Publications by authors named "Abid Azhar"

32 Publications

Comparative analysis of chicken cecal microbial diversity and taxonomic composition in response to dietary variation using 16S rRNA amplicon sequencing.

Mol Biol Rep 2021 Sep 24. Epub 2021 Sep 24.

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, 75270, Pakistan.

Background: Antibiotic resistance poses a grave threat to One-Health. By replacing antibiotics with non-antibiotic additives (are alternatives to antibiotics, ATAs) like phytogenic feed additives and organic acids in poultry feed. ATAs are a potential alternative as these decline the proliferation of pathogenic bacteria and strengthen gut function in broiler chickens. In this study, we use 16S rRNA amplicon sequencing of the V3-V4 region to evaluate phytogenic feed additives and organic acids on the cecal microbial diversity of broiler chickens.

Methods And Results: Two hundred & forty broiler chicks were divided into five treatments comprising: a controlled basal diet (CON), antibiotic group (AB), phytogenic feed additives (PHY), organic acids (ORG), and a combination of PHY + ORG (COM). A distinctive microbial community structure was observed amongst different treatments with increased microbial diversity in AB, ORG, and COM (p < 0.05). The synergistic effects of PHY and ORG increased bacterial population of phyla: Firmicutes, Bacteroides, and Proteobacteria in the cecum. The presence of species, Akkermansia muciniphila (involved in mucin degradation) and Bacillus safensis (a probiotic bacterium) were noticed in COM and PHY, respectively. Clustering analysis revealed a higher relative abundance of similar microbial community composition between AB and ORG groups.

Conclusions: Treatments with PHY and ORG modified the relative abundance and presence/absence of specific microbiota in the chicken cecum. Hence, cecal microbiota modulation through diet is a promising strategy to reduce cross-contamination of zoonotic poultry pathogens, led to healthy and economical broiler meat.
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http://dx.doi.org/10.1007/s11033-021-06712-3DOI Listing
September 2021

Growth performance, intestinal histomorphology, gut microflora and ghrelin gene expression analysis of broiler by supplementing natural growth promoters: A nutrigenomics approach.

Saudi J Biol Sci 2021 Jun 13;28(6):3438-3447. Epub 2021 Mar 13.

Department of Zoology, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.

In an epoch of escalating number of antibiotic-resistance bacteria, there is a dire need to develop efficient and novel feeding strategies for animal nutrition as alternatives to antibiotics. Here, implicating nutrigenomic approach, phytobiotics and organic acids were used to evaluate ghrelin gene expression levels, gut microflora composition, performance parameters and intestinal histomorphological changes in broiler chickens. One-day-old chicks (n = 315) were reared for 42 days and distributed randomly into five experimental groups; each with three replicates (21 birds per replicate). Experimental groups were control: basal diet only, antimicrobial growth promoter: 40 g/metric ton of basal diet (virginiamycin), organic acids: 4 kg/metric ton of basal diet, phytobiotics: 3 kg/metric ton of basal diet, combination: 7 kg/metric ton of basal diet (organic acids 4 kg and phytobiotics 3 kg metric ton of feed). Growth performance, histological and ghrelin gene expression analysis were executed on 21 and 42 days while, quantitative bacterial analysis of cecum and ileum was performed on day 42. Increased feed intake and body weight () were noticed in phytobiotics group. Addition of phytobiotics significantly improved () villus height and ratio of villus height/crypt depth in ileum, jejunum, and duodenum and down-regulated ghrelin gene expression levels. Total coliform and in cecal and ileal digesta were decreased significantly () in organic acids group. Correlation analysis revealed spp. were positively correlated to villus height/crypt depth ration in duodenum. The findings indicated the importance of gene-nutrient-microbiota interactions based on nutrigenomics approach. Hence, phytobiotics and organic acids might be suitable alternatives to antibiotics for improved performance and immunity, along with healthier meat production in poultry.
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http://dx.doi.org/10.1016/j.sjbs.2021.03.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8176037PMC
June 2021

Enhanced modulation of gut microbial dynamics affecting body weight in birds triggered by natural growth promoters administered in conventional feed.

Saudi J Biol Sci 2020 Oct 24;27(10):2747-2755. Epub 2020 Jun 24.

Karachi Institute of Biotechnology and Genetic Engineering, University of Karachi, Karachi, Pakistan.

This study explored the effects of natural growth promoters (phytogenic feed additives and organic acids) on animal performance, carcass characteristics, blood parameters, gut microflora composition, and microbe-host interactions in broiler chickens over a 42-day feeding period. Two-hundred-fifty-day-old chicks were randomly assigned to one of five treatments: (i) control diets (CON); (ii) control diets + 40 g/tons antibiotic growth promoter (AB); (iii) control diets + 3 kg/tons organic acids (ORG); (iv) control diets + 3 kg/tons phytogenic feed additives (PHY); (v) control diets + 3 kg/tons organic acids + phytogenic feed additive combination (COM). A non-significant differences ( > ) were observed in broiler performance among treatments at 21 days of age; however, a gradually increasing body weight gain and reduced feed conversion ratio were observed at 42 days in treatments versus control group. Biochemical indices were non-significant ( > ) except for decreased cholesterol ( < ) and increased A/G ratio ( < ) recorded in the treatment groups. The addition of PHY and ORG improved total counts of spp. and spp. ( < ) as well as reduced caecal and ileal spp. and ( < ). Correlation analysis elucidated beneficial bacteria ( spp. and spp.) were positively and pathogenic bacteria ( spp. and ) were negatively correlated ( < ) with host weight gain. The findings indicated that dietary supplementation of PHY and ORG sustained balanced gut microflora, which in turn improved body weight. This study broadens the significance of using PHY and ORG as safe alternatives to antibiotic growth promoters for achieving healthier and economical broiler production.
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http://dx.doi.org/10.1016/j.sjbs.2020.06.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7499368PMC
October 2020

Association between obesity and risk of knee osteoarthritis.

