Role of cytochrome epoxygenase (CYP2J2) in the pathophysiology of coronary artery disease in South Indian population

Background The cytochrome P-450 2J2 (CYP2J2) is known to be one of the major enzymes of epoxygenase pathway of arachidonic acid in extrahepatic tissues, which produces series of regioisomeric cis-epoxyeicosatrienoic acids (EETs) such as 5,6-, 8,9-, 11,12-, and 14,15-EETs. In the present study, we analyzed the impact of a genetic variant in CYP2J2 on coronary artery disease (CAD) in the Telangana region of Indian population. Material and methods The case–control study consisted of 100 CAD cases and 110 healthy controls. The deoxyribonucleic acid was extracted using the salting out method. Genotyping and gene expression was performed by polymerase chain reaction (PCR)-restriction fragment length polymorphism and real-time-PCR methods. Results In the present study, the percentage of smokers, alcoholics, hypertensive patients, and diabetics was high. Increase in fasting glucose, urea, creatinine, fasting triglycerides, total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C), total cholesterol/high-density lipoprotein (TC/HDL), LDL/HDL, homocysteine, and C-reactive protein levels were significantly higher in patients with CAD than in controls (p < 0.001). CYP2J2 G-50T was associated with CAD (p = 0.04). The mRNA expression of CYP2J2 showed altered gene expression in this study among CAD patients in comparison with control (p = 0.01). Conclusions A functionally relevant polymorphism of the CYP2J2 gene was independently associated with an increased risk of CAD.

Association of CYP3A4*1B and CYP3A5*3 Genetic Polymorphisms with Lung Cancer and its Impact on Taxane Metabolism in Indian Population.

Background: The purpose of this study was to investigate the genotype frequencies of CYP3A4*1B and CYP3A5*3 in lung cancer patients which may be useful in determining the patients’ predisposition to platinum based therapies, and may be helpful for individualized drug dosing and improved therapeutics and disease management. Results: We evaluated these two common polymorphisms in south Indian population, based on case-control study of 126 lung cancer cases and 111 controls using a PCR-RFLP-based assay. The investigation of the CYP3A4*1B gene polymorphism revealed, no significant difference in distribution between the lung cancer patients and the controls (p=0.65) . The distribution of CYP3A5*3 homozygous genotypes and heterozygous plus homozygous genotypes were significantly associated (p=0.0004 & p=0.0001) with lung cancer patients, and the *3/*3 genotype had a 4.38 fold increased risk for lung cancer. In our study *1A/*1B heterozygous genotype patients were found to constitute a major percentage of patients receiving Gemcitabine plus carboplatin therapy. Conclusions: In conclusion, the results of our study indicated a relationship between CYP 3A4*1B and CYP3A5*3 polymorphisms and genetic predispositions to lung cancer. Thus detection of CYP3A4*1B/CYP3A4 and CYP3A5*3/CYP3A5 genotype frequencies in Indian population may be important in view of interindividualized drug dosing, improved therapeutics and disease management.

ANTIMICROBIAL DRUG RESISTANCE IN STRAINS OF Escherichia coli ISOLATED FROM FOOD SOURCES / Resistência microbiana a drogas em linhagens de Escherichia coli isoladas de fontes alimentares

