Association of vascular endothelial growth factor a gene (VEGFA) polymorphisms, rs699947 and rs1570360, with diabetic retinopathy and altered VEGF secretion in the Pakistani patients with type 2 diabetes mellitus: a casecontrol study.

OBJECTIVE: To investigate the association of vascular endothelial growth factor A gene polymorphisms 2578C/A (rs699947) and 1154G/A (rs1570360) with type 2 diabetes mellitus, diabetic retinopathy and serum vascular endothelial growth factor levels in Pakistani patients. METHODS: The case-control study was conducted from Jan 2017 to Dec 2018 after approval from the ethics review board of Riphah International University, Islamabad, Pakistan, and comprised type 2 diabetes mellitus patients of either gender with diabetic retinopathy in group A, and without diabetic retinopathy in group B. Non-diabetic healthy individuals were enrolled in control group C. Genotyping was done by amplification refractory mutation system-polymerase chain reaction and serum vascular endothelial growth factor levels were measured using enzyme-linked immunosorbent assay. Data was analysed using SPSS 22. RESULTS: Of the 450 subjects, 150(33.3%) were in each of the 3 groups. The mean age in group A was 58.16±9.42, in group B 56.25±8.5 years and in group C it was 55.90±10.90. The proportion of Punjabi ethnicity was significantly high in group B compared to other groups (p<0.05). There was no significant association of rs699947 and rs1570360 genotypic and allelic frequencies in group B compared to group A. Further, rs699947 AA genotype was significantly associated with proliferative diabetic retinopathy compared to group A (p<0.05). Minor allele A showed significant association in groups A and B compared to group C (p<0.05). Significantly raised serum vascular endothelial growth factor levels were found in group B compared to group A (p<0.05), and were associated with rs699947 and rs1570360 heterozygosity in group A (p<0.05). Also, rs699947 genotype showed significant association with groups A and B in Punjabi and Pathan ethnicities (p<0.05) and with Kashmiri ethnicity in group B (p<0.05). CONCLUSIONS: There was a strong association of vascular endothelial growth factor 2578C/A (rs699947) gene polymorphism with proliferative diabetic retinopathy in type 2 diabetic Pakistani patients, suggesting its role in the pathogenesis of this condition.

Association of Pro-Inflammatory Cytokines with Vitamin D in Hashimoto's Thyroid Autoimmune Disease.

Background and objectives: Hashimoto's thyroiditis is an important autoimmune thyroid condition. It is characterized by lymphocytic congestion of the thyroid gland followed by progressive deterioration and fibrous substitution of the thyroid in the parenchymal structure. This study has provided insight into the variations of blood pro-inflammatory cytokine levels in patients with Hashimoto's disease and the key role of vitamin D levels among selected patients. Materials and Methods: A total of 144 participants including healthy controls and patients were studied in the current study in which 118 were female and 26 were male. The thyroid profile was evaluated in patients with Hashimoto's thyroiditis and healthy controls. Results: The mean ± SD Free T4 in the patients was recorded as 14.0 ± 4.9 pg/mL, and TSH was 7.6 ± 2.5 IU/L, whereas the median ± IQR thyroglobulin antibodies (anti-TG) were 285 ± 142. Thyroid peroxidase antibodies (anti-TPO) were 160 ± 63.5, whereas in the healthy controls, the mean ± SD Free T4 was recorded as 17.2 ± 2.1 pg/mL, and TSH was 2.1 ± 1.4 IU/L, whereas the median ± IQR anti-TGs were 56.30 ± 46.06, and anti-TPO was 5.6 ± 5.12. The assessment of pro-inflammatory cytokines (pg/mL) and total Vitamin D levels (nmol/L) in patients with Hashimoto's thyroiditis was recorded with values IL-1B 6.2 ± 0.8, IL-6 9.4 ± 0.4, IL-8 7.5 ± 0.5, IL-10 4.3 ± 0.1, IL-12 3.8 ± 0.5, TNF-α 7.6 ± 1.1, and total vitamin D 21.89 ± 3.5, whereas in healthy controls the mean ± SD IL-1B was 0.6 ± 0.1, IL-6 2.6 ± 0.5, IL-8 3.0 ± 1.2, IL-10 3.3 ± 1.3, IL-12 3.4 ± 0.4, TNF-α 1.4 ± 0.3 and total vitamin D was 42.26 ± 5.5. Conclusions: It was found that individuals with Hashimoto's thyroiditis had raised serum levels of IL-1B, IL-6, IL-8, IL-10, IL-12, and TNF-α as compared to the healthy controls, whereas the total vitamin D levels were remarkably low as compared to health controls. Serum TSH, anti-TG, and anti-TPO levels were typically lower in controls and much higher in individuals with Hashimoto's thyroiditis. The current study's findings might aid in future studies and in the diagnosis and management of autoimmune thyroid disease.

