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U R Rao

Publications and source records attributed to U R Rao.

At least 19 recordsLinked to original sources
PubMed2005

Kinetics of T cell cytokine gene expression in gerbils after a primary subcutaneous Brugia pahangi infection.

The majority of patients infected with lymphatic filariae are microfilaremic but tend to manifest little obvious pathology because of the infections. Data collected from the Mongolian gerbil-Brugia spp. model for human lymphatic filariasis suggest this experimental animal model system most closely represents this patient group and will be useful in studying immunological parameters associated with chronic infections. This article reports the quantitation of interleukin (IL)-4, IL-5, IL-10, IL-13, and interferon (IFN)-gamma messenger RNA (mRNA) in gerbils after a primary subcutaneous infection with Brugia pahangi. Chronically infected gerbils showed elevated IL-4 in all tissues, compared with earlier time points, linking this Th2 cytokine to the downregulation of responsiveness, which develops in gerbils and humans. Both IL-5 and IL-13 mRNA expression were transient in all tissues. The peak in IL-5 at 14-28 days postinfection reflects the peak of peripheral eosinophilia observed in B. pahangi-infected gerbils. Little IFN-gamma mRNA was reported from chronically infected gerbils. The data collected thus far suggest that the expression profile of many of the measured cytokines in B. pahangi-infected gerbils reflects what is seen in an important subset of humans infected with lymphatic filariae, the microfilaremic, asymptomatic patient.

Animals↗
PubMed2005

Profiling the cellular immune response to multiple Brugia pahangi infections in a susceptible host.

Human lymphatic filariasis is caused primarily by Brugia malayi and Wuchereria bancroffi. Unraveling this disease is complex, as people living in endemic areas exhibit a vast array of clinical states and immune responses. The Mongolian gerbil (Meriones unguiculatus)-B. pahangi model of human lymphatic filariasis has provided much information on immune parameters associated with filarial infection. Prior investigations in our laboratory have shown that gerbils closely mimic a subset of patients classified as microfilaremic but asymptomatic, a group that comprises the majority of people living in endemic areas. Worm recovery data suggest that gerbils carrying current B. pahangi infections do not show any resistance to subsequent subcutaneous B. pahangi infections. The aim of the present studies was to investigate the T cell cytokine response in gerbils receiving multiple infections of B. pahangi as a means of mimicking the conditions experienced by people in endemic areas. The T cell cytokine profile generated by multiply infected gerbils was not different from that previously generated by gerbils infected only once with B. pahangi. Gerbils infected multiple times with B. pahangi showed a transient increase in IL-5, which corresponded to the increased eosinophil levels previously reported from multiply infected gerbils. Chronically infected gerbils showed elevated IL-4 mRNA levels, as has been reported from gerbils infected only once with B. pahangi. Chronic infections were also associated with a state of immune hyporesponsiveness, as determined by the characterization of lymphatic thrombi and lymphoproliferation of spleen and renal lymph node cells to worm antigen.

Animals↗
PubMed2002

Quantification of PCR amplification products of Brugia HhaI repeat DNA using a semiautomated Q-PCR system.

The sensitive, rapid and species-specific diagnosis of Brugia infections in humans or animal models is important in determining the level of parasitemia and the efficacy of chemotherapy or vaccinations. The HhaI family of highly repeated DNA sequences from Brugia have been useful in polymerase chain reaction (PCR)-based diagnosis of brugian filarial infections in blood samples and in mosquitoes. A PCR assay was developed using a biotinylated primer, a non-biotinylated primer and a species-specific chemiluminescent probe [tris[2,2'bipyridine] ruthenium (II) chelate, TBR] to detect PCR amplified Hhal family repeats. Individual blood samples from jirds infected with Brugia malayi or B. pahangi and with different levels of microfilaremia were tested in this assay. Known concentrations of Brugia DNA and DNA from the blood of uninfected control jirds were used as positive and negative controls, respectively. The PCR products generated by this method were analyzed using a semi-automated quantitative (Q)-PCR system. The levels of parasite DNA can be calculated from the luminosity units generated. Significant amounts of parasite DNA were detected in blood samples from infected jirds, and these values were correlated with the levels of microfilaremia. In contrast, reductions in circulating microfilaria following treatment with ivermectin correlated with low levels of measurable DNA. Using this system, we were also able to detect HhaI repeat DNA in the spleens of B. pahangi- infected jirds at 56 days post-infection when circulating microfilariae were not readily detectable. The results indicate that the species-specific Hhal Q-PCR detection and quantification method is rapid and sensitive, is useful in the detection of Brugia DNA in blood and other tissues and is suited for use in clinical settings because it does not require radioactive isotopes and gel-based protocols.

