Association of SP-D, MBL and I-NOS genetic variants with pulmonary tuberculosis.

Background: Pulmonary tuberculosis is caused by Mycobacterium tuberculosis . It is a multifactorial disease with both host as well as pathogen factors contributing to susceptibility and protection from the disease. Various reports have highlighted important roles of lung surfactant protein D (SP-D), mannan-binding lectin (MBL) and I-NOS in innate immune defense against M. tuberculosis Aims : The present study investigated the role of polymorphisms in three candidate genes encoding Lung surfactant protein D, Mannan binding lectin and Inducible Nitric oxide synthase, in susceptibility and protection to pulmonary tuberculosis. Settings and Design : A case-control association study of SNP's in lung surfactant protein D (SP-D), mannan-binding lectin (MBL) and I-NOS with pulmonary tuberculosis in Indian population was carried out. This involved sequencing of all the coding exons of lung surfactant protein D (SP-D) , while, exon 1 (collagen region) and exon 4 (carbohydrate recognition domain) of mannan-binding lectin (MBL) and exons 2, 8 and 16 of I-NOS and their flanking intronic regions for single nucleotide polymorphisms in DNA samples isolated from 30 pulmonary tuberculosis patients and 30 controls of Indian population. Statistical analysis: Various allele frequencies were calculated using online two by two table (home.clara.net/sisa/). Odds ratio and P values were calculated at 95% confidence interval (CI). Results : A total of fourteen single nucleotide polymorphisms (5 in SP-D , 5 in MBL and 4 in I-NOS ) were observed of which four (G459A SP-D , G274T I-NOS , G1011A and T357G MBL ) have not been reported earlier. Four single nucleotide polymorphisms viz. G459A of exon 7 of SP-D ( P =0.00, odds ratio (OR) = 4.96, 2.18 P = 0.00 or= 3.85 1.66 P =0.00 or=4.04, 2.20< OR<7.42) and G274T of intron 16 of I-NOS ( P =0.00 or=4.46, 2.40 Conclusion: The present study has led to identification of 4 SNP's in SP-D , MBL and I-NOS associated with pulmonary tuberculosis in Indian population.

Tuberculin specific T cell responses in BCG vaccinated children.

The effector mechanisms of BCG protection were examined 5-7 years after vaccination. The in vitro lymphoproliferation, following stimulation with tuberculin, in normal, (A) vaccinated and (B) unvaccinated children and children with tuberculosis (C), were assayed. The mean stimulation index (SI) of lymphocyte transformation in normal subjects were significantly (P < 0.05) higher than those with tuberculosis. The ratio of tuberculin-specific CD4 to CD8 cells in short-term cultures were significantly (P less than 0.05) higher in the vaccinees. In group (A), 70 % had positive ratios as against 20 %and 0 %in groups (B) and (C), respectively. Secretion of IL-2 by the cells was significantly (P < 0.05) high in the vaccinated. None of the unvaccinated children had positive levels of IL-2. The vaccinees also had highly significant (P < 0.01) levels of IFN-)in the supernatants of cell-cultures, following tuberculin stimulation. In majority of the BCG vaccinated children, the stimulation of specific TH1 cells seem to be considerably high, in short-term in vitro cultures. While these responses were not so marked in the unvaccinated, they were almost totally absent in the patients.

Post-BCG vaccination check based on serum IgG to culture filtrate antigens.

Since there are no reliable methods to demonstrate the effect of BCG vaccination in children, culture filtrates of BCG were evaluated for their specificity and sensitivity. BCG culture filtrate (BCG-CF), BCG sonicate and tuberculin were used as antigens and tested against serum, for the presence of IgG class of antibodies by ELISA and Western blot. Methodology: Children in the age-group of 1 to 10 years, were categorized as: (a) normal, and vaccinated, n=35; (b) normal, without a scar and with no evident history of vaccination, n=15; and (c) children with tuberculosis (meningitis, miliary and lymphadenitis) n=15. Results: The mean values of optical density (OD) in group (a), 4.0+0.08, were significantly high (P<0.001) to BCG-CF, compared to that of groups (b) (1.0+0.02) and (c) (1.4+0.03). The Western blot results revealed that a greater number of children (71%) in the vaccinated group reacted to low molecular weight proteins (10-30kDa) compared to other groups (unvaccinated: 17% and TB: 20%). The overall specificity, sensitivity, positive predictive value and negative predictive value of BCG-CF were higher in the vaccinated group. Conclusions: The results of the study suggest that the secreted antigens of BCG induce antibody formation, which are specific and are directed mostly towards the low molecular weight proteins. The presence of these antibodies could probably be exploited in an assay to distinguish children immunized by BCG from the unvaccinated and those having tuberculosis.

Pulmonary function tests in Indian girls--prediction equations.