Differential expression of cancer associated plasma miRNAs in the ordnance factory workers, exposed to 2,4,6-Trinitrotoluene.

Objectives: To determine differential expression of microRNAs (miRNAs) in plasma of 2, 4, 6-Trinitrotoluene (TNT) exposed ordnance factory workers. Methods: A case control study was conducted at the Department of Toxicology, Armed Forces Institute of Pathology, Rawalpindi from July to December 2020. A total 30 subjects were recruited from an ordnance factory that were directly exposed to TNT and 120 non-exposed individuals from non-factory healthy population. Plasma levels of five miRNAs including miRNA-let-7a-2, miRNA-34a-1, miRNA-21-2, miRNA-106b-1, miRNA-122a-1 were measured by quantitative real-time polymerase chain reaction (RT-PCR). Results: Micro RNAs showed a wide range of Ct (cycle threshold) values ranging from 23.48 to 41.94. Among the five miRNAs let-7a-2 and miRNA-122a-1 displayed relatively high expression with Ct values ranging from 26.58 ± 2.25 to 27.18 ± 0.80 respectively. Relative fold change expression for all five miRNAs of exposed individuals were found high (p <0.0001) vs non-exposed. Dividing fold change expression of exposed individuals into two groups as ≤ 10 and > 10, the individuals having ≤ 10-fold change expression were19 (63.3%) in miRNA-let-7a-2, 30 (100%) in miRNA-34a-1 and 23 (76.7%) in miRNA-122a-1 while in miRNA-21-2 and miR-106b-1, 23 (76.7%) and 18(60%) individuals had > 10-fold change expression respectively. Among the five miRNAs in exposed individuals, miRNA-let-7a-2, miR-21-2, miR-106b-1 and miR-122a-1 were found highly expressed with fold change expression > 10 (p <0.0001). No significant association was found between miRNAs expression levels with age and working duration. Conclusion: The study shows upregulation of all five miRNAs in TNT exposed subjects with no significant association of expression levels with age and working duration.

Association analysis of -429T/C receptor for advanced glycation end products (RAGE) gene polymorphism with type 2 diabetic retinopathy and serum soluble RAGE levels in Pakistani patients.

