Biomedical subjects
K Pai
Publications and source records attributed to K Pai.
Immunochromatographic strip-test detection of anti-K39 antibody in Indian visceral leishmaniasis.
Stored sera from 429 Indian subjects were assayed to extend the analysis of the accuracy of immunochromatographic strip-test detection of anti-K39 antibody in the non-invasive diagnosis of visceral leishmaniasis (VL). All 225 samples from patients with proven Leishmania infection tested positive [estimated sensitivity = 100%; 95% confidence interval (CI)=98%-100%]. Sera from 99 of the 100 symptomatic patients with other diseases were non-reactive (estimated specificity = 99%; CI = 94%-100%). However, samples from 13 of the 104 apparently healthy controls showed positive strip-test results (estimated specificity = 88%; CI = 79%-93%), yielding an overall specificity of 93% (190/204; CI = 88%-96%). If applied in a practical clinical setting (on symptomatic patients in whom active VL is suspected and other common infections have been excluded), strip testing of serum for anti-K39 antibody should be both sensitive and specific for diagnosing VL in India.
Identification of four novel mutations in classical Menkes disease and successful prenatal DNA diagnosis.
Menkes disease is an X-linked recessive disorder of the copper metabolism and affected males suffer a systemic copper deficiency due to malabsorption and defective distribution of dietary copper. It is caused by a defect in the Menkes (ATP7A) gene, which encodes a transmembrane copper-transporting P-type ATPase. A variety of mutations were reported; however, only a few mutations were reported in Asian patients. We identified four novel mutations and one known mutation in five Korean patients. Arg646Ter in exon 8, a novel mutation transmitted from his carrier mother, was identified in one patient. Prenatal DNA diagnosis on an unaffected fetus in this carrier mother was successfully accomplished. An additional three novel mutations, Leu706Arg in exon 9, Gly1118Asp in exon 17, and Gly1255Arg in exon 19, were identified. Splicing mutation was not identified. Menkes disease in Korean patients appears to be caused by heterogeneous mutations with different spectrums from Caucasian patients.
Reactive oxygen intermediates, nitrite and IFN-gamma in Indian visceral leishmaniasis.
Reactive oxygen intermediates (ROI), nitrite and interferon-gamma (IFN-gamma) production were investigated at different times during treatment in 10 patients with visceral leishmaniasis (VL). Hydrogen peroxide (H2O2), superoxide (O2-) and IFN-gamma production by cultured monocytes from patients with active VL were significantly lower compared with the healthy controls. In contrast, nitrite levels in the supernatants from monocyte cultures of VL patients were comparable to healthy controls and increased significantly during antileishmanial therapy. On day 20 of treatment, a significant increase in the release of H2O2, O2- and IFN-gamma was observed. However, at follow-up, 4 months after the end of treatment, the production of H2O2, O2-, IFN-gamma and nitrite had declined significantly. Thus, the impairment in hydrogen peroxide and superoxide production suggests that down-regulation of these mediators may be involved in the reduced killing of parasites by monocytes of active VL patients. Furthermore, the monocytes regained respiratory burst activity as the antileishmanial therapy progressed, suggesting that an immune-based mechanism is involved in successful drug therapy.
Enzyme-linked immunosorbent assay for recombinant K39 antigen in diagnosis and prognosis of Indian visceral leishmaniasis.
The recombinant product (rK39) of the 39-amino-acid repeats encoded by a kinesin-like protein-encoding gene of Leishmania chagasi was evaluated by enzyme-linked immunosorbent assay (ELISA) for diagnostic potential and the ability to predict the response to therapy in Indian kala-azar or visceral leishmaniasis (VL); we also compared its performance with that of crude soluble antigen (CSA). At the diagnosis of VL, the anti-rK39 antibody titer was 59-fold higher than the anti-CSA antibody titer. With successful therapy, antibody titers declined steeply at the end of treatment and during follow-up. In contrast, patients who relapsed showed increased titers of antibodies to rK39. The extremely high levels of anti-rK39 antibodies in VL cases suggest the application of rK39 for sensitive and specific serodiagnosis, and rK39 ELISA is also valuable in monitoring drug therapy and detecting relapse of the disease.