Animals↗
PubMed2000

Identification and localization of glutathione S-transferase as a potential target enzyme in Brugia species.

Brugia filarial nematodes are pathogenic lymphatic-dwelling parasites that, like other helminths, may modify the host's defense mechanisms by a major detoxification process involving glutathione-binding proteins such as glutathione S-transferases (GSTs). In the present study, soluble extracts of third-stage larvae, adult male and female worms, microfilariae of either B. pahangi or B. malayi or the adult worm excretory-secretory products of B. malayi were used to determine GST activity. These extracts and affinity-purified fractions of B. pahangi adult worms had a specific enzymatic activity when 1-chloro-2,4-dinitrobenzene was used as a substrate. The observance of this enzyme in all life cycle stages of Brugia spp. demonstrates its ubiquitous nature. Lavage of intraperitoneally infected jirds, but not that of uninfected jirds, also showed increased enzymatic activity, suggesting that GST is secreted in vivo. Soluble proteins of both Brugia spp. were strongly recognized by antibodies in sera from rabbits immunized with affinity-purified native GST of Onchocerca volvulus. Immunohistochemical studies localized these proteins in adult worms, demonstrating cross-reactivity between the GST of these two filarial nematodes. The effect of this enzyme on the motility and viability of adult worms, microfilariae, and larvae was tested in vitro using a battery of known GST inhibitors. Of all those tested, ethacrynic acid, N-ethylmalemide, 4-nitropyridine-oxide, or 1-chloro-2,4-dinitrobenzene at micromolar concentrations reduced the viability and motility of microfilariae, third-stage larvae, and adult worms. These results suggest that Brugia GSTs are major metabolic enzymes and may play an important role in the parasite's survival.

Animals↗
PubMed1999

Transglutaminase activity in equine strongyles and its potential role in growth and development.

Transglutaminases (E.C. 2.3.3.13) are a family of Ca(2+)-dependent enzymes that stabilize protein structure by catalyzing the formation of isopeptide bonds. A novel form of transglutaminase has been identified and characterized that seem to play an important role in growth, development, and molting in adult and larval stages of filarial nematodes. The aim of this study was to identify the ubiquitous nature of this enzyme in other nematodes and to measure its significance to larval growth, molting, and development. For this purpose, equine Strongylus spp. were used. Activity of this enzyme was identified in extracts of larvae and adults of Strongylus vulgaris, S. edentatus, Parascaris equorum and Cylicocyclus insigne. The significance of transglutaminase in the early growth and development of Strongylus vulgaris, S. edentatus and S. equinus was tested by adding specific inhibitors, monodansylcadaverine (MDC) or cystamine (CS), to in vitro cultures of third (L3) and fourth stage larvae (L4). The viability, molting and growth of these nematode species were affected by both inhibitors. Cystamine promoted abnormal development of Strongylus edentatus L3, resulting in an aberrant expansion of the anterior end. Addition of these inhibitors to cultures of L4 also reduced growth of the three species. The results indicated that transglutaminase is present in a wide array of nematode parasites and may be important in growth and development of their larval stages.

Animals↗
PubMed1999

Granulomatous inflammatory response to recombinant filarial proteins of Brugia species.

The lymphatic inflammatory response in Brugia-infected jirds peaks early during primary infections and then decreases in severity as judged by the numbers of lymph thrombi present within these vessels. Antigen-specific hypersensitivity reactions in these animals was measured by a pulmonary granulomatous inflammatory response (PGRN) induced by somatic adult worm antigen (SAWA)-coated beads, and by cellular proliferative responses of renal lymph node cells. The kinetics of these responses temporally correspond to lymphatic lesion formation. The importance of any single antigen to the induction of this inflammatory response has not been elucidated. In this study, the PGRN was used to measure the cellular immune response to four recombinant filarial proteins during the course of a primary B. pahangi infection. These proteins were BpL4, glycoprotein (glutathione peroxidase) gp29, heat shock protein (hsp) 70, and filarial chitinase. All were fusion proteins of maltose-binding protein (MBP). Control beads included those coated with diethanolamine (DEA), SAWA, or MBP. The measurements of PRGN were made at 14, 28, 56, and > 150 days postinfection (PI) in infected jirds, in jirds sensitized with SAWA, and in uninfected jirds. The secretory homolog of glutathione peroxidase gp29 was the only recombinant protein tested that induced a significantly greater PGRN (P < 0.05) than controls. This was seen at 28 days PI. These observations indicate that gp29 may be part of the worm antigen complex that induces an early inflammatory response, a response similar to that observed with SAWA. These studies indicate that this approach is useful in investigating the functional ability of specific proteins in the induction and down-regulation of immune-mediated inflammatory responses elicited by filarial parasites. Absence of a granulomatous response to the other recombinant proteins used may be related to the nature and sensitivity of the assay used or the character of recombinant proteins tested.

