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Biomedical subjects

R Chandrashekar

Publications and source records attributed to R Chandrashekar.

At least 55 records · Page 3Linked to original sources
PubMed1990

IgG response of rats to the excretory-secretory products of Litomosoides carinii.

The nature of antibody responses to the excretory-secretory (ES) products of adult worms and microfilariae (Mf) of Litomosoides carinii in albino rats and their possible role in protection have been studied. Rats were immunized with ES products derived from in vitro incubation of adults or Mf. The sera from these rats promoted neutrophil-mediated killing of Mf in vitro. The antibody responsible for the cytocidal activity was identified as IgG isotype. Indirect fluorescent antibody test showed the presence of IgG on the surface of Mf incubated in either of the immune sera. The immune sera were effective in clearing the circulating Mf in Mastomys natalensis, indicating the protective nature of the antibody. Thus, L. carinii in culture liberate functional antigens that seem to have protective potential.

Animals↗
PubMed1990

Antibody-mediated cytotoxic effects in vitro and in vivo of rat cells on infective larvae of Brugia malayi.

Albino rat macrophages and neutrophils in the presence of immune serum adhered to and promoted killing of Brugia malayi infective larvae in vitro. At a similar cell-target ratio, macrophages were more potent than neutrophils in inducing cytotoxic response to the larvae. Eosinophils were also effective in killing but only at a high cell-target ratio. The activity in the immune serum could be absorbed to and eluted from a Protein A-Sepharose column suggesting involvement of IgG antibody in the reaction. An indirect fluorescent antibody test confirmed the presence of IgG on the surface of larvae incubated in immune serum. Infective larvae were attacked by host cells within micropore chambers 16-24 h after implantation into immunized rats. Further, a strong cytotoxic response to the larvae was seen when they were introduced intraperitoneally into immune rats indicating the role of antibody and cells in vivo. We suggest that antibody-dependent cellular cytotoxicity may represent an important mechanism of parasite killing in an immune host.

Animals↗
PubMed1990

Immune response to Acanthocheilonema viteae infection in multimammate rats (Mastomys natalensis).

The multimammate rat Mastomys natalensis, when infected with the filarial parasite Acanthocheilonema viteae, develops amicrofilaraemia. Worm recovery and the duration and intensity of microfilaraemia were analysed and related to the humoral and cellular immune responses of the host by using an antibody-dependent cell-mediated cytotoxicity (ADCC) assay towards microfilariae (Mf). Mf were detected in the peripheral blood at 7 weeks post-infection (p.i.), reaching maximum levels by 20 weeks p.i., and then gradually decreasing to undetectable levels during the next 36 weeks. The cytotoxic antibodies appeared around 15-18 weeks p.i., and the serum at 36 weeks p.i. induced 70% cytotoxicity to the Mf in vitro in the presence of host cells. The IgM fraction of the immune serum from amicrofilaraemic Mastomys promoted ADCC to Mf both in vitro and in vivo. Macrophages were more potent in inducing cytotoxic effect than eosinophils and neutrophils. Platelets were ineffective in killing the Mf in the presence of immune serum. IgM antibody cleared the circulating Mf from the blood when given passively to infected Mastomys. The average recovery of adult worms was about 20% of the inoculated larvae. No live females could be recovered 56 weeks p.i. Thus protective immune responses built up over an extended period of time are elicited against the Mf and perhaps even to adults in Mastomys infected with A. viteae.

Animals↗
PubMed1990

IgG4 subclass antibody serology for onchocerciasis.

Antibodies to phosphorylcholine and carbohydrate determinants responsible for much of the cross-reactivity among nematodes are subclass restricted in humans and absent in the IgG4 subclass. Total IgG and IgG4 antibody responses to Onchocerca volvulus were examined by enzyme immunoassay and by immunoblot. Significant background IgG reactivity was detected in both assays in US control sera and sera from patients with intestinal nematode infections, but background reactivity was negligible in the IgG4 assays. IgG4 antibodies were detected by enzyme immunoassay in 17 of 18 Nigerian onchocerciasis serum samples and in 2 of 9 endemic control serum samples. Cross-reactive IgG4 antibodies were present in serum pools from patients infected with other filariae. IgG4 antibodies recognized a restricted subset of O. volvulus antigens in immunoblots relative to IgG. These results confirm the previously reported enhanced specificity of IgG4 antibody assays for filariasis and extend the observation to include onchocerciasis. Whereas IgG4 subclass antibody serology is more specific than measurement of total IgG antibodies for onchocerciasis, cross-reactivity among filariae limits the utility of this approach.