Pak J Pharm Sci 2020 Jan;33(1(Supplementary)):295-298

Dr. A. Q. Khan Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

The study was designed to investigate the association between obesity and the risk of knee osteoarthritis, recruiting 400 knee osteoarthritis patients and an equal number of controls. After the informed consent, diagnosed patients from Jinnah Post Graduate Medical Centre, Karachi were included as "cases". Age-matched individuals without the disease were included as "controls". Sociodemographic data were taken from each participant. Characteristics were compared by odds ratio and chi-square using SPSS 20 software. Obesity (OR 3.29; 95% CI 2.40-4.51), female gender (OR 2.87; 95% CI 1.94-4.25) and family history (OR 3.61; 95% CI 2.69-4.85) were found to be significantly associated with osteoarthritis (p<0.001). Highest OR was found in case of stair climbing >10 flights/d (OR 6.08; 95% CI 4.16-8.89; p<0.001), whereas heavy lifting (>25 kg/d for > 4 hr) was observed as another major factor with OR of 5.24 (95% CI 3.54-7.75; p<0.001) that elevates the risk. The study concluded that obesity is significantly associated with osteoarthritis and obese individuals (BMI>25 kg/m2) are at high risk of disease development. Furthermore, family history, prolonged standing (>2 h/d for >1 yr), heavy lifting (>25 kg/d for > 4 hr), stair climbing (>10 flights/d) and sitting on the floor (>5 h/d) might also be associated with knee osteoarthritis.
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January 2020

Relationship of Serum Leptin and Reproductive Hormones in Unexplained Infertile and Fertile Females.

Cureus 2019 Dec 31;11(12):e6524. Epub 2019 Dec 31.

Internal Medicine, King Abdulaziz University, Jeddah, SAU.

Objective: To investigate the relationship between serum leptin and reproductive hormones in females with unexplained infertility (UI).

Methodology: It was a case-control study conducted in the Gynecology and Obstetrics Department and Infertility Clinic, Jinnah Postgraduate Medical Center (JPMC), Karachi, Pakistan. A total of 235 primary infertile females with an unidentified cause of infertility were selected from the Infertility Clinics. The patients were excluded if they were found to have polycystic ovary syndrome, endometriosis, tubal blockage, irregular menstrual cycles, hyperthyroidism, hypothyroidism, hyperprolactinemia, hyperandrogenemia, fasting blood sugar >110 mg/dl, and male factor infertility. A total of 205 healthy, fertile females were selected from the general population. The blood samples of both groups were collected on the 12th and 21st day of their menstrual cycle. Serum leptin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), and E2 levels were measured. Statistical analysis was executed using the SPSS version 16 (SPSS Inc., Chicago, IL).

Results: No significant difference was observed in leptin values of fertile and UI females, 37.110±1.19 vs. 35.321±0.901. In the preovulatory phase (12th day) of the cycle, infertile subjects with body mass index (BMI) <20 and 20-24.9 had significantly higher values for leptin (p<0.05), whereas, with an increase in BMI, leptin levels were reduced in these females. Leptin was reduced further in the luteal phase of infertile females with BMI 25-30, with a significantly lower value for FSH (p<0.005), LH (p<0.005), and estradiol (p<0.005. In infertile subjects, it correlated with estradiol (r=0.501, p<0.005), BMI (r=0.903, p<0.001), and progesterone (r=0.146, p<0.05).

Conclusion: Low levels of leptin observed to have an increase in the BMI of UI females were associated with a reduced estradiol and progesterone production in the luteal phase of the cycle.
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http://dx.doi.org/10.7759/cureus.6524DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6991145PMC
December 2019

TNF gene promoter region polymorphisms and association with young-onset rheumatoid arthritis.

Pak J Pharm Sci 2019 Sep;32(5(Supplementary)):2295-2297

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Rheumatoid arthritis (RA) is an inflammatory autoimmune disease that shares a major global economic burden due to disabilities and mortality risk. It affects all age groups with a female predominance. Tumor Necrosis Factor (TNF) a proinflammatory cytokine is one of the key players in etiology of autoimmune diseases such as RA. TNF gene promoter polymorphisms predict disease susceptibility, severity and therapeutic response. Therefore, the current case-control study was designed to evaluate the possible association of TNF gene promoter polymorphisms (-238 and -308) with susceptibility to young-onset RA. The study involves 102 individuals (50 young-onset RA patients, 52 healthy individuals). Genomic DNA was extracted using a standard phenol-chloroform method followed by PCR-RFLP for the screening of TNF gene promoter polymorphisms (-238 and -308). The study resulted in the association of TNF -238G/A polymorphism with susceptibility to young-onset RA in the homozygous form GG (Odds Ratio = 3.23, p-value= <0.05), though no significant difference was observed for -308G/A polymorphism among young-onset RA patients and controls. Thus concludes; TNF -238/G/A contributes to the risk of susceptibility to young-onset RA, conversely, TNF -308 G/A protects against the disease. Consequently, the study has demonstrated a possible relationship of studied TNF polymorphism with young-onset RA.
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September 2019

Association analysis and allelic distribution of deletion in CC chemokine receptor 5 gene (CCR5Δ32) among breast cancer patients of Pakistan.

Mol Biol Rep 2019 Apr 8;46(2):2387-2394. Epub 2019 Mar 8.

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, 75270, Karachi, Sindh, Pakistan.

Chemokine CC receptor type 5 (CCR5) is a cell surface receptor that has high affinity for chemotropic cytokines called chemokines. The CCR5 gene contains a 32 base pairs (bp) deletion (CCR5Δ32). This deletion may result in a malformed and nonfunctional receptor, reported to be responsible for the development and dissemination of different cancers. CCR5Δ32 exists in two allelic forms i.e. deletion (D) and wild type (WT). This study aims to detect the role of CCR5Δ32 in breast cancer development. Blood samples were collected from breast cancer patients (330) and controls of same gender (306). Along with this histopathologically diagnosed malignant tissue samples were also excised from breast lesions of 100 patients. Genetic variations within the blood and tissue samples were examined by PCR then observed through gel electrophoresis and confirmed by direct DNA sequencing. Obtained DNA sequences were aligned and analyzed by MEGA6 software. Genotypic and association analyses were done by SPSS software version 17.0. Deletion of 32 bp in CCR5 gene has been analyzed. Genotypic variations of CCR5Δ32 are; homozygous wild type (WT/WT), heterozygous deletion (WT/D) and homozygous deletion (D/D). Statistical analyses of CCR5Δ32 data revealed that WT/D was significantly higher in blood samples of breast cancer patients (7.27% (24/330)) as compare to controls (1.30% (4/306)). In tumor tissue samples WT/WT being the most frequent genotype (99.00% (99/100)) with 1.00 (1/100) of D/D which suggested that it may be acquired. Hence, association analysis showed that CCR5Δ32 is positively associated with breast cancer in Pakistan (p < 0.001). The risk ratio of CCR5Δ32 was 5.6610 (95% confidence interval: 2.0377 to 15.7267) and odds ratio was calculated to be 6.0335 (95% confidence interval: 2.1288 to 17.0999) which signifies that deletion also increases the risk of breast cancer development. Moreover, association analyses also revealed that clinicopathological features do not have any impact on the CCR5Δ32 genotype of breast cancer. This suggests that deletion of 32 bp in CCR5 gene may be associated with breast cancer. CCR5 signals the activation and migration of immune cells at the site of tumor formation. Because of deletion; deformed CCR5 receptor might be unable to express and function properly which may subdue the immunity against cancer hence, leading to its progression.
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http://dx.doi.org/10.1007/s11033-019-04699-6DOI Listing
April 2019

Association and implications in triple negative and triple positive breast cancer: Relationship with sociodemographic and reproductive factors in Pakistan.

