Identification and Molecular Simulation of Genetic Variants in ABCA1 Gene Associated with Susceptibility to Dyslipidemia in Type 2 Diabetes.

Genetic insights help us to investigate disease pathogenesis and risk. The ABCA1 protein encoded by ABCA1 is involved in transporting cholesterol across the cell membrane. Genetic variations in the ABCA1 gene are well documented; however, their role in the development of diabetic dyslipidemia still needs to be explored. This study aimed to identify the associations of rs757194699 (K1587Q) and rs2066714 (I883M) with dyslipidemia in type 2 diabetes and performed molecular simulations. In our case-control study, 330 individuals were divided equally into a diabetic dyslipidemia cases and a healthy controls. Allele-specific polymerase chain reaction and restriction fragment length polymorphism were performed to screen selected variants of the ABCA1 gene. Sanger sequencing was also performed to find genetic mutations in exon 5 of the ABCA1 gene. The C allele of rs757194699 was observed at a high frequency in cases compared to controls and followed the overdominant genetic model (p < 0.0001, OR:3.84; CI:1.67-8.82). The frequency of G allele of rs2066714 was significantly higher in cases compared to controls and followed the genetic model of codominant (p< 0.0001, OR: 39.61; CI:9.97-157.32), dominant (p < 0.0001,OR:59.59; CI:15.19-233.81), overdominant (p< 0.0001, OR:9.75; CI:3.16-30.11), and log-additive (p< 0.0001, OR:42.15; CI:11.08-160.40). In silico modeling and docking revealed that rs2066714 and rs757194699 produced deleterious conformational changes in the ABCA1 protein, resulting in alterations in the binding of the apoA1 protein. There were no genetic variations found in exon-5 in Sanger sequencing. The G allele of rs2066714 and C allele of rs757194699 in the ABCA1 gene were found to be risk alleles in the development of dyslipidemia in type 2 diabetes. These polymorphisms could alter the binding site of ABCA1 with apoA1 thus disturbs the reverse cholesterol transport.

Association of single nucleotide polymorphism rs3213119 variant of IL-12B gene in diagnosed Rheumatoid Arthritis patients.

Objective: To identify the IL12B gene variant (rs3213119) and to find its association in Pakistani clinical population of Rheumatoid Arthritis. Methods: It was a population association (unrelated) case control study, performed from January - December 2022 at Laboratory of Department of Biochemistry and Molecular Biology, Army Medical College, Rawalpindi. Blood samples were collected from all 150 study participants, followed by DNA extraction and Allele-specific polymerase chain reaction performed at Center for Research in Experimental and Applied Medicine (CREAM) Laboratory of Department of Biochemistry and Molecular Biology, Army Medical College Rawalpindi. Statistical analysis was done using 'SPSS' (version-22), followed by gene analysis on 'SNPstat'. Results: About 28.0% of RA patients were smokers, 38.7% had history of RA in a first degree relative and 70.7% had positive history of consanguinity. Considering rs3213119 variant of IL12B gene, frequency of major allele C was 100%, minor allele A was 21%, genotype C/C was 79% and C/A was 21%. Applying the log additive model, the odds ratio of the genotype C/C was 1.00 (adjusted by age and gender with 95 % CI) and the odds ratio of the genotype C/A was 0.00, 52.0% of RA patients originated from four predominant ethnic groups, namely Awaans (18.7%), Rajputs (14.7%), Pathans (12.0%) and Araeens (6.7%). Conclusion: The study findings suggest the role of minor allele 'A' as risk allele in our clinical population. CA genotype confers susceptibility towards the RA development.

Association of serum IL12 with clinical and biochemical parameters in a cohort of diagnosed rheumatoid arthritis patients on oral conventional synthetic disease modifying anti-rheumatic drugs.

