IL-12 reverses the suppressive effect of the CD40 ligand blockade on experimental autoimmune encephalomyelitis (EAE).

Blockade of the CD40 ligand (CD40L)-CD40 interaction suppresses experimental autoimmune encephalomyelitis (EAE). Since this interaction induces IL-12, an essential cytokine for EAE induction, we hypothesized that CD40L blockade may suppress EAE through IL-12 inhibition. Here we show that exogenous IL-12 abolishes the ability of anti-CD40L monoclonal antibodies to prevent EAE. Anti-IL-12 antibodies prevent this reversal and protect from EAE. These results show that IL-12 is sufficient to overcome CD40L blockade and suggest that, of the multiple consequences of the CD40L-CD40 interaction, IL-12 induction is an essential one for induction of EAE.

Pathogenesis of neuroimmunologic diseases. Experimental models.

Animal models of autoimmune diseases have greatly improved our current understanding of the pathogenesis of human autoimmunity and have provided the potential for therapies based on manipulation of the immune system. In our laboratory, we have investigated the immunopathogenesis of autoimmune diseases of the nervous system and muscle. We have developed immune-based approaches for the suppression of experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis (MS), and experimental autoimmune neuritis (EAN), a model for the Guillain-Barré syndrome (GBS). These approaches included induction of peripheral tolerance, immunotoxin targeting of activated T cells, and cytokine manipulations. In addition, we identified the antigen and characterized immunopathologically an autoimmune inflammatory disease of skeletal muscle, experimental autoimmune myositis (EAM), a model for the human inflammatory muscle disease polymyositis.

In vitro assays elucidate peculiar kinetics of clindamycin action against Toxoplasma gondii.

In order to characterize the delayed effect of clindamycin and macrolide antibiotics against Toxoplasma gondii tachyzoites (E. R. Pfefferkorn and S. E. Borotz, Antimicrob. Agents Chemother. 38:31-37, 1994), we have carefully examined the replication of parasites as a function of time after drug addition. Intracellular tachyzoites treated with up to 20 microM clindamycin (> 1,000 times the 50% inhibitory concentration) exhibit doubling times indistinguishable from those of controls (approximately 7 h). Drug-treated parasites emerge from infected cells and establish parasitophorous vacuoles inside new host cells as efficiently as untreated controls, but replication within the second vacuole is dramatically slowed. Growth inhibition in the second vacuole does not require continued presence of drug, but it is dependent solely on the concentration and duration of drug treatment in the first (previous) vacuole. The susceptibility of intracellular parasites to nanomolar concentrations of clindamycin contrasts with that of extracellular tachyzoites, which are completely resistant to treatment, even through several cycles of subsequent intracellular replication. This peculiar phenotype, in which drug effects are observed only in the second infectious cycle, also characterizes azithromycin and chloramphenicol treatment, but not treatment with cycloheximide, tetracycline, or anisomycin. These findings provide new insights into the mode of clindamycin and macrolide action against T. gondii, although the relevant target for their action remains unknown.

Scanning electron microscopy of Setaria cervi adult male worms.

Scanning electron microscopy studies of adult Setaria cervi male worms showed the presence of horn-like lateral appendages and characteristic striated bands on the ventral side of the tail. All other features were indistinguishable from other members of the genus Setaria.

Evaluation of Setaria cervi antigen in the diagnosis of human filariasis.

Bovine filariid worm (Setaria cervi) antigen was evaluated for the immunodiagnosis of human filariasis. Patients with manifestations of filarial infection; [1) microfilaremia cases, (2) chronic clinical cases, (3) tropical pulmonary eosinophilia (TPE) cases) were taken for the investigations along with (4) normal endemic and non-endemic human subjects. CIEP, IFAT and ELISA tests were employed for detection of serum antibodies. There was a high degree of sensitivity shown by IFAT (1:64-1:256) and ELISA (1:200-1:20,800) in microfilaraemia cases and higher in chronic clinical cases with IFAT (1:128-1:1,024) and ELISA (1:12,800-1:102,400). Only one TPE case showed positive titre by IFAT (1:64) and ELISA (1:200) whereas sera from controls and patients with helminth infections did not show positive titres (1:100). In the case of CIEP, positive reaction was seen in only one case from group I.

Liver function and plasma protein metabolism in rodent model of filariasis.

Rodent model of filariasis was developed by infecting Wistar rats with Litomosoides carinii. Liver function tests, plasma protein concentrations, and synthesis rates of liver-formed proteins were estimated in these rats at 63 and 90 days post-infection. At 63 days post-infection, aspartate aminotransferase and alkaline phosphatase were significantly increased. Alanine aminotransferase, plasma total proteins and plasma albumin were in the normal range. However, at 90 days post-infection all these parameters were affected, reflecting progressive liver involvement. Hypoalbuminemia at 90 days post-infection did not appear to be due to decreased synthesis rate, indicating higher catabolism and/or altered distribution in pools.

Experimental chemotherapy of Angiostrongylus cantonensis infection in mice with flubendazole, levamisole, and their combination.

The efficacy of the drugs flubendazole, levamisole, and their combination was studied against Angiostrongylus cantonensis infection in mice. The drugs were evaluated on day 15 post infection. The efficacy was assessed by using single-dose and 5-dose treatment schedules. Flubendazole was effective at 25 mg/kg, 50 mg/kg, 100 mg/kg, 200 mg/kg when given each day for five days. Levamisole at a dose rate of 200 mg/kg was completely effective in a single treatment. The combined therapy of levamisole (25 mg/kg) and flubendazole (15 mg/kg) produced 100% efficacy. Both drugs given prior to experimental infection showed no prophylactic activity.

