Lack of association of HFE gene polymorphism with high body iron status in Pakistani patients with type 2 diabetes mellitus.

OBJECTIVE: The Aim of this study was to investigate the relationship of 3 common polymorphisms in the HFE gene (C282Y, H63D and S65C) with high body iron status in a population of Pakistani subjects with type 2 diabetes mellitus (DM) and to explore if there is any novel mutation in HFE gene in a sample of Pakistani subjects with type 2 DM. METHODS: In a case-control design, 200 healthy controls and 200 consecutive adult subjects with type 2 DM (both gender; age range of 30-70 years) were enrolled with informed consent. Their serum samples were analyzed for body iron status (ratio of concentration of soluble transferrin receptor to ferritin concentration). DNA from blood was screened for HFE gene polymorphisms via polymerase chain reaction, followed by restriction fragment length polymorphism or via Sanger sequencing to identify any novel mutation(s) in HFE gene. RESULTS: We found that there was lack of any association between HFE polymorphism and body iron status in Pakistani subjects with type 2 DM and healthy controls. H63D was the most common polymorphism found in this population. Single base substitution of G nucleotide instead of C at the codon position 187 in the HFE gene exon 2 was discovered in one subject with DM. There was also a lack of association between D allele (variant allele of H63D) and type 2 DM. A significant relationship was found between CG genotype and abnormal albuminuria in subjects with type 2 DM (p = 0.036). CONCLUSION: In conclusion, HFE gene polymorphism is not associated either with high body iron status or type 2 DM in a hospital based Pakistani population and variant allele of H63D polymorphism appears to be associated with diabetic nephropathy.

Relationship of sociodemographic factors with serum levels of vitamin D in a healthy population of Pakistan.

High prevalence of vitamin D deficiency has been reported from Pakistan. Association of sociodemographic factors with vitamin D status has received little attention in this region. Therefore, we embarked on investigating the relationship of sociodemographic factors with vitamin D levels in a healthy Pakistani population. Venous blood from 226 healthy participants (age range 19-69 years) was collected and analyzed for serum concentrations of 25(OH) vitamin D [25(OH)D] and other related biomarkers. Demographic characteristics of the study participants were collected. Vitamin D deficiency (25(OH)D levels less than 20 ng/ml) was found to be 75% in this cohort. Gender, sunlight exposure and monthly household income emerged as predictors of hypovitaminosis D. Mean serum 25(OH)D levels in the groups with monthly household income less than Pakistani Rupees (PKR) 20,000, between PKR 20,000-50,000 and above PKR 50,000 were found to be 11.0±7.5, 13.9±9.6 and16.9±11.7 ng/ml, respectively. Using logistic regression the odds of having vitamin D deficiency was 3.22 (95% CI, 1.65-6.28) in the group with household income less than PKR 50,000 per month compared to the group with household income more than PKR 50,000 per month when the model was adjusted for gender and exposure to sunlight. There is an association between household income and hypovitaminosis D in a healthy Pakistani population.

Lack of association of statin use with vitamin D levels in a hospital based population of type 2 diabetes mellitus patients.

OBJECTIVE: To investigate the relationship of statins (drug given to reduce serum levels of LDL-cholesterol) on vitamin D levels of Pakistani type 2 diabetes mellitus (DM) patients in a hospital in Karachi. METHODS: In a cross-sectional survey, 312 consecutive patients with type 2 DM (219 males and 93 females, age 22-70 years) were recruited with informed consent. A questionnaire was administered to find out whether they were statin users or non-users. Serum was analyzed for concentrations of 25(OH) vitamin D [25(OH)D] and other related biomarkers such as serum cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol, phosphate and calcium using kit methods. Multiple Linear Regression was used to evaluate association of statin use with serum levels of vitamin D while adjusting for related covariates including duration of statin use, duration of type 2 DM and smoking. RESULTS: Mean concentrations of serum cholesterol, and LDL-cholesterol were lower among statin users compared to statin non-users (P < 0.01), while HDL-cholesterol levels were higher (P<0.01). No relationship was observed between statin use and serum levels of vitamin D (P=0.768), when adjusted for age, gender, BMI, duration of type 2 DM, smoking, serum cholesterol and LDL-cholesterol. The adjusted regression coefficient (ß) and standard error [SE(ß)] for statin use duration were 0.012 (0.042), when serum levels of vitamin D was taken as an outcome. CONCLUSION: Lack of association was found between statin use and vitamin D levels in a hospital-based population of Pakistani patients with type 2 DM.

