Cryptolepine-Induced Cell Death of Leishmania donovani Promastigotes Is Augmented by Inhibition of Autophagy

Leishmania donovani are the causative agents of visceral leishmaniasis worldwide. Lack of vaccines and emergence of drug resistance warrants the need for improved drug therapy and newer therapeutic intervention strategies against leishmaniasis. In the present study, we have investigated the effect of the natural indoloquinoline alkaloid cryptolepine on L. donovani AG83 promastigotes. Our results show that cryptolepine induces cellular dysfunction in L. donovani promastigotes, which leads to the death of this unicellular parasite. Interestingly, our study suggest that cryptolepine-induced cell death of L. donovani is counteracted by initial autophagic features elicited by the cells. For the first time, we show that autophagy serves as a survival mechanism in response to cryptolepine treatment in L. donovani promastigotes and inhibition of autophagy causes an early increase in the amount of cell death. This study can be exploited for designing better drugs and better therapeutic strategies against leishmaniasis in future

Novel Betulin Derivatives as Antileishmanial Agents with Mode of Action Targeting Type IB DNA Topoisomerase

ABSTRACT Toward developing antileishmanial agents with mode of action targeted to DNA topoisomerases of Leishmania donovani, we have synthesized a large number of derivatives of betulin. The compound, a natural triterpene isolated from the cork layer of Betula spp. plants exhibits several pharmacological properties. Three compounds (disuccinyl betulin, diglutaryl dihydrobetulin, and disuccinyl dihydrobetulin) inhibit growth of the parasite as well as relaxation activity of the enzyme type IB topoisomerase [Leishmania donovani topoisomerase I (LdTOP1LS)] of the parasite. Mechanistic studies suggest that these compounds interact with the enzyme in a reversible manner. The stoichiometry of these compounds binding to LdTOP1LS is 1:1 (mole/mole) with a dissociation constant on the order of ϳ10 Ϫ6 M. Unlike CPT, these compounds do not stabilize the cleavage complex; rather, they abrogate the covalent complex formation. In processive mode of relaxation assay condition, these compounds slow down the strand rotation event, which ultimately affects the relaxation of supercoiled DNA. It is noteworthy that these compounds reduce the intracellular parasite burden in macrophages infected with wild-type L. donovani as well as with sodium antimony gluconate resistant parasite (GE1). Taken together, our data suggest that these betulin derivatives can be exploited as potential drug candidates against threatening drug resistant leishmaniasis

Virulence of Cholera Toxin Gene-Positive Vibrio cholerae Non-O1/non-O139 Strains Isolated From Environmental Water in Kolkata, India

Cholera toxin (CT)-producing Vibrio cholerae O1 and O139 cause acute diarrheal disease and are proven etiological agents of cholera epidemics and pandemics. On the other hand, V. cholerae non-O1/non-O139 are designated as non-agglutinable (NAG) vibrios and are not associated with epidemic cholera. The majority of NAG vibrios do not possess the gene for CT (ctx). In this study, we isolated three NAG strains (strains No. 1, 2, and 3) with ctx from pond water in Kolkata, India, and examined their pathogenic properties. The enterotoxicity of the three NAG strains in vivo was examined using the rabbit ileal intestinal loop test. Strain No. 1 induced the accumulation of fluid in the loop, and the volume of fluid was reduced by simultaneous administration of anti-CT antiserum into the loop. The volume of fluid in the loop caused by strains No. 2 and 3 was small and undetectable, respectively. Then, we cultured these three strains in liquid medium in vitro at two temperatures, 25 degrees C and 37 degrees C, and examined the amount of CT accumulated in the culture supernatant. CT was accumulated in the culture supernatant of strain No.1 when the strain was cultured at 25 degrees C, but that was low when cultured at 37 degrees C. The CT amount accumulated in the culture supernatants of the No. 2 and No. 3 strains was extremely low at both temperature under culture conditions examined. In order to clarify the virulence properties of these strains, genome sequences of the three strains were analyzed. The analysis showed that there was no noticeable difference among three isolates both in the genes for virulence factors and regulatory genes of ctx. However, vibrio seventh pandemic island-II (VSP-II) was retained in strain No. 1, but not in strains No. 2 or 3. Furthermore, it was revealed that the genotype of the B subunit of CT in strain No. 1 was type 1 and those of strains No. 2 and 3 were type 8. Histopathological examination showed the disappearance of villi in intestinal tissue exposed to strain No. 1. In addition, fluid accumulated in the loop due to the action of strain No. 1 had hemolytic activity. This indicated that strain No. 1 may possesses virulence factors to induce severe syndrome when the strain infects humans, and that some strains of NAG vibrio inhabiting pond water in Kolkata have already acquired virulence, which can cause illness in humans. There is a possibility that these virulent NAG vibrios, which have acquired genes encoding factors involved in virulence of V. cholerae O1, may emerge in various parts of the world and cause epidemics in the future