OBJECTIVE: To investigate the association of receptor for advanced glycation end products gene polymorphism 429T/C (rs1800625) with diabetic retinopathy and serum soluble receptor for advanced glycation end products levels in patients with type 2 diabetes. METHODS: The case-control study was conducted from January 2017 to December 2018 at Pakistan Railway Hospital, Rawalpindi, and the Multidisciplinary Laboratories of Islamic International Medical College, Riphah International University (RIU), Islamabad, Pakistan. Those included were healthy controls in group A, diabetics without retinopathy in group B and patients having diabetic retinopathy in group C. Genotyping for 429T/C was done by tetra-primer amplification refractory mutation system-polymerase chain reaction. Serum soluble receptor for advanced glycation end products levels were measured using enzyme-linked immunosorbent assay. Data was analysed using SPSS 22. RESULTS: Of the 450 subjects, 150(33.3%) were in each of the three groups. The frequency of TT, TC and CC genotypes of 429T/C polymorphism were 137(91.3%), 10(6.7%) and 3(2%) in group A; 133(88.6%), 13(8.7%) and 4(2.7%) in group B; and 127(84.7%), 18(12%) and 5(3.3%) in group C. No significant association of 429T/C genotypic and allelic frequencies were found with groups B and C (p>0.05). Serum soluble receptor for advanced glycation end products levels were significantly high in patients with proliferative diabetic retinopathy and were positively correlated with fasting plasma glucose in group C (p<0.05). TC and CC genotypes were significantly associated with raised serum soluble receptor for advanced glycation end products, and TC with raised fasting plasma glucose in group C. CONCLUSIONS: The 429T/C receptor for advanced glycation end products gene polymorphism was found to be associated with severe non-proliferative diabetic retinopathy, and serum soluble receptor for advanced glycation end products levels had a positive correlation with severity of diabetic retinopathy.

Association analysis of 374T/A (rs1800624) receptor for advanced glycation end-products (RAGE) gene polymorphism with diabetic retinopathy in Pakistani patients.

OBJECTIVES: to determine the relationship of 374T/A (rs1800624) polymorphism in the gene encoding RAGE with Type-2 diabetes mellitus (T2DM), diabetic retinopathy (DR) and serum soluble RAGE (sRAGE) level in Pakistani patients. METHODS: A case-control study, conducted from January 2017 to December 2018, involving 150 healthy controls (HC), 150 T2DM patients with no retinopathy (DNR) and 150 DR patients diagnosed by coloured fundus photography. Tetra-primer amplification refractory mutation system - polymerase chain reaction (T-ARMS-PCR) was used for genotyping. Serum sRAGE levels were measured by enzyme-linked immunosorbent assays (ELIZA). RESULTS: The frequency of TT, TA and AA genotypes of rs1800624 polymorphism were: 92.7%, 6%, 1.3% in HC, 80%, 17.3%, 2.7% in DNR and 76.7%, 19.3%, 4.3% in DR groups. Heterozygous TA genotype and mutant A allele showed significant association with diabetes and DR vs HC. In dominant model, mutant allele showed significant association with DNR and DR vs HC. No significant association of rs1800624 was detected with DR and its sub-groups, non-proliferative DR (NPDR) and proliferative DR (PDR) vs DNR. Dividing NPDR into mild, moderate and severe, heterozygous TA genotype showed significant association with moderate and severe NPDR vs DNR. In DNR and DR groups, TA genotype was significantly associated with raised sRAGE. CONCLUSION: rs1800624 RAGE gene polymorphism might be a risk factor for T2DM and NPDR in Pakistani patients. Raised sRAGE levels have a positive correlation with PDR and are associated with heterozygosity of rs1800624 polymorphism in DNR and DR groups.

Association of miR-146α gene polymorphism and expression levels of its target protein RhoA in Pakistani breast cancer patients.

OBJECTIVE: To assess the association of miR-146a and its target protein RhoA expression levels in breast cancer. METHODS: The case-control study was conducted at Riphah International University, Islamabad, Pakistan, from March 2017 to November 2018, and comprised confirmed breast cancer cases and controls who were matched for age and ethnicity. Genotyping and expression profiling of archived samples was performed. Data was analysed using SPSS 22. RESULTS: Of the 590 subjects, 295(50%) each were cases and controls. Among the cases, there were 195(66%) Punjabis, 59(20%) Pathans and 41(14%) Kashmiris. The corresponding numbers among the controls were 198(67%), 58(19.7%) and 39(13.2%). The association between genotypes of the cases and controls was significant (p<0.05). Strong association was seen in dominant, recessive and allelic models (p=0.05). In Punjabi group the association was (p<0.00) significant, but this association was not significant in Kashmiri and Pathan groups (p>0.05). No association was found with the receptor status and miR-146a polymorphism. CONCLUSIONS: The miR-146a gene polymorphism rs2910164 G/C was found to have increased susceptibility to breast cancer at genotype and allelic levels.