Animals↗
PubMed1990

Circulating immune complex-associated parasite antigens in human onchocerciasis.

This study identified and characterized parasite antigens in sera from humans infected with Onchocerca volvulus. Immune complexes were precipitated from human sera with polyethylene glycol and analyzed by immunoblot with rabbit antibodies to O. volvulus. A 23-kDa parasite antigen was detected in sera from 17 of 23 Nigerian onchocerciasis patients and 5 of 10 endemic controls. Other parasite antigens with apparent molecular masses of 62, 66, and 70 kDa were less consistently observed. These antigens were not present in Nigerian or US nonendemic control sera, in sera from patients with various other parasitic infections, or in sera from US patients with autoimmune diseases. Biochemical studies indicated that these antigens are nonglycosylated acidic proteins that do not contain phosphorylcholine. These antigens may be useful as targets for improved diagnostic tests for onchocerciasis based on parasite antigen detection.

Adult↗
PubMed1990

Use of parasite antigen detection to monitor macrofilaricidal therapy in Brugia malayi-infected jirds.

Improved methods are needed to evaluate new treatments for filarial infections. We have recently developed a monoclonal antibody-based enzyme immunoassay to detect circulating parasite antigen in sera from Brugia malayi-infected jirds. In the present study, parasite antigen levels were compared to parasitological parameters after treatment of B. malayi-infected jirds with CGP 20376 that has been reported to be active against both microfilariae and adult worms of this parasite. Microfilariae were cleared promptly and permanently after CGP 20376 treatment, and no adult worm was recovered in jirds at necropsy 20 wk after treatment. In contrast, untreated animals had sustained microfilaremia throughout the course of the study, and adult worms were recovered in all control animals (mean worm recovery; 24.3 +/- 7.8 SE). Parasite antigen was present in sera from all infected animals before treatment. Parasite antigen titers in sera were unchanged 5 wk after treatment but fell to undetectable levels in 4 of 6 animals by 20 wk after treatment. Low-level antigenemia was detected in 2 of 6 animals at 20 wk, perhaps suggesting incomplete killing of parasites or incomplete clearance of antigen. Parasite antigen levels were stable throughout the study in control animals. These preliminary results suggest that parasite antigen detection is useful as a means of noninvasively monitoring the efficacy of anti-filarial drug therapy.

Animals↗
PubMed1990

Circulating parasite antigen in Brugia pahangi-infected jirds.

The Mongolian jird is used widely in filariasis research for studies of protective immunity, pathogenesis, and therapy. The purpose of this study was to evaluate parasite antigen detection as a means of noninvasively monitoring Brugia pahangi infection in jirds. A parasite antigen with Mr of 105-110 kDa was identified in sera from i.p.- and s.c.-infected jirds by immunoblot with a monoclonal antibody to phosphorylcholine. The same antibody was used in a direct sandwich enzyme immunoassay to measure antigen in jird sera. Parasite antigen was detectable as early as 2 wk after i.p. or s.c. injection of L3. Antigen titers increased between 2 and 12 wk and stabilized between 12 and 36 wk after infection in s.c.-infected animals. A different pattern was seen in i.p.-infected jirds with antigen titers peaking at 16 wk and falling significantly between 16 and 32 wk after infection. Parasite antigen titers correlated significantly with adult worm infection intensities in jirds with mature i.p. and s.c. infections. Antigenemia was also detectable in sera from jirds after i.p. implantation of adult parasites of either sex. However, antigen was not detected in sera from infant offspring of antigenemic infected mothers. We conclude that parasite antigen detection allows B. pahangi development and survival as well as infection intensity to be monitored in living animals with unprecedented sensitivity and accuracy. This technique should facilitate drug and vaccine studies in this important experimental filariasis model.

Animals↗
PubMed1988

Characterization of a monoclonal antibody against infective larvae of Brugia malayi.

