Hepatitis B virus subgenotypes D1 and D3 are prevalent in Pakistan

<p>Abstract</p> <p>Background</p> <p>As the hepatitis B genotyping is important for assessing its clinical implications and geographical distribution, the sub-genotypes have been found useful for determination of specific genomic markers related to hepatocarcinogenesis. In Pakistan, there is no reported data on molecular evolutionary analysis of HBV. A study was, therefore, much needed to evaluate the spectra of mutations present in the strains prevalent here.</p> <p>Findings</p> <p>to confirm specificity of PCR typing, phylogenetic analysis of the pre-S1 region and the divergence was studied through 13 sequences of 362 bp (accession number <ext-link ext-link-type="gen" ext-link-id="EF432765">EF432765</ext-link> – <ext-link ext-link-type="gen" ext-link-id="EF432777">EF432777</ext-link>). A total of 315 serum samples, selected from HBsAg positive patients representing the major ethnic groups, residing in Karachi, Sindh were tested for genotyping. Genotype D (219/315) was found to be the most prevalent (70%) amongst our patients. The rest of the genotypes A and a mixture of A and D (AD) were distributed as 20%, and 10% respectively. Phylogenetic tree demonstrated clustering of 11 samples with subgenotype D1 sequences and the remaining two strains on a branch within D3 samples. All samples intermixed with strains from other countries and were found to be closely related to Indian, Iranian and Egyptian HBV strains with 98.7 – 99.0% homology.</p> <p>Conclusion</p> <p>This study confirms the predominance of genotype D in southeastern Asia and presence of subgenotypes DI and D3 in the Pakistani infected patients. More studies are required to investigate the reason for fewer inclusions of D3 compared to the D1 in Pakistani HBV strains.</p

A Novel Variant Of Regenerating Iα Gene (REG) In Type II Diabetics Among Pakistani Targeted Population

Abstract Objective: Regeneration of pancreatic β-cells, is an essential step towards diabetes management. The regenerating (REG) Iα gene is secreted from damaged β-cell for the synthesis of β-cell. This study aimed to identify REG Iα gene polymorphisms and their association with Type II diabetes (T2DM). Methods: Patients (110) with T2DM and age-related controls were selected from PNS Shifa Hospital, Karachi. DNA was extracted PCR was performed and amplified products were sequenced to identify polymorphisms. For six exons of the REG 1a gene, 6 sets of primers were designed. The selected (51) samples were amplified and sequenced for 306 (51x6) times. Odds ratios were calculated through logistic regression analysis. The correlation was used to find an association between REG Iα and diseases. p&lt; 0.05 was considered significant. Results: Blood samples were drawn from 90 finalized patients, including 70 diabetics and 20 controls with an M: F ratio of 12:8. Twenty patients opted to withdraw. The REG Iα and disease duration in type II diabetics showed a negative correlation (r= -0.355, p=0.005). The single nucleotide polymorphisms (SNPs) of eight sites were detected: g.-385T&gt;C, g.-243T&gt;G, g.-145G&gt;A, g.+142A, g.+209G&gt;T, g.+226A&gt;G, g.+2199G&gt;A, g.+2360A&gt;G.  The novel SNP g.-145G&gt;A was found in all patients (controls, T2DM). Among all SNPs, only g.+209G&gt;T showed a positive association (OR= 2.4, p=0.01) with T2DM. Whereas, g.-243T&gt;G showed a positive association (OR=8.06, p=0.0003) with smoking. Conclusion: A novel variant g.-145G&gt;A REG Iα gene was found among all participants. The SNPs g.+209G&gt;T had a significant positive association with T2DM and SNP g.-243T&gt;G showed an increased risk of the disease among smokers

Hepatitis B virus interacts with albumin precursor in vivo

A range of proteins has been recognized as mediators/hypothetical receptors for hepatitis B virus (HBV), but the results are conflicting and inconclusive especially regarding their biological significance. This study was aimed at identifying a novel HBV-interacting protein which would provide a better understanding of its transport in blood, attachment, fusion and entry into hepatocytes. Serum samples positive for HBV confirmed by PCR were subjected to ammonium sulfate fractionations at 50, 75, 100%, saturation. PCR of each fraction demonstrated amplification of HBV in 100% fraction. Protein analysis by SDS PAGE of fractions showed one band of approximately 69 kDa protein, in 100% fraction. The 100% fraction band was excised from gel and sequence was determined by MALDI TOF which showed mass values from 705 to 3722. Mass spectrometry of trypsinized 69 kDa species revealed peptide sequences that covered 54% of the serum albumin precursor amino acid sequence, with pI of 5.9. Western blot, carried out using primary anti-albumin precursor antibody, further validated this protein. This study establishes that Hepatitis B Virus binds to albumin precursor, suggesting its role in the initiation of HBV infection and hence may offer new therapeutic strategies against HBV infection

The association of complex liver disorders with HBV genotypes prevalent in Pakistan