Pak J Med Sci 2018 Nov-Dec;34(6):1468-1472

Saima Saleem, Ph.D. The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Background & Objectives: Triple negative and triple positive breast cancer have adverse effects than other types of breast cancer. However, triple negative has poor prognosis with short survival as compared with triple positive breast cancer. Good prognosis is one of the key factors for successful treatment trial. This study aimed to find out the association of sociodemographic and reproductive features like parity, menopause, number of child bearing as risk factors in the development and prognosis of triple negative and triple positive breast cancer.

Methods: This study is a part of an ongoing project which is being conducted in Karachi from 2013 to 2020. Informed consent from triple negative breast cancer (n=134) and triple positive breast cancer (n=87) patients were taken prior to their recruitment into the study. Demographic, anthropometric, reproductive and disease history of patients were recorded. Means, frequency distribution, data classification and association analyses were done by SPSS version 17.0.

Results: Statistical analyses revealed that delayed first child bearing age and lower number of children are associated with the development of triple negative breast cancer. However, no significant effect of these parameters has been observed on the outcomes of triple positive breast cancer.

Conclusions: Reproductive factors have more pathological implications than sociodemographic factors in both triple positive and triple negative breast cancer development. These findings might prove to be beneficial for effective and better breast cancer management.
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http://dx.doi.org/10.12669/pjms.346.15763DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6290238PMC
December 2018

Glutathione S-Transferase M1 and T1 Gene Deletions and Susceptibility to Acute Lymphoblastic Leukemia (ALL) in adults.

Pak J Med Sci 2018 May-Jun;34(3):666-670

Abid Azhar, DSc, PhD. The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Objective: Biotransformation of xenobiotics are critical for their metabolism and removal from the body which is carried out by xenobiotic metabolizing enzymes. Individuals carrying variants of genes that encode these enzymes have an altered ability to metabolize xenobiotics which may lead to an increased risk of acute lymphoblastic leukemia. The current study aimed to investigate the impact of and gene deletions in causing predisposition to adult ALL.

Methods: The current case-control study involved 62 adult ALL patients and 62 age and gender matched healthy controls. Whole blood samples processed with standard phenol chloroform protocol for DNA isolation were genotyped using multiplex PCR approach for simultaneous identification of and deletions. The genotype frequency obtained for patients was compared to controls using odds ratio and chi-square.

Results: The null genotype frequency of and in a group of adult ALL patients from Pakistan were 47% and 11% respectively. Deletion of and did not show statistically significant association with adult ALL (=0.86 and =0.35 respectively). The combined deletion was observed in 2% patients and was not significantly associated with ALL in adults (=0.85).

Conclusions: The results reveal that homozygous null polymorphism of and genes does not influence ALL susceptibility among adult patients. Cancer susceptibility associated with polymorphism varies with ethnic and geographic differences. Therefore, further investigation on different populations is needed to understand the role of these genetic variations in modifying adult ALL risk.
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http://dx.doi.org/10.12669/pjms.343.14911DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6041525PMC
July 2018

Vitamin D Status and Its Associated Risk Factors among Adults in the Southwest Region of Cameroon.

J Nutr Metab 2018 19;2018:4742574. Epub 2018 Mar 19.

Faculty of Science, University of Buea, Buea, Cameroon.

Background: Vitamin D has been shown to exert its actions on the musculoskeletal, gastrointestinal, prostate, renal, endocrine, immune, and cardiovascular systems. Current reported data of hypovitaminosis D reveals a global pandemic, with an estimated one billion people worldwide presenting with hypovitaminosis D.

Objective: This study aimed at investigating the vitamin D status and its associated risk factors in Cameroonians from the South West Region.

Method: The study was a community- and hospital-based prospective longitudinal study. It was carried out during the dry and rainy seasons between the months of July and December 2015 in the South West Region of Cameroon involving 372 participants aged 35 years and above. After obtaining informed consent, a structured questionnaire was used to capture demographic data and risk factors of vitamin D deficiency. Blood samples were collected from the volunteer participants in the peak months of the rainy season and dry season, and the serum used to analyse for vitamin D by ELISA and calcium by spectrophotometry. 25(OH)D levels ≥75 nmol/L (≥30 ng/mL) were considered sufficient while levels <75 nmol/L were considered as hypovitaminosis D (insufficiency/deficiency).

Results: Hypovitaminosis D (deficiency/insufficiency) was prevalent in 25.8% (96) of the study population, with only 3.2% (12) deficiency and 22.6% (84) insufficiency. There was a significant inverse relationship (=-0.119, =0.02) between age and 25(OH)D levels; however, this relationship was not significant when controlled for gender, number of hours spent outdoors, and percentage of body covered. Gender, ethnic origin, percentage of body covered, time spent outdoors, and season did not influence serum vitamin D levels.

Conclusion: Results of this study suggest that the prevalence of hypovitaminosis D is relatively low in this study population and only age is a risk factor of vitamin D deficiency.
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http://dx.doi.org/10.1155/2018/4742574DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5884209PMC
March 2018

A Study on Gene Promoter Polymorphism in Healthy Pakistani population.

Pak J Med Sci 2017 Nov-Dec;33(6):1521-1524

Abid Azhar, PhD.Director General, The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University Of Karachi, Karachi, Pakistan.

Background & Objective: Catalase (CAT) is an important endogenous antioxidant enzyme that detoxifies HO into water and oxygen, consequently limiting the deleterious effects of reactive oxygen species. It has suggested that OMIM: gene promoter polymorphism is predominantly associated with different human disorders such as hypertension, cancers, diabetes, nephropathy, and other diseases accompanied by oxidative stress. This study was designed to investigate the prevalence of mutant T allele frequency in healthy individuals.

Methods: The study group consisted of 110 healthy individuals were enrolled from Baqai Institute of Diabetology and Endocrinology (BIDE), Karachi, Pakistan, during the period of April 2010 to May 2013. DNA was isolated from leukocytes. Genotyping of gene promoter polymorphism was carried out using thermal cycler followed by RFLP. Blast N analysis was performed for the confirmation of gene sequences.