Objective: To determine the association of serum interleukin-12 levels with disease progression in active rheumatoid arthritis patients on oral conventional synthetic disease-modifying anti-rheumatic drugs. METHODS: The case-control study was conducted at the Army Medical College, Rawalpindi, in collaboration with the Pak Emirates Military Hospital, Rawalpindi, Pakistan, from January to December 2022, and comprised rheumatoid arthritis patients or either gender aged 18-75 years who were placed in group I, while group II comprised healthy controls. Demographic and clinical data was noted, and 2ml blood samples were drawn from each subject. The serum was separated and analysed using sandwich enzyme-linked immunosorbent assay to quantify serum interleukin-12 levels. Data was analysed using SPSS 22. RESULTS: Of the 150 subjects, 75(50%) were in group I; 27(36%) males and 48(64%) females with overall mean age 45.70±11.70 years. There were 75(50%) subjects in group II; 37(49.3%) males and 38(50.7%) females with overall mean age 31.70±7.70 years. Serum interleukin-12, erythrocyte sedimentation rate and C-reactive proteinquantitative levels were significantly higher in group I compared to group II (p<0.05). Smoking, positive family history of rheumatoid arthritis in a first-degree relative and history of consanguinity were identified as risk factors though they were not statistically significant (p>0.05). In group I (n=75), out of total study subjects, only 55(73.3%) cases belonged to the predominant castes, namely Awan, Rajput, Pathan, Araeen, Bhatti, Malik, Mughal, Sudhan, Chaudary, and Jutt. These individuals showed significantly higher mean serum interleukin-12 levels compared to patients of other castes in the same group. Conclusion: Mean serum interleukin-12 levels were higher in rheumatoid arthritis patients despite being on oral conventional synthetic disease-modifying anti-rheumatic drugs.

An insight into genetic landscape of inflammatory bowel disease.

Inflammatory bowel disease has been regarded to be chronic intestinal inflammation characterised by a dsyregulatory immune response. The disease pathophysiology is known to be complex. Growing pieces of evidences underpin the involvement of various environmental and genetic determinants in the disease onset. The current narrative review was planned to manifest the contribution of genetic drivers for disease onset and to target signalling pathways that might present a therapeutic potential for further research. The factors of the disease that provide the genetic nature and understanding of the pathways involved have been researched in recent times. Also, numerous diseasedeveloping factors have been studied and assessed. Among them genetic determinants of disease onset have further improved the understanding of disease development. Genetic contributors to the onset of disease as well as important therapeutic targets need to be understood as predictive genetic risk factors have a potential implication for personalised treatment.

Oxidative Stress and Lipid Peroxidation in NAFLD with and without Type 2 Diabetes Mellitus.

OBJECTIVE: To compare superoxide dismutase 1 (SOD 1) and malondialdehyde (MDA) levels along with biochemical parameters in patients of non-alcoholic fatty liver disease (NAFLD) with and without Type 2 diabetes mellitus. STUDY DESIGN: Cross-sectional comparative study. Place and Duration of the Study: Centre for Research in Experimental and Applied Medicine, AMC, in collaboration with the Department of Radiology, Combined Military Hospital, Rawalpindi, from February to November 2022. METHODOLOGY: Two hundred and ten patients were selected by non-probability purposive sampling and divided into 3 groups. Healthy individuals were labelled as Group Ι, Group II included patients of NAFLD without diabetes mellitus, and Group III had patients of NAFLD with diabetes mellitus. Fasting blood glucose levels and lipid profile were measured. ELISA (enzyme-linked immunoassay) was done for the assessment of SOD 1 and MDA levels. The data was analysed by version 22.0 of SPSS and expressed in mean ± SD and percentage. One-way ANOVA was done for all groups and grade comparison was followed by the post-hoc Tukey test. RESULTS: When compared to control groups, the mean SOD 1 level in diseased groups was significantly lower (p<0.001). There was a statistically significant difference between each group (p<0.001). Mean levels of MDA were significantly increased in diseased groups as compared to controls with a statistically significant difference between all groups except between Group II and III. CONCLUSION: In patients having NAFLD with and without diabetes mellitus, SOD 1 levels were considerably lower compared to controls whereas MDA levels were significantly higher. This decrease in SOD 1 and raise in MDA levels was indicative of increased oxidative stress in patients and can be viewed as a biomarker for oxidative stress. KEY WORDS: NAFLD, ELISA, Oxidative stress, SOD 1, MDA, Lipid peroxidation.

Expression analysis of ABCA1 in type 2 diabetic Pakistani patients with and without dyslipidemia and correlation with glycemic index and lipid profile.

