUND: Polycystic Ovary Syndrome (PCOS) affects a significant proportion of human females worldwide and is characterized by hormonal, metabolic, and reproductive dysfunctions, including infertility, irregular menstrual cycles, acanthosis nigricans, and hirsutism. Mutations in the estrogen receptor genes ESR1 and ESR2, involved in normal follicular development and ovulation, can contribute to development of the PCOS. The present study focuses on investigating the potential correlation between single nucleotide polymorphisms (SNPs) of ESR1 and ESR2 genes and the incidence of this syndrome. METHODS: For this study, SNPs in ESR1 and ESR2 genes were retrieved from the ENSEMBL database and analyzed for their effect on mutated proteins using different bioinformatics tools including SIFT, PolyPhen, CADD, REVEL, MetaLR, I-Mutant, CELLO2GO, ProtParam, SOPMA, SWISS-MODEL and HDDOCK. RESULTS: All the SNPs documented in the present study were deleterious. All the SNPs except rs1583384537, rs1450198518, and rs78255744 decreased protein stability. Two variants rs1463893698 and rs766843910 in the ESR2 gene altered the localization of mutated proteins i.e. in addition to the nucleus, proteins were also found in mitochondria and extracellular, respectively. SNPs rs104893956 in ESR1 and rs140630557, rs140630557, rs1596423459, rs766843910, rs1596405923, rs762454979 and rs1384121511 in ESR2 gene significantly changed the secondary structure of proteins (2D). SNPs that markedly changed 3D configuration included rs1554259481, rs188957694 and rs755667747 in ESR1 gene and rs1463893698, rs140630557, rs1596423459, rs766843910, rs1596405923, rs762454979 and rs1384121511 in ESR2 gene. Variants rs1467954450 (ESR1) and rs140630557 (ESR2) were identified to reduce the binding tendency of ESRα and ß receptors with estradiol as reflected by the docking scores i.e. -164.97 and -173.23, respectively. CONCLUSION: Due to the significant impact on the encoded proteins, these variants might be proposed as biomarkers to predict the likelihood of developing PCOS in the future and for diagnostic purposes.

Mitigative potential of rhoifolin against cisplatin prompted testicular toxicity: biochemical, spermatogenic and histological based analysis.

Rhoifolin (ROF) is a naturally occurring flavonoid compound with diverse pharmacological and therapeutic benefits. The current investigation was designed to evaluate the curative potential of Rhoifolin (ROF) against Cisplatin (CP) induced testicular damage. Mature male albino rats (n = 48) were randomly distributed into 4 equal groups: control, CP (10 mg/kg), CP + ROF (10 mg/kg + 20 mg/kg) and ROF (20 mg/kg) supplemented group. Following 56 days of the trial, biochemical, inflammatory markers, spermatogenic, steroidogenic, hormonal, apoptotic, anti-apoptotic, and histopathological parameters were evaluated. The exposure to CP markedly (p < 0.05) lowered the activities of anti-oxidant enzymes, glutathione reductase (GSR), catalase (CAT), and glutathione peroxidase (GPx) as well as superoxide dismutase (SOD) in testicular tissues of male albino rats. Besides the levels of reactive oxygen species (ROS) and thiobarbituric acid reactive substances (TBARS) were considerably augmented in CP exposed rats. The administration of CP also increased the level of inflammatory cytokines i.e. IL-6, TNF-α, 1L-1ß and NF-κß as well as COX-2 activity. Additionally, a notable (p < 0.05) upsurge was observed in dead sperms count, abnormality in the tail, midpiece as well as head of sperms along with a notable decline in sperm motility in CP treated rats. Moreover, the expressions of steroidogenic enzymes were also lowered in CP administered group. The levels of follicle stimulating hormone (FSH) and plasma testosterone as well as luteinizing hormone (LH) were decreased in CP treated group. Moreover, the expression of Bax as well as Caspase-3 (apoptotic markers) were increased. On the other hand, Bcl-2 expression (anti-apoptotic marker) was reduced. Furthermore, the histopathological analysis showed that CP considerably (p < 0.05) damaged the testicular tissues. However, the administration of ROF significantly reduced the damaging effects of CP in testicular tissues. The results of our study suggested that ROF can potentially alleviate CP-induced testicular damages due to its androgenic, anti-oxidant and anti-inflammatory as well as anti-apoptotic nature.

Determination of biochemical and histopathological changes on testicular and epididymis tissues induced by exposure to insecticide Imidacloprid during postnatal development in rats.