Background Genotyping of HBV is generally used for determining the epidemiological relationship between various virus strains and origin of infection mostly in research studies. The utility of genotyping for clinical applications is only beginning to gain importance. Whether HBV genotyping will constitute part of the clinical evaluation of Hepatitis B patients depends largely on the availability of the relevance of the evidence based information. Since Pakistan has a HBV genotype distribution which has been considered less virulent as investigated by earlier studies from south East Asian countries, a study on correlation between HBV genotypes and risk of progression to further complex hepatic infection was much needed Methods A total of 295 patients with HBsAg positive were selected from the Pakistan Medical Research Council\u27s (PMRC) out patient clinics. Two hundred and twenty six (77%) were males, sixty nine (23%) were females (M to F ratio 3.3:1). Results Out of 295 patients, 156 (53.2%) had Acute(CAH), 71 (24.2%) were HBV Carriers, 54 (18.4%) had Chronic liver disease (CLD) Hepatitis. 14 (4.7%) were Cirrhosis and HCC patients. Genotype D was the most prevalent genotype in all categories of HBV patients, Acute (108), Chronic (39), and Carrier (53). Cirrhosis/HCC (7) were HBV/D positive. Genotype A was the second most prevalent with 28 (13%) in acute cases, 12 (22.2%) in chronics, 14 (19.7%) in carriers and 5 (41.7) in Cirrhosis/HCC patients. Mixed genotype (A/D) was found in 20 (12.8%) of Acute patients, 3 (5.6%) of Chronic and 4 (5.6%) of carriers, none in case of severe liver conditions. Conclusion Mixed HBV genotypes A, D and A/D combination were present in all categories of patients except that no A/D combination was detected in severe conditions. Genotype D was the dominant genotype. However, genotype A was found to be more strongly associated with severe liver disease. Mixed genotype (A/D) did not significantly appear to influence the clinical outcome

BONE IS NOT JUST CALCIUM AND VITAMIN DA REVIEW OF ESSENTIAL NUTRIENTS REQUIRED FOR BONE HEALTH

Bone formation is a constant procedure in which osteoblasts lay new bone and osteoclasts resorb it. Mineralization takes place at active bone formation sites where the extracellular matrix vesicles, the major sites of apatite mineral deposition, are present. Turnover of bone, which maintains its structure and integrity, has several events such as activation, resorption and osteogenesis by osteoblasts. The bone crystals proliferation depends on the presence of collagen, hormones, minerals and vitamins such as phosphorus, magnesium, calcium, zinc, fluoride, potassium, manganese, boron, copper, iron, calcium and a number of vitamins such as B,K,C,A,D, etc. Information regarding the elements involved in bone formation was retrieved through studies published up to 2017 in PubMed, Medline and other authentic search engines available in the University. This review highlights the individual roles of specific vitamins and minerals at the respective steps of bone formation, insufficiency at all particular stages result in various bone pathologies, leading to deficiency disorders, fractures and poor friable bones. Since the role of vitamin D and calcium is well established therefore these were not included in this review

Expression Of Nephrin In Early Diagnosis Of Diabetic Nephropathy

Background & Objectives: Diabetic nephropathy occurs as a result of proximal tubule dysfunction with podocyte damage due to increased advanced glycation end-products’ insult in diabetes. Nephrin, one of the three proteins that make up the podocyte architecture, is excreted foremost following renal damage. The aim of this study was to evaluate the efficiency and reliability of nephrin for an early biomarker of kidney damage in diabetic patients. Methods: Urine samples (78) were collected from diabetic center. Protein and glucose were determined by Dipstick. The patients were grouped on the basis of Albumin/creatinine ratio (UACR) as normoalbuminuric, Microalbuminuric and Macroalbuminuric with UACR less than 30 mg/g, from 30-300mg/g, and above 300mg/g respectively. ELISA KIT (Exocell USA) was used for Nephrin estimation. Statistical evaluation was done on SPSS version 20. Results: Nephrinuria was present in 70(89.7%) out of 78 diabetic patients including 35(81.4%) of 43(55.1%) normoalbuminurics, 5(6.4%) of 30(38.5%) microalbuminurics and all 5 of macroalbuminurics (p=0.027). Nephrinuria was found associated with duration of diabetes, 21(91.3%)/23 were positive with less than three years of disease, 24(92.3%)/26 with three to seven years and 25(86.2%)/29 in more than seven years (p=0.039). Nephrin levels were found increasing from normo- (0.86ug/ml) to Micro- (11.6μg/ml) to Macroalbuminuria group (47.6μg/ml), compared to 0.15 μg/ml in comparison group. \ud Conclusion: The increase in Nephrin levels from 0.86μg/ml in patients with normal albuminuria to 47.6μg/ml in patients with macroalbuminuria suggests that Nephrin precedes albumin in urine predicting early signs of kidney damage. Nephrin biomarker can be used as predictor for early diagnosis of Nephropathy in diabetics. Key Words: Diabetic Nephropathy; Albuminuria; Podocyte

Complications of Obesity in Polycystic Ovary Syndrome: Insulin Resistance and Inflammation