OBJECTIVE: The present study was conducted to evaluate pulmonary functions and develop prediction equations in Indian girls. METHODS: 1038 normal healthy schoolgirls in the age group of 5 to 15 years were selected for the present study. The anthropometrical parameters like height, weight, sitting height and chest circumference were measured and body surface area (BSA) and fat free mass (FFM) were derived using equations. The pulmonary functions such as FEV1, FVC, FEV1/FVC% and PEFR were studied. RESULTS: The height, sitting height, weight, BSA, chest circumference, body fat as well as FEV1, FVC, FEV1/FVC% and PEFR were comparable with Indian standards. Multiple regression equations were developed to predict FEV1, FVC and PEFR using anthropometrical indices like height, fat free mass and age or chest circumference in view of significantly high correlation of these parameters with lung function variables [height and FEV1 (r-0.90), height and FVC (r-0.899), height and PEFR (r-0.891), chest and FEV1 (r-0.868), chest and FVC (r-0.867), chest and PEFR (r-0.83)]. CONCLUSION: The regression equations to predict the pulmonary functions were presented using the independent variables like height, fat free mass and chest circumference or age, since these variables have shown very strong predictability for FEV1, FVC and PEFR. The equations presented in this study can be considered as referral standards for Indian girls.

Study on lung function tests and prediction equations in Indian male children.

OBJECTIVE: The present study was carried out to evaluate lung functions and develop prediction equations in Indian boys. SUBJECTS: 1555 normal healthy schoolboys from Hyderabad city who were in the age group of 5 to 15 years were selected for the present study. DESIGN: The anthropometric parameters such as height, sitting height, weight, and chest circumference were measured and body surface area (BSA) and percent body fat (% Fat) were derived. The lung functions studied were FEV1, FVC, FEV1% and PEFR. RESULTS: The height, sitting height, weight, BSA, chest circumference, body fat as well as FEV1, FVC, FEV1 % and PEFR were comparable with Indian boys. The height for age, weight for age and weight for height were found to be lower than 50th percentile of NCHS standards in the subjects studied. Similarly the lung function values of the study population were found to be lower than the values of corresponding western population. CONCLUSION: Regression equations were derived to predict FEV1, FVC and PEFR using physical characteristics. Height, chest circumference and fat free mass were the best predictors for FEV1, FVC, and PEFR. Age, height, sitting height, weight, chest circumference and fat free mass showed significant association with lung functions.

BCG vaccine--current status.

Tuberculosis incidence has made re-emergence due to multidrug resistance of the bacilli and for acquired immunodeficiency syndrome. Of all vaccines BCG is most widely used. It protects children from secondary forms of tuberculosis. Several authors recommend a second dose to boost the waning effect of BCG. The vaccine at present needs to be improved to make it more efficacious. Vaccines which involve the protective immune responses are ideal.

Cell mediated immunity in children with scar-failure following BCG vaccination.

OBJECTIVE: To find out the incidence of BCG-scar failure, in BCG vaccinated children and assess their in vitro cellular response. DESIGN: Four year prospective cohort observational study. SETTING: Immunization centers at: (a) State Tuberculosis Center; (b) Tuberculosis Association of Andhra Pradesh; and (c) Niloufer Hospital for Women and Children in Hyderabad. METHODS: Healthy children brought to the immunization centers for BCG vaccination and were followed up till 6 months of age for scar failure. These 655 BCG vaccinated children were classified into three groups based on the age at vaccination: (i) 0 day-1 day; (ii) 2 days-30 days; and (iii) 31 days-90 days. Of these children, in vitro leukocyte migration inhibition (LMI) levels against PHA/PPD were investigated in 228 of them. RESULTS: Of the 655 children, 591 (90.2%) showed presence of scar. Out of the three groups, number of children belonging to the first group in whom the scar was absent, was highest. Of 591 children with scar, LMI was performed in 34, 110 and 43 of them in the three different age groups, respectively out of whom 88.2%, 87.2% and 86% had positive response (> or = 20%) to PPD. Of 64 children who failed to develop a scar, LMI was performed in 17, 19 and 5 in three different age groups out of whom 88.2%, 94.7% and 80% had positive (> or = 20%) in vitro response to PPD. CONCLUSION: Scar failure may occur in 10% of BCG vaccinated and is more common with immunization within 48 hours of life. Failure of formation of BCG-scar at the site of BCG vaccination may not necessarily imply failure of immunization because majority of them do elicit positive in vitro LMI response.

Crossmatching considerations in renal transplantation.

During the year 1993-1994, 73 renal transplant cases have been screened for the presence of anti-HLA antibodies using the standard lymphocytotoxicity assay. Amongst the 9 related transplantations with 100% negative crossmatch 6 were successful. About 8.2% of the patients had a shift from positive to negative crossmatch. It was observed that an increased number of transfusions (ranging from 3 to 21) in males and females yielded negative crossmatches. In females, however, owing to various factors such as pregnancies, parity and infections, varied percentages were observed with different donors. The crossmatches in diabetics and hypertensive patients suggest no particular correlation and probably have no role in the outcome of the assay.