Resistance to treatment in Kala-azar: speciation of isolates from northeast India.
Kala-azar in India is becoming increasingly difficult to treat, which may be due to the presence of species other than Leishmania donovani; Leishmania tropica was reported to cause the same clinical syndrome in the area. Over the past 3 years, we have collected samples from 241 patients with visceral leishmaniasis from across the region. Of the 189 isolates that grew on diphasic medium, 159 were successfully transferred to liquid medium for typing. Clinically, 80% of these were resistant to antimony. Lipophosphoglycan-specific monoclonal antibodies were used to distinguish the 2 species by agglutination of promastigotes; all 159 were shown to be L. donovani. Eighty-three isolates were confirmed to be L. donovani by isoenzyme analysis, by amplification of kinetoplast DNA, or both, in comparison with multiple reference strains; none were L. tropica. Thus, the rising incidence of clinical resistance to treatment is unlikely to be due to a different species causing kala-azar in north Bihar.
A simple and inexpensive "cell dissociation sieve-tissue grinder" apparatus.
A simple and inexpensive cell dissociation sieve-tissue grinder apparatus consisting essentially of stainless steel sieve (the one popularly used for sieving tea leaves) and a glass syringe plunger acting as pestle, is described for making single cell suspension.
Kala-azar epidemic in Varanasi district, India.
Reports at the Sir Sunder Lal Hospital, Banaras Hindu University, of a large number of kala-azar cases from one particular village in Varanasi district, Uttar Pradesh, led us to carry out an epidemiological study of the situation using standard techniques. The overall prevalence and case fatality of the disease were 12.9% and 10.5%, respectively. A history of fever and hepatosplenomegaly was noted for all the cases. The case definition was the presence of parasites in bone marrow or splenic aspirate smears. The disease was more prevalent among adults, but occurred also among children. However, there was no clear linear relationship between the prevalence of the disease and age group. Kala-azar occurred among males and females, and its prevalence did not correlate significantly with income. Since the disease vector continues to be present in the study area, the health authorities should take strong steps to control the disease.
The role of cyclosporin A on antibody-dependent monocyte-mediated cytotoxicity against human multidrug-resistant cancer cells.
A P-glycoprotein (P-gp) inhibitor, cyclosporin A (CsA) was found to enhance the susceptibility of multidrug resistant (MDR) cancer cells to anti-P-gp antibody-dependent cellular cytolysis (ADCC) by monocytes, but the exact mechanism is unknown. In this study, we examined whether CsA enhanced the susceptibility of MDR cells through its inhibitory effect of P-gp function by using anti-ganglioside GM2 (GM2) monoclonal antibody (Ab), KM966, instead of anti-P-gp Ab, MRK16. Monocyte-ADCC induced by both KM966 and MRK16 against P-gp positive human MDR ovarian cancer cells was significantly augmented by addition of CsA. KM966, but not MRK16, induced monocyte-ADCC against P-gp negative human ovarian cancer cells and CsA enhanced this ADCC activity, indicating that suppressive effect of P-gp function by CsA was not essential to the enhancement of ADCC. Moreover, pretreatment of tumor cells with CsA augmented their susceptibility to monocyte-ADCC irrespective of P-gp expression. Interestingly, KM966 or MRK16 induced monocyte-ADCC against various human lung cancer cells expressing either GM2 or P-gp, but CsA did not affect these ADCC. These findings suggest that CsA may enhance the susceptibility to the monocyte-ADCC of ovarian cancer cells, but not of lung cancer cells, irrespective of its suppressive effect of P-gp function.
Differential effect of interleukin 10 on interleukin 12- and interleukin 2-mediated killer induction from blood mononuclear cells.