Results: In gene promoter polymorphism, wild type genotype (AA) was observed in 18.26% and alterered genotype (AT/TT) found in 81.74% cases.

Conclusions: Data demonstrates that frequency and distribution of mutant T allele was more prevalent as compared to wild type A allele in the study group.
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http://dx.doi.org/10.12669/pjms.336.13188DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5768856PMC
March 2018

Linear Energy Transfer Modulates Radiation-Induced NF-kappa B Activation and Expression of its Downstream Target Genes.

Radiat Res 2018 04 25;189(4):354-370. Epub 2018 Jan 25.

a   German Aerospace Centre (DLR), Institute of Aerospace Medicine, Radiation Biology Department, Linder Höhe, D-51147 Köln, Germany.

Nuclear factor kappaB (NF-κB) is a central transcription factor in the immune system and modulates cell survival in response to radiotherapy. Activation of NF-κB was shown to be an early step in the cellular response to ultraviolet A (UVA) and ionizing radiation exposure in human cells. NF-κB activation by the genotoxic stress-dependent sub-pathway after exposure to different radiation qualities had been evaluated to a very limited extent. In addition, the resulting gene expression profile, which shapes the cellular and tissue response, is unknown. Therefore, in this study the activation of NF-κB after exposure to low- and high-linear energy transfer (LET) radiation and the expression of its target genes were analyzed in human embryonic kidney (HEK) cells. The activation of NF-κB via canonical and genotoxic stress-induced pathways was visualized by the cell line HEK-pNF-κB-d2EGFP/Neo L2 carrying the destabilized enhanced green fluorescent protein (d2EGFP) as reporter. The NF-κB-dependent d2EGFP expression after irradiation with X rays and heavy ions was evaluated by flow cytometry. Because of differences in the extent of NF-κB activation after irradiation with X rays (significant NF-κB activation for doses >4 Gy) and heavy ions (significant NF-κB activation at doses as low as 1 Gy), it was expected that radiation quality (LET) played an important role in the cellular radiation response. In addition, the relative biological effectiveness (RBE) of NF-κB activation and reduction of cellular survival were compared for heavy ions having a broad LET range (∼0.3-9,674 keV/μm). Furthermore, the effect of LET on NF-κB target gene expression was analyzed by real-time reverse transcriptase quantitative PCR (RT-qPCR). The maximal RBE for NF-κB activation and cell killing occurred at an LET value of 80 and 175 keV/μm, respectively. There was a dose-dependent increase in expression of NF-κB target genes NF-κB1A and CXCL8. A qPCR array of 84 NF-κB target genes revealed that TNF and a set of CXCL genes (CXCL1, CXCL2, CXCL8, CXCL10), CCL2, VCAM1, CD83, NF-κB1, NF-κB2 and NF-κBIA were strongly upregulated after exposure to X rays and neon ions (LET 92 keV/μm). After heavy-ion irradiations, it was noted that the expression of NF-κB target genes such as chemokines and CD83 was highest at an LET value that coincided with the LET resulting in maximal NF-κB activation, whereas expression of the NF-κB inhibitory gene NFKBIA was induced transiently by all radiation qualities investigated. Taken together, these findings clearly demonstrate that NF-κB activation and NF-κB-dependent gene expression by heavy ions are highest in the LET range of ∼50-200 keV/μm. The upregulated chemokines and cytokines (CXCL1, CXCL2, CXCL10, CXCL8/IL-8 and TNF) could be important for cell-cell communication among hit as well as nonhit cells (bystander effect).
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http://dx.doi.org/10.1667/RR14905.1DOI Listing
April 2018

Mutations in Hnrnpa1 cause congenital heart defects.

JCI Insight 2018 01 25;3(2). Epub 2018 Jan 25.

School of Biomedical Sciences, Joint Laboratories of Matrix Biology and Diseases, The University of Hong Kong, Hong Kong, China.

Incomplete penetrance of congenital heart defects (CHDs) was observed in a mouse model. We hypothesized that the contribution of a major genetic locus modulates the manifestation of the CHDs. After genome-wide linkage mapping, fine mapping, and high-throughput targeted sequencing, a recessive frameshift mutation of the heterogeneous nuclear ribonucleoprotein A1 (Hnrnpa1) gene was confirmed (Hnrnpa1ct). Hnrnpa1 was expressed in both the first heart field (FHF) and second heart field (SHF) at the cardiac crescent stage but was only maintained in SHF progenitors after heart tube formation. Hnrnpa1ct/ct homozygous mutants displayed complete CHD penetrance, including truncated and incomplete looped heart tube at E9.5, ventricular septal defect (VSD) and persistent truncus arteriosus (PTA) at E13.5, and VSD and double outlet right ventricle at P0. Impaired development of the dorsal mesocardium and sinoatrial node progenitors was also observed. Loss of Hnrnpa1 expression leads to dysregulation of cardiac transcription networks and multiple signaling pathways, including BMP, FGF, and Notch in the SHF. Finally, two rare heterozygous mutations of HNRNPA1 were detected in human CHDs. These findings suggest a role of Hnrnpa1 in embryonic heart development in mice and humans.
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http://dx.doi.org/10.1172/jci.insight.98555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5821217PMC
January 2018

Association of single Nucleotide Missence Polymorphism Val109Asp of Omentin-1 gene and coronary artery disease in Pakistani population: Multicenter study.

Pak J Med Sci 2017 Sep-Oct;33(5):1128-1133

Dr. Abid Azhar, Ph.D. Director General, Dr. A. Q. Khan Institute of Biotechnology and Genetic Engineering (KIBGE), University Of Karachi, Karachi, Pakistan.

Background & Objective: Coronary artery disease (CAD) is a most important cause of morbidity and mortality worldwide as well as in Pakistan. Recent studies have shown that the combination of obesity, insulin resistance and fluctuation in circulating adipocytokines levels is associated with the pathogenesis of coronary artery disease. Omentin-1 is recently found adipocytokine that is highly expressed in visceral adipose tissue. It has anti- inflammatory properties and is negatively correlated with ischemic heart disease. Therefore, this study was designed to investigate the relationship between omentin-1 Val109Asp polymorphism and CAD in Pakistani population.