Association of Vitamin D binding protein polymorphism with risk of type 2 diabetes mellitus in a Pakistani urban population: A case control study.

OBJECTIVE: To assess if genotypes/diplotypes of vitamin D binding protein have any association with type 2 diabetes mellitus. METHODS: This case-control study was conducted from January 2013 to July 2015 at the endocrinology clinics of the Aga Khan University Hospital, Karachi, and comprised adult patients with type 2 diabetes and their age- and gender-matched healthy controls. Venous blood was obtained and assessed for serum/plasma 25 hydroxyvitamin D, parathyroid hormone, calcium, alkaline phosphatase and creatinine. Deoxyribonucleic acid was isolated and genotyping was done by polymerase chain reaction-restriction fragment length polymorphism procedures. RESULTS: Of the 330 participants, there were 165(50%) cases and as many controls. There were 116(70.3%) males and 49(29.7%) females in each group. The mean age of the patients was 48.82±9.23 years and that of the controls was 46.27±8.77 years (range: 22-70 years) (p=0.010) Mean serum concentration of 25 hydroxy vitamin D was significantly higher among the patients compared to the controls (p<0.001), but not significantly different by genotypes or diplotypes (p>0.05). Multiple conditional logistic regression revealed an association of group-specific 1-2 genotype with patients when adjusted for age, body mass index, and serum levels of 25 hydroxy vitamin D with matched adjusted odds ratio (95% confidence interval) being 3.1(1.22-7.88). CONCLUSIONS: Group-specific 1-2 genotype of vitamin D binding protein gene was associated with the risk of type 2 diabetes.

Association of vitamin D deficiency and VDBP gene polymorphism with the risk of AMI in a Pakistani population.

OBJECTIVE: To investigate the relationship of vitamin D deficiency and risk of AMI in a Pakistani population, and to find out any association between vitamin D binding protein (VDBP) genotypes and risk of AMI in this population. METHODS: In a comparative cross-sectional study, 246 patients (age: 20-70 years; 171 males and 75 females) with first AMI were enrolled with informed consent. Similarly, 345 healthy adults (230 males and 115 females) were enrolled as controls. Their fasting serum samples were analyzed for 25 (OH) vitamin D, lipids and other biomarkers using kit methods, while DNA was analyzed for VDBP genotypes using PCR-RFLP based methods. Chi-squared test and logistic regression were used for association of vitamin D deficiency and VDBP genotypes with AMI. RESULTS: Mean serum concentration of 25(OH) vitamin D was significantly lower in AMI patients compared to healthy subjects (p=0.015) and percent vitamin D deficiency was higher in AMI patients compared to healthy subjects (p=0.003). VDBP IF-IF genotype was positively associated with the risk of AMI in subject above 45 years after adjusting for potential confounders [OR = 9.86; 95% CI=1.16 to 83.43]. CONCLUSION: Vitamin D deficiency and VDBP IF-IF genotype are associated with AMI in Pakistani adults.

Association of vitamin D deficiency with poor glycaemic control in diabetic patients.

OBJECTIVE: An association between serum levels of vitamin D and glycaemic control in type-2 diabetes mellitus (DM) patients has been reported in some of the studies carried out in the West. However, there are no reports on this relationship in Pakistani diabetic patients. The aim of this study was to ascertain whether vitamin D levels have any influence on glycaemic control in Pakistani patients with type-2 DM. METHODS: In a cross-sectional survey, relationship between serum levels of 25-hydroxy vitamin D (25(OH)D) and glycated haemoglobin (HbA1C) was examined in 141 type-2 diabetic patients including 102 males and 39 females; age range 22 to 70 years, visiting the Aga Khan University Hospital during July 2013-April 2014. Venous blood was collected and analyzed for serum/plasma levels of 25(OH)D and related biomarkers using kit methods. HbA1C levels <7.0% and >7.0% were taken as indicators of good and poor glycaemic control, respectively. An association between 25(OH)D and HbA1C was investigated using regression analysis. RESULTS: Percent vitamin D deficiency (serum level of 25(OH)D < 20 ng/ml) was significantly higher in patients with poor glycaemic control compared to patients with good glycaemic control (58.7% vs. 30.6%; p-value=0.006). Binary logistic regression analysis revealed positive association between vitamin D deficiency and poor glycaemic control while adjusting for BMI, serum levels of albumin, alanine aminotransferase and alkaline phosphatase (OR, 4.86 (95% CI, 1.9-11.9; p-value<0.001). CONCLUSIONS: The association between vitamin D deficiency and abnormal HbA1C in Pakistani diabetic patients is suggestive that patients with hypovitaminosis D could benefit from vitamin D supplementation.