Association of single nucleotide polymorphism of transforming growth factor ß1 (T29C) in breast cancer patients: a case control study in Rawalpindi.

OBJECTIVE: To determine the association of single nucleotide polymorphism in three CC, TT and TC genotypes of transforming growth factor ß1 T29C in breast cancer patients. METHODS: The case-control study was conducted from April 2017 to April 2018 at the Islamic International Medical College, Rawalpindi, Pakistan, in collaboration with Nuclear Oncology Medicine and Radiotherapy Institute and Holy Family Hospital, Rawalpindi. Using convenience sampling, breast cancer cases and healthy controls were enrolled. All investigations were done using standardized laboratory protocols. The outcomes were determined in terms of association of single nucleotide polymorphism of transforming growth factor ß1with breast cancer. Data was analysed using SPSS 21. RESULTS: Of the 150 subjects, 80(53.3%) were cases and 70(47.7%) were healthy controls. Among the cases, the most frequent genotype was CC 38(47.5%) followed by TC 26(32.5%) and TT 16(20%). Among the controls, the corrsesponding values were 50(71.42%), 13(18.5%) and 7(10%). Transforming growth factor ß1 TC genotype was strongly associated with the increased risk of developing breast cancer (odds ratio: 3.79). CONCLUSIONS: The incidence of breast cancer was markedly lower among women with CC genotype compared to those with CT or TT genotypes.

Genetic polymorphism of miRNA-196a and its target gene annexin-A1 expression based on ethnicity in Pakistani female breast cancer patients.

OBJECTIVE: To evaluate the association of miR-196a rs11614913 C/T genetic variation and its target gene annexin A1 mRNA expression with breast cancer risk in Pakistani female ethnicities. METHODS: This case control study, conducted from March 2017 to November 2018 included 295 breast cancer patients, 295 controls of three Pakistani ethnicities and archived 100 samples of cohort group for genotyping and expression profiling. Genotyping of miR-196a (rs11614913 C/T) was done by ARMS PCR technique. Annexin-A1 (ANXA1) mRNA expression was measured with qRT-PCR and detection of protein expression of ANXA1 was done by immunohistochemistry. RESULTS: CC homozygous genotype of miR-196a rs11614913 was present in 81.4% of cases and 73.9% controls. C/T polymorphism was found to be significantly associated with decrease risk of breast cancer (OR=0.25 (0.11- 0.58, p <0.05). Similar trend was seen with the minor T allele (OR=0.55 (0.39-0.77, p <0.05, and both dominant and recessive models (OR=0.64; p=0.02 and OR=0.26, p=0.00). In the KPK ethnic group significant decrease association with breast cancer risk was observed (OR= 0.22 (0.09-0.53, p < 0.05). Immunohistochemical staining showed loss of ANXA1 protein expression in 72 samples, and significant association was observed with pathological type p=0. 00 and triple negative receptor status p=0.03 and with genotypes of miR-196a p=0.00. Increase relative expression of 2.81± .88 by qPCR analysis of ANXA1 mRNA was noted with TT genotype. CONCLUSIONS: Our results demonstrate that miR-196a rs11614913 C/T polymorphism is associated with a decreased risk and loss of protein expression in breast cancer in the Pakistani population.

Relationship of S/R warfarin ratio with CYP2C9 genotypes in Pakistani population.

Warfarin is administered as a racemic preparation of R- and S-enantiomers. S-warfarin is more potent than R-warfarin, so changes in blood levels of S-warfarin affect the anticoagulant response. This study was carried out to determine the effect of CYP2C9*2 and CYP2C9*3 polymorphisms on S/R warfarin ratio. A single blood sample was collected 12-16 hours after drug administration from 170 stable patients fulfilling the criteria. Genotyping of the CYP2C9 polymorphisms was done by polymerase chain reaction-restriction fragment length polymorphism assay. S- and R-warfarin enantiomers extraction from plasma was accomplished by a validated HPLC method. The concentration of S-warfarin was significantly different among CYP2C9 genotypes (p =0.018) whereas there was no effect on R-warfarin (p =0.134). There was statistically significant effect of different CYP2C9 genotypes on S/R warfarin ratio (p=0.000). It is concluded that CYP2C9 polymorphisms influence CYP2C9 enzymatic activity in turn affecting S-warfari levels but not R-warfarin, thus leading to different S/R warfarin enantiomers ratio among different CYP2C9 genotypes.

