Increased levels of erythrocyte glutathione in acute myocardial infarction: An antioxidant defence

Objective: Glutathione (GSH) has a central role in the defence against oxidative damage. This study was carried out to investigate any change in erythrocyte GSH levels in a population of patients with acute myocardial infarction (AMI) and compare them with levels in normal healthy subjects.Method: GSH levels were determined in erythrocytes of one hundred and seventy six patients with AMI (age: 30-70 years; 131 males and 45 females) admitted to the National Institute of Cardiovascular Diseases, Karachi. These levels were compared with eryrocyte GSH levels obtained from 95 normal healthy subjects (controls).Results: Mean +/- SD erythrocyte GSH levels in AMI patients and controls were found to be 2.34 +/- 0.62 micromol/ml of packed cells and 2.08+/- 0.62 micromol/ml of packed cells, respectively. The two values when compared with one way ANOVA were found to be significantly different (p=0.001). Age had little effect on erythrocyte GSH levels in both AMI patients and normal healthy subjects.Conclusion: Increased production of reactive oxygen species is a feature of cardiovascular disease, such as AMI and cells can respond to mild oxidative stress by upregulating antioxidant defence in terms of increased production of GSH

Heterogeneity of methotrexate binding in human colon tumor cells

[3H]-methotrexate binding at pH 5.0 and pH 7.2 by the cytosol of tumor tissues and the surrounding normal areas of the gastrointestinal tract of patients suffering from colon or gastric cancer has been used to identify in these cells the presence of a binder of methotrexate having low-affinity for this drug in addition to the enzyme dihydrofolate reductase. Scatchard analysis of the [3H]-methotrexate binding data by a colon tumor sample also reveals that there are two binders of this drug present in the cytosol of these cells. The association constant (Kass) for one binder of methotrexate is = 5.6 x 10(7) M-1 while the Kass for the second binder is = 1.0 x 10(6) M-1. The two binders do not differ very much in their apparent molecular weight. Upon isoelectric focusing, the tumor cell cytosol resolves into 4 major isoproteins each having the ability not only to bind [3H]-methotrexate but also reduce [3H]-pteroylglutamic acid to [3H]-tetrahydropteroylglutamic acid. This suggests that the two binders of methotrexate may be the two forms of dihydrofolate reductase having different affinities for this anticancer drug

Induction of white cell proliferation due to haematopoietic growth factors is associated with an increase in multiple forms of dihydrofolate reductase in non-neutropenic cancer patients

Objective: Granulocyte-colony stimulating factor (G-CSF) and granulocyte macrophage-colony stimulating factor (GM-CSF) are frequently used in cancer patients to overcome the granulocytopenic effects of chemotherapy, and also to mobilize the stem cells. The mobilized stem cells are collected from the peripheral blood and used for transplantation following high doses of chemotherapy. However, the molecular mechanism by which these colony stimulating factors (CSFs) bring about proliferation of myeloid precursor cells is not clearly known. Dihydrofolate reductase (DHFR), which has an established role in DNA synthesis, could be a link between administration of CSF and stem cell proliferation. The purpose of this study was to investigate whether CSFs induce white cell proliferation by producing multiple forms of DHFR.Methods: Twelve patients with non-haematological malignancies were treated with either G-CSF or GM-CSF to mobilize stem cells. Nine healthy subjects were treated with placebo as controls. Blood samples were obtained before and after stimulation with CSFs or placebo. White blood cells were separated and concentrations of both active DHFR and immunoreactive nonfunctional form of DHFR were determined in their cytoplasm using methotrexate-binding assay and enzyme-linked immunosorbent assay, respectively. Total leucocytes count (TLC) was also monitored before and after stimulation with CSFs or placebo.Results: There was a significant (P \u3c 0.05) increase in concentration of immunoreactive nonfunctional form of DHFR and TLC following stimulation with CSFs. There was an increase in concentration of active DHFR as well, however, this did not reach statistical significance. In the placebo-treated subjects, no significant increase in active DHFR, immunoreactive nonfunctional form of enzyme or TLC was observed. However, it was noticed that the base-line values of active DHFR and immunoreactive nonfunctional form of enzyme in leucocytes of cancer patients were higher than the base-line values in leukocytes of normal healthy subjects.Conclusion: Our data suggest that colony stimulating factors induce white cell proliferation by increasing levels of multiple forms of DHFR

