Efficacy of Withania somnifera chemotypes NMITLI - 101R, 118R and Withaferin A against experimental visceral leishmaniasis.

The immunoprophylactic and therapeutic potentials of root extracts of Withania somnifera chemotypes (NMITLI-118, NMITLI-101) and pure withanolide-withaferin A was investigated against Leishmania donovani infection in hamsters. The naive animals, fed orally with immunostimulatory doses of chemotypes 101R, 118R (10 and 3 mg/kg) and withaferin A (9 and 3 mg/kg) for five consecutive days and challenged with Leishmania parasites on day 6, were euthanized on days 30 and 45 p.c. for the assessment of parasite clearance, real-time analysis of mRNAs of Th1/Th2 cytokines (IFN-γ, IL-12, TNF-α, iNOS/IL-4, IL-10 and TGF-ß), NO production, reactive oxygen species (ROS) generation, lymphocyte transformation test and antibody responses. By day 45 p.c., there was a significant increase in the mRNA expression of iNOS, IFN-γ, IL-12 and TNF-α but decrease in IL-4, IL-10 and TGF-ß, an enhanced Leishmania-specific LTT response as well as ROS, NO and antileishmanial IgG2 levels in 101R-treated hamsters followed by 118R- and withaferin A-treated ones, respectively. When these chemotypes were given to L. donovani-infected hamsters at different doses, there was moderate therapeutic efficacy of chemotype 101R (~50%) at 30 mg/kg × 5 followed by the other two. The results established that the 101R is the most potential chemotype and can be evaluated for combination therapy along with available antileishmanials.

Withania somnifera chemotypes NMITLI 101R, NMITLI 118R, NMITLI 128R and withaferin A protect Mastomys coucha from Brugia malayi infection.

Withania somnifera is an ayurvedic Indian medicinal plant whose immunomodulatory activities have been widely used as a home remedy for several ailments. We recently observed immunostimulatory properties in the root extracts of chemotypes NMITLI-101, NMITLI-118, NMITLI-128 and pure withanolide, withaferin A. In the present study, we evaluated the potential immunoprophylactic efficacies of these extracts against an infective pathogen. Our results show that administration of aqueous ethanol extracts (10 mg/kg) and withaferin A (0·3 mg/kg), 7 days before and after challenge with human filarial parasite Brugia malayi, offers differential protection in Mastomys coucha with chemotype 101R offering best protection (53·57%) as compared to other chemotypes. Our findings also demonstrate that establishment of B. malayi larvae was adversely affected by pretreatment with withaferin A as evidenced by 63·6% reduction in adult worm establishment. Moreover, a large percentage of the established female worms (66·2%) also showed defective embryogenesis. While the filaria-specific immunological response induced by withaferin A and NMITLI-101 showed a mixed Th1/Th2 phenotype, 118R stimulated production of IFN-γ and 128R increased levels of IL-4. Taken together, our findings reveal potential immunoprophylactic properties of W. somnifera, and further studies are needed to ascertain the benefits of this plant against other pathogens as well.

Adult Brugia malayi mitochondrial and nuclear fractions impart Th1-associated sizeable protection against infective larval challenges in Mastomys coucha.

Protective immunity to the subperiodic human filariid, Brugia malayi, was explored in the rodent host, Mastomys coucha after vaccination with subcellular fractions derived from the adult stage of the parasite. The highest level of protection was conferred in animals vaccinated with the 'mitochondria rich' (MT) fraction, in which microfilaraemia and worm burden were markedly reduced by 67.2 and 65.9%, respectively, followed by the 'nucleus rich' (NR) fraction, showing reductions of 62 and 52.3%, respectively, over the non-immunized control group. Mastomys vaccinated with MT and NR, displayed a significant increase in the level of antigen-specific serum immunoglobulin G (IgG). The levels of IgG2a, IgG2b and IgM antibody isotypes were remarkably elevated in both the MT and NR immunized groups, while IgG1 and IgG3 levels were low. Apart from antibodies, both these fractions also led to marked antigen-specific lymphoproliferation in vitro, along with enhanced release of nitric oxide by peritoneal macrophages. There was an increased population of CD4+ and CD8a+T-cells in MT immunized animals, as measured by flow cytometry, accompanied by elevated levels of proinflammatory cytokines; interferon gamma (IFN-gamma), tumour necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1beta) in the culture supernatants of the activated splenocytes. The results suggest that both NR and MT contain proinflammatory molecules which evoke a protective Th1 type of immune response.

7-O-[4-methyl piperazine-1-(2-acetyl)]-2H-1-benzopyran-2-one: a novel antifilarial lead compound.

In preliminary studies we found that benzopyrones (coumarins), which are known to exert many biological activities including anti-inflammatory effect, possess promising macrofilaricidal action as well. In order to explore the possibility of combining such a macrofilaricidal activity with the microfilaricidal potential of the known piperazine pharmacophore, we synthesized a series of compounds and evaluated their antifilarial effect. In the present study, one of these compounds, 7-O-[4-methyl piperazine-1-(2-acetyl)]-2H-1-benzopyran-2-one (2), which has shown promising macrofilaricidal action against rodent filariid Litomosoides carinii in cotton rats, was evaluated against infection with Brugia malayi in Mastomys coucha and jird (Meriones unguiculatus). In the B. malayi-M. coucha system, the compound at a dose of 300 mg/kg, oral (p.o.) x5 days showed 53.6% adulticidal and 46.0% microfilaricidal activity along with 46.3% sterilization effect on the female worms. In addition, the compound interfered with the establishment of infective larvae (L(3))-induced infection to an extent of 50% at the same dose level. At 1 microM concentration it inhibited protease activity of B. malayi to 82%. The compound thus provides a novel lead for further synthesis and development of antifilarial agents with macrofilaricidal, microfilaricidal, female-sterilizing and possible larvicidal efficacy.