Background: The polycystic ovarian syndrome (PCOS), a well-known endocrine-metabolic disease, is caused by the interaction of environmental, genetic and metabolic factors. A cross-sectional, comparative study was planned to evaluate high sensitivity C-reactive protein (hs-CRP), insulin and lipid profile in non-obese and obese PCOS patients. Methods: Seventy-two women diagnosed with PCOS as per Rotterdam criteria, were placed in three groups in accordance with the body mass index: Group 1 (normal weight), Group 11 (overweight), and Group 111 (obese). Blood glucose, hs-CRP, serum insulin, lipid profile was estimated and insulin resistance was calculated. Data was analyzed using version 20 of SPSS. Analysis of variance (ANOVA) was used to compare the numeric variables among three groups of PCOS women and p-value < 0.05 was considered statistically significant. Results: Significantly, higher levels of insulin (13.03 ± 0.22), triglyceride (1.74 ± 0.96) and hs-CRP (7.24±4.11) were detected in obese PCOS women. The levels of fasting blood glucose (4.61±0.54) were also raised. Homeostatic Model Assessment for Insulin Resistance (HOMA-IR) (2.69±0.79) showed insulin resistance in obese PCOS women (p=0.004). The obese group had significantly lower HDL-Cholesterol levels (0.82±0.09). Levels of total cholesterol (4.69±0.97) and LDL-Cholesterol (3.08±0.64) were increased but results were non-significant. These results indicate obesity leads to insulin resistance, dyslipidemia with low grade inflammation in PCOS. Conclusion: Frequency of insulin resistance, dyslipidemia and low-grade inflammation was found significantly higher in young obese PCOS women. Obesity may have an important role in the aetiology as well as the complications associated with PCOS. Keywords: Polycystic Ovarian Syndrome; hs-CRP; Insulin Resistance; Dyslipidemia; Body Mass Index

HIGH FREQUENCY OF OSTEOPENIA IN YOUNG ADULTS OF KARACHI

Background: Osteopenia or low bone density, generally considered the disease of the old, is now sneaking around the younger generation. The increase in frequency of low trauma fractures is raising an alarm. This study was aimed to explore the frequency of osteopenia amongst young adults in Karachi, Pakistan. Methods: Non-probability consecutive sampling was used to choose 116 subjects at Ziauddin University, Karachi, in January 2018. After informed consent participants underwent bone scans for measurement of bone mineral density on their calcaneus (bone in heel). Tocategorize osteopenia T-score between -1.0 and -2.5 was evaluated. All participants with T-score of -2.5 or below were identifiedas osteoporotic. The data were entered on IBM SPSS statistics version 20.0 and descriptive analysis was done. Results: Osteopenia was detected in 57 (49.1%) of the participants [42 (36.2%) male and 15 (12.9%) female] of which 38 (32.8%) belonged to aged 21 to 35 years. Osteoporosis was found in 20 (17.2%) of this group. In the older participants’ frequency of osteopenia and osteoporosis was 12.2% and 42.9% respectively. Chi square test indicated no significant association between age and bone scan results (p=0.432). Frequency of osteopenia was higher in males (56%) compared to females (36.6%) whereas, osteoporosis was higher in females (53.7%) than males (12%). Results showed statistically significant association (p>0.01) with gender and bone scan results. Conclusion: Almost half the young adults in our study classified for osteopenia. Lifestyle modification factors are hypothesized to play an important role towards this high frequency. Further studies should evaluate risk factors for osteopenia in younger population

Association of FTO variant with parental history of type 2 diabetes mellitus in adults

Objectives: To find out the association between fat mass and obesity-associated gene polymorphism and risk factors frequently associated with type 2 diabetes mellitus. Method: The case-control study was conducted January 2020 to March 2021 at the Ziauddin University, Karachi, and comprised deoxyribonucleic acid samples for fat mass and obesity-associated gene polymorphism from non-diabetic Pakistani population. Group A comprised non-diabetics with parental history of type 2 diabetes mellitus and Group B had controls without parental history of type 2 diabetes mellitus. Analysis was based on restriction fragment length polymorphism and polymerase chain reaction. Data was analysed using SPSS 25. Results: Of the 150 subjects, 75(50%) each were in Group A and Group B. There were 53.3% males and 46.7% females in Group A compared to 46.7% males and 53.3% females in Group B. Overall, 48% subjects were single and 52 % were married. A difference in frequency of fat mass and obesity-associated gene (rs9939609) alleles, such as TT, AA TA, was noted between the groups (p>0.999). TA allele was found to be associated with Group A (33) 44% (p=0.40), while TT allele was associated with Group B (41) 54% (p=0.414). AA allele was equally distributed between the groups (6) 8% (p=1.00). Conclusion: The TT allele of fat mass and obesity-associated gene was found to be an independent allele associated with the risk of developing type 2 diabetes mellitus. Key Words: Type 2 diabetes mellitus, FTO gene, BMI, SNP, Multifactorial inheritance, Family history, Genetic variant PCR and RFLP