Efficacy of UV-irradiated larval vaccine of Ancylostoma ceylanicum (Looss, 1911) in golden hamsters (Mesocricetus auratus).

A vaccination trial in golden hamsters with UV-irradiated infective larvae of Ancylostoma ceylanicum was attempted. One oral vaccination of hamsters with 100 infective larvae irradiated by means of UV-tube (390 nm) at different time intervals induced the development of resistance. As the time exposure of irradiation was increased, there was a corresponding decrease in the subsequent worm establishment. A high level of protection afforded by larvae irradiated for 15 min UV-exposure was recorded giving 99.0% and 95.0% worm reduction against the challenge doses of 100 and 1000 normal larvae respectively. There was no marked difference in worm establishment in hamsters vaccinated either orally or subcutaneously, followed by oral challenge. In the vaccinated hamsters, the manifestations of resistance at 15 min UV-exposure were shown by marked reduction in worm establishment and highly reduced epg in pellets with significantly higher blood haemoglobin levels compared with those given normal larvae as vaccine and challenge controls.

Immunological evidence of chemotherapeutic action of mebendazole against Ancylostoma ceylanicum (Looss, 1911) in hamsters (Mesocricetus auratus).

The immunological responses in hamsters during treatment with mebendazole against Ancylostoma ceylanicum are studied. Indirect haemagglutination (IHA) counterimmunoelectrophoresis (CIEP), passive cutaneous anaphylaxis (PCA), foot pad swelling for immediate (ITH) and delayed (DTH) types of hypersensitivities were employed for measuring the responses. Serum antibody which was 1:32 before treatment increased to the maximum of 1:512 (control 1:128) on the 10th day and it declined subsequently. The CIEP test was positive for 10 days and then became negative. The PCA test was positive throughout the observation period both in the treated and untreated groups. Foot pad swelling for ITH and DTH responses were comparatively more prominent than in the untreated control up to the 20th day and then both decreased simultaneously. The immunological responses remained prominent for a longer period and decreased more slowly in the untreated control group.

Ancylostoma ceylanicum (Looss, 1911) in golden hamsters (Mesocricetus auratus): pathogenicity and humoral immune response to a primary infection.

The response of hamsters to Ancylostoma ceylanicum was studied between days 13 and 200 after infection. The infected hamsters showed signs of anaemia, particularly between the 13th and 60th days. A significant loss in body-weight in the infected animals was also observed during this period. Serological studies with agar gel-diffusion, counterimmunoelectrophoresis and indirect haemagglutination tests revealed positive reactions between the 20th and 60th days followed by a negative response in the latent period between the 60th and 200th days of infection, counterimmunoelectrophoresis test remained positive for a period of 150 days in this experimental model.

Studies on suspected clinical and experimental angiostrongyliasis: serological responses.

Serum antibodies in suspected angiostrongyliasis patient were detected by ELISA. The antibody titre was 1:51,200 in the serum and 1:6,400 in CSF with preadult A. cantonensis antigen. Other tests like AGD and CIEP failed to show any positive reaction with both preadult and adult worm antigens. Experimental infection with 100 A. cantonensis larvae in albino rats indicated positive CIEP reaction in serum from the day 5 to 375 after infection. No precipitin line was seen on the other hand, in AGD during observation period. Different rat groups infected with larval doses of 100, 500, 2,000, and 5,000 showed positive CIEP reaction, on the 21st day of infection when preadult worms were seen in CNS. There was no CIEP reaction when a low dose of 15 larvae was used. Cerebral fluid of rats infected with heavy dose of 5,000 larvae showed positive CIEP reaction on the 21st day.

Pulmonary superoxide dismutase activity in the mouse infected with Ancylostoma caninum larvae.

Albino mice were infected per os with 1000 infective larvae of Ancylostoma caninum and pulmonary superoxide dismutase activity was studied after completion of lung migration. The total superoxide dismutase activity per mg of lung protein was elevated by 15% in infected mice. Of the two forms of superoxide dismutases studied, that containing Cu-Zn was elevated by 44%. This suggests that A. caninum in mice may result in histopathological changes leading to transient pulmonary oxygen toxicity.

The distribution of Angiostrongylus cantonensis (Chen) in the central nervous system of laboratory rats.

The distribution of preadult Angiostrongylus cantonensis helminths in various regions of the central nervous system of rats, infected with varying doses of larvae, was studied. A dose of 10 000 larvae proved to be lethal; the rats showed paralytic symptoms and loss of vision prior to death. A very low dose of 15 larvae also proved to be infective to the central nervous system. Incidence of eye involvement was observed in two rats receiving a dose of 500 larvae. Maximum number of helminths were recovered from the cerebral hemisphere region but their involvement per unit weight of the tissue was found to be highest in the region of medulla oblongata.

Necator americanus in infant rabbits: complete development, humoral antibody, leukocyte response and serum protein changes following infection.

Necator americanus, originally isolated from man, developed completely in infant rabbits (RSG-1). This infection was serially passed in infant rabbits up to the 6th generation without using any immunosuppressant. Two day old infant rabbits demonstrated a high degree of susceptibility to N. americanus. Studies revealed the retention of worms in the intestine for more than 150 days during which egg production was high. In this experimental model the leukocyte, antibody and serum protein responses due to N. americanus infection were monitored. Results showed that eosinophil numbers rose significantly from day 28 to 170. A maximum was observed on day 80 (41%) in rabbits of the fourth generation. Antibodies to N. americanus were detectable in infected rabbits by counterimmunoelectrophoresis using third stage larval antigen (Rabbit strain). Generally, total globulins increased with decrease in albumin. Serum protein increase was associated with alpha-2, beta and gamma globulin components.