Immunomodulator tuftsin augments antifilarial activity of diethylcarbamazine against experimental brugian filariasis.

In the present study, we evaluated the potential of an immunomodulator tuftsin in increasing the efficacy of liposomised diethylcarbamazine (DEC) against experimental filarial infection of Brugia malayi. The liposomised form of DEC, when used at sub-optimal dose of 25 mg/kg body weight, successfully eliminated filarial parasite from systemic circulation in animals inflicted with B. malayi infection. However, the formulation was effective upto 60 days post infection only, followed by recurrence of the infection. In contrast, the co-administration of liposomal formulation of DEC along with an immunomodulator tuftsin was found to be competent enough to suppress microfilarial stage of parasite till 90 days post treatment. Interestingly, tuftsin bearing DEC liposomes were found to be effective against adult parasite as well.

Transmission efficiency of Culex quinquefasciatus and Aedes aegypti to Wuchereria bancrofti infection: an experimental study.

Present study was undertaken to evaluate the suitability of natural (Culex quinquefasciatus) and experimental (Aedes aegypti) vectors for supporting the development of W. bancrofti larvae for onward transmission. Both the species permitted development of W. bancrofti mf to infective larvae (L3) within 11 to 13 days. The mf intake by both the species of mosquitoes was directly related to mf density in donor's blood. Culex exhibited higher L3 recovery than Aedes. In Aedes maximum percent L3 development occurred after ingesting 4 mf whereas Culex exhibited best establishment at an average mf intake of 11.5. Nevertheless wide variation in mf density in donor's blood did not significantly affect the larval establishment in Aedes mosquito while in Culex very high (> 400 mf/40 microliters) or low (< 50 mf/40 microliters) mf counts in donor's blood adversely affected the L3 recovery. The results reveals that A. aegypti has an edge over the natural vector, Culex in being a voracious feeder, their easy laboratory maintenance and better transmission potential.

Macrophages in the development of protective immunity against experimental Brugia malayi infection.

The present report compares the macrophage function in rodent hosts susceptible and resistant to the human lymphatic filariid Brugia malayi. Macrophages from both mastomys (resistant) and gerbil (susceptible) infected intraperitoneally (i.p.) with the infective larvae (L3) of B. malayi were isolated from peritoneal lavage at different time-intervals and formation rate of NO, H2O2, O2-, TNF-alpha, glutathione peroxidase and reductase was assayed. NO release was found to be significantly increased in resistant mastomys as compared to gerbils and the release was markedly suppressed by i.p. administration of the NOS inhibitor aminoguanidine (AG). The AG-treated mastomys also demonstrated significantly greater establishment of larvae which correlated well with suppressed formation of NO. Nitric oxide synergizes with superoxide to form peroxynitrite radical (potent oxidant), which is known to be more toxic per se than NO. Results indicate the possible involvement of peroxynitrite in the rapid killing of larvae in the peritoneal cavity of mastomys. In contrast, the production of H2O2 was found to be enhanced in both species indicating that B. malayi L3 could withstand the toxic effects of H2O2. The higher level of glutathione peroxidase and reductase, as observed in mastomys compared with the gerbil after larval introduction, possibly protects the cell against the injurious effect of H2O2. The TNF-alpha level remained virtually unchanged in both the hosts, suggesting an insignificant role for this cytokine in parasite establishment.

The antifilarial activity of a marine red alga, Botryocladia leptopoda, against experimental infections with animal and human filariae.

The antifilarial activity of the marine red alga Botryocladia leptopoda against rodent and human lymphatic filarial parasites is described. The animal filarial species included Litomosoides sigmodontis and Acanthocheilonema viteae maintained in cotton rats and Mastomys coucha, respectively, while a subperiodic strain of the human lymphatic filarial parasite Brugia malayi was maintained in M. coucha. The crude extract and its hexane fraction brought about a marked reduction in the peripheral microfilarial level in both of the rodent filarial parasites L. sigmodontis and A. viteae. The microfilaricidal effect began slowly from day 8 or 15 after initiation of treatment and increased with time with a very high efficacy at the end of the observation period against both rodent filariids. The microfilaricidal efficacy was, however, not as prominent in the case of B. malayi. The antifilarial activity, which occurred in the hexane fraction, exerted action at a much lower dose. The product killed a significant proportion of A. viteae and L. sigmodontis adult parasites. In the case of B. malayi, although the macrofilaricidal efficacy was much less than that of the rodent parasites, it (hexane fraction) caused sterilization of a significant proportion of the surviving female parasites. The present findings indicate the possibility of developing an adulticidal and female sterilizing agent against filarial parasites from a marine red alga.

Immunomodulatory effect of albizzia lebbeck.

The immunomodulatory effect of the bark of Albizzia lebbeck (Sirisha) was evaluated by studying humoral and cell mediated immune responses. The hot aqueous extract and its butanolic fraction were administered once daily for one week in mice, immunised previously with sheep red blood cells (SRBC). At the dose levels tested (6.25, 12.5 and 25 mg/kg, p.o.), A. lebbeck treated mice developed higher serum antibody titres compared to the vehicle treated group and the effect was comparable to the standard drug muramyl dipeptide (MDP). Delayed type hypersensitivity response was suppressed in SRBC immunised mice treated with A. lebbeck extract. The macrophage migration index remained unaltered in both mice and rats. These results are discussed in the light of possible immunopotentiating effects of A. lebbeck.