Serum Albumin and C3 Complement Levels in Endometriosis.

OBJECTIVE: To compare the efficacy of albumin and C3 complement as serum marker for the early diagnosis of endometriosis in women of reproductive age. STUDY DESIGN: Cross-sectional comparative study. PLACE AND DURATION OF STUDY: Biochemistry Department, Islamic International Medical College, Rawalpindi, in collaboration with the Gynecology and Obstetrics Department, Pakistan Railways Hospital, Rawalpindi, from March 2017 to February 2018. METHODOLOGY: Forty-four patients with endometriosis and 44 controls of reproductive age group were enrolled for the study. Those having other comorbid systemic or endocrine diseases were excluded from the study. Levels of albumin and C3 complement were measured by using Micro Lab 300 and ELISA, respectively; and the results were compared by using SPSS version 21. groups, respectively (p=0.201). The mean serum C3 levels were 1120.9±265 µg/ml and 2241.0±293 µg/ml in the control and endometriosis patient groups, respectively (p <0.001). RESULTS: The mean serum albumin levels were 4.62±0.88 g/dl and 4.42±0.52 g/dl in the control and endometriosis Conclusion: Serum C3 complement were significantly increased in endometriosis patients but not in controls; whereas, serum albumin levels were not significantly different in endometriosis cases as compared to the controls.

Genotoxic And Cytotoxic Effects Of Oral Vanadyl Sulphate.

BACKGROUND: Vanadyl sulphate is available as herbal medicine against diabetes mellitus and body building supplement, over the counter worldwide. The available data on its safety is controversial and inadequate. The objective of this study was to analyse its safety in usual therapeutic dose range. METHODS: It was an experimental study carried out at the Department of Biochemistry & Molecular Biology, Army Medical College, National University of Medical Sciences (NUMS), Rawalpindi, Pakistan, from Jun 2014 to Oct 2018. The study was carried out on 105 Sprague Dawley rats for duration of 24 weeks. The animals were randomly distributed in three groups of 35 each. The group I rats were marked as control while rats of group II & III were administered vanadyl sulphate 0.06mg/day and 0.3mg/day respectively. Alanine amino transferase (ALT) and Malondialdehyde (MDA) were measured in serum while comet assay was performed on WBCs. RESULTS: The plasma levels of ALT and MDA were significantly raised in group II and III subjects. Single cell gel electrophoresis (SCGE) / comet assay showed minimal "tail moment" in control group and increased tail moment in group II and III in a dose dependent manner which indicates dsDNA breaks. CONCLUSIONS: It was observed that vanadyl sulphate causes hepatocellular toxicity, oxidative stress and damage to the DNA in usual therapeutic/ supplemental doses. Due to hazardous effects, its use in humans as alternate medicine may be reviewed.

Exploring NS3/4A, NS5A and NS5B proteins to design conserved subunit multi-epitope vaccine against HCV utilizing immunoinformatics approaches.