Monoclonal antibodies were produced following immunization of mice with live infective larvae of Brugia malayi. One of these, 46.08.76, is an antibody that promotes adherence of mouse peritoneal macrophages and human peripheral blood leucocytes to the infective larvae of B. malayi and Wuchereria bancrofti, respectively, and kills them. Fresh normal serum, as a source of complement, augments this effect. The same monoclonal antibody conferred 89% protection to jirds (Meriones unguiculatus) against challenge infection of B. malayi stage-three larvae. This monoclonal antibody recognizes antigens of 80,000, 67,000, 52,000 and 36,000 MW proteins present among the antigens of larvae, as detected by an immunoblotting technique. The antibody also reacts with antigens of infective larvae of Litomosoides carinii, Dipetalonema viteae and B. pahangi, but to a smaller extent.

Animals↗
PubMed1987

Complement activation by eggs and microfilariae of filarial parasites.

The complement of fresh normal rat serum was activated by filarial eggs and microfilariae (mf). C3 was deposited on the surface of Litomosoides carinii, Brugia pahangi, Brugia malayi and Dipetalonema viteae as seen by immunofluorescence. Intra-uterine and in vitro-derived mf did not bind C3. In contrast, C3 bound to the blood-derived mf of B. pahangi and B. malayi as well as exsheathed mf of L. carinii and B. malayi. Significant consumption of complement was observed with eggs of all filarial species, as well as sheathed mf of B. pahangi, B. malayi and exsheathed mf of L. carinii and B. malayi. These experiments indicated that complement was activated by filarial parasites via the alternative pathway. The bound complement promoted neutrophil-mediated adherence and cytotoxicity.

Animals↗
PubMed1987

Lectin-binding characteristics of Wuchereria bancrofti microfilariae.

The binding of 10 different lectins to the surface of microfilariae of Wuchereria bancrofti has been investigated. Wheat germ agglutinin (WGA) and Helix pomatia lectin (HPA) bound specifically to the sheathed microfilariae indicating the presence of N-acetyl-D-glucosamine and N-acetyl-D-galactosamine respectively on the surface. Exsheathed microfilariae did not react with any of the lectins. Treatment of sheathed microfilariae with proteases resulted in increased binding of WGA and HPA. Such treated microfilariae showed a weak binding of Concanavalin A (Con A), and lectins of lentil (LCH) and of Limulus polyphemus (LPA). Sheathed microfilariae incubated with sera of people living in endemic zones of filariasis but with no apparent evidence of infection (endemic normals), or with sera of chronic elephantiasis patients, or with their respective gamma globulin fractions, bound Con A and LCH. These lectins bound weakly to exsheathed microfilariae under the same conditions. Binding was due to the mannose components of the specific immunoglobulins of the sera which coated the microfilariae. However, microfilariae when incubated with sera or their globulin fractions from non-endemic normals (NEN), or from microfilarial carriers, did not bind Con A and LCH, suggesting that specific immunoglobulins were neither present in NEN sera nor in significant amounts in sera of microfilarial carriers.

Acetylgalactosamine↗
PubMed1987

Litomosoides carinii: characterization of surface carbohydrates of microfilariae and infective larvae.

The carbohydrate moieties of microfilariae (Mf) and infective larvae (L3) have been investigated by lectin-binding technique. Mf derived from three sources, namely, uteri (in utero), released in vitro from adults and from blood of rodents infected with Litomosoides carinii were examined by using fluoresceinated lectins. Wheat germ agglutinin (WGA) bound to these Mf and the binding was inhibited by N-acetyl glucosamine. In addition to WGA, Concanavalin A (Con A) and lentil lectin (LCH) bound to in vitro-released and in utero-derived Mf showing the presence of mannose moieties on their surface. In utero-derived Mf also showed binding with the agglutinins of Limulus polyphemus (LPA), peanut (PNA), Ricinus communis (RCA), Helix pomatia (HPA), Soyabean (SBA) and Dolichos biflorus (DBA) but not to that of Ulex europaeus (UEA) indicating the presence of additional carbohydrate molecules like sialic acid, galactose and N-acetyl galactosamine on their sheath. None of the lectins bound to the cuticle of exsheathed Mf. Treatment of blood-derived and in vitro-released Mf with certain proteases exposed additional binding sites for SBA, HPA, Con A and LCH. In case of L3, only PNA bound to the larvae isolated from infective mites Bdellonyssus bacoti, and the binding was inhibited by D-galactose. No such binding of the lectins was seen to the larvae that migrated to the pleural cavity of jirds indicating that there is considerable change on the parasite surface during their migration in the vertebrate host. Sheathed Mf and mite-derived L3 when incubated with immune rat sera, bind Con A and LCH lectins possibly due to the mannose components of the specific immunoglobulins that coat onto the Mf and L3.