BACKGROUND: Imidacloprid is a neonicotinoid insecticide belonging to the chloronicotinyl nitroguanidine chemical family. Toxicity of IMD for mammals in scientific studies has shown high mutagenic, immunotoxic, teratogenic and neurotoxic effects. The present study was designed to assess the toxic effects of imidacloprid (IMD) on the testicular and epididymis tissues as well as testosterone levels of neonatal male rats. METHODS: Neonatal male rats from postnatal day (PND) 1 to PND 26 were consecutively administered with different concentrations of IMD (1, 5 and 10 mg/kg) subcutaneously. The effect of IMD on body and organ weight, lipid profile, histopathological alterations, oxidative stress and altered testosterone levels were assessed in the testis and plasma. RESULTS: The results of body weight gain showed a significant difference in group 4 (10 mg/kg) animals as compared to the control. A significant increase in total cholesterol and triglycerides, while a decrease in high-density lipoprotein concentrations was evident. Similarly, a significant decrease in concentrations of antioxidant enzymes (CAT and SOD) among all the IMD-treated groups was evident, when compared to the control. Increased production of ROS was also noticed in the highest-dose treatment group. Further, we observed that IMD-treated rats indicated histopathological changes in the testis and epididymis along with a significant decrease in the plasma testosterone concentrations among IMI-treated groups in contrast to the control. Histological examination of the testis of IMD-treated neonatal male rats also showed decreased spermatogenesis in the treated groups when compared to the control. Furthermore, an increase in lumen diameter and a decrease in epithelial height of seminiferous tubules were also observed in IMD-treated rats in comparison with the control. CONCLUSION: It is concluded that sub-chronic exposure to IMD in neonatal male rats may induce histopathological changes in reproductive tissues and damage normal testicular functions via inducing oxidative stress, decrease in body weight, disturbing normal blood lipid profile and testosterone concentration. IMD exposure can induce pathophysiological effects calls for further evaluation of this widely used insecticide.

Alleviative effects of pinostrobin against cadmium-induced renal toxicity in rats by reducing oxidative stress, apoptosis, inflammation, and mitochondrial dysfunction.

Introduction: Cadmium (Cd) is a highly toxic heavy metal that can be found everywhere in the environment and can have harmful effects on both human and animal health. Pinostrobin (PSB) is a bioactive natural flavonoid isolated from Boesenbergia rotunda with several pharmacological properties, such as antiinflammatory, anticancer, antioxidant, and antiviral. This investigation was intended to assess the therapeutic potential of PSB against Cd-induced kidney damage in rats. Methods: In total, 48 Sprague Dawley rats were divided into four groups: a control, a Cd (5 mg/kg), a Cd + PSB group (5 mg/kg Cd and 10 mg/kg PSB), and a PSB group (10 mg/kg) that received supplementation for 30 days. Results: Exposure to Cd led to a decrease in the activities of catalase (CAT), glutathione reductase (GSR), superoxide dismutase (SOD), and glutathione peroxidase (GSH-PX), whereas levels of reactive oxygen species (ROS) and malondialdehyde (MDA) increased. Cd exposure also caused a substantial increase in urea, kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), and creatinine levels. Moreover, a noticeable decline was noticed in creatinine clearance. Moreover, Cd exposure considerably increased the levels of inflammatory indices, including interleukin-1b (IL-1b), tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), nuclear factor kappa-B (NF-kB), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) activity. Cd treatment decreased the expression of the antiapoptotic markers (Bcl-2) while increasing the expression of apoptotic markers (Bax and Caspase-3). Furthermore, Cd treatment substantially reduced the TCA cycle enzyme activity, such as alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, and isocitrate dehydrogenase. Moreover, mitochondrial electron transport chain enzymes, succinatedehydrogenase, NADH dehydrogenase, cytochrome c-oxidase, and coenzyme Q-cytochrome reductase activities were also decreased following Cd exposure. PSB administration substantially reduced the mitochondrial membrane potential while inducing significant histological damage. However, PSB treatment significantly reduced Cd-mediated renal damage in rats. Conclusion: Thus, the present investigation discovered that PSB has ameliorative potential against Cd-induced renal dysfunction in rats.

Molecular Screening of Bioactive Compounds of Garlic for Therapeutic Effects against COVID-19.