We investigated the regulatory effect of interleukin (IL) 10 on IL-12-inducible killer activity of blood mononuclear cells (MNC) of healthy subjects. IL-10 suppressed non-MHC-restricted killer activity induced by IL-12 in a dose-dependent manner. Cytotoxicity against Daudi cells of blood MNC activated with optimal dose (100 U/ml) of IL-12 was significantly suppressed from 36.6 +/- 6.8% (mean +/- SE) to 26.0 +/- 5.0% by 100 ng/ml of IL-10 (n = 5; P < 0.05) at an E/T ratio of 20. IL-10, however, did not suppress killer induction by IL-2 from blood MNC. On the other hand, IL-4, a known inhibitor of IL-2-mediated killer induction, had no effect on IL-12-mediated killer induction from blood MNC. Moreover, production of interferon-gamma by IL-12-stimulated MNC was suppressed by IL-10. Because the presence of IL-10 is reported in the tumor-growing site, these observations suggest that IL-10 acts as a key cytokine in the suppression of IL-12-mediated antitumor response in situ.
Activation of P388D1 macrophage cell line by chemotherapeutic drugs.
It has been found that certain antineoplastic drugs impart their function with a distinct duality. Besides being tumoricidal, they are capable of acting as immunopotentiator. This led us to investigate the effect of cytosine arabinoside (CA), vincristine sulphate (VS), cyclophosphamide (CS), mitomycin C (MMC), hydroxy urea (HU) and lipopolysaccharide (LPS) on a macrophage cell line P388D1. Supernatants collected from P388D1 cells treated with CA, VS, CS, MMC, HU or LPS demonstrated enhanced production of tumor necrosis factor (TNF) confirmed by bioassay on L929 tumor target cells and increased interleukin-1 (IL-1) production by standard thymocyte proliferation bioassay. Also, supernatants showed increased amounts of nitric oxide and lysozyme using Griess reaction and reduction in turbidity of Micrococcus lysodeikticus, respectively. The above findings demonstrate that these drugs may be used not only as chemotherapeutic agents but also as macrophage-activating agents.
Novel and frequent mutations of hepatitis B virus coincide with a major histocompatibility complex class I-restricted T-cell epitope of the surface antigen.
We examined the full-length hepatitis B virus (HBV) envelope (surface antigen or HBV small surface antigen [HBsAg]) sequences of 12 different liver samples from 10 different hepatoma-containing chronic carriers. Surprisingly, novel and frequent mutations occurred predominantly at amino acids 40 and 47 of HBsAg, in addition to within a known protective B-cell epitope (so-called group a determinant of HBsAg 124-148). Approximately 58% of chronic carriers contain mutations at the group a determinant. The mutation frequency at the hotspot codons 40 and 47 is approximately 83%, 1 order of magnitude higher than at the known polymorphic sites of subtype-specific determinants at codons 122 and 160, which is approximately 4%. This new mutational domain is found to coincide with a major histocompatibility complex class I-restricted T-cell epitope. The potential biological significance of this novel mutation in the immunopathogenesis of HBV chronic carriers is discussed.
Pediatric germ cell tumor. An experience with BEP.
PURPOSE: This study is an analysis of our experience with bleomycin, etoposide, and cisplatin (BEP) chemotherapy, in pediatric germ cell tumors (GCTs). PATIENTS AND METHODS: The study included all children (age < 16 years) who were registered between May 1988 and May 1993 with a histologically confirmed diagnosis of GCT and received BEP chemotherapy. In addition to the clinicopathological features, the response rate, survival rate, and toxicity were analyzed. RESULTS: There was a total of 56 patients, of whom 22 had an ovarian tumor and 17 each had a testicular or an extragonadal tumor. Histologically, endodermal sinus tumor was the most common type (62%). Tumor markers were increased in 89% (50 of 56). Complete responses were observed in 89.1% (49 of 55) and partial responses in 10.9% (6 of 55) of the evaluable patients. Five-year actuarial survival was 83% and progression free survival was 93%. Median follow-up was 18 months. Median survival is not yet reached. The chemotherapy was well tolerated. CONCLUSIONS: From the present report, it is apparent that BEP chemotherapy is very effective and well tolerated in children with GCT. The data probably suggests that conservative surgery, when combined with effective chemotherapy, can result in cure of the majority of children with GCTs.