Animals↗
PubMed1987

Effect of ivermectin on serum dependent cellular interactions to Dipetalonema viteae microfilariae.

The effects of ivermectin (Iv) on the filarial infection has been investigated using Dipetalonema viteae--Mastomys natalensis model system. In vitro, Iv (3 micrograms/ml) had no effect on the microfilariae (mf) in the presence or absence of serum from normal or infected Mastomys (NMS, IMS). However, the sera promoted cell-mediated cytotoxicity with macrophages or eosinophils to the mf when Iv was added at 3 ng/ml. Sera isolated from normal or infected Mastomys after subcutaneous treatment with Iv (100 micrograms/kg) (Iv-NMS, Iv-IMS) also induced cellular cytotoxicity to the mf in vitro. Available evidence suggests that Iv mediates the cellular cytotoxicity by complement activation by alternative pathway. Iv treatment of D. viteae infected Mastomys resulted in an initial mobilization of the mf into circulation followed by the death of both the mf and adults in vivo. There was enhanced antibody level in Iv treated infected animals and antibody dependent cellular cytotoxicity mediated by IgM seems to be predominantly involved in elimination of the mf. The serum containing these antibodies was effective in clearing the circulating mf from Mastomys on passive immunization. Thus, Iv seems to synergize host immune factors against D. viteae infection.

Animals↗
PubMed1986

Brugia malayi: rat cell interactions with infective larvae mediated by complement.

Albino rat macrophages and neutrophils, in the presence of fresh normal rat serum as a source of complement, adhered to and promoted killing of Brugia malayi infective larvae in vitro. Eosinophils, by themselves, were marginally cytotoxic at a high cell-target ratio but promoted cytotoxicity when mixed with macrophages. Eosinophil culture supernatants enhanced the macrophage mediated killing of infective larvae. The complement of fresh normal rat serum was found to act by the alternate pathway. Fresh normal rat serum depleted of alternate pathway complement activity by treatment with zymosan A, or of Factor B by heating at 50 C for 20 min, or of Factor D by passing through Sephadex G75 column, failed to promote cell adherence to the parasite. C3 molecules were detected on the surface of infective larvae by immunofluorescence. There was a significant consumption of complement when Brugia malayi infective larvae were incubated in fresh normal rat serum. Albino rat cells were more potent in inducing cytotoxicity to infective larvae in vitro than those from jird or Mastomys natalensis, which may reflect the greater resistance offered by the rat to B. malayi infection. There was much less cellular infiltration on introduction of Brugia malayi infective larvae into the peritoneal cavity of jirds compared to rats and Mastomys natalensis indicating the greater susceptibility of jirds to intraperitoneally induced infections.

Animals↗
PubMed1985

Serum dependent cell-mediated immune reactions to Brugia pahangi infective larvae.

Fresh normal rat serum (fNRS) promoted adherence and cytotoxicity of albino rat neutrophils and macrophages to Brugia pahangi infective larvae (L3) in vitro. EDTA and not EGTA abolished the adherence activity suggesting the involvement of complement components via the alternate pathway. C3 molecules were detected on the surface of the parasite by immunofluorescence. fNRS depleted of complement by treatment with Zymosan A or of factor B by heating at 50 degrees C for 20 min, failed to promote cell adherence to the parasite. fNRS and cells from albino rat were more potent in inducing cytotoxicity to L3 than those from jird or Mastomys which may reflect the greater resistance offered by the albino rat to B. pahangi infection. In the presence of IgG and a heat labile factor, possibly complement, of immune serum, neutrophils and macrophages and to a lesser extent eosinophils adhered to and killed the larvae. Immune sera raised against microfilariae of different filarial parasites promoted cell-mediated cytotoxicity to B. pahangi L3 suggesting sharing of antigens between the two stages.

Animals↗