Adult↗
PubMed1997

Brugia pahangi: differential induction and regulation of jird inflammatory responses by life-cycle stages.

It has been hypothesized that different life-cycle stages of filarial nematodes induce different host responses. This concept was examined in the Brugia pahangi-jird model of lymphatic filariasis by measuring the kinetics of inflammatory responses to parasite antigens following intraperitoneal inoculation of different life-cycle stages. For this purpose, viable female or male worms, L3, L4, or microfilarial stage, were used. Dead worms served as controls. Worm and microfilarial burdens, pulmonary granulomatous inflammation (PGRN) to soluble adult worm antigen (SAWA)-coated beads, and peritoneal eosinophil and macrophage numbers were assessed at different days post-inoculation. All jirds inoculated with any of these life-cycle stages developed an early PGRN to SAWA which was later significantly reduced. Only viable worms induced down-regulation of the PGRN response. These results indicate that the hyporesponsive state is induced and maintained by all life-cycle stages. Also, the degree of granulomatous response was influenced by worm burden, with larger worm burdens inducing lower initial levels of PGRN to SAWA. Peritoneal inflammatory responses differed from the systemic response in that numbers of macrophages increased with time and microfilarial accumulation. No correlation was observed between peritoneal inflammatory responses measured by eosinophil and macrophage numbers and PGRN to SAWA.

Animals↗
PubMed1997

Adoptive transfer of granulomatous inflammation to Brugia antigens in jirds.

Brugia pahangi infections of jirds (Meriones unguiculatus) produce a granulomatous inflammatory response within the lymphatic vessels. Granulomas that form around beads coated with soluble adult antigens embolized in the lungs have been used to measure this response. Similar lesions were observed in naive jirds receiving lymph node cells and splenocytes from animals with acute infections. This was not the case with cells from chronically infected jirds that were hyporesponsive to implanted antigen-coated beads. Passively transferred immune sera collected during the acute and chronic periods of infection did not transfer this response. Lymph node cells but not splenocytes obtained from chronically infected jirds induced a down regulation of this response in animals during the acute period. These results indicate that this inflammatory response in the jird is cell mediated and that adoptive transfer in jirds is feasible. The induction of the down-regulated state may also be mediated by cells, but not serum factors.

Acute Disease↗
PubMed1997

Effect of gamma radiation on Brugia L3 development in vivo and the kinetics of granulomatous inflammation induced by these parasites.

Previous studies have shown that the downregulation of parasite-specific cellular immune response in Brugia-infected jirds requires viable worms but is not dependent on microfilariae (MF) for either induction or maintenance of this phenomenon. To clarify further which life cycle stages induce filarial hyporesponsiveness, jirds were infected intraperitoneally with third stage larvae (L3) exposed to 0, 15, 25, 35, 45, or 90 krad of gamma radiation to differentially alter L3 development. Necropsies were performed at 7, 14, 28, and 118 days postinoculation (DPI). The degree of parasite development, intraperitoneal inflammation, and pulmonary granulomatous inflammation (PGRN) to parasite antigen-coated beads embolized in the lungs were monitored at the time of necropsy. Parasite survival and worm lengths were inversely related to the irradiation dose. Gamma radiation at 35, 45, or 90 krad prevented larval molt to the adult stage. Some parasites irradiated with 15 or 25 krad developed beyond fourth stage larvae (L4) to infertile adult females. The PGRN peaked at 14 DPI in all infected groups. Downregulation of the PGRN occurred after 14 DPI in groups that received nonirradiated L3 or L3 irradiated with 15 krad. No significant decrease of the PGRN occurred in groups that received parasites irradiated with more than 15 krad. Significant peritoneal inflammation as indicated by an increase in macrophages occurred only in jirds that received nonirradiated L3. These data demonstrate the importance of the adult stages in inducing downmodulation in the absence of MF and suggest that the L4 may also play a role in the induction of this phenomenon. An alternate conclusion is that parasite burden and not developmental stage is important in the induction of this hyporesponsive state.

