The first successful simultaneous pancreas-kidney transplant in India.

Insulin-dependent diabetes mellitus is associated with renal failure, diabetic retinopathy, neuropathy and vasculopathy. We report the first successful simultaneous pancreas-kidney transplant in India in a young diabetic with renal failure. The dual transplant has cured his diabetes and renal failure and has had a beneficial effect on his neuropathy, retinopathy and quality of life. Obstacles to dual transplant in India include a lack of suitable recipients and a cadaver donor programme that is still in its infancy.

Distribution of C282Y and H63D mutations in the HFE gene in healthy Asian Indians and patients with thalassaemia major.

BACKGROUND: Mutations in the HFE gene have been shown to be strongly associated with hereditary haemochromatosis, an autosomal recessive disease of iron overloading. The majority of patients with hereditary haemochromatosis possess a homozygous mutation C282Y that disrupts the binding of the HFE gene with beta2 microglobulin and prevents its surface expression. Another HFE mutation H63D is known to increase the relative risk of developing hereditary haemochromatosis. This disease is rare in India although secondary haemochromatosis is commonly seen among children suffering from thalassaemia major. The status of HFE mutations has not been explored among Indians, particularly in patients with thalassaemia major. It is also possible that in India clinical haemochromatosis could be masked by iron deficiency. METHODS: We examined a cohort of 59 unrelated, healthy individuals from north India, 57 from south India and 75 thalassaemia major patients from north India for HFE mutations (C282Y and H63D) in cis/trans by the polymerase chain reaction sequence-specific primer method. RESULTS: The C282Y and H63D mutations in the HFE gene were rare among Indians. Although the HFE mutations were increased among patients of thalassaemia their effect on iron burden or disease pathogenesis remains unclear. CONCLUSIONS: Hereditary haemochromatosis is rarely observed among Indians and so are the C282Y and H63D mutations in the HFE gene. Long-term follow up studies would be required to determine whether the relatively higher frequency of these mutations among patients of thalassaemia has any influence on iron accumulation.

Biodegradation of wastewater of Najafgarh drain, Delhi using autochthonous microbial consortia : a laboratory study.

There are seventeen drains, which discharge their untreated urban and industrial wastewaters into the Delhi segment of river Yamuna. The Najafgarh drain is the first and the largest drain, and it alone contributes 1667.84 mld i.e. 60% of the total wastewater discharge into the river Yamuna and as such add 81.36 tons of BOD load per day. As per the available data approximately 95% of the wastewater of this drain is biodegradable. In the present study, an attempt has been made to reduce the BOD load and COD levels of wastewater of Najafgarh drain using autochthonous microbial consortium. During this study the raw wastewater samples were treated for 6 h time interval with different concentration of consortium. It was observed that by increasing the existing microbial population in the wastewater sample by 150-200% there is a significant decrease in BOD and COD levels. Finally, BOD/COD ratios before and after biotreatment have been analyzed to assess the efficacy of the natural consortium.

Influence of HLA compatibility on renal graft survival using live unrelated & cadaver donors in India.

BACKGROUND & OBJECTIVES: Living unrelated donor (LURD) renal transplantation has shown a rising trend over the last 5 yr at our center following the passing of The Transplantation of Human Organs Act by the Government of India in 1994. In this paper, the results of LURD and cadaver (CAD) donor renal transplantation are compared. We have also looked into factors that have a bearing on graft survival such as the extent of HLA mismatch (MM), infections, acute rejections (AR), donor age and sex. METHODS: A total of 42 LURD and 25 CAD renal transplants performed between March 1994 and February 1999 has been included in the study. HLA typing, panel reactive antibody (PRA) screening and T and B cell cross match assay were performed by the complement dependent cytotoxicity (CDC) method for all patients. RESULTS: The graft survival rates were generally higher in the LURD category as compared to the CAD group and were significant at 6 month period (90 vs 56%, P = 0.002). A follow up of the patients up to 60 months revealed a matching effect since the 3, 4 allele MM group had better survival rates as compared to the 5, 6 MM group. Twenty six of the 67 recipients (39%) experienced episodes of acute rejection (AR). Patients with 3, 4 MM had fewer such episodes than those with 5, 6 allele MM (P < 0.05). Of the 32 deaths, 20 were those with a functional kidney, of which 15 were caused by severe infections. INTERPRETATION & CONCLUSION: Better HLA matching ensures fewer episodes of rejection and better long term graft survival in comparison to the poorly matched grafts. The graft survival for LURD recipients was appreciably higher than that of CAD recipients.

Polymorphism in heat-shock protein 70-1 (HSP70-1) gene promoter region and susceptibility to tuberculoid leprosy and pulmonary tuberculosis in Asian Indians.