Development of Derivatives of 3, 3′-Diindolylmethane as Potent Leishmania donovani Bi-Subunit Topoisomerase IB Poisons

Background: The development of 3, 39-diindolyl methane (DIM) resistant parasite Leishmania donovani (LdDR50) by adaptation with increasing concentrations of the drug generates random mutations in the large and small subunits of heterodimeric DNA topoisomerase I of Leishmania (LdTOP1LS). Mutation of large subunit of LdTOP1LS at F270L is responsible for resistance to DIM up to 50 mM concentration. Methodology/Principal Findings: In search of compounds that inhibit the growth of the DIM resistant parasite and inhibit the catalytic activity of mutated topoisomerase I (F270L), we have prepared three derivatives of DIM namely DPDIM (2,29diphenyl 3,39-diindolyl methane), DMDIM (2,29-dimethyl 3,39-diindolyl methane) and DMODIM (5,59-dimethoxy 3,39diindolyl methane) from parent compound DIM. All the compounds inhibit the growth of DIM resistant parasites, induce DNA fragmentation and stabilize topo1-DNA cleavable complex with the wild type and mutant enzyme. Conclusion: The results suggest that the three derivatives of DIM can act as promising lead molecules for the generation of new anti-leishmanial agents

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Domestic Environmental Experience Design

The term ‘domestic environmental experience’ was defined as users’ experiences of cognitive perceptions and physical responses to their domestic built environments. Domestic environments can be enriched through the implementation of environmental experience design (EXD) by combining users’ environmental, spatial and contextual factors that may accommodate occupants’ needs and demands as well as their health and wellbeing. Here, an EXD theoretical concept has been developed based on the ‘User-Centred Design’ thematical framework

DNA topoisomerases in life and death: implications in kinetoplastid protozoa

Current biomedical research has its focus on the search for newer intervention strategies to control public health impact of parasitic diseases. The dramatic advances of molecular and cellular biology in recent times have provided opportunities for discovering and evaluating molecular targets for drug designing, which now form a rational basis for the development of improved anti parasitic therapy. DNA topoisomerases, the "cellular magicians" involved in nearly all biological processes governing DNA, have emerged as one such biological target. Over the last two decades, interest in topoisomerases has expanded beyond the realm of the basic science laboratory into the clinical arena. This review aims at providing a comprehensive insight into the biology of DNA topoisomerases and also focus on its evolution as a drug target in the unicellular kinetoplastids

Novel Betulin Derivatives as Antileishmanial Agents with Mode of Action Targeting Type

M. Unlike CPT, these compounds do not stabilize the cleavage complex, rather they abrogate the covalent complex formation. In processive mode of relaxation assay condition, these compounds slow down the strand rotation event, which ultimately affect the relaxation of supercoiled DNA. Interestingly, these compounds reduce the intracellular parasite burden in macrophages infected with wild type Leishmania as well as with sodium antimony gluconate resistant parasite (GE1). Taken together, our data suggest that these betulin derivatives can be exploited as potential drug candidates against threatening drug resistant leishmaniasis. MOL #72785 5 Introduction

Methodological Approach of <i>Environmental Experience Design</i> to Enhancing Occupants’ Well-Being, Bangladesh

Bangladesh is an emerging nation that is urbanizing the fastest. Generally, middle-income families (as the main economic contributors) reside in high-density urban housing due to their socioeconomic disparities in Dhaka, Bangladesh, where physical design aspects focus on current housing sectors. The congested living situation worsens indoor environmental quality and has a negative impact on occupants’ mental well-being in their dwellings, resulting in a direct or indirect adverse effect on their productivity. Occupants’ living conditions can be improved by adjusting individual perceptions and experiences in their domestic environments. This study has developed an “Environmental Experience Design (EXD)” methodological approach that reflects a user-centered design theoretical framework. A field study on dwelling units (±1000 sq ft) conducted throughout the selected housings in Dhaka, Bangladesh, was carried out to examine occupants’ domestic experiences using semi-structured and structured interviews. After binary and thematic coding with significance, “Association Rules and Cluster Analysis” were used to ascertain relationships between three aspects (spatial, environmental, and user context) to explore and customize outcomes. This EXD methodological approach can be utilized to create an environmental (architectural) design solution that will enhance mental well-being by considering occupants’ needs and demands in household settings locally and worldwide