Methods: A total of 350 subjects were included in the study. Two hundred fifty were diagnosed with coronary artery disease while 100 served as healthy controls. PCR-RFLP was performed at Dr. A Q. Khan Institute of Biotechnology (KIBGE) to analyze Val109Asp polymorphism. In this, case control study SPSS software version 16 (Chicago, IL, USA) was used for data analysis. Continuous variables and categorical variables were presented as mean±SD or in percentage. Independent sample test and chi-square test was performed to compare the differences in means between cases and controls. Genotype distribution was analyzed by chi-square test and results were presented as percentage and frequency. Multivarible regression analysis indicated that Val109Asp SNP might be an independent risk factor for CAD susceptibility after adjustment for some well- known CAD risk factors including age, gender, body mass index, smoking, hypertension, diabetes mellitus and lipid abnormalities. There was estimation of odd ratios (OR) and 95% confidence intervals (CIs) to determine the correlation between genotypes and the risk of CAD. (p> 0.05). Genotype frequencies were compared by Chi-square test.

Results: There was prevalence of Omentin-1 Val109Asp polymorphism in both case and control groups. However, Val/Asp (heterozygous mutant) genotype was detected more frequently in patients with CAD, OR(95%)=1.921; CI=1.173-3.1469 in comparison of Asp/Asp and Val/Val genotypes.

Conclusion: Individuals having Val/Asp heterozygous gemotype of omentin-1 gene polymorphism are at more risk of developing CAD in Pakistani population, further studies are required in different populations and ethnicities to confirm our findings.
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http://dx.doi.org/10.12669/pjms.335.13110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5673720PMC
November 2017

New anthrarobin acyl derivatives as butyrylcholinesterase inhibitors: synthesis, and studies.

Heliyon 2017 Jul 10;3(7):e00350. Epub 2017 Jul 10.

Department of Pharmacognocy, College of Pharmacy, King Saud University, PO Box 2457, Riyadh11451, Saudi Arabia.

To treat Alzheimer's disease (AD), the available candidates are effective only against mild AD or have side effects. So, a study was planned to synthesis new candidates that may have good potential to treat AD. A series of new anthrarobin acyl derivatives (-) were synthesized by the reaction of anthrarobin () and acetic anhydride/acyl chlorides. The product were characterized by H NMR and EI-MS, and evaluated for butyrylcholinesterase (BuChE) inhibition activity. Compounds and showed notable BuChE inhibitory potential with IC 5.3 ± 1.23 and 17.2 ± 0.47 μM, respectively when compared with the standard eserine (IC 7.8 ± 0.27 μM), compound showed potent BuChE inhibition potential than the standard eserine. The active compounds and have acyl groups at 2-OH and 10-OH positions which may be responsible for inhibitory potential as this orientation is absent in other products. studies of and products revealed the high inhibitory potential due to stable binding of ligand with the BuChE active sites with docking energy score -18.8779 kcal/mol and -23.1159 kcal/mol, respectively. Subsequently, compound that have potent BuChE inhibitory activity could be the potential candidate for drug development for Alzheimer's disease.
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http://dx.doi.org/10.1016/j.heliyon.2017.e00350DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5506883PMC
July 2017

Cheminformatics-Based Anticoagulant Study of Traditionally Used Medicinal Plants

Iran Biomed J 2017 11 29;21(6):400-5. Epub 2017 Apr 29.

Department of Clinical Research, Pasteur Institute of Iran, Tehran 13164, Iran.

Backgroung: Medicinal plants, as a complementary medicine, have been used to treat various diseases since ancient times. These plants have numerous beneficial applications and are the source of certain conventional drugs. In diseases such as stroke and ischemia, which are caused by several factors, abnormal coagulation is an important causative factor. Accordingly, novel and effective therapies such as herbal remedies should be practiced to prevent such lethal diseases.

Methods: Using the available databases such as Google Scholar and PubMed, the previously reported anticoagulant compounds and plants possessing anticoagulant activity were identified and collected in two separate lists. Next, the fast and cost-effective cheminformatics methods incorporated in PubChem were applied to detect some compounds similar to reported anticoagulants. Subsequently, 15 native medical plants of Iran containing the potential anticoagulants were selected. The selected plants were purchased and chopped, and the potential compounds were extracted by ethanol. Then three concentrations of extracts (1, 10, and 100 µg per ml) were made. Finally, anticoagulant effect of the selected plants was evaluated by in vitro prothrombin time and activated partial thromboplastin time coagulation tests.

Results: Among the 15 selected medicinal plants, three plants, including Terminalia bellirica (P=0.0019), Astragalus arbusculinus (P=0.0021), and Origanum vulgare (P=0.0014) showed a more promising anticoagulant effect in comparison to the control.

Conclusion: The anticoagulant activity was identified for the first time in these three plants. Further in vivo study and mechanism of action assay are required to be performed on these three plants, which could be suitable candidates for use as natural anticoagulant medicines.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5572436PMC
http://dx.doi.org/10.18869/acadpub.ibj.21.6.400DOI Listing
November 2017

SLC11A1 polymorphisms and host susceptibility to cutaneous leishmaniasis in Pakistan.

Parasit Vectors 2017 01 7;10(1):12. Epub 2017 Jan 7.

Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi, 75270, Pakistan.

Background: The vector-borne cutaneous leishmaniasis (CL) is endemic in several regions of Pakistan mainly affecting poor populations. Host genetic factors, particularly SLC11A1 (solute carrier transmembrane protein) within macrophages, play a crucial role in disease pathology and susceptibility. Association of SLC11A1 with cutaneous leishmaniasis, a neglected tropical disease, is not well established. Inconsistencies have been observed within different populations worldwide with respect to genetic susceptibility. This study was designed to investigate genetic variation(s) in SLC11A1 and to assess possible association with cutaneous leishmaniasis in Pakistan.

Results: Eight polymorphisms (rs2276631, rs3731864, rs2290708, rs2695342, rs201565523, rs17215556, rs17235409, rs17235416) were genotyped across SLC11A1 in 274 patients and 119 healthy controls. Six polymorphisms were studied by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequencing. Two single nucleotide polymorphisms were analyzed with newly designed semi-nested PCR assays. Case-control analysis showed no association between selected polymorphisms in SLC11A1 and cutaneous leishmaniasis. No significant difference was observed in the distribution of alleles between leishmaniasis patients and healthy individuals. Strong pairwise linkage disequilibrium was observed between rs2276631 and rs2290708 (r  = 64); and rs17235409 and rs17235416 (r  = 78).

Conclusions: This study shows that genetic variations in the candidate gene SLC11A1 do not affect susceptibility to cutaneous leishmaniasis in the sample population from Pakistan.
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http://dx.doi.org/10.1186/s13071-016-1934-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5219684PMC
January 2017

CC chemokine receptor 5 Δ32 polymorphism: association analysis and allele distribution among cutaneous leishmaniasis patients from Pakistan.

J Cutan Pathol 2016 Jul 6;43(7):564-70. Epub 2016 Apr 6.

The Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Background: Human immunodeficiency virus (HIV)/leishmaniasis coinfection is a matter of deep concern worldwide. CC chemokine receptor 5 (CCR5) functions as a co-receptor for HIV entry into host immune cells with an elevated expression observed during leishmaniasis, promoting parasite persistence. A 32 bp deletion (Δ32) in the CCR5 gene provides protection against HIV infection and increased resistance to Leishmania infection.

Methods: In this study, CCR5-Δ32 distribution within Pakistani population with cutaneous leishmaniasis was investigated to evaluate genetic susceptibility to HIV infection. CCR5-Δ32 polymorphism was analyzed in 276 leishmaniasis patients and 119 uninfected healthy controls. Genotypic and allelic frequencies were evaluated and tested for Hardy-Weinberg equilibrium (HWE).

Results: The overall Δ32 allele frequency was 6.58% of the population (n = 395). There was a significant difference (p < 0.05) in the geographical distribution of Δ32 allele which was higher in the northern region of the country when compared with the south. Five individuals were identified to be homozygous for the Δ32 allele which has not been reported before from Pakistan. However, no significant association was observed between CCR5-Δ32 and cutaneous leishmaniasis.

Conclusion: The higher frequency of CCR5 wild-type allele among leishmaniasis patients may suggest an increased risk of HIV infection and also support its facilitative role in Leishmania infection.
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http://dx.doi.org/10.1111/cup.12712DOI Listing
July 2016

The association of GSTM1 and GSTT1 polymorphisms with squamous cell carcinoma of cervix in Pakistan.

Tumour Biol 2015 Jul 9;36(7):5195-9. Epub 2015 Feb 9.

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan,

Gene deletions in GSTM1 and GSTT1 may result in tempering the activation and detoxification of several carcinogens and thereby may increase the risk of cancer pre-disposition. This study aims to investigate the clinical impact of glutathione-S-transferase GSTM1 and GSTT1 polymorphisms on squamous cell carcinoma of cervix (SCCA).The GSTM1 and GSTT1 polymorphisms were analyzed in cervical cancer patients and healthy controls. Touch down multiplex polymerase chain reaction (PCR) strategy was adopted for genotyping of GSTM1 and GSTT1 polymorphisms. The null genotype of GSTM1 exhibited a significantly higher percentage in patients with SCCA (74 %) than in the control group (34.0 %). However, no significant difference was observed in the null genotype of GSTT1 among SCC patients and healthy subjects, respectively. GSTM1 exhibited a significant association with increased risk of squamous cell carcinoma (p < 0.001). The odds ratio for the GSTM1 null genotype was also calculated (odds ratio 3.7484; 95 % confidence interval 1.6562-84834). This suggests that GSTM1 null genotype in cervical cell samples may be associated with more severe precancerous lesions of the cervix.
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http://dx.doi.org/10.1007/s13277-015-3175-yDOI Listing
July 2015

Fresh and aged human lymphocyte metaphase slides are equally usable for GTG banding.

Pak J Pharm Sci 2014 Sep;27(5):1255-9

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

The identification of chromosomes for routine cytogenetic analysis is based on quality of metaphases and good banding pattern. Fresh slides of human lymphocytes have been shown to produce good bands for the identification of chromosomes morphology. G-bands by Trypsin using Giemsa (GTG) banding of aged slides is generally considered hard to get desired band pattern of chromosomes persistently. The current study is focused on GTG banding of aged slides. A total of 340 subjects including 290 primary infertile and 50 fertile were selected. The blood samples were drawn aseptically for cytogenetic analysis. Lymphocytes were cultured and GTG banding was done on 1440 glass slides. Giemsa trypsin banding of aged slides were done by adjusting average trypsin time for each month according to the slide age and metaphase concentration. Correlation analyses showed a significant and positive correlation between slide ageing and trypsin pre-treatment time. The results of this study suggest that, the fresh and aged human lymphocyte metaphases are equally usable for GTG banding.
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September 2014

Genetic variants of ACE (Insertion/Deletion) and AGT (M268T) genes in patients with diabetes and nephropathy.

J Renin Angiotensin Aldosterone Syst 2014 Jun 15;15(2):124-30. Epub 2014 Apr 15.

The Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Introduction: Diabetes mellitus (DM) has been a growing epidemic worldwide and poses a major socio-economic challenge. The leading cause of DM death is nephropathy due to end-stage renal disease (ESRD). This study aims to identify the possible association of I/D variants of the ACE gene and M268T (rs699) of the AGT gene of renin-angiotensin-aldosterone system (RAAS).

Materials And Methods: Study subjects include 115 patients with DM, 110 with diabetic nephropathy (DN) and 110 controls. Fasting blood samples were collected for biochemical analyses and PCR amplification of specific regions of the ACE and AGT genes using primers.

Results: The distribution of ACE (I/D) II 28.8%, ID 35.6% and DD 35.6% while in DN II 24.5%, ID 41% and DD 34.5%. The AGT (M268T) genotypes were distributed in DM as TT 30.4%, MT 66.9% and MM 2.6% while in DN subjects TT 56.4%, MT 42.7% and MM 0.9%.

Conclusion: Significant differences were observed in the DD genotype and D allele of the ACE gene and the TT genotype and T allele of AGT genes between diabetic patients with and without nephropathy. The study may conclude that the D allele polymorphism in the ACE gene and the T allele polymorphism in AGT gene may be considered as genetic risk factors for the development of nephropathy in diabetes.
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http://dx.doi.org/10.1177/1470320313512390DOI Listing
June 2014

Protein expression profiling of nuclear membrane protein reveals potential biomarker of human hepatocellular carcinoma.

Clin Proteomics 2013 Jun 1;10(1). Epub 2013 Jun 1.

Neurochemistry Research Unit Laboratory, Department of Biochemistry, University of Karachi, Karachi, Pakistan.

Background: Complex molecular events lead to development and progression of liver cirrhosis to HCC. Differentially expressed nuclear membrane associated proteins are responsible for the functional and structural alteration during the progression from cirrhosis to carcinoma. Although alterations/ post translational modifications in protein expression have been extensively quantified, complementary analysis of nuclear membrane proteome changes have been limited. Deciphering the molecular mechanism that differentiate between normal and disease state may lead to identification of biomarkers for carcinoma.