Heat shock proteins (HSP) act as immunological target structures either by themselves because of an unusual expression pattern, or they are carrier proteins for immunogenic peptides. A three-allele polymorphism of HSP70-1 promoter region was analysed in random patients with pulmonary tuberculosis (PTB), or with tuberculoid (TT) leprosy and healthy controls from North India. HSP70-1A and HSP70-1C occurred more frequently (> 60%) while HSP70-1B occurred infrequently in this population. Only HSP70-1A allele was significantly increased in TT leprosy as compared to healthy controls (91.8% Vs 71.1%, Pc < 0.03, RR = 4.58). Although a strong association of HLA-DR15 was observed with both of these patient groups in earlier studies, no correlation was found between HSP70-1 promoter alleles with any of the HLA allotypes. Amongst six possible genotype combinations of HSP70-1 promoter allele, only four (A/A, A/B, A/C, C/C) were encountered in Asian Indians. A significant increase of HSP70-1 A/C genotype was observed among DR15 negative PTB patients as compared to DR15 negative controls (87.5% Vs 35.7%, X2 = 8.6, Pc < 0.02) giving highest relative risk of 12.6. These findings suggest that HSP70-1 genes may play a secondary role to HLA-DR in governing susceptibility to mycobacterial infectious diseases.

Cold reactive lymphocytotoxic antibodies in pulmonary tuberculosis: correlation with disease activity and HLA.

Cold reactive lymphocytotoxic antibodies (LCA) are more reactive in cold than at 37 degrees C and occur following infection, immunization or vaccination and in various autoimmune diseases. In the present study, LCA activity against T and B-lymphocytes has been investigated in patients with pulmonary tuberculosis (PTB), their various clinical sub-groups and consanguineous relatives. Further, the relevance of HLA factors in LCA activity was analyzed. The sera from 144 PTB patients, 52 family contacts and 52 healthy individuals were tested for presence of LCAs by a modified two-stage NIH microlymphocytotoxicity assay. A significant increase in LCA activity against both T (32.6% vs 5.7%, P < 0.0001) and B (59.7% vs 13.4%, P < 0.0000001) cells was observed in PTB patients as compared to healthy controls. There was no correlation between serum LCA activity and sputum acid-fast bacilli status. However, only B cell LCAs revealed significant increase in parallel to disease advancement as assessed by X-ray chest examination. Further, LCA activity was more pronounced in drug responders than drug failure group of patients. No significant difference in the distribution of HLA class I and class II antigens was observed between LCA positive and LCA negative patients. However, panel cells carrying HLA-A1, -A11 and -DR3 were often found reactive in LCA positive patient sera. In household family contacts, LCAs were significantly increased only against B cells as compared to healthy controls (38.4% vs 13.4%, P < 0.01). This study suggests that Mycobacterium tuberculosis infection/exposure could account for the occurrence of LCAs in pulmonary tuberculosis and the strength of these antibodies is related to disease severity and the extent of lung involvement.

Genetic Diversity Of HLA Polymorphism And New Genes.

DNA level studies, based on SSOP hybridization of HLA class II PCR products have revealed considerable diversity in HLA among Asian Indians. High resolution typing of specific alleles such as DR2 and DR4 in the HLA class II region and their DR-DQ haplotypes have helped to detect unique haplotypes and novel alleles which have subsequently been confirmed by sequencing. Incidentally, remarkable stability is maintained in several other DRBI alleles, viz DR1, DR7, DR9 and DR10. Further characterization of new alleles will be carried out by sequencing of mRNA. The ARMS-PCR technology has been found to be particularly useful for typing difficult HLA-A, HLA-B and all HLA-Cw alleles. These technologies are remarkably superior over serological methods. Our studies have shown appreciable heterogeneity of common HLA-A and B alleles in Asian Indians. Molecular subtypes of HLA-A2 revealed that subtype A*0211 is found only in the Indian population and may be the result of a selection pressure in this population. Investigations into polymorphism in the HLA-B27 gene revealed that subtypes common both to the western Caucasoids and orientals occur in the Indian population. It is apparent that the population of the Indian subcontinent, placed as it is between the Caucasoids and Negroids on one hand and Australoids and Mongoloids on the other, provides a rich source of many HLA haplotypes. While the most frequent Caucasian haplotypes occurred with a reasonable frequency in Asian Indians, those found predominantly in other ethnic groups (e.g., Australian Aborigines and populations of Oceania, China and Japan) are also detected. Knowledge on this is most important for donor selection during organ and bone marrow transplantation and for designing MHC targeted vaccines in specific diseases. The major histocompatibility complex (MHC) encompasses two major classes of molecules: the MHC classes I and class II, both of which appear to have originated from a common ancestor gene. The major biological function of the MHC is to bind peptide fragments derived from protein antigens (viruses, peptides etc) and display them on the surface of antigen presenting cells (APCs), evoking effector responses upon recognition by the antigen specific receptors on T lymphocytes. This process requires an efficient intracellular machinery to fragment the protein antigens into smaller peptides capable of binding to a host MHC molecule. Since the number of peptides that can theoretically be generated is very large, there is need for an extensive MHC gene pool. Thus the HLA system which is the MHC of man is extremely polymorphic. Although the general rules for peptide-MHC interactions for both classes of MHC molecules are essentially similar; there are two fundamental differences: i) MHC class I ligands originate from endogenous sources, mainly from proteins of the cytosol or the nucleus and are delivered by the 'endogenous processing pathway: In contrast, the MHC class II ligands are generated by the degradation of proteins from the extracellular compartment. These distinctions are also reflected in the responding T cells: CD8 positive T cells being restricted by class I-peptide complexes, and CD4 positive T cells by class II-peptide complexes. ii) In accordance with the distribution of hydrogen bonds in the peptide binding groove of the MHC, the anchor residues are placed at the terminal ends of the class I groove. Contrarily, the binding forces are distributed throughout the class II groove and ensure bonds between the peptide's backbone and class II molecule (Madden et al, 1993; Stern et al, 1994; Stern and Wiley 1994). The specificity of the interaction is determined by pockets in the MHC groove that have a fixed spacing from each other and which also have specificity for anchoring particular side chains of the peptide's amino acids.