Influence of systemic chemotherapy on anti-P-glycoprotein antibody-dependent cell-mediated cytotoxicity in patients with small cell lung cancer.
Anti-P-glycoprotein antibody (MRK-16)-dependent cell-mediated cytotoxicity (ADCC) by blood mononuclear cells (MNC) was examined in patients with small cell lung cancer (SCLC) before and after systemic chemotherapy. The effect of in vitro treatment of MNC with interleukin (IL)-2 and macrophage-colony-stimulating factor (M-CSF) was also examined. The ADCC reaction was assessed by a 6 h 51Cr-release assay using a multidrug-resistant (MDR) SCLC cell line (H69/VP cells). The MRK-16 monoclonal antibody was able to augment spontaneous cytotoxicity by MNC, even in SCLC patients. Pretreatment of MNC with IL-2 significantly augmented their ADCC ability in SCLC patients, while M-CSF had no effect on ADCC activity. After the first cycle of systemic chemotherapy, the ADCC activity tended to decline, but ADCC of MNC pretreated with IL-2 was not affected. The results suggest that anti-P-glycoprotein antibody, in combination with a cytokine such as IL-2, may be therapeutically useful against human SCLC resistant to chemotherapeutic drugs.
Increased production of interleukin-1 and tumor necrosis factor by human monocytes treated in vitro with cisplatin or other biological response modifiers.
Supernatants collected from cisplatin, lipopolysaccharide (LPS), muramyl dipeptide (MDP) or interferon-gamma (IFN-gamma) treated human monocytes enhance the thymocyte proliferation by a submitogenic concentration of concanavalin A. Also supernatants collected from cisplatin or IFN-gamma treated monocytes demonstrated enhanced cytotoxicity against actinomycin-D treated L 929 cells, suggesting that cisplatin or rIFN-gamma treated monocytes release tumor necrosis factor (TNF) into the culture medium. The supernatant collected from untreated monocytes showed only little IL-1 and TNF activity.
Studies on the release of interleukin-1 (IL-1), tumor necrosis factor (TNF) and lysozyme from human non-adherent mononuclear cells (nMNC) in vitro after treatment with cisplatin and other biological response modifiers.
The supernatants collected from cisplatin, lipopolysaccharide (LPS), muramyl dipeptide (MDP) or recombinant interferon gamma (rIFN-gamma) treated non-adherent mononuclear cells (with NK cell activity) enhanced thymocyte proliferation by a submitogenic concentration of concanavalin A as compared to untreated nMNC. Supernatants collected from cisplatin or rIFN-gamma treated nMNC also demonstrated enhanced cytotoxicity against actinomycin-D treated L929 cells, suggesting that cisplatin or rIFN-gamma treated nMNC release tumor necrosis factor (TNF) into the culture supernatant. On the other hand, supernatant collected from untreated nMNC showed little TNF activity. Treatment of nMNC with cisplatin, LPS, MDP or rIFN-gamma resulted in enhanced release of lysozyme into the culture medium as compared to untreated nMNC.
Studies on the natural killer cell activity of human nonadherent mononuclear cells (nMNC) with tumor necrosis factor, interleukin-1, interferon-gamma and cisplatin.
The potential for additive or synergistic augmentation of NK cell activity of nMNC with recombinant tumor necrosis factor (TNF-alpha), interleukin-1 (IL-1), interferon-gamma (IFN-gamma) and cisplatin in different combinations was examined. Significant augmentation in NK activity of nMNC was observed when treated with cisplatin, rIFN-gamma, rIL-1 or rTNF-alpha in vitro. Combined treatment of nMNC with rIL-1 and rTNF-alpha resulted in moderate increase in NK cell activity. However, killing of K 562 target cells was markedly augmented by co-stimulation of nMNC with rIL-1 plus rIFN-gamma and rIL-1 plus cisplatin. Further, only a moderate augmentation of NK activity was observed when nMNC were treated with cisplatin plus TNF or IFN-gamma. These observations demonstrated the individual and co-stimulative effects on NK function of nMNC in vitro by rTNF-alpha, rIL-1, cisplatin and rIFN-gamma.