Comparison of the immune responses in children vaccinated with three strains of BCG vaccine.

The present study was conducted to evaluate and compare the specific cellular responses of children vaccinated with three different strains of BCG. The study comprised of normal children with normal weight and normal general responses (PHA) to in vitro leukocyte migration inhibition test (LMIT). The three strains of BCG under study were Japan-BCG, Glaxo-BCG and Madras-BCG. One hundred children were selected at random from each group. The mean ages of these infants were 9.9 +/- 9.5, 9.8 +/- 7.6 and 9.8 +/- 8.3 weeks, respectively. Six weeks after vaccination, the diameter (in mm) of induration at the vaccination site was measured. Three months after vaccination, in vitro LMIT was performed against PPD tuberculin antigen. This test was done again after 3 months in all the children who tested negative. The mean value of the diameter of the Glaxo-BCG group (10.0 +/- 13.5 mm) was significantly higher (p < 0.05) than the mean values of Japan-BCG (9.10 +/- 3.9 mm) and Madras-BCG (8.38 +/- 4.1 mm). The mean LMI values were similar in all the three groups. There was no correlation between the in vitro and in vivo parameters. The number of children positive to LMI (PPD) were 59, 58 and 63, for the Madras, Japan and Glaxo-BCG groups, respectively. A total number of 91, 91 and 95 were positive to LMIT at the end of 6 months after BCG in the Madras, Japan and Glaxo-BCG groups, respectively. The observations suggested that there were no major differences between the three strains of BCG in their capacity to induce cellular responses.

Optimum age of a child for BCG vaccination.

The objectives of this study were to evaluate whether a newborn or a neonate is capable of responding immunologically after BCG vaccination and to find out if this immunity persists for one year. Normal infants aged between 0 days-3 months brought to immunization centre were included in the study. In vitro leukocyte migration inhibition test was performed in these children using Phytohemagglutinin and purified protein derivative (PPD). They were grouped based on their age at vaccination, their LMI values and on the time interval after vaccination. The mean values of % LMI (PPD) in all the age groups were positive and there were no significant differences between the newborns, the neonates and other groups. The values were positive and comparable even after 12 months in all the groups. The percentage of infants with positive or negative values to LMI (PHA) and negative values to LMI (PPD) were also comparable at different time intervals in different age groups. The results suggest that newborns or neonates are as capable of eliciting a positive immune response after BCG vaccination, as older infants and the practise of vaccinating a child at birth could be continued.

Comparison of effects of yoga & physical exercise in athletes.

The effect of pranayama a controlled breathing practice, on exercise tests was studied in athletes in two phases; sub-maximal and maximal exercise tests. At the end of phase I (one year) both the groups (control and experimental) achieved significantly higher work rate and reduction in oxygen consumption per unit work. There was a significant reduction in blood lactate and an increase in P/L ratio in the experimental group, at rest. At the end of phase II (two years), the oxygen consumption per unit work was found to be significantly reduced and the work rate significantly increased in the experimental group. Blood lactate decreased significantly at rest in the experimental group only. Pyruvate and pyruvate-lactate ratio increased significantly in both the groups after exercise and at rest in the experimental group. The results in both phases showed that the subjects who practised pranayama could achieve higher work rates with reduced oxygen consumption per unit work and without increase in blood lactate levels. The blood lactate levels were significantly low at rest.

Cell mediated immune responses in BCG vaccinated children.

The immunological status of BCG vaccinated and unvaccinated healthy children was evaluated to assess the efficacy of BCG. The duration of immunity conferred by the vaccine was also investigated. Of the 326 children studied, 170 (52%) had the BCG scar and only 24 (14%) showed a positive Mantoux response. Among the unvaccinated group, 14 of 156 (9%) showed a positive response. All cases had normal proportions of T and B cells in the peripheral blood. The mean values of the leukocyte migration inhibition (LMI) test with PHA were also normal. The per cent LMI values against PPD were compared in the children classified into groups based on their vaccination status and response to Mantoux test. A higher number of the vaccinated children had positive LMI values compared to those unvaccinated (p < 0.01). The LMI values of children classified into three age groups decreased significantly (p < 0.01) with increase in age. Hence, BCG seems to afford some protection in children and has to be administered at birth. Revaccination at the age of eight years, may boost the waning immunity and, may be considered in this age-group.

Excretion studies of nitrofurantoin and nitrofurantoin with deglycyrrhizinated liquorice.

Nitrofurantoin and nitrofurantoin with liquorice were given to healthy volunteers and patients suffering from urinary tract infections. The excretion rates of the drug, colony counts and side effects were studied in patients and excretion rates in the volunteers. The excretion rates of the drug were significantly higher in patients receiving the drug with liquorice and also side effects were minimal. There was no significant difference in the excretion rates of the drug with addition of liquorice in healthy volunteers.