Biomedical subjects
Publications and source records attributed to Shrimati Shetty.
Explore the source record for details and available documents.
During the last 10 years (1995-2005) we have offered diagnosis in 438 families out of 502 families referred, by chorionic villus sampling procedure between 10 and 12 weeks of pregnancy in severe haemophilia A and B families. Sixty four families could not be offered a diagnosis in the first trimester either due to non-informativeness with the available techniques or due to the absence of affected members in the family and were subsequently offered diagnosis in the second trimester by direct analysis of factor VIII/IX clotting activity and antigen in the fetal blood samples. For first-trimester diagnosis in the chorionic villus samples, we have used both direct and indirect methods, that is, intron 22 and 1 inversions in the factor VIII gene, a multiplex PCR for the detection of gross deletions in the factor IX gene and RFLP analysis using a battery of markers within and outside the factor VIII/IX gene. The informativeness of all these techniques was found to be 92.21% in case of haemophilia A and 83.91% in case of haemophilia B. We followed up 122 children out of 326 (diagnosed unaffected prenatally) after birth and the diagnosis was adequately reconfirmed by both factor assays and by DNA analysis. Only one case of misdiagnosis was detected so far (0.22%), where the diagnosis was based only on the extragenic marker of the factor VIII gene.
Explore the source record for details and available documents.
von Willebrand disease (VWD) is caused by qualitative or quantitative defects, or both, of the von Willebrand factor, a multimeric, high-molecular-weight glycoprotein. Type 2N VWD differs from other subtypes in that it shows normal primary haemostasis as all the routine tests performed for the diagnosis of VWD are normal. They present with defective coagulation, due to a moderate or severe deficiency of factor VIII, and consequently often they are misdiagnosed as either mild haemophilia A or type 1 VWD. Thus the actual prevalence of this subtype is not known in many of the populations. The differential diagnosis of this subtype is mainly based on the in-vitro capacity of plasma von Willebrand factor to bind exogenous factor VIII by immunoassay. However, most of the routine laboratories in developing countries do not have the testing facilities for type 2N VWD, mainly because of the cost of the reagents involved and the complexity of various assays reported. We report here the prevalence of type 2N VWD in an Indian population by developing a simple, not too expensive, user-friendly enzyme-linked immunosorbent assay technique and compared it with the reported technique of the factor VIII binding assay. Nine out of 97 VWD patients (10%) were found to be type 2N VWD using both the reported assay and the in-house ELISA; thus indicating that the prevalence of type 2N VWD is high in our population. The simple ELISA technique described here should be available in all routine laboratories in developing countries.
The efficacy of the two von Willebrand factor (vWF) intron 40 variable number of tandem repeat (VNTR) markers, vWF1 and vWF2, in the genetic diagnosis of von Willebrand disease (vWD) in Indian patients was studied. Three hundred and sixty-five unrelated normal individuals and 100 vWD patients (type 1: 18; type 2: 21; type 3: 61) were analyzed for the two vWF intron 40 VNTR markers. Polymerase chain reaction of the two markers vWF1 and vWF2 was done using specific primers followed by electrophoresis on 10% polyacrylamide gel. VNTR analysis revealed the presence of VNTR9 and VNTR15 along with the eight alleles VNTR6 to VNTR14 in the vWF1 marker. Furthermore, apart from the six alleles, VNTR1 to VNTR6 of the vWF2 marker, two new alleles, VNTR7 and VNTR8, were also found. The heterozygosity rates were 75 and 74% for vWF1 and vWF2, respectively. Overall, the heterozygosity rate, i.e. when both vWF1 and vWF2 were considered in combination, was 81%. These data were successfully applied for the detection of carriers in 10 severe type 3 vWD families. The high heterozygosity of the two vWF intron 40 VNTR markers and the simplicity of the technique without much cost being involved suggest the practical feasibility of this technique in developing countries like India.
Explore the source record for details and available documents.