Polymorphisms in MTHFR, MS and CBS genes and premature acute myocardial infarction in a Pakistani population.

High prevalence of premature coronary heart disease in Pakistanis compared to other populations points towards the genetic predisposition of this population to develop this disease. Since no investigations have been carried out in Pakistan to study the relationship of polymorphisms in genes involved in homocysteine cycle, the objective of the present study was to find out if there is any association of methylenetetrahydrofolate reductase (MTHFR) C677T, A1298C; methionine synthase (MS) A2756G; cystathionine-ß-synthase (CBS) 844ins68, G919A polymorphisms with premature acute myocardial infarction (AMI) in a population of Pakistani patients with this disease. In a cross-sectional study, DNA samples of 143 AMI patients (age <45 years) and 153 healthy controls were genotyped for the above mentioned polymorphisms using PCR-RFLP methods. Plasma/serum samples of both patients and healthy controls were screened for homocysteine, folate and vitamin B12. One way ANOVA and chi-squared test were used for analysis of data. Mean plasma homocysteine levels in premature AMI patients and healthy controls were found to be 23±17.2 and 23±13.4 µmol/l, respectively which are higher than the upper normal limit of this biomarker (15µmol/l). MTHFR 677 CT genotype in healthy controls and MTHFR 677 TT genotype in AMI patients were found to have significantly increased levels of plasma homocysteine (p value <0.05), while all other polymorphisms did not show any significant difference in mean levels of homocysteine between AMI patients and healthy controls. Moreover, no association was observed between MTHFR C677T, A1298C; MS A2756C; CBS844ins68 polymorphisms and premature AMI in this population. This indicates that common polymorphisms in MTHFR, MS and CBS genes have no role in premature AMI in Pakistani population.

High prevalence of vitamin D deficiency and insufficiency in a low income peri-urban community in Karachi.

OBJECTIVE: To determine the frequency of vitamin D deficiency and insufficiency in a low-income peri-urban population. METHODS: The cross-sectional study was conducted in a low-income, unplanned settlement in Karachi, and comprised apparently healthy adults who were recruited randomly with informed consent. Serum levels of 25-hydroxy vitamin D were measured using a kit obtained from Roche Diagnostics. One-way analysis of variance and logistic regression were used for statistical analysis. RESULTS: Of the 858 subjects, 507(59%) were females and 351(41%) males (age range: 18-60 years). Prevalence estimates of vitamin D deficiency (<20ng/ml) and insufficiency (20.0-29.9ng/ml) were found to be 501(58.4%) and 269(31.4%), respectively. Odds ratio of vitamin D deficiency was significantly higher in females compared to males after adjusting for education and smoking status (p=0.001). CONCLUSIONS: High prevalence of vitamin D deficiency among females in one of the peri-urban areas points towards a public health problem which requires attention of the medical community.

Short Communication: Lack of association between MTHFR gene polymorphisms and response to methotrexate treatment in Pakistani patients with rheumatoid arthritis.

Methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms have been reported to be associated with response to methotrexate (MTX) in certain populations of patients with rheumatoid arthritis (RA). This study aims at investigating any relationship of two single nucleotide polymorphisms (SNPs) in MTHFR gene, C677T and A1298C with response to therapy with MTX in Pakistani RA patients. Allelic frequencies of the two polymorphisms (C677T and A1298C) were determined in 67 RA patients (9 males and 58 females; mean age 42.87 ± 13.5 years) who had previously participated in a prospective clinical trial. Fifty-one patients had received MTX and were followed up for response up to 6 months. Genotyping of the two MTHFR polymorphisms was carried out using PCR-RFLP, while fasting concentration of plasma homocysteine was determined using a kit method. Twenty-eight patients were found to be "good responders", while twenty-three were "poor responders". MTHFR 1298C and MTHFR 677T alleles' frequencies in "good responders" were not different from frequencies in "poor responders" (0.574 vs. 0.521; p=0.6 and 0.197 vs. 0.196; p=0.75, respectively). Plasma homocysteine levels in female RA patients were significantly higher compared to general population in Karachi (13.1 ± 6.7 µmol/l vs. 11.4 ± 5.3 µmol/l; p<0.001). MTHFR C677T and A1298C polymorphisms are not associated with response to MTX in a population of Pakistani RA patients.

Positive Association of Vitamin E Supplementation with Hemoglobin Levels in Mildly Anemic Healthy Pakistani Adults.

BACKGROUND: Hemoglobin levels slightly below the lower limit of normal are common in adults in the general population in developing countries. A few human studies have suggested the use of antioxidant vitamins in the correction of mild anemia. The objective of the present study was to investigate the association of vitamin E supplementation in mildly anemic healthy adults with post-supplemental blood hemoglobin levels in the general population of Karachi, Pakistan. METHOD: In a single-blinded and placebo-controlled randomized trial, 124 mildly anemic subjects from the General Practitioners' Clinics and personnel of the Aga Khan University were randomized into intervention (n = 82) and control (n = 42) group. In the intervention group, each subject was given vitamin E (400 mg) everyday for a period of three months, while control group subjects received a placebo. Eighty six subjects completed the trial. Fasting venous blood was collected at baseline and after three months of supplementation. Hemoglobin levels and serum/plasma concentrations of vitamin E, vitamin B12, folate, ferritin, serum transferrin receptor (sTfR), glucose, total cholesterol, triglycerides, LDL-cholesterol, HDL-cholesterol, creatinine, total-antioxidant-status and erythropoietin were measured and analyzed using repeated measures ANOVA and multiple linear regression. RESULTS: The adjusted regression coefficients (ß) and standard error [SE(ß)] of the significant predictors of post-supplemental hemoglobin levels were serum concentration of vitamin E (0.983[0.095]), gender (- 0.656[0.244]), sTfR (- 0.06[0.02]) and baseline hemoglobin levels (0.768[0.077]). CONCLUSION: The study showed a positive association between vitamin E supplementation and enhanced hemoglobin levels in mildly anemic adults.

Association of alkaline phosphatase with acute myocardial infarction in a population with high prevalence of hypovitaminosis D.

BACKGROUND: Since Pakistanis have high prevalence of hypovitaminosis-D as well as acute myocardial infarction (AMI), the objective of the study was to investigate the relationship between vitamin-D deficiency and risk of AMI in a hospital-based population and to identify major risk factors for this disease. METHODS: Fasting serum samples from 66 consecutive AMI patients [age 30-70 y] and 132 gender and age-matched (within 5 y) healthy controls were analyzed for concentrations of glucose, total-cholesterol, LDL-cholesterol, HDL-cholesterol, triglycerides, calcium, inorganic phosphate, alkaline phosphatase (ALP), bone-ALP, parathyroid hormone (PTH), 25(OH) vitamin-D (25(OH)D) and alanine aminotransferase. RESULTS: Mean concentrations of serum 25(OH)D, PTH, total-ALP, bone-ALP, LDL-cholesterol, HDL-cholesterol and glucose were significantly different compared to healthy controls (p<0.05). Percent vitamin-D deficiency/insufficiency (levels<30 ng/ml) was significantly greater in AMI patients compared to controls (93.9% vs.75.8%; p=0.001). Multiple conditional logistic regression analysis revealed that increased levels of 25(OH)D were associated with decreased risk of AMI [MAOR (95% CI)=0.821 (0.718, 0.940); p=0.004]. Hypertension and smoking were positively associated with AMI. CONCLUSIONS: Increased vitamin-D levels were associated with decreased risk of AMI, while serum glucose, bone-ALP, hypertension and smoking were positively associated with it. Association of bone-ALP with AMI in hypovitaminosis-D is a novel finding of this study.