Urinary N-acetyl-beta-D-glucosaminidase in rheumatoid arthritis

Excretion of urinary N-acetyl beta-D-glucosaminidase (NAG) and its isoenzyme patterns were studied in two groups of patients with rheumatoid arthritis (RA) and in normal control subjects. Urine samples were collected from 30 seropositive RA patients, 19 seronegative RA patients, and 15 normal healthy subjects. All the patients and normal subjects were assessed to have normal liver and kidney functions. A small portion of the urine sample was dialyzed against 0.01 M phosphate buffer, pH 7.0 and NAG activity was monitored. Mean +/- SD values of urinary NAG in seropositive RA patients, in seronegative RA patients and in normal healthy subjects were found to be 4.20 +/- 3.73 U/g creatinine, 2.96 +/- 2.11 U/gm creatinine, and 1.71 +/- 0.6 U/g creatinine, respectively. The mean urinary, NAG value in RA patients was found to be significantly higher (P \u3c 0.05) in seropositive RA compared to the mean NAG value in normal healthy subjects and patients with seronegative RA when analyzed by one way ANOVA and Tukey-HSD test. The mean proportion of isoenzyme form B to isoenzyme form A in seropositive RA patients was also found to be significantly different (P \u3c 0.05) from the mean proportion of these forms in normal healthy subjects and seronegative RA patients. There also appears to be a correlation between the concentration of urinary NAG and severity of the disease in seropositive RA

Variability in lipid profile in patients with acute myocardial infarction from two tertiary care hospitals in Pakistan

Objective: To investigate changes in total cholesterol, high density lipoprotein (HDL)-cholesterol, low density lipoprotein (LDL)-cholesterol and triglycerides in serum of Pakistani patients with AMI due to age, gender, body mass index (BMI), diabetes, hypertension, and smoking, and also find out the prevalence of hypercholesterolemia, hypertriglyceridemia, low HDL-cholesterol and isolated low-HDL cholesterol in them.Patients and Methods: Serum samples from 451 consecutive AMI patients (250 from National Institute of Cardiovascular Diseases, Karachi and 201 from Armed Forces Institute of Cardiology, Rawalpindi) were analyzed for total cholesterol, HDL-cholesterol and triglycerides using kit methods. LDL-cholesterol was determined using the Friedewald formula.Results: Mean serum concentrations of total cholesterol, HDL-cholesterol, LDL-cholesterol and triglycerides in AMI patients were found to be 181 +/- 50 mg/dl, 35.7 +/- 11.3 mg/dl, 110 +/- 47 mg/dl and 177 +/- 127 mg/dl, respectively. Mean levels of total cholesterol and HDL-cholesterol were not significantly affected by age, gender, BMI, diabetes mellitus, hypertension and smoking. Mean LDL-cholesterol concentration, however, was found to be significantly increased in diabetes mellitus (p=0.047), while age, gender, BMI, hypertension and smoking had no significant effect on the levels of this lipoprotein. Mean levels of triglycerides were significantly decreased in older patients (\u3e50 years) compared to younger (\u3c50 \u3eyears) ones (p=0.019). Gender, BMI, diabetes mellitus, hypertension and smoking, however, had no effect on triglyceride levels The frequencies of hypercholesterolemia, hypertriglyceridemia, low HDL-cholesterol and isolated low-HDL-cholesterol were found to be 30.6%, 30.1%, 48.6% and 34.1%, respectively.Conclusion: High prevalence of hypertriglyceridemia and low HDL-cholesterol (which constitute a component of metabolic syndrome) in Pakistani AMI patients is suggestive that these two lipid abnormalities could be playing a major role in the development of atherosclerosis in Pakistani population

Increased levels of multiple forms of dihydrofolate reductase in peripheral blood leucocytes of cancer patients receiving haematopoietic colony-stimulating factors: Interim analysis