Combined deficiency of factors V (FV) and factor VIII (FVIII) (F5F8D) is an autosomal recessive bleeding disorder caused by simultaneous moderate-to-mild decrease of both clotting proteins. Mutations in two components of the ER-Golgi intermediate compartment (ERGIC-53), i.e., lectin mannose binding protein (LMAN1) and multiple coagulation factor deficiency 2 (MCFD2), have been found to be responsible for this dual deficiency in most of the cases reported in literature. Three Indian families with F5F8D were analyzed for the presence of mutations in their LMAN1 and MCFD2 genes. One of the three families showed the presence of a G to A substitution in exon 2 of the MCFD2 gene, whereas another family showed a nonsense mutation, i.e., G to T substitution, in exon 2 of the LMAN1 gene, the latter being a novel mutation not previously reported. The third family did not show mutations in either of the two genes, suggesting that a significant subset of F5F8D cases may be due to additional genes resulting in a similar phenotype.
BACKGROUND AND OBJECTIVE: von Willebrand disease (VWD) is one of the most common inherited bleeding disorders in the west. Limited studies from India showed a prevalence of approximately 10 per cent of VWD among the cases with hereditary bleeding disorders. VWD remains an underdiagnosed entity in India. The prevalence of different subtypes of VWD is also not known which is essential for a proper management of these cases. The present study was thus undertaken to know the prevalence of VWD and its various subtypes in the western part of our country. METHODS: A total of 796 consecutive patients presented with various bleeding manifestations were analysed. The initial screening and confirmation tests for the diagnosis of VWD included bleeding time (BT), screening coagulation tests i.e., prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), factor VIII: C assay, ristocetin-induced platelet aggregation (RIPA) and VWF antigen (VWF:Ag) estimations. VWF multimer analysis, ristocetin cofactor activity (RCOF), VWF collagen binding assay (VWF: CBA), factor VIII : VWF binding assay were also done to classify and subtype these cases. RESULTS: The patients were subtyped as per the International Society of Thrombosis and Haemostasis (ISTH) criteria. Of the 796 patients screened, 58 were diagnosed as VWD. Of the 15 families with a positive family history of bleeding, 26 additional cases were diagnosed as VWD. Majority of the patients were type 3 (59.5%) with severe clinical manifestations, about 18 per cent of type 1 VWD patients were detected in this group while the prevalence of the qualitative variants of VWD i.e., type 2 VWD was found to be 19 per cent and the prevalence of various subtypes were type 2A (9.52%), type 2B (4.76%), type 2M (1.2%), type 2N (3.6%). INTERPRETATION AND CONCLUSION: The high prevalence of type 3 and a low prevalence of type 1 VWD which is in contrast to the western reports, suggests the low awareness of the disease as also the underdiagnosis of the mild cases in our country.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
BACKGROUND: The pathogenesis of arterial thrombotic disease involves multiple genetic and environmental factors related to atherosclerosis and thrombosis. But, there have been very few studies in India which have investigated some of the thrombophilia markers. AIM: To look for combined thrombophilia in MI patients. SETTINGS AND DESIGN: One hundred twenty patients of myocardial infarction (age below 40 yrs.) were recruited 8-10 weeks after stabilization. Hundred age and sex-matched healthy controls were also recruited in the present study. METHODS AND MATERIAL: Following thrombophilia markers were screened in these patients--plasma fibrinogen, protein C, protein S, antithrombin III, factor V Leiden, PT G20210A polymorphism, MTHFR C677T, homocysteine, fibrinogen b448 Arg/Lys polymorphism and CBS T833C mutation. STATISTICAL ANALYSIS: Statistical analysis was done using Statistical Package for Social Sciences (SPSS) version 10.0, SPSS Inc., Chicago, USA. RESULTS AND CONCLUSION: Elevated fibrinogen levels, homocysteine (p< 0.001 and homocysteine with odds ratio 6.26) and factor V Leiden (p=0.038) were independently associated with MI in our patients. A total of 37 patients (42.5%) had the presence of more than one thrombophilia markers in combination. Out of these, 10 had the presence of three markers in combination and 1 had five thrombophilia markers in combination. Only 2 controls had prothrombotic markers in combination. Combined prothrombotic risk factors were significant in cases in comparison to controls (p< 0.001). Further larger studies on a nationwide basis recruiting a large number of young MI patients should be done to substantiate these findings.