Association of body iron status with the risk of premature acute myocardial infarction in a Pakistani population.

BACKGROUND: Coronary artery disease is very common in Pakistani population. Some of the studies carried out on Western populations have shown a relationship between body iron status as determined by the ratio of concentrations of serum soluble transferrin receptor (sTfR) to ferritin and the risk of acute myocardial infarction (AMI). In order to investigate whether increased body iron status has any relationship with the risk of premature AMI in Pakistani population, a case-control study was carried out. METHODOLOGY/PRINCIPAL FINDINGS: In this case-control study, 203 consecutive AMI patients [146 males and 57 females; age range 18-45 years] admitted to the National Institute for Cardiovascular Diseases, Karachi, were enrolled with informed consent. In addition, 205 healthy controls whose gender and age (within 3 years) matched the patients, and who had a similar socio-economic background were recruited. Fasting venous blood was obtained and assessed for plasma/serum folate, vitamin B12, homocysteine, total cholesterol, triglycerides, LDL-cholesterol, HDL-cholesterol, sTfR and ferritin and blood lead. It was found that serum concentration of ferritin and blood lead levels were significantly higher in AMI patients compared to their age and gender-matched healthy controls (p value <0.05), while the concentrations of vitamin B12 and HDL-cholesterol were significantly lower in AMI patients compared to controls (p value <0.01). The ratio of sTfR to ferritin was significantly lower in AMI patients compared to controls [mean ± SD/median (IQR) values 84.7 ± 295/28.9 (38.4) vs 255 ± 836/49.4 (83.8), respectively; p value <0.001]. Compared with the highest quartile of sTfR/ferritin (low body iron status), the OR for the risk of AMI was 3.29(95% CI, 1.54-7.03) for the lowest quartile (quartile 1) when the model was adjusted for vitamin B12 and HDL-cholesterol (p value for trend <0.01). CONCLUSIONS/SIGNIFICANCE: This study shows a positive association between total body iron status and risk of premature AMI in a Pakistani population.

Effects of betel nut on cardiovascular risk factors in a rat model.

BACKGROUND: Areca nut (commonly known as betel nut) chewing has been shown to be associated with metabolic syndrome and cardiovascular disease (CVD). The mechanism by which betel nut ingestion could lead to development of CVD is not precisely known; however, dyslipidemia, hyperhomocysteinemia, hypertriglyceridemia and inflammation could be some of the potential risk factors. This study was undertaken to investigate the effects of two dosages of betel nut on homocysteinemia, inflammation and some of the components of metabolic syndrome, such as hypertriglyceridemia, low HDL-cholesterol, obesity and fasting hyperglycemia in a rat model. METHODS: Thirty-six adult female Sprague Dawley rats, aged 10-12 weeks were divided into three equal groups. Group-1 served as the control group (n = 12) and received water, whereas groups 2 and 3 were given water suspension of betel nut orally in two dosages, 30 mg and 60 mg, respectively for a period of 5 weeks. At the end of the fifth week, the animals were weighed and sacrificed, blood was collected and liver, kidney, spleen and stomach were removed for histological examination.Plasma/serum was analyzed for glucose, total cholesterol, HDL-cholesterol, LDL-cholesterol, triglycerides, homocysteine, folate, vitamin B12 and N-acetyl-ß-D-glucosaminidase (NAG) - a marker of inflammation. RESULTS: When the mean concentration values of 3 groups were compared using one way ANOVA followed by Tukey's HSD-test, there was a significant increase in the concentration of total cholesterol (p = 0.04) in the group receiving 30 mg/day betel nut compared to the control group. However, administration of a higher dose of betel nut (60 mg/day) had no significant effect on the serum concentrations of glucose, total cholesterol, HDL-cholesterol, LDL-cholesterol, and NAG. Histological examination of spleen revealed a dose-dependent extramedullary hematopoiesis. No other remarkable change in the tissues (liver, kidney and stomach) was observed.Mean serum/plasma levels of folate, vitamin B12 and homocysteine were not found to be significantly different in all the groups. Betel nut ingestion had no effect on the mean body weights of rats. CONCLUSIONS: Low dosage of betel nut is found to be associated with hypercholesterolemia. However, betel nut ingestion is not associated with hyperhomocysteinemia, hypertriglyceridemia, hyperglycemia, inflammation and increase in body weight in a rat model.