HLA class I profile in Asian Indian patients with pulmonary tuberculosis.

HLA class I antigen profile was studied in 153 unrelated patients with pulmonary tuberculosis (PTB), 40 family contacts and 289 healthy individuals by the NIH microlymphocytotoxicity test to find out the role of HLA-A, -B, -C alleles in influencing susceptibility to PTB and its various clinical groups. HLA-A2 was found to be significantly increased in the total patient group as compared to controls (38.6% vs 26.3%, p < 0.01, RR = 1.76). The increase of HLA-A2 was more pronounced in the sputum negative patients (59.4%, pc < 0.001, RR = 4.1) suggesting its possible role in the mediation of CD8+ suppressor T cell activity against Mycobacterium tuberculosis, resulting in the development of limited disease in these patients. Further, HLA-B18 was found to be decreased in patients as compared to controls (2.6% vs 7.3%, p < 0.05, RR = 0.34). None of the class I antigens was associated with the dynamics of chemotherapy or disease severity as assessed by the extent of lung involvement on chest X-ray examination.

Restriction fragment length polymorphisms in HLA-DR4-DQ3 haplotypes associated with rheumatoid arthritis.

Restriction fragment length polymorphism (RFLP) patterns were studied in serologically confirmed DR4-DQ3 positive patients with rheumatoid arthritis by Southern blot analysis using full length cDNA probes specific for DRB, DQA and DQB hybridized with genomic DNA digested with informative restriction endonucleases. The RFLP patterns correlated with serology confirming all patients to be DR4+ve. The DQB1*0302 (DQ8) allele identified by 12.0kb BamHI, 3.3kb Hind III and 1.8kb Taql fragments was present in all patients suggesting them to be DR4-DQB1*0302. Hybridization of Taq 1 and PVU II digested genomic DNA with DQA cDNA probe revealed four informative RFLP patterns. While three of them correlated with known DR4 subtypes, one was a new polymorphism observed specifically in Indian patients with rheumatoid arthritis. The study further indicated that two of the several known subtypes of DR4, viz., DRB1*0401-DW4-DQB1*0302 and DRB1*0404-DW14-DQB1*0302 may be implicated in susceptibility to rheumatoid arthritis in the Indian population.

Effect of D-penicillamine on lymphocyte subsets: correlation with clinical response in rheumatoid arthritis.

We studied the effects of D-penicillamine (DP) on the clinical response, immunoinflammatory parameters and the lymphocyte subsets in 46 patients with rheumatoid arthritis (RA). Patients were evaluated before the start of the drug and then at 3 and 9 months during the follow up. 38 of 46 (82.6%) patients could continue DP treatment for over 9 months, while in 8 the drug was withdrawn due to adverse effects. Improvement in the various disease activity indices of more than 50% (responders) was seen in 25 of 38 (65.8%) patients. Responders showed a significant decrease in the serum IgA and IgM at 9 months, and in IgM only at 3 months. The serum levels of C3 and C4 did not show any significant change. Serum levels of C-reactive protein and rheumatoid factor (RF) showed a significant decrease at 3 and 9 months. A significant decrease in CD3+ and CD4+ lymphocytes along with a fall in CD4+/CD8+ lymphocyte ratio was also seen in responders at 3 and 9 months, compared to the baseline. Our results suggest that DP may have immunomodulatory action in RA.