We report a novel 465T-->C (S123P) mutation in exon 3 of the GPIIIalpha gene in a patient with type III or variant Glanzmann's thrombasthenia (GT). Though this mutation did not affect fibrinogen binding to GPIIb-IIIalpha in activated platelets, it interfered with the platelet aggregation in a manner similar to GT.
Chronic synovitis affects about 10% of patients with severe haemophilia in India. This disease has some features in common with ankylosing spondylitis, which has been linked to HLA B27. We therefore aimed to test whether there is an association between HLA B27 and chronic synovitis. We studied 473 patients with severe haemophilia (33 of whom had chronic synovitis), and 1175 healthy controls using a standard serological technique and the reverse line strip assay. 64% (21 of 33) of patients with haemophilia and chronic synovitis were positive for HLA B27, compared with 5% (23 of 440) of those with severe haemophilia, but not chronic synovitis (odds ratio 31.6 [95% CI 9.28-39.38], p<0.0001), and 9% (100 of 1175) of healthy controls (18.81 [9.6-27.7], p<0.0001). We conclude that there is a strong association between HLA B27 and chronic synovitis in Indian patients with severe haemophilia and screening in this population could allow treatment and prevention of the complication.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Large numbers of activated partial thromboplastin time (aPTT) reagents are sold in the market. The phospholipid content and its source, nature and the amount of activators are highly varied in different aPTT reagents. The present study was undertaken to evaluate which of the four aPTT reagents commonly used is suitable as an all-purpose reagent for a modest haemostasis laboratory. Four aPTT reagents (reagent A, Platelin LS; reagent B, Silimat; reagent C, Actin FSL; reagent D, CK Prest) were tested against 75 different plasmas obtained from normal patients as well as from patients with different haemostatic problems. All the tests were conducted by one of us (S.S.) in duplicates. Different aPTT reagents missed different proportions of mild factor VIII and factor IX deficiency (36.4, 18.2, 4.6 and 13.6% for reagents A, B, C and D, respectively) and showed abnormal results with normal plasmas (i.e. more than 5 s prolongation) (29.2, 25, 8.3 and 12.5% for reagents A, B, C and D, respectively). All the reagents faithfully picked up moderate and severe factor VIII and factor IX deficiency. There was no difference among the four aPTT reagents regarding their ability to prolong aPTT to therapeutic dosage of heparin or in their ability to give comparable factor VIII or factor IX levels in one-stage aPTT-based assays. There were differences in aPTT reagents in their ability to pick up mild deficiency of coagulation factor VIII and factor IX. Some reagents showed abnormal aPTT results in mild cases of factor VIII and factor IX deficiency without producing a large number of falsely prolonged aPTT with normal plasma.
An enzyme-linked immunosorbent assay (ELISA) has been developed to measure factor VIII antibodies in haemophilia patients. The assay utilizes binding of the antibodies in the plasma to solid phase antigen, i.e. recombinant factor VIII which was subsequently detected by a human polyclonal IgG labelled with the alkaline phosphatase-p-nitrophenyl phosphate substrate system. Comparisons were made with the Bethesda assay for the quantitation of factor VIIII inhibitors. Dose response curves for the reference standards were consistently linear and reproducible. The assay was specific for factor VIII antibodies, showing a negative reaction for antibodies to other coagulation factors, antinuclear factors and antiphospholipid antibodies. Using this method, 312 samples from haemophilia A patients and 31 samples from healthy controls were screened for the presence of inhibitors and compared with the conventional Bethesda assay. Twenty-four cases were found to be positive for inhibitors in both the ELISA and Bethesda assay. Five additional cases were also found to be positive in the ELISA assay, which, however, were negative in Bethesda assay. One patient who was initially positive for factor VIII inhibitors both by the Bethesda assay and ELISA eventually became negative for the Bethesda assay (<0.5 BU/ml) but was still positive for the ELISA assay. The ELISA thus described had a specificity of 97.8% and a sensitivity of 100% when tested against a large cohort of haemophilia A samples.