Relationship between vitamin B12, folate and homocysteine levels and H. pylori infection in patients with functional dyspepsia: a cross-section study.

BACKGROUND: H. pylori infection has been associated with many micronutrient deficiencies. There is a dearth of data from communities with nutritional deficiencies and high prevalence of H. pylori infection. The aim of this study was to determine the impact of H. pylori infection on serum levels of vitamin B(12), folate and homocysteine in patients with functional dyspepsia (FD). METHODS: One hundred and thirty-two patients with FD undergoing gastroscopy were enrolled. The serum was analyzed for B(12), folate and homocysteine levels before gastroscopy. H. pylori infection was diagnosed by histopathological examination of gastric biopsies and urea breath test. An independent sample t-test and the Mann-Whitney test were used to compare mean serum concentrations of biomarkers between H. pylori-positive and H. pylori-negative groups of patients. A Chi-square test was performed to assess the differences among proportions, while Spearman's rho was used for correlation analysis between levels of B(12) and homocysteine. RESULTS: The mean age of the group was 40.3 ± 11.5 (19-72) years. Folate deficiency was seen in 43 (34.6%), B(12) deficiency in 30 (23.1%) and hyperhomocysteinemia in 60 (46.2%) patients. H. pylori was present in 80 (61.5%) patients with FD while it was absent in 50 (38.5%). Mean serum levels of B(12), folate and homocysteine in the H. pylori-positive group of patients were not significantly different from the levels in the H. pylori-negative group (357 ± 170 vs. 313 ± 136 pg/mL; p = 0.13), (4.35 ± 1.89 vs. 4.42 ± 1.93 ng/mL; p = 0.84); (15.88 ± 8.97 vs. 16.62 ± 7.82 µmol/L; p = 0.24); respectively.B(12) deficiency (≤200 pg/mL) was 23.8% in the H. pylori-positive patients versus 22.0% in the H. pylori-negative patients. Folate deficiency (≤3.5 ng/mL) was 33.8% in the H. pylori-positive group versus 36% in the H. pylori-negative group. Hyperhomocysteinemia (>15 µmol/L) was present in 46.2% of H. pylori-positive patients compared to 44% in the H. pylori-negative group. Correlation analysis indicated that serum B(12) levels were inversely associated with serum levels of homocysteine in patients with FD (rho = -0.192; p = 0.028). CONCLUSIONS: This study demonstrated an inverse relationship between serum levels of B(12) and homocysteine in patients with FD. Moreover, no impact of the presence of H. pylori was found on B(12), folate and homocysteine levels in such patients.

Folic acid and vitamin B6 deficiencies related hyperhomocysteinemia in apparently healthy Pakistani adults; is mass micronutrient supplementation indicated in this population?

OBJECTIVE: To determine the plasma/serum levels of homocysteine, and vitamins folate, B6 and B12, in Pakistani healthy adults. STUDY DESIGN: Cross-sectional study. PLACE AND DURATION OF STUDY: The Aga Khan University, from October 2006 to April 2008. METHODOLOGY: Fasting levels of plasma/serum folic acid, pyridoxal phosphate (PLP), vitamin B12 and homocysteine were determined in 290 apparently healthy hospital personnel from institutions in two cities of Pakistan. Spearman correlation test and linear regression analysis was conducted. RESULTS: There were 219 males and 71 females with mean age of 46+/-10.5 years and mean body mass index of 23.5 +/-3.8. Mean plasma homocysteine levels in Pakistani normal adults were found to be 17.95+/-8.4 micromol/l. Mean concentrations of plasma/serum folate, vitamin B12 and PLP were found to be 5+/-3.9 ng/ml, 522+/-296 pg/ml and 21.6+/-14 nmol/l, respectively. Serum/plasma levels of folate, vitamin B12 and PLP were negatively correlated with plasma homocysteine (rho coefficient=-0.367, p<0.001; -0.173, p=0.004; -0.185, p=0.002, respectively). Serum folate and plasma PLP levels were inversely related with plasma homocysteine, adjusted for gender, age, smoking and body mass index (p<0.001 and p=0.003, respectively). Percent deficiency values of folate, vitamin B6 and vitamin B12 were 39.7%, 52.8% and 6.6% respectively. CONCLUSION: The high levels of plasma homocysteine could indicate a reason for mass micronutrient supplementation to prevent the high incidence of cardiovascular disease observed in Pakistani population.