An outbreak of pneumonia occurred on December 2019 in Wuhan, China, which caused a serious public health emergency by spreading around the globe. Globally, natural products are being focused on more than synthetic ones. So, keeping that in view, the current study was conducted to discover potential antiviral compounds from Allium sativum. Twenty-five phytocompounds of this plant were selected from the literature and databases including 3-(Allylsulphinyl)-L-alanine, Allicin, Diallyl sulfide, Diallyl disulfide, Diallyl trisulfide, Glutathione, L-Cysteine, S-allyl-mercapto-glutathione, Quercetin, Myricetin, Thiocysteine, Gamma-glutamyl-Lcysteine, Gamma-glutamylallyl-cysteine, Fructan, Lauricacid, Linoleicacid, Allixin, Ajoene, Diazinon Kaempferol, Levamisole, Caffeicacid, Ethyl linoleate, Scutellarein, and S-allylcysteine methyl-ester. Virtual screening of these selected ligands was carried out against drug target 3CL protease by CB-dock. Pharmacokinetic and pharmacodynamic properties defined the final destiny of compounds as drug or non-drug molecules. The best five compounds screened were Allicin, Diallyl Sulfide, Diallyl Disulfide, Diallyl Trisulfide, Ajoene, and Levamisole, which showed themselves as hit compounds. Further refining by screening filters represented Levamisole as a lead compound. All the interaction visualization analysis studies were performed using the PyMol molecular visualization tool and LigPlot+. Conclusively, Levamisole was screened as a likely antiviral compound which might be a drug candidate to treat SARS-CoV-2 in the future. Nevertheless, further research needs to be carried out to study their potential medicinal use.

Mutation pattern of cancer suppressor p53 gene in factory workers exposed to polypropylene and impact on their reproductive hormones.

BACKGROUND: Polypropylene is a thermoplastic polymer playing the role of an endocrine disruptor that interferes with the union, emission, transport or elimination of normal hormones. Epidemiological information indicated the relation of endocrine-disturbing chemicals with prostate cancer, testis tumor and diminished fertility. p53 is a key tumor silencer gene. The present study aimed to evaluate luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels and the risk of p53 mutations as a result of exposure to polypropylene in non-tumorous adult male factory workers. METHODS: In total, 150 (controls = 35, workers = 115) subjects were recruited. Groups were maintained according to the tenure of exposure G1 (1-5 years), G2 (6-10 years), G3 (11-15 years) and G4 (16-20 years). Concentrations of LH and FSH were determined through an enzyme-linked immunosorbent assay. Genotyping analysis was performed by polymerase chain reaction based gel electrophoresis followed by DNA sequencing. The structural and functional impact of the mutation on the p53 structure was evaluated using 50-ns molecular dynamics (MD) simulations and protein-DNA docking. RESULTS: Mean plasma LH levels were significantly decreased in G1 (p > 0.05) as well as the G2, G3 and G4 (p > 0.001) groups. Similarly, FSH levels were significant decrease in G1 (p > 0.05), G2 (p > 0.01), G3 (p > 0.001) and G4 (p > 0.001) compared to the control group. Sequencing results found three variants i.e. g.13450 T>G, g.13430C>T and g.13737G>A. One of them was predicted to be disease-causing others are polymorphisms. MD simulation of missense mutation R273H showed no structural impact on the protein structure in MD simulation, but it resulted in weaker binding of p53 with the DNA that might lower the gene expression of cell cycle regulatory proteins. CONCLUSIONS: These findings predict decreased fertility and risk of malignancies in the future. The spectrum of p53 mutations as a result of polypropylene exposure in the Pakistani population has not been investigated before. Further studies and meta-analyses are required to elucidate the role of different plasticizers in reproduction and cancer-causing risk factors in a larger population.

Discovering the anti-diabetic potential of pomegranate peel metabolites by examining molecular interplay with the thioredoxin-interacting protein.