A variety of foods and environmental sources harbor bacteria that are resistant to one or more antimicrobial drugs used in medicine and agriculture. Antibiotic resistance in Escherichia coli is of particular concern because it is the most common Gram-negative pathogen in humans. Hence this study was conducted to determine the antibiotic sensitivity pattern of E. coli isolated from different types of food items collected randomly from twelve localities of Hyderabad, India. A total of 150 samples comprising; vegetable salad, raw egg-surface, raw chicken, unpasteurized milk, and raw meat were processed microbiologically to isolate E. coli and to study their antibiotic susceptibility pattern by the Kirby-Bauer method. The highest percentages of drug resistance in isolates of E. coli were detected from raw chicken (23.3%) followed by vegetable salad (20%), raw meat (13.3%), raw egg-surface (10%) and unpasteurized milk (6.7%). The overall incidence of drug resistant E. coli was 14.7%. A total of six (4%) Extended Spectrum β-Lactamase (ESBL) producers were detected, two each from vegetable salads and raw chicken, and one each from raw egg-surface and raw meat. Multidrug resistant strains of E. coli are a matter of concern as resistance genes are easily transferable to other strains. Pathogen cycling through food is very common and might pose a potential health risk to the consumer. Therefore, in order to avoid this, good hygienic practices are necessary in the abattoirs to prevent contamination of cattle and poultry products with intestinal content as well as forbidding the use of untreated sewage in irrigating vegetables.

Variedade de alimentos e fontes ambientais contem bactérias resistentes a uma ou mais drogas antimicrobianas usadas em medicina e agricultura. Resistência antibiótica pela Escherichia coli é particularmente preocupante porque ela é o patógeno mais comum Gram negativo em humanos. Portanto este estudo foi conduzido para determinar o aspecto de sensibilidade antibiótica da E. coli isolados de diferentes tipos de alimentos obtidos ao acaso de 12 localidades de Hyderabad, India. Um total de 150 amostras compreendendo saladas, vegetais, superfícies de ovos crus, galinhas cruas, leite não pasteurizado e carne crua foram processados microbiologicamente para isolar E. coli e estudar o quadro de sensibilidade antibiótica pelo método de Kirby-Bauer. A maior percentagem de resistência à droga foi isolada de E. coli obtidos de galinha crua (23,3%) seguido de saladas e vegetais (20%), carne crua (13,3%), superfície do ovo cru (10%) e leite não pasteurizado (6,7%). Incidência total de E. coli resistente foi de 14,7%. Um total de seis (4%) Extended Spectrum β-Lactamase (ESBL) produtores foram detectados, dois cada de salada de vegetais e galinha crua e um cada de superfície de ovo cru e carne crua. Espécies resistentes a múltiplas drogas de E. coli são matéria de preocupação uma vez que os genes de resistência podem facilmente ser transferidos para outras linhagens. O ciclo do patógeno é muito comum nos alimentos e pode ser risco potencial para a saúde do consumidor. Portanto, para evitar isto boas práticas de higiene são necessárias nos abatedouros para prevenir a contaminação de gado e aves com conteúdo intestinal assim como proibir o uso de águas de esgoto não tratadas para irrigar vegetais.

Pluripotent lineage of CD133 stem cells isolated from human skin samples.

Skin stem cells are very important in cosmetics, pharmacological and regenerative medicine and burn cases. Foreskin samples surgically removed after circumcision from boys below 7 years were collected and primary epidermal cells were prepared by enzymatic and mechanical tituration method. Selecting CD133 (prominin-1) multipotent stem cell marker, enriched stem cells were analyzed by MACS using CD133 antibodies conjugated with magnetic beads. CD133 positive and negative cells with specific skin stem cells markers like - CD34 (Universal stem cells marker), CD29 (integrin beta-1) and CD49f (integrin alpha-6) immunophenotypes were screened and sorted in flowcytometer. Further the expression of four embryonic genes or transcription factors of pluripotent stem cells were analyzed for pluripotent character of sorted cells. It was found that skin stem cell markers associated with CD133 cells, differentially expressed CD34, CD29 and CD49f immunophenotyes on both positive and negative CD133 cells in FACS analysis. The embryonic stem cell markers (induced pluripotent stem cell markers) like Oct4, SOX2, Notch-2 and K19 genes were expressed in CD133 positive epidermal cells. It is therefore evident that foreskin derived epidermal stem cells showed pluripotent or multipotent nature. This finding opens up avenues for new uses of these stem cells for direct cell seeding in wound healing, surgical suturing and drug screening.