Results: Many proteins displayed differential expression when nuclear membrane proteome of hepatocellular carcinoma (HCC), fibrotic liver, and HepG2 cell line were assessed using 2-DE and ESI-Q-TOF MS/MS. From the down regulated set in HCC, we have identified for the first time a 15 KDa cytochrome b5A (CYB5A), ATP synthase subunit delta (ATPD) and Hemoglobin subunit beta (HBB) with 11, 5 and 22 peptide matches respectively. Furthermore, nitrosylation studies with S-nitrosocysteine followed by immunoblotting with anti SNO-cysteine demonstrated a novel and biologically relevant post translational modification of thiols of CYB5A in HCC specimens only. Immunofluorescence images demonstrated increased protein S-nitrosylation signals in the tumor cells and fibrotic region of HCC tissues. The two other nuclear membrane proteins which were only found to be nitrosylated in case of HCC were up regulated ATP synthase subunit beta (ATPB) and down regulated HBB. The decrease in expression of CYB5A in HCC suggests their possible role in disease progression. Further insight of the functional association of the identified proteins was obtained through KEGG/ REACTOME pathway analysis databases. String 8.3 interaction network shows strong interactions with proteins at high confidence score, which is helpful in characterization of functional abnormalities that may be a causative factor of liver pathology.

Conclusion: These findings may have broader implications for understanding the mechanism of development of carcinoma. However, large scale studies will be required for further verification of their critical role in development and progression of HCC.
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http://dx.doi.org/10.1186/1559-0275-10-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3691657PMC
June 2013

P53 (Pro72Arg) polymorphism associated with the risk of oral squamous cell carcinoma in gutka, niswar and manpuri addicted patients of Pakistan.

Oral Oncol 2013 Aug 14;49(8):818-23. Epub 2013 May 14.

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Objectives: The chewing habit of paan, chhaliya, and tobacco is common in the traditional culture of Pakistan. Currently, niswar, gutka and manpuri are also commercially available in the Pakistani market. Epidemiologic evidences and increased rate of oral squamous cell carcinoma (OSCC) cases may indicate a direct relationship of these chewing habits with oral carcinogenesis. The p53 gene has been known to be a tumor suppressor gene that is found mutated in common human cancers. The p53 gene contains a single nucleotide polymorphism at codon 72 of exon 4 which encodes either proline (Pro) or arginine (Arg). The aim of the present study was to investigate association of p53 gene codon 72 polymorphism with patients of oral squamous cell carcinoma consuming these carcinogenic chewable materials.

Materials And Methods: Blood and tissue samples of 260 OSCC patients were collected with informed consent from the local hospitals of Karachi. The patients were compared with controls of similar age and sex. The exon 4 of p53 gene was examined by PCR-SSCP. The tumor samples showing mobility shift were purified and sequenced.

Results: The C>G missense mutation at nucleotide position 215 of the coding sequence was identified which substitutes proline with arginine at codon 72 of p53 protein. When the data for CCC72CGC polymorphism was analyzed statistically, a significant difference was observed between OSCC and control samples. The Pro allelic frequencies were significantly higher in OSCC patients as compare to controls. The current study indicated the Pro form of p53 codon 72 increases the risk of developing OSCC in Pakistani population. The risk ratio for Pro allele was 1.5004 (95% confidence interval: 1.2559 to 1.7924) and odds ratio of Pro allele was 2.389 (95% confidence interval: 1.5591 to 2.8137) in comparison with the Arg and Pro alleles in the OSCC group.

Conclusion: These evidences suggest that there may be specific genetic targets with these chewing ingredients that are responsible for causing OSCC. The p53 codon 72 polymorphism is associated with OSCC at somatic cell level but the polymorphism was not associated at inherited level.
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http://dx.doi.org/10.1016/j.oraloncology.2013.04.004DOI Listing
August 2013

Phylogenetic analysis of HDV isolates from HBsAg positive patients in Karachi, Pakistan.

Virol J 2012 Aug 15;9:162. Epub 2012 Aug 15.

The Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi, Pakistan.

Background: In spite of a high occurrence of Hepatitis Delta in the province of Sindh in Pakistan, no genetic study of Hepatitis Delta virus (HDV) isolates from this region was carried out. The aim of this study is to analyze the genetic proximity within local HDV strains, and relationship with other clades of HDV, using phylogenetic analysis.

Results: Phylogenetic analysis of nucleotide sequences of the Hepatitis Delta Antigen (HDAg) R0 region obtained in this study, showed considerable diversity among the local strains with a potential subgroup formation within clade I. The multiple sequence alignment of predicted amino acids within clade I showed many uncommon amino acid substitutions within some conserved regions that are crucial for replication and assembly of HDV.

Conclusions: The studied strains showed a range of genetic diversity within HDV clade I. There is clustering of sequences into more than one group, along with formation of potential subgroup within clade I. Clustering shows the genetic closeness of strains and indicates a common origin of spread of HDV infection. Further phylogeny-based studies may provide more information about subgroup formation within clade I and may be used as an effective tool in checking and/or preventing the spread of hepatitis D virus infection in this region.
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http://dx.doi.org/10.1186/1743-422X-9-162DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3493343PMC
August 2012

Protective effects of salivary factors in dental caries in diabetic patients of Pakistan.

Exp Diabetes Res 2012 24;2012:947304. Epub 2012 Jun 24.

Department of Biochemistry, Liaquat College of Medicine and Dentistry, Karachi 75290, Pakistan.

Salivary factors have been studied for their effects on the process of dental caries in patients of diabetes mellitus type 2. In this study, protective role of salivary pH, salivary flow rate, and salivary calcium is assessed in the patients of diabetes mellitus type 2 with dental caries. The samples of saliva were collected from 400 patients of diabetes mellitus type 2 and 300 age- and sex- matched controls after getting informed consent. All the subjects were classified into four groups according to age. The severity of dental caries was counted by decayed, missed, and filled teeth (DMFT) score. The salivary pH, flow rate, and calcium levels were found to be low in patients as compared to controls. The levels of fasting blood sugar, HbA1c, and DMFT score were found to be significantly high in patients than controls. The glycemic factors were significantly correlated with salivary factors indicating their influence on progression of caries in diabetes. On the basis of findings, it is concluded that the suitable salivary pH and flow rate may be regarded as main protective factors against dental caries in diabetes. Optimum level of salivary calcium may be responsible for continuous supply of calcium to arrest the demineralization and help reduce the occurrence of dental caries.
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http://dx.doi.org/10.1155/2012/947304DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3388590PMC
October 2012

Continuous Production of Dextran from Immobilized Cells of Leuconostoc mesenteroides KIBGE HA1 Using Acrylamide as a Support.

Indian J Microbiol 2011 Jul 26;51(3):279-82. Epub 2011 Jan 26.

The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, 75270 Pakistan.