Hepatitis C virus (HCV) vaccines, designed to augment specific T-cell responses, have been designated as an important aspect of effective antiviral treatment. However, despite the current satisfactory progress of these vaccines, extensive past efforts largely remained unsuccessful in mediating clinically relevant anti-HCV activity in humans. In this study, we used a series of immunoinformatics approaches to propose a multiepitope vaccine against HCV by prioritizing 16 conserved epitopes from three viral proteins (i.e., NS34A, NS5A, and NS5B). The prioritised epitopes were tested for their possible antigenic combinations with each other along with linker AAY using structural modelling and epitope-epitope interactions analysis. An adjuvant (ß-defensin) at the N-terminal of the construct was added to enhance the immunogenicity of the vaccine construct. Molecular dynamics (MD) simulation revealed the most stable structure of the proposed vaccine. The designed vaccine is potentially antigenic in nature and can form stable and significant interactions with Toll-like receptor 3 and Toll-like receptor 8. The proposed vaccine was also subjected to an in silico cloning approach, which confirmed its expression efficiency. These analyses suggest that the proposed vaccine can elicit specific immune responses against HCV; however, experimental validation is required to confirm the safety and immunogenicity profile of the proposed vaccine construct.

Association of RETN C-420G single nucleotide polymorphism with type 2 diabetes mellitus in Pakistani Punjabi Rajput population.

OBJECTIVE: To determine the association of human resistin gene RETN C-420G single nucleotide polymorphism with type 2 diabetes mellitus in a specific ethnic population.. METHODS: The controlled study was conducted from June 2012 to January 2015 at Military Hospital, Rawalpindi, Army Medical College, Rawalpindi, and the Institute of Biomedical and Genetic Engineering, Islamabad, Pakistan. Patients with type 2 diabetes and healthy controls belonging to Pakistani Punjabi Rajput ethnic group were genotyped for human resistin gene RETNC-420G single nucleotide polymorphism. Serum resistin, serum insulin, fasting blood sugar, lipid profile, body mass index and insulin resistance was determined and correlated with genotypes. SPSS 18 was used for data analysis. RESULTS: Of the 789 subjects, 539(68%) were diabetics and 250(32%) were controls. Serum resistin levels were significantly higher in diabetics than controls (p<0.05). The frequency of GG, GC and CC was 15(2.8%), 322(59.75%) and 202(37.5%) in diabtics. This single nucleotide polymorphism was associated with diabetes (p<0.02).Human resistin gene RETN C-420G single nucleotide polymorphism was not associated with serum resistin, insulin, body mass index, insulin resistance and dyslipidaemia in both groups (p<0.05 each). CONCLUSIONS: Human resistin gene RETN C-420G single nucleotide polymorphism was found to be a risk factor for type 2 diabetes in Pakistani Punjabi Rajput population..

Correlation between Serum Amyloid A-Low Density Lipoprotein and Genotoxicity in Smokers.

OBJECTIVE: To investigate the relation between serum amyloid A-low density lipoprotein (SAA-LDL) and genotoxicity in smokers. STUDY DESIGN: An experimental study. PLACE AND DURATION OF STUDY: Army Medical College, Rawalpindi and National Institute of Health (NIH), Islamabad, from June 2014 to February 2015. METHODOLOGY: Seventy healthy Sprague Dawley rats were purchased from NIH and exposed to cigarette smoke in smoke chamber for three months. Blood samples were drawn from each rat at the end of the study period. SAA-LDL was determined by enzyme-linked immunosorbent assay (ELISA). Genotoxicity was assessed by cytokinesis block micronucleus (CBMN) assay. Pearson correlation was used to find correlation between SAA-LDL and genotoxicity. RESULTS: Strong positive correlation was found between SAA-LDL and micronuclei frequency in smoke-exposed rats (r=0.799, N=70, p <0.01). CONCLUSION: Statistically significant strong positive correlation between SAA-LDL and genotoxicity in smoke-exposed rats shows that changes in one is associated with changes in other and vice versa.

Frequency of Common VKORC1 Polymorphisms and Their Impact on Warfarin Dose Requirement in Pakistani Population.