The precise mechanism whereby granulocytes proliferate when haematopoietic colony stimulating factors (CSFs) are used in neutropenic cancer patients is poorly understood. The purpose of this study was to investigate whether these cytokines bring about leucocyte proliferation by increasing the levels of multiple forms of dihydrofolate reductase (DHFR). Blood samples were collected from 36 cancer patients (25 males and 11 females) with chemotherapy-induced neutropenia. One sample of blood from each patient was obtained before therapy either with CSF, such as granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF) or with placebo, and another one at the time of resolution of neutropenia. Peripheral blood leucocytes in these blood samples were counted, separated and lysed. From lysates, cytoplasmic samples were prepared and analyzed for active DHFR by a methotrexate-binding assay and for total immunoreactive DHFR by an enzyme linked immunosorbent assay. The increase in total leucocyte count (TLC) was most prominent (P \u3c 0.005) in the CSF group and less so (P \u3c 0.05) in the placebo group. The mean +/- SD concentration values of active DHFR before and after stimulation with GM-CSF found were to be 0.34 +/- 0.4 ng/mg protein and 0.99 +/- 0.82 ng/mg protein, respectively, and in the group treated with G-CSF, 0.24 +/- 0.32 ng/mg protein and 1.18 +/- 2.4 ng/mg protein, respectively. This increase in active DHFR after stimulation with CSF was statistically significant (P \u3c 0.05). Similarly, concentration values of immunoreactive but nonfunctional form of DHFR (IRE) were 110 +/- 97 ng/mg protein and 605 +/- 475 ng/mg protein before and after stimulation with GM-CSF, and 115 +/- 165 ng/mg protein and 1,054 +/- 1,095 ng/ mg protein before and after stimulation with G-CSF. This increase in concentration of IRE after stimulation with GM-CSF or G-CSF was statistically significant (P \u3c 0.005). In the control group, there was an increase in the concentration of both active DHFR and IRE after treatment with placebo. However, this was not statistically significant. Resolution of neutropenia was quicker in the groups treated with CSF compared to the control group. Results of this study indicate that colony stimulating factors (G-CSF and GM-CSF) induce white cell proliferation by increasing the levels of multiple forms of DHFR

Methotrexate in rheumatoid arthritis: a 2 year experience at a university hospital in Pakistan

In this study we report our two years experience of methotrexate (MTX) in the management of rheumatoid arthritis (RA) at the Aga Khan University Hospital, Karachi. We studied the clinical course of 124 RA patients. The mean age was 44 +/- 11 years (range 19-72) and mean duration of RA was 5 +/- 4 years (range 0.3-25). Female to male ratio was 10:2.4 (100F:24M). All of them were diagnosed according to the criteria set by American Rheumatism Association. The mean value of ESR was 60 +/- 30 (Range 3-128). Fifty one percent had severe disease (\u3e 10 joints involved and evidence of erosions and deformities). Twenty-one patients had extra-articular manifestations. None of them had received MTX previously. Their kidney and liver functions were assessed to be normal. Patients were divided into two groups. One group (n = 92) received MTX (7.5-10 mg/week) as initial treatment, while the other group (n = 32) was given other disease modifying anti-rheumatic drugs (penicillamine, salazopyrin, gold, or chloroquine) followed by MTX. Assessment of the treatment outcome and development of any adverse reactions was carried out at 3-month interval over an average period of 1 year. Assessment of the treatment outcome in the group which received MTX as initial drug revealed the response to be excellent in 13%, good in 70%, fair in 11% and variable in 4%. In the group which received MTX as second-line of therapy, 59% of the patients had the response from good to excellent, while 25% of the patients exhibited poor to fair response. Regarding side-effects of MTX treatment, 57% exhibited none, while 35% had nausea and vomiting. Alopecia was the next common toxicity in these patients. Two individuals had abnormal liver function tests (value twice more than normal), while one developed lung fibrosis. MTX despite its adverse effects in some of the patients is still an effective, well tolerated and inexpensive disease modifying drug in RA

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-β-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

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\u3c0.05). Percent vitamin-D deficiency/insufficiency (levels\u3c30 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