The cells of L. mesenteroides KIBGE HA1 were immobilized for the production of dextran on acrylamide gel and gel concentration was optimized for maximum entrapment. Sucrose at substrate concentration of 10% produced high yield of dextran at 25°C with a percent conversion of 5.82 while at 35°C it was 3.5. However, increasing levels of sucrose diminished dextran yields. The free cells stopped producing dextran after 144 h, while immobilized cells continued to produce dextran even after 480 h. Molecular mass distribution of dextran from free cells indicate that it is identical to that of blue dextran while the molecular mass of dextran from immobilized cells is lower than that of free cells.
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http://dx.doi.org/10.1007/s12088-011-0130-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3209923PMC
July 2011

Homozygous frame shift mutation in ECM1 gene in two siblings with lipoid proteinosis.

J Dermatol Case Rep 2010 Dec;4(4):66-70

Department of Dermatology, Jinnah Postgraduate Medical Center, Karachi, Pakistan.

Background: The extracellular matrix protein 1 (ECM1) is a glycoprotein, expressed in skin and other tissues. Loss-of-function mutation in ECM1 causes a rare autosomal recessive disorder called lipoid proteinosis. Lipoid proteinosis is presented by varying degrees of skin scars, beaded papules along the eyelid margins, variable signs of hoarseness of voice and respiratory disorders. More than 250 cases of this disorder have been described in the literature, but occurrence of lipoid proteinosis in siblings is very rare. This study was designed to investigate the possible mutation causing lipoid proteinosis in a Pakistani family and to elaborate the scope of possible genetic changes, causing the genodermatosis in Pakistan.

Main Observations: In this study, two siblings (12 and 9-years sisters) were presented with scaly itchy lesions on whole body, hoarse voice and macroglossia. Their deceased father had similar clinical manifestations but mother and younger brother were unaffected. Blood samples from clinically affected and unaffected family members were collected with informed consent. The coding region of ECM1 gene containing 10 exons were amplified and sequenced. Both the affected siblings were shown to have homozygous frame shift mutation by deletion of the nucleotide T at 507, codon 169, exon 6. This resulted in a frame shift from codon 169 and appearance of a premature stop codon at 177, causing formation of a mutated protein (176 amino acids) instead of normal ECM1 protein (540 amino acids).

Conclusion: A case of homozygous 62-bp insertion in ECM1 gene causing lipoid proteinosis has been reported in another Pakistani family. The current study presents a homozygous frame shift mutation supporting an unusual function of ECM1 protein and broadens the spectrum of disease-linked mutations in this rare case of genodermatosis in this region.
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http://dx.doi.org/10.3315/jdcr.2010.1056DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3157823PMC
December 2010

Purification and characterization of novel α-amylase from Bacillus subtilis KIBGE HAS.

AAPS PharmSciTech 2011 Mar 14;12(1):255-61. Epub 2011 Jan 14.

Pharmaceutical Research Center, PCSIR Laboratories Complex, Karachi, Pakistan.

Purification of extracellular α-amylase from Bacillus subtilis KIBGE HAS was carried out by ultrafiltration, ammonium sulfate precipitation and gel filtration chromatography. The enzyme was purified to homogeneity with 96.3-fold purification with specific activity of 13011 U/mg. The molecular weight of purified α-amylase was found to be 56,000 Da by SDS-PAGE. Characteristics of extracellular α-amylase showed that the enzyme had a Km and V (max) value of 2.68 mg/ml and 1773 U/ml, respectively. The optimum activity was observed at pH 7.5 in 0.1 M phosphate buffer at 50 °C. The amino acid composition of the enzyme showed that the enzyme is rich in neutral/non polar amino acids and less in acidic/polar and basic amino acids. The N-terminal protein sequence of 10 residues was found to be as Ser-Ser-Asn-Lys-Leu-Thr-Thr-Ser-Trp-Gly (S-S-N-K-L-T-T-S-W-G). Furthermore, the protein was not N-terminally blocked. The sequence of α-amylase from B. subtilis KIBGE HAS was a novel sequence and showed no homology to other reported α-amylases from Bacillus strain.
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http://dx.doi.org/10.1208/s12249-011-9586-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3066380PMC
March 2011

Dental caries in diabetes mellitus: role of salivary flow rate and minerals.

J Diabetes Complications 2011 May-Jun;25(3):183-6. Epub 2010 Aug 30.

Department of Biochemistry, Liaquat College of Medicine & Dentistry, Karachi, Pakistan.

This study was designed to evaluate the possible protective role of salivary factors like salivary flow rate and adequate level of calcium, phosphate, and fluoride in diabetes mellitus type 2 patients with dental caries. A total of 398 diabetes mellitus type 2 patients with dental caries and 395 age- and sex-matched non-diabetic subjects with dental caries were included as controls, all of whom gave informed consent. All subjects were divided into four groups according to their age. Decayed, missed, and filled teeth (DMFT) were scored to indicate the severity of dental caries. Saliva was collected, flow rate was noted, and calcium, phosphate, and fluoride were analyzed. The blood glucose, HbA1c, and DMFT indices were found to be significantly high in diabetic patients as compared to controls. The salivary flow rate, calcium, phosphate, and fluoride were found to be significantly low whereas no significant difference was found in salivary magnesium in patients as compared to controls. Optimum salivary flow rate is responsible for establishing protective environment against dental caries. Adequate level of salivary calcium, phosphate, and fluoride is also involved in significant deposition of these minerals in plaque, which greatly reduces the development of caries in the adjacent enamel of teeth.
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http://dx.doi.org/10.1016/j.jdiacomp.2010.07.001DOI Listing
August 2011

Partial purification and some properties of alpha-amylase from Bacillus subtilis KIBGE-HAS.

Indian J Biochem Biophys 2009 Oct;46(5):401-4

Pharmaceutical Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.

An extracellular alpha-amylase from Bacillus subtilis KIBGE-HAS was partially purified by ultrafiltration and ammonium sulphate precipitation with 19.2-fold purification and specific activity of 4195 U/mg. The enzyme showed relatively high thermostability and retained 62% of its activity when kept at 70 degrees C for 15 min. alpha-Amylase was highly stable at -18 degrees C and loss of activity was very low during stability study. Metal ions like Mn2+ Ca2+, Co2+, K+, Mg2+, and Fe3+ activated the enzyme, while Hg2+ Ba2+, Cu2+, Na+ and Al3+ strongly inhibited the activity. The a-amylase was highly stable in various surfactants and detergents. In the presence of surfactants such as SDS and Triton X-100 the enzyme activity was found 2.9 and 1.8-fold higher respectively than control. The non-ionic detergents (Tween 20 and Tween 80) exhibited slightly inhibitory effect on the enzyme activity.
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October 2009
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