Diabetes Mellitus type II, earlier considered as an endocrinological disorder is now more regarded as an inflammatory disorder along with lipid aberrations. It demands for regular monitoring, healthy dietary habits and lifestyle modification. This study was focused on gene expression of ATP binding cassette protein 1 (ABCA1) in diabetic dyslipidemia patients in comparison with control groups of only diabetics and healthy individuals. Blood samples and data were collected from recruited 390 patients who were further divided into three groups (130 each). Glycemic index and lipid profile was assessed. Delta Delta Ct method was used that revealed downregulation of the studied gene more in diabetic dyslipidemia patients as compared to only diabetics and healthy controls. The Ct values of ABCA1 were associated with glycemic index and lipid profile using Pearson's correlation. A negative correlation with fasting blood sugar and a positive correlation with HbA1cwas observed in only diabetics group. While in diabetic dyslipidemia and normal healthy controls, a negative correlation was found with both. As far as the lipid profile is concerned a positive correlation was observed among only diabetics with whole lipid profile. In diabetics with dyslipidemia, a negative correlation with all parameters except the TAGs was observed. A positive correlation with all except HDL was observed in healthy controls. The Ct values and fold change were compared among diseased and healthy individuals by applying independent t test. The cycle threshold in only diabetics was p = 0.000018 and in diabetic dyslipdemia individuals was p = 0.00251 while fold change in only diabetics (p = 0.000230) and in diabetics with dyslipidemia (p = 0.001137) was observed to be as statistically significant.

Association of R1939W and P1987R variants of Otoferlin (OTOF) gene with severe to profound nonsyndromic sensorineural hearing loss in Pakistani subjects.

Objective: To find possible association of R1939W and P1987R variants of OTOF gene with severe to profound NSSHL in cochlear implant subjects. Methods: It was a case control study, conducted from June 2021 to February 2022, comprising 50 cases of severe to profound NSSHL who had received cochlear implant from ENT Department, CMH Rawalpindi and 50 age-matched healthy controls from PEMH Rawalpindi. Blood samples were collected from all the subjects, followed by DNA extraction and allele-specific polymerase chain reaction, performed at Multi-disciplinary Laboratory of Department of Biochemistry and Molecular Biology, Army Medical College Rawalpindi. Statistical analysis was done using 'SPSS' and 'XLSTAT', followed by genetic analysis using 'SNPstat'. Results: Mean age of the cases was 5.96 ± 4.62 years (N=50), comprising 58% males and 42% females. All had bilateral and prelingual HL. Parental consanguinity was 72%, whereas 62% cases had a positive family history of deafness. Alleles of R1939W and P1987R were not associated with NSSHL, as shown by their p values of 0.56 and 0.89 respectively. For R1939W ORs were 0.71 (dominant model) and 0.80 (overdominant model), indicating negative association with NSSHL. Regarding P1987R OR was 0.96 (log-additive model). Genotypes of both variants were not in HW Equilibrium (p <0.0001), whereas their alleles showed high LD (D'=0.92). Conclusion: High percentage of parental consanguinity was observed among cochlear implant candidates. The OTOF variants R1939W and P1987R were found to have protective roles against NSSHL in study population.

Risk Analysis and Assessment of Lipid Abnormalities as the Earliest Complication in Newly Diagnosed Diabetic and Non-Diabetic Individuals of a Local Population.

Lipid variations have been frequently observed in global populations that can affect health status. Mainly studies have been conducted on the type 2 diabetic population, but limited data is available on newly diagnosed ones to unravel complications and risk predictors independent of disease progression. This study comprising 244 individuals was carried out to assess the lipid abnormalities in newly diagnosed diabetics and non-diabetics. The clinical and socio-demographic data were collected and analyzed using independent samples t-test and linear regression. Serum lipid variations were observed individually and in combination. The individuals in group I (diabetics with dyslipidemia) revealed elevated levels of low-density lipoprotein and serum triglycerides higher than in group II (non-diabetics with dyslipidemia). The frequency of deranged total cholesterol in group I was observed to be higher than in group II. Independent samples t-test showed a significant mean difference in variables between the two groups. Linear regression analysis showed a significant variable outcome for predictors between high-density lipoprotein (HDL) and physical activity (B= -0.043, 95% CI: -0.80, -0.006) and total cholesterol (TC) with family history (B= -0.062, 95% CI: -0.123, -0.001). The findings conclude that lipid levels deranged independently regardless of type 2 diabetes mellitus and present as an early onset in type 2 diabetes instead of later stage complication. These derangements of lipid levels are an independent risk factor for future cardiovascular pathology.