Mutation in mexR-gene leading to drug resistance in corneal keratitis in human.

The present study deals with the genetic polymorphism of the mexR gene which is involved in the resistance to drugs like ciprofloxacin. Mutations in mexR result in increased resistance to multiple antibiotics due to overexpression of this efflux system. The MexR product contains 147 amino acids with a molecular mass of 16,964 Da. We detected 28 point mutations in 14 samples from corneal scraping infected with Pseudomonas aeruginosa, which were screened for ciprofloxacin resistance. Twenty four were silent mutations and four missense mutations. Mapping these mutations was done by using in silico methods on the protein 3D- structure obtained from PDB database, localized at 3 specific sites. Single amino acid changes (mutations) may influence MexR stability or its ability to dimerise, and thus result in the conformation changes at the DNA-binding domain, of the structure. Hence it is concluded that the mutations in the DNA binding domain of mexR gene could be one of the factors contributing to the possible drug resistance in these patients.

In vitro cytogenetic studies of cypermethrin on human lymphocytes.

Assessment of cytotoxicity and response to external factors like pesticides were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide (MTT) or MTT assay, which measures mitochondrial metabolism in the entire cell culture and provides information about the percentage of cell survival. Utilizing the MTT assay, the cytotoxicity of cypermethrin was determined on lymphocyte cultures from human peripheral blood samples, the short-term lymphocyte cultures were incubated with various aliquots of the cypermethrin and the LC50 was found to be 33.6 microM. Lymphocytes treated with low-doses (1/10 of LC50) of cypermethrin showed an increase in the frequency of chromosomal aberrations and found to be significant. Karyotype analysis revealed more satellite associations and chromosomal breaks in cypermethrin treated samples. Low-doses of the pesticide also induced single-strand breaks in the DNA as assessed by comet assay. The pesticide caused increase in the comet tail length with increase in pesticide concentration, implicating genotoxicity in somatic cells. It is concluded that In vitro assays could give important information of the mechanism of toxicity at low dosages and impact on genetic material of human origin.

Acute and sub-acute effects of 2-butenoic acid-3-(diethoxy phosphinothioyl) methyl ester (RPR-II) on testis of albino rat.

Acute and sub-acute toxic effects of a novel phosphorothionate coded as RPR-II on testis of albino rats were studied. In acute study rats received a single dose of 12.3 mg/kg of RPR-II and sacrificed after 24 hr. For sub-acute study 0.58 mg/kg/day was administered orally to rats for 10 and 21 days. Acute exposure of rats to RPR-II brought no change either in the gonadosomatic index (GSI) or in the structure of testis or in the serum levels of testosterone. Testis glutathione (GSH) level and glutathione S-transferase (GST) activity was significantly decreased whereas, acid phosphatase (AcP) levels increased significantly at 24 hr post-treatment. On 7th day (withdrawal period) after the cessation of the treatment the GSH, GST, AcP, and AkP levels reached to near control. The sub-acute study revealed a significant decrease in GSI on 10th and 21st day of the treatment. In contrast, a time-dependent and significant increased in GSH level and GST activity was observed on 100th and 21st day of post-treatment, except GSH level on 10th day, which was declined. Due to RPR-II treatment the testis AcP and alkaline phosphatase (AkP) levels were significant at both 10th and 21st day of medication but AcP levels were increased whereas AkP levels decreased. The histopathological studies on day 10th showed considerable loss of spermatozoids in testis and at 21st day complete derangement of cellular organization was observed. Testosterone levels decreased significantly on 10th day and remained significantly low at 21st day. However, withdrawal studies showed a recovery in testis of rat treated with RPR-II. GST, GSH, GSI, AcP and AkP values recovered, testosterone levels were also well recovered but recovery in testis structure remained at a low profile. The present study suggests that RPR-II may cause testicular toxicity in rats affecting the normal functioning of testis and it also gave some new information in withdrawal studies.