Polymorphisms in vitamin K epoxide reductase complex subunit 1 (VKORC1) gene lead to interindividual variability in warfarin dose requirement. The characterization of genotype frequency distribution is required in different populations for construction of customized dosing algorithms to enhance the efficacy and reduce the toxicity of warfarin therapy. This study was carried out in Pakistani population to evaluate the contribution of common VKORC1 polymorphisms to warfarin therapy. A total of 550 stable patients taking warfarin were enrolled after medical history, physical examination, and laboratory investigations. Single blood sample was collected after informed consent. Genomic DNA was extracted and genotype analysis for VKORC1 1173C>T and VKORC1-1639G>A polymorphisms was done by polymerase chain reaction-restriction fragment length polymorphism assay. A number of samples were also analyzed by direct DNA sequencing for validation of results. Data were analyzed using SPSS version 20. Genotype frequency distributions of VKORC1 1173C>T and VKORC1-1639G>A were found to be different from other populations. Both of these polymorphisms did not demonstrate significant effect on warfarin dose requirement. Although Cytochrome P450 2C9 (CYP2C9) and VKORC1 polymorphisms together attributed only 3.8% variability in warfarin dose but it was statistically significant ( p value = .004). It is concluded that there is a need to study genotype frequency distribution and their effect on warfarin dose variability among different populations due to diversity in outcome. At the same time, no effect on warfarin dose variation explained by VKORC1 polymorphisms and small variability explained by studied genotypes stresses the need for exploration of more genetic and nongenetic factors in Pakistani population.

Association of ADIPOQ C-11377G (rs266729), TNF-α αG-308A(rs1800629) and TNF-α αG-238A(rs361525) Single Nucleotide Polymorphisms with Type 2 Diabetes Mellitus in Pakistani Population.

OBJECTIVE: To determine the frequencies of adiponectin (ADIPOQ) C-11377G, tumor necrosis factor-alpha (TNF-α) G-308A and TNF-αG-238Asingle nucleotide polymorphisms (SNP) and their association with serum levels in Pakistani T2DM and healthy population. STUDY DESIGN: Case control study. PLACE AND DURATION OF STUDY: Military Hospital, Rawalpindi, Army Medical College and Institute of Biomedical and Genetic Engineering, Islamabad, Pakistan, from June 2012 till 2014. METHODOLOGY: Cases (n=539) and controls (n=250) comprising of T2DM and healthy subjects, respectively, belonging to Pakistani Punjabi Rajput ethnicity were genotyped for SNPs. Serum adiponectin, TNF-α, insulin, blood sugar fasting (BSF), lipid profile, body mass index (BMI), and insulin resistance (IR) was determined and correlated with genotypes. RESULTS: Serum TNF-αwas significantly higher and adiponectin was lower in T2DM than healthy controls (p < 0.003 and 0.0001, respectively, Mann-Whitney U-test). The frequency of ADIPOQ CC, GC and GG was 340 (63.1%), 167 (31%) and 32 (5.9%) in T2DM patients. ADIPOQ -11377 SNPwas not significantly associated with T2DM [OR = 1.116 (95% CI 0.811.53), p = 0.27- Fisher's exact test]. Genotypes deviated from Hardy-Weinberg equilibrium. Minor alleles of TNF-αG-308A and TNF-αG-238Awere not found in either groups. CONCLUSION: Frequency of ADIPOQ -11377 risk allele is low and does not functionally affect the serum adiponectin levels; hence, ADIPOQ C-11377G SNPis not a risk factor for T2DM in Pakistani Punjabi Rajput patients. Moreover, TNF-αG-308A and TNF-αG-238ASNPs are not prevalent in this ethnic group.

Effects of two functionally important SLCO1B1 gene polymorphisms on pharmacokinetics of atorvastatin.

Organic anion transporter polypeptide 1B1 (OATP1B1) encoded by (SLCO1B1) gene, an uptake transporter involved in the transport of drugs and endogenous compounds and located in hepatocyte sinusoidal membrane. Objective of study was to investigate the effects of two functionally significant SNPs (388A>G and 521T>C) and their respective genotypes of SLCO1B1 gene encoding OATP1B1 on the pharmacokinetics of atorvastatin. A total of 100 subjects divided into 6 groups as per their genotype profile were recruited. A single dose of 80mg atorvastatin was orally administered and plasma concentration measured up to 48 hours. The 388A>G and 521T>C genotypes were significantly associated with each other when compared for AUC and Cmax but exhibited no significant variations in Tmax and t1/2. 521 SNP is rather more strongly associated with altered pharmacokinetics of atorvastatin when compared with the 388 SNP, though the homozygous bi-allelic variant of 388 SNP also exhibited a fairly significant variation along with homozygous bi-allelic variant of 521 SNP. The inter-individual variation in pharmacokinetics can be explained by SLCO1B1 polymorphism.