Animals↗
PubMed1996

Cellular immune responses of jirds to extracts of life cycle stages and adult excretory secretory products during the early development of Brugia pahangi.

The Brugia-jird model of lymphatic filariasis was used to examine the induction of cellular immune responses during the early premicrofilaremic phases of the infection. The intensity of the pulmonary granulomatous inflammatory response (PGRN) was determined by measuring granuloma areas around Sepharose beads coated with parasite extracts which were embolized in the lungs of jirds prior to necropsy. Necropsies were performed at 7, 14, 28, 56, and 150 days postinfection (DPI). These time points correspond to specific developmental changes in the life cycle. Lymphocyte blastogenesis assays were performed using cells from draining renal lymph nodes and splenocytes at 14 and 150 DPI. Soluble extracts of third stage larvae (L3), fourth stage larvae (L4), adult females, adult males, microfilariae (MF), and excretory secretory products (ES) of males and females were used in both measurements of cellular responsiveness. A marked granulomatous response to parasite extracts peaked at 7 DPI or 14 DPI followed by a gradual decrease to a hyporesponsive state at 120 DPI. The response of renal lymph node cells also was significantly elevated at 14 DPI and significantly decreased at > 150 DPI. The splenocyte responses were erratic and did not follow this pattern. Significant differences in PGRN responses to somatic extract preparations were not seen during the early stages of the infection (7, 14, 28 DPI), but those to MF and L3 were significantly less at 56 and 120 DPI. Although PGRN responses to ES followed a similar pattern, these were less than those to the somatic extract. The data indicated that a rapid, intense cell-mediated inflammatory response is induced early during a primary infection and that this response is rapidly downregulated. This downregulation begins prior to the maturation of adult parasites and microfilarial production. The early phase of the cellular response appears to be compartmentalized in that this response was consistently observed in the renal lymph nodes but not in the spleen. Soluble protein components of the parasites responsible for these responses are likely multiple and shared by all life cycle stages.

Animals↗
PubMed1996

Ocular complications of adult rheumatoid arthritis.

In a prospective study of 325 patients with adult rheumatoid arthritis, ocular complications were seen in 73 patients (22.4%). Keratoconjunctivitis sicca was the most common ocular lesion. Other lesions were episcleritis, scleritis, marginal thinning of the cornea with keratolysis, stromal corneal opacities with peripheral vascularisation, and iridocyclitis. The mean duration of the arthritis and the mean duration of seropositivity were found to be significantly higher in patients with ocular complications. Physicians should include ophthalmic examination as a routine in their protocol for patients with rheumatoid arthritis to facilitate early diagnosis and treatment of ocular complications.

Adolescent↗
PubMed1996

Regulatory cytokines in the lymphatic pathology of athymic mice infected with Brugia malayi.

To investigate whether Brugia malayi-induced lymphatic inflammation is due to production of pro-inflammatory cytokines, we determined the lymph and serum levels of interleukin (IL)-1, IL-6, tumor necrosis factor (TNF)-alpha and granulocyte-macrophage colony stimulating factor (GM-CSF) using enzyme immunoassays. Serum from normal and infected mice did not show elevated cytokine concentrations. Samples of lymph from parasitized lymphatics had significantly increased levels of IL-1 (range = 6-1620 pg/ml), IL-6 (19-17,800 pg/ml), TNF-alpha (19-2000 pg/ml) and GM-CSF (4-275 pg/ml). The anti-inflammatory cytokine IL-4 (7-12 pg/ml) was in the normal range and no increase in interferon (INF)-gamma was detected in lymph samples. The data suggest that increased levels of mediators or cytokines localized in the lymphatics may be important contributors to massive lymphatic dilation and inflammation.

Animals↗
PubMed1996

Effect of Brugia malayi on the growth and proliferation of endothelial cells in vitro.