Dermatoglyphic patterns of patients with rheumatoid arthritis.

Finger tip and palmar dermatoglyphics were studied in 31 patients (22 females and 9 males) with rheumatoid arthritis (RA) and 38 matched controls (20 females and 18 males) from North India. While not many differences were observed in palmar patterns, a low ending of line A was found on both hands of two patients. Finger tip patterns were significantly different in patients compared to controls. No association with any dermatoglyphic feature and HLA antigens was observed.

Protective & risk DR phenotypes in Asian Indian patients with rheumatoid arthritis.

This study of 168 north Indian patients with rheumatoid arthritis (RA) confirms the significant association of susceptibility to RA with DR4 specificity (P less than 0.0001). This association was observed equally in familial as well as sporadic patients. The HLA-DR2 and DR5 alleles were identified to be conferring protection in RA, DR5 being reduced significantly in the non-familial patients only. None of the other DR antigens revealed any association with RA in this population, including the DR4 negative group of patients. An analysis of the DR phenotypes in patients and controls revealed that DR4 in combination with DR1 provided the highest relative risk (71.9) followed by DR4, DR4 (RR = 4.1). These results demonstrate that susceptibility to RA is not due to a single HLA specificity but the effect of a group of related epitopes occurring in common among subtypes of DR4 as well as in some DR1 alleles.

HLA-B5 and B21 antigens in aortoarteritis.

The distribution of class I HLA antigens (HLA-A, B) were determined in 50 patients of Aortoarteritis in an Indian population. This included 29 females and 21 males. The difference in antigen frequency was observed between patients and controls with reference to HLA-A19, B5 and B21 antigens. A decreased frequency of HLA-A19 was observed in the patients as compared to controls (14% vs 33.25%, X2 = 6.81, P less than 0.025). Of the B locus antigens, an increased frequency of HLA B5 was observed in the patients as compared to controls (48% vs 29.5%, X2 = 6.2, P less than 0.025). HLA-B21 was also increased in the patients as compared to the controls (18% vs 6.5%, X2 = 6.67, P less than 0.025). These data suggest the involvement of genetic factor (s) in the aetiopathogenesis of this disease. Further, the observations indicate that HLA-B5 and B21 may be associated with Aortoarteritis.

Immunologic and immunogenetic studies in rheumatic fever and rheumatic heart disease.

In order to evaluate all the important limbs of the immune system in the same patient population with rheumatic fever (RF) and rheumatic heart disease (RHD) cellular and humoral immune parameters as well as the immunogenetic profile in 265 North Indian patients with RHD were evaluated. They were studied for class in HLA antigens and 165 of them were also evaluated for the class II (DR locus) antigen profile. Data obtained was compared with 400 and 134 healthy controls respectively of the same ethnicity. Humoral immune parameters (Serum immunoglobulins IgG, IgA; Serum complement fractions C3, C4, C3d; circulating immune complexes and B lymphocyte numbers) and cellular immune parameters (total leucocyte and lymphocyte counts; T lymphocyte sub-populations--CD4, CD8 counts; lymphocyte migration inhibition to an extracellular streptococcal antigen, streptolysin 'O') were studied in 23 patients with RF, 21 patients with "inactive" RHD and 20 normal controls. Patients of RHD were noted to have an increased frequency of DR3 (P less than 0.001; Relative risk = 2.3) and a decreased frequency of DR2 (P less than 0.001; Relative risk = 0.3) as compared to the controls. Patients of RF had evidence of an altered regulatory T cell function (increased CD4/CD8 ratio) and decreased cell mediated immunity to streptolysin 'O'. An increased humoral immune response (increased B cell counts, elevated serum IgG, circulating immune complexes and C3d) was noted in patients of RF as well as "inactive" RHD. An integrated pathogenetic model with immune response associated antigens of the DR locus influencing selection of cardiac cross-reactive antigens by the antigen processing macrophages, an altered regulatory T cell function with decreased suppressor T cell activity leading to an abnormal immune response is proposed to explain the pathogenesis of RF.

Lymphocytotoxic antibodies in patients with systemic lupus erythematosus & their household contacts.

Twenty patients with systemic lupus erythematosus (SLE) and their 51 household contacts were screened for the presence of lymphocytotoxic antibodies. A high prevalence of lymphocytotoxic antibodies (LCTAB) was observed in the patients (80% against T cells and 100% against B cells). The antibodies carried specificity for HLA-B5/35 and HLA-DR2/DR3. A high prevalence of LCTAB was also observed in contacts (55% LCTAB-T cells, 88% LCTAB-B cells) with no difference being seen between consanguineous and non-consanguineous male or female relatives.