MicroRNA pharmacogenomics based integrated model of miR-17-92 cluster in sorafenib resistant HCC cells reveals a strategy to forestall drug resistance.

Among solid tumors, hepatocellular carcinoma (HCC) emerges as a prototypical therapy-resistant tumor. Considering the emerging sorafenib resistance crisis in HCC, future studies are urgently required to overcome resistance. Recently noncoding RNAs (ncRNAs) have emerged as significant regulators in signalling pathways involved in cancer drug resistance and pharmacologically targeting these ncRNAs might be a novel stratagem to reverse drug resistance. In the current study, using a hybrid Petri net based computational model, we have investigated the harmonious effect of miR-17-92 cluster inhibitors/mimics and circular RNAs on sorafenib resistant HCC cells in order to explore potential resistance mechanisms and to identify putative targets for sorafenib-resistant HCC cells. An integrated model was developed that incorporates seven miRNAs belonging to miR-17-92 cluster (hsa-miR-17-5p, hsa-miR-17-3p, hsa-miR-19a, hsa-miR-19b, hsa-miR-18a, hsa-miR-20a and hsa-miR-92) and crosstalk of two signaling pathways (EGFR and IL-6) that are differentially regulated by these miRNAs. The mechanistic connection was proposed by the correlation between members belonging to miR-17-92 cluster and corresponding changes in the protein levels of their targets in HCC, specifically those targets that have verified importance in sorafenib resistance. Current findings uncovered potential pathway features, underlining the significance of developing modulators of this cluster to combat drug resistance in HCC.

Association of IL-6 C-174G (rs 1800795) single nucleotide polymorphism with type 2 diabetes mellitus in Pakistani population.

OBJECTIVE: To determine the association of interleukin-6 C-174G single nucleotide polymorphism with type 2 diabetes mellitus and metabolic parameters. METHODS: This case-control study was conducted from June 2012 to December 2013 at the Military Hospital Rawalpindi, the Centre for Research in Experimental and Applied Medicine, Army Medical College, Rawalpindi, and the Institute of Biomedical and Genetic Engineering, Islamabad, Pakistan. Two cohorts of subjects were genotyped for the single nucleotide polymorphism. One cohort comprised type 2 diabetics and other included healthy subjects. In these groups, serum interleukin-6, serum insulin, blood sugar fasting, lipid profile, body mass index and insulin resistance was determined and correlated with genotypes. RESULTS: Of the 789 participants, 539(68.3%) were in the study group and 250(31.7%) in the control group. Serum interleukin-6 was significantly higher in diabetics than healthy controls (p<0.0001). The frequency of GG, GC and CC was 267(49.5%), 235(43.6%) and 37(6.9%) in diabetic patients and 128(51.2%), 74(29.6%) and 48(19.2%) in healthy controls, respectively. Interleukin-6 C-174G single nucleotide polymorphism was significantly associated with diabetes [odds ratio = 3.22 (95% confidence interval: 2.04-5.1; p<0.0001). Genotypes were within Hardy-Weinberg equilibrium. Interleukin-6 C-174G single nucleotide polymorphism was significantly associated with serum interleukin-6 in the order of GC>GG>CC but was not associated with body mass index, insulin resistance, serum insulin and dyslipidaemia in diabetic patients (p>0.05 each). CONCLUSIONS: Interleukin-6 C-174G single nucleotide polymorphism was a risk factor in type 2 diabetes and contributed to higher serum interleukin-6 levels among the participants.