Introduction: Medicinal plants like Punica granatum (pomegranate) have traditional uses against diabetes, inflammation and other diseases. The study was initiated to get an insight into the interaction tendency of P. granatum derived compounds with diabetes associated human thioredoxin-interacting protein (TXNIP). High glucose in diabetes induces production of TXNIP resulting in ß-cells apoptosis. Its inhibition might reduce the diabetes incidence. Methods: To elucidate the therapeutic potential of P. granatum peel against diabetes through GC-MS based identification of extracted compounds followed by application of computational algorithms. P. granatum peel extracts were screened for antioxidant, anti-inflammatory, anti-diabetic, antimicrobial and wound healing properties. Phytochemical and GC-MS based analysis were performed to identify the bioactive compounds. Molecular docking analysis was performed by Auto Dock Vina to predict the binding tendency of P. granatum derived compounds with TXNIP. Results and Discussion: The peel exhibited antioxidant, anti-inflammatory and anti-diabetic activities, which were attributed to phytochemicals like phenols, tannins and steroids. GC-MS analysis identified 3,5-octadien-2-one, 1H-pyrrole -2,5-dione, Beta-D-lyxofuranoside, 5-O-(beta-D-lyxofuranosyl)-decyl, diethyl phthalate, 9-octadecenoic acid (Z)-, methyl ester, hexadecanoic acid, methyl ester, n-hexadecanoic acid, tetradecane, 2,6,10-trimethyl, bis (2-ethylhexyl) phthalate, decane, 3,8-dimethyl, 9-octadecenoic acid (Z)-, methyl ester and bis (2-ethylhexyl) phthalate in P. granatum peel extracts. Docking analysis revealed high binding affinities of bis (2-ethylhexyl) phthalate and 9-octadecenoic acid with TXNIP i.e., -4.5 and -5.0 kcal/mol, respectively, reflecting these compounds as potent antidiabetic agents. This study validates the traditional uses of P. granatum peel and demonstrates how computational approaches can uncover pharmacologically active phytochemicals. The results suggest P. granatum peel is a promising source of novel therapeutics against diabetes, inflammation, and oxidation. Further studies on the optimization of identified ligands are warranted.

Unraveling Catechol-O-Methyltransferase rs4680 SNP's Role in Patients' Response to Tramadol and Its Adverse Effects: A Pharmacogenetics Insight into Postoperative Pain Management.

Effective postoperative pain management is essential for patient well-being and an efficient healthcare system. Variations in the Catechol O-Methyltransferase (COMT) gene, specifically rs4680, play a crucial role in pain perception and opioid response. This study seeks to elucidate the impact of rs4680 polymorphism on tramadol efficacy and adverse reactions in post-surgical patients. We performed an uncontrolled cohort pharmacogenetics study in which participants underwent postoperative tramadol administration. The frequencies of rs4680 alleles were determined and the association between rs4680 genotypes and the efficacy of tramadol analgesic as pain relief, measured by the Numeric Rating Scale (NRS), was analyzed. Secondary outcomes included tramadol-induced sedation levels, opioid-induced nausea and vomiting, and other adverse effects of tramadol. Data analysis, using IBM SPSS Statistics 23, focused on pain and side effect differences across genotypes, with statistical significance set to p ≤ 0.05. The COMT (rs4680) genotype distribution exhibited a 'G' allele frequency of 41.5% and an 'A' allele frequency of 58.5%, with the AA genotype present in 44% of individuals, adhering to the Hardy-Weinberg equilibrium (p = 0.788). Patients with the AA genotype reported lower pain scores post-tramadol administration across all times examined (p < 0.001), but also experienced statistically significant (p < 0.001) higher incidences of tramadol-induced nausea, vomiting, and sedation. However, GG genotype individuals experienced poor pain relief from tramadol, requiring more supplemental analgesia. These significant findings underscore the critical role of COMT rs4680 polymorphism in response to tramadol and the necessity of a personalized approach to postoperative pain management.

Determination of possible contraceptive potential of methanolic leaf extract of Mentha longifolia L. in adult male rats: a biochemical and histological study.

Mentha longifolia L (ML) is locally used as an herbal contraceptive. We aim to assess the toxicity profile and validate the antifertility effects of its leaves in male rats. Adult male Sprague Dawley rats (n = 10 animals/group) were divided into four groups. The Control group received .9% saline, while groups II, III, and IV received 50, 75, and 100 mg/kg/day ML doses for 28 days. ML induced a dose-dependent decrease in percent fertility as well as the number of pups born in ML preexposed pairs, with maximum change observed in the ML3 group (60%, 3.75 ± 1.25) as compared to control (100%, 8.75 ± .48) group. A significant (P < .01) decrease in the rate of daily sperm production was observed in the ML3 group (.86 ± .16) in contrast to the control (2.93 ± .05). The levels of catalase and superoxide dismutase declined significantly in treatment groups (ML1-P < .001, ML2-P < .05, and ML3-P < .01). A significant increase in the production of reactive oxygen species (P < .001) was found in all treatment groups. The concentrations of LH (P < .001), FSH (P < .001), and testosterone (P < .001) were significantly reduced in a dose-dependent manner among ML-treated groups. Decreases in spermatogonial populations, mature spermatids, seminiferous tubule diameter, lumen diameter, and epithelial height were noticed at higher doses. The reversibility study depicted a reduction in both pregnancy outcomes and litter size. Besides contraceptive effects, M. longifolia L usage is associated with oxidative stress; therefore, it is important to consume this herb appropriately and its excessive usage should be prohibited.