Athymic mice (C3H/HeN) parasitized by Brugia malayi develop massively dilated lymphatics. The lymphatic endothelial lining is perturbed, and numerous mononuclear and giant cells are closely apposed to the endothelium. The hyperplastic endothelial cells and low opening pressure of the lymphatics suggest abnormal multiplication of these cells may be important in the dilation. We studied the in vitro growth rate of human umbilical vein endothelial cells cultured with adult worms and microfilariae of B. malayi. The tetrazolium salt reduction assays were used to quantify possible direct mitogenic or inhibitory effects. The growth factor-induced proliferation of endothelial cells was significantly suppressed by 44-51% on day 1, 46-81% on day 3, and 45-79% on day 5 in cultures containing adult female worms, which had greater suppressor activity on endothelial cell proliferation than male worms, microfilariae, or soluble adult worm extract. Culture supernatant containing female worm excretory-secretory products significantly inhibited the growth and multiplication of cells, suggesting that adult female worms release antigens or proteins that have inhibitory activity on growth factors necessary for endothelial cell proliferation in vitro. Excess human recombinant epidermal growth factor and bovine brain extract partly reversed the inhibitory activity of worms in culture and restored the endothelial cell proliferation when incubated with worm culture supernatant. Indomethacin and BW 775Hcl failed to restore normal endothelial proliferation in the presence of female worms, suggesting that parasite-derived prostanoids and cyclooxygenase products did not cause the inhibition. Lymph from dilated lymphatics, but not serum from infected mice, increased the proliferation of cells in vitro. Together, these data demonstrate that excretory-secretory products of B. malayi parasites suppress vascular endothelial proliferation in vitro. Furthermore, increases in the number of these cells in vitro in the presence of lymph suggest that parasite-induced host factors may be important in modulating the degree of proliferation.

Animals↗
PubMed1996

Effect of Brugia malayi infections on endothelial cells: a morphological study.

Athymic mice (C3H/HeN) parasitized by Brugia malayi develop gross lymphatic dilations at the chronic stage of infection. The hyperplastic endothelial cells and low fluid pressure of the lymphatics, characteristic of these infections, suggest that abnormal changes in these cells may play an important role in the dilation. We studied the lymphatic and vascular endothelium of parasitized mice for morphological changes by scanning and/or transmission electron microscopy. The lymphatic endothelium of dilated lymphatics was perturbed, scalloped, bulbous and highly indented. Numerous mononuclear and giant cells were closely apposed to the endothelial wall. Endothelial cells of both the lymphatics and the adjacent venules revealed no focal cytoplasmic lesions. Growth factor-dependent cell proliferation was significantly suppressed in vitro in endothelial cell cultures containing adult female worms, male worms or microfilariae. The actin cytoskeletal network appeared intact in these cells, and no gross changes in distribution were evident. Although the lymphatic walls were highly tortuous, our examination revealed no significant alterations in their morphology. Perivascular infiltration of activated mast cells, lymphocytes and monocytes/macrophages indicated polarization of inflammatory cells into the lymphatic tissue. It is possible that these inflammatory cells might induce temporal functional changes in the lymphatics of infected athymic mice.

Animals↗
PubMed1995

Comparison of phenytoin with auranofin and chloroquine in rheumatoid arthritis--a double blind study.

OBJECTIVE: To evaluate efficacy of phenytoin in modifying the course of rheumatoid arthritis (RA) by comparing it to gold (auranofin) and chloroquine. METHODS: A double blind, randomized study of 6 months' duration was conducted at the Nizam Institute of Medical Sciences, Hyderabad, India. One hundred and thirty-two patients with active RA (defined by the 1987 ARA criteria) were entered into the study and randomized into 3 groups: phenytoin, chloroquine, or auranofin. RESULTS: Full data were evaluable in 100 patients who satisfactorily completed the protocol (phenytoin, 35; auranofin, 30; and chloroquine, 35). Twenty-four patients were noncompliant and did not take medication or return for evaluation; 8 patients had the drug withdrawn because of side effects before study completion. For each of the 3 drugs all clinical and laboratory variables improved when pre and posttreatment values (p < 0.05 to 0.001) were compared. There was a greater reduction in posttreatment mean morning stiffness in the chloroquine group than in the phenytoin and auranofin groups (p < 0.05). Posttreatment grip strength was also greatest in the chloroquine group. On the other hand, there were statistically significant decreases in IgM levels in both the phenytoin and auranofin groups (p < 0.001), but not with chloroquine. Among the 53 patients with a disease history of 3-6 months, global outcome was best with phenytoin (16/17), compared to chloroquine (12/18) and auranofin (12/18) (p < 0.03). However, there was no such difference in the 47 patients in all 3 groups with a disease history longer than 6 months. Eight patients had side effects (phenytoin, auranofin, 2; chloroquine, 1) requiring withdrawal of the drug. However, the incidence of side effects was not significantly different for the 3 drugs. CONCLUSION: Our data indicate that phenytoin is comparable to auranofin and chloroquine in its efficacy in RA and may be considered an alternative disease modifying agent for RA.

Adult↗