N-acetyl-B-D-glucosaminidase and inflammatory response after cardiopulmonary bypass.

OBJECTIVE: To determine the changes in activity of plasma N-acetyl-beta-D-glucosaminidase, a marker for inflammation as well as renal, pulmonary and cardiac damage and proinflammatory cytokines in patients undergoing coronary artery bypass grafting and find out the relationship between their plasma levels with clinical outcome of patients. STUDY DESIGN: Cross-sectional study. PLACE AND DURATION OF STUDY: The Aga Khan University, Karachi, from January to June 2003. PATIENTS AND METHODS: N-acetyl-beta-D-glucosaminidase (NAG) activity and concentrations of tumor necrosis factor-alpha of (TNFalpha), interleukin 6 (IL-6), interleukin 8 (IL8) and granulocyte-macrophage colony stimulating factor (GM-CSF) were monitored in plasma samples of 12 angina patients undergoing coronary artery bypass grafting (CABG), before, immediately after and 5 days post-surgical procedure. Serum glucose concentrations were also monitored in those patients. Patient's clinical condition was monitored during this time period. RESULTS: No significant increase was observed in plasma NAG activity (a marker of inflammation) or in plasma levels of TNFalpha, IL-6, IL-8 and GM-CSF immediately after surgery, indicating that cardiopulmonary bypass itself does not produce any significant amount of inflammation immediately after CABG. However, 5 days post surgery, there was a significant increase in plasma NAG activity (p=0.001), TNFalpha (p=0.047) and GM-CSF (p=0.045). There was no relationship between plasma NAG activity and clinical outcome because various parameters of renal, cardiac and pulmonary functions, though slightly affected, remained within the normal limits. CONCLUSION: Increased levels of NAG and TNFalpha did not affect clinical outcome. However, data suggest that NAG can be a potential marker for inflammation and end organ damage following CABG. An increase in GM-CSF on day 5 following CABG indicates enhanced body's defense mechanism against infection.

Ascorbic acid contents of Pakistani fruits and vegetables.

Fresh fruits and vegetables are good sources of vitamin C which is known for its antioxidant and immune-enhancing effects. The objective of this study was to determine ascorbic acid (vitamin C) contents of regularly consumed fruits and vegetables available in Pakistani markets. Most commonly used fresh fruits and vegetables were homogenized in 5% trichloroacetic acid, and ascorbic acid contents in the extracts were determined using a spectrophotometric method. Banana, custard apple, orange, lemon, guava and papaya were found to be very rich in ascorbic acid. Among vegetables, capsicum (green sweet pepper), cauliflower, bittergourd, roundgourd, beetroot, spinach, cabbage and radish contained high concentrations of ascorbic acid. Chikoo, grapes, pear, apricot, peach, carrot, cucumber, lettuce and "kakri" were found to be poor sources of ascorbic acid. Several Pakistani fruits and vegetables (pear, melon, onion, sweet green pepper, spinach, cucumber) had ascorbic acid values similar to those reported by US Department of Agriculture in these fruits and vegetables in USA. However, wide differences in vitamin C contents were also observed in certain other fruits and vegetables from these two countries. This indicates that regional varieties of fruits and vegetables could vary in their ascorbic acid contents. Since subclinical deficiency of vitamin C appears to be quite common in developing countries like Pakistan, there is a need to develop awareness among masses to consume fresh fruits and vegetables with high contents of vitamin C.

Plaque formation reduction with glutathione monoester in mice fed on atherogenic diet.