Isorhamnetin: a flavonoid, attenuated doxorubicin-induced testicular injury via regulation of steroidogenic enzymes and apoptotic signaling gene expression in male rats.

Background: Male reproductive damage is one of the most adverse side effects of doxorubicin (DOX). Isorhamnetin is a natural flavonoid, which displays remarkable antioxidant potential. Objective: The current research was designed to assess the protective effects of Isorhamnetin against DOX-instigated testicular damages. Methods: Adult male Wistar rats (n=32) were divided into 4 groups: control, DOX (3 mg/kg i.p. 3 doses each after 1 week), DOX + Isorhamnetin (3 mg/kg 3 doses each after 1 week +10 mg/kg i.p. daily for 28 days, respectively), and Isorhamnetin (10 mg/kg i.p. per day). After 28 days of treatment, biochemical, spermatogenic, steroidogenic, hormonal, proapoptotic, antiapoptotic, and histopathological parameters were estimated. Results: DOX exposure significantly decreased the activity of acid phosphatase, lactate dehydrogenase, and gamma-glutamyl transferase. Furthermore, DOX substantially decreased the activities of antioxidant enzymes, i.e. catalase, superoxide dismutase, glutathione reductase, and glutathione peroxidase along with protein content, whereas it increased the malondialdehyde level. It also reduced sperm progressive motility, viability, the number of hypoosmotic tail swelled spermatozoa, and epididymis sperm count and increased the sperm morphological anomalies (head, midpiece, and tail). Besides, it decreased the levels of follicle-stimulating hormone, luteinizing hormone, and plasma testosterone and lowered the expression of steroidogenic enzymes (3ß-hydroxysteroid dehydrogenase, 17ß-hydroxysteroid dehydrogenase, and steroidogenic acute regulatory protein) and testicular antiapoptotic marker (B-cell lymphoma 2) but increased the expression of proapoptotic markers (BCL2-associated X protein and caspase-3) along with histopathological impairments. However, isorhamnetin prevented all the damages caused by DOX. Conclusion: Conclusively, Isorhamnetin can be used as a powerful mitigating agent to avert DOX-induced testicular damages.

Determination of Pleiotropic Effect of Warfarin in VKORC1 and CYP2C9 Genotypes in Patients With Heart Valve Replacement.

Warfarin has been widely used as an oral anticoagulant agent. In past, efforts have been done to study the contribution of genetic variation on warfarin dose requirements. The possible therapeutic dose determination of warfarin is very challenging, i.e., extremely low dose leading to unusable antithrombotic therapy or high dose causes particularly bleeding complications. Our study aimed to investigate these observations in more detail, we determined the correlation of interleukin-6 (IL-6), cyclooxygenase-2 (COX-2), and tumor necrosis factor-α (TNF-α) among VKORC1 and CYP2C9 genetic variants in patients with heart valve replacement who were treated with a range of warfarin doses and compared with levels in healthy controls. A total of 107 human subjects were recruited with low < 5 mg, medium 5-10 mg/day, and high > 10 mg/day warfarin doses. The genetic study of VKORC1-1639G/A, C1173T, 3730G > A, CYP2C9*2, and CYP2C9*3 was performed using TaqMan genotyping and DNA sequencing. The gene expression of IL-6, TNF-α, and COX-2 mRNA was analyzed. IL-6, TNF-α, and COX-2 protein expressions were determined by ELISA and Western blot analysis to evaluate the pro- and anti-inflammatory effects of warfarin. A statistically significant difference was found among the haplotypes of VKORC1 rs9934438 (C1173T), rs9923231 (-1639G > A), rs7294 (3730G > A) and CYP2C9 *2 p. Arg144 Cys (rs28371674), CYP2C9 *3 p. Ile359Leu (rs1057910) genotypes with warfarin dose requirements (p = 0.001). The increased levels of COX-2, IL-6, and TNF-α proteins were observed when a high dose of warfarin (>10 mg/ml) was administered. However, a lower concentration (1.0 mg/ml) was observed with decreased warfarin dose (<5 mg/day). The present study reported that in addition to its anticoagulant action, the genetic variants of warfarin may have a pleiotropic effect by influencing IL-6 depending on the dosing regimen and inducing the expression of COX-2.