OBJECTIVE: To determine the role of glutathione monoester on reducing the development of plaque formation in an animal model. DESIGN: Laboratory control trial. PLACE AND DURATION OF STUDY: Aga Khan University, Karachi, from January 2004 to December, 2004. MATERIALS AND METHODS: Twenty-four Balb/c mice were divided into 3 equal groups. First group was fed on atherogenic diet alone, while the second group received atherogenic diet plus twice weekly injections of glutathione monoester. The third group was fed on normal diet for mice. After one year, the animals were sacrificed. Blood was analyzed for lipid levels, while liver, kidney, spleen, heart and aorta were removed to study morphological changes. RESULTS: In the groups of mice receiving atherogenic diet (with and without glutathione monoesters), there was significant increase in levels of total cholesterol (p=0.011) and LDL cholesterol (p=0.001) compared to levels of these lipids in mice on normal diet. However, a significant decrease in levels of triglycerides (p=0.01) was observed in the group receiving atherogenic diet along with glutathione monoester. Supplementation with glutathione monoester had the most pronounced effect only on triglyceride levels. Atherosclerotic plaques were seen in heart and/or aorta of mice receiving atherogenic diet. However, such plaques were either totally absent or if seen in an animal, were extremely small and diffuse in the group receiving glutathione monoester along with atherogenic diet. Mice on normal diet had no evidence of any plaque formation. Cholesterol granuloma was seen in liver of mice on atherogenic diet alone. In mice receiving atherogenic diet plus glutathione monoester, no cholesterol granuloma was found in liver. There were no remarkable morphological changes in spleen and kidney in the three groups of mice. CONCLUSION: Glutathione monoester appears to inhibit or reduce the development of plaque formation in mice.

N-acetyl-beta-D-glucosaminidase--a marker of reperfusion and a prognostic indicator in patients with acute myocardial infarction.

OBJECTIVE: N-acetyl-beta-D-glucosaminidase (NAG) is a lysosomal enzyme of which the activity in plasma is increased in a number of conditions including myocardial infarction. Plasma levels of cardiac proteins, such as myoglobin, troponin and creatine kinase, have been used as markers of myocardial reperfusion as well as for the prognosis of the disease. The aim of this study is to investigate whether NAG could be used as an additional biochemical marker to predict myocardial reperfusion and to find out if its release following thrombolysis has a prognostic value as well. METHOD: Streptokinase (SK) in a dose of 1.5 million units was administered intravenously to 75 patients with acute myocardial infarction (AMI) and the response to SK was assessed over a period of 90 minutes. Plasma NAG activity was monitored before (0 min) and 90 min after SK treatment. RESULTS: The mean NAG activity values were found to be 8.6 +/- 4.8 U/I and 10.95 +/- 7.3 U/I, respectively, and when compared using paired samples t-test revealed a significant increase (p = 0.0001) following thrombolytic therapy. The increase appears perfusion related as rabbits injected with SK failed to show any increase in plasma NAG activity. There was an association between plasma NAG activity and clinical response to SK treatment (p = 0.045). A follow-up of 66 patients over a period of 18 months, revealed increased survival in AMI patients having significantly more activity/release of plasma NAG after thrombolytic treatment (p = 0.001). CONCLUSION: NAG appears to be another potential biochemical marker of reperfusion. Moreover, NAG release profile during thrombolysis could be of value in predicting prognosis of the disease.

Variability in lipid profile before and after coronary artery bypass grafting (CABG).

OBJECTIVE: To investigate changes in total cholesterol, low density lipoprotein (LDL) cholesterol, high density lipoprotein (HDL) cholesterol, and triglycerides in serum of Pakistani patients before, immediately after and 5 days post CABG. METHOD: Serum samples from 31 consecutive Pakistani angina patients undergoing CABG at the Aga Khan University Hospital were analyzed for total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides using kit methods. RESULTS: Immediately after CABG, there is a significant decline in the mean levels of serum cholesterol, LDL cholesterol, HDL cholesterol and triglycerides. However, 5 days post CABG, there is a significant increase in the concentrations of total cholesterol (P = 0.01) and LDL cholesterol (P = 0.001) in nondiabetic angina patients (n = 13). Among the diabetic group of patients (n = 18), the levels of total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides went back to the pre-operative levels within 5 days post CABG. Compared to European patients, Pakistani patients tend to have very low levels of HDL cholesterol (24.9 +/- 7.1 mg/dl) and high levels of triglycerides (185 +/- 50 mg/dl) on day 5 post CABG. CONCLUSION: Since risk of mortality following CABG increases with low level of HDL cholesterol and high level of triglycerides, close monitoring and treatment of high lipid levels of Pakistani patients following CABG is necessary to prevent further coronary events.