Facile Synthesis of Benzimidazoles via Oxidative Cyclization of Acyclic Monoterpene Aldehyde with Diamines: Studies on Antimicrobial and in Vivo Evaluation of Zebrafish

Citral ( 1a ), a bioactive component of Cymbopogon citratus (lemongrass) could be isolated and semi-synthetic analogs synthesized with improved therapeutic properties. Herein we first report describes citral ( 1a ) as a primary material for the synthesis of benzimidazole derivatives between various o-phenylenediamines ( 2a – l ) in the presence of Diisopropylethylamine (DIPEA) as a commercially available environmentally benign base, ethanol as a green solvent and the yield of all benzimidazole derivatives ( 3a – l ) was between 68–76 %; The semi-synthetically prepared benzimidazole derivatives ( 3a – l ) were assessed for their anti-bacterial and anti-fungal properties. The benzimidazole compounds ( 3a – b , and 3g – j ) exhibit good anti-microbial activity. In addition, in silico study was carried out to determine the specific binding affinity of the diamine halogen substituted benzimidazole derivatives to the specific target proteins. In silico analysis revealed a high correlation between docking results and experimental results. Finally, benzimidazole demonstrated significant antibacterial and antifungal activity. Zebrafish embryos were subjected to In vivo toxicological test found that all of the benzimidazole compounds ( 3a – l ) were non-toxic and had low embryotoxicity after 96 h, with an LC₅₀ of 36.425 μg, which could facilitate the design of novel antimicrobial agents using a cost-effective method.     

Antifertility effect of Andrographis paniculata (Nees) in male albino rat

Dry leaf powder of A. paniculata, when fed orally to male albino rats, at a dose level of 20 mg powder per day for 60 days, resulted in cessation of spermatogenesis, degenerative changes in the seminiferous tubules, regression of Leydig cells and regressive and/or degenerative changes in the epididymis, seminal vesicle, ventral prostate and coagulating gland. There was reduction in the weight and fluid content of the accessory glands. The treatment also resulted in accumulation of glycogen and cholesterol in the testis, and increased activities of lactate dehydrogenase in testis and alkaline phosphatase in testis and ventral prostate. The results suggest antispermatogenic and/or antiandrogenic effect of the plant.     

A delayed rectifier current is modulated by the circadian pacemaker in Bulla

Basal retinal neurons of the marine mollusc Bulla gouldiana continue to express a circadian modulation of their membrane conductance for at least two cycles in cell culture. Voltage-dependent currents of these pacemaker cells were recorded using the whole-cell perforated patch-clamp technique to characterize outward currents and investigate their putative circadian modulation. Three components of the outward potassium current were identified. A transient outward current (IA) was activated after depolarization from holding potentials greater than -30 mV, inactivated with a time constant of 50 ms, and partially blocked by 4-aminopyridine (1-5 mM). A Ca(2+)-dependent potassium current (IK(Ca)) was activated by depolarization to potentials more positive than -10 mV and was blocked by removing Ca2+ from the bath or by applying the Ca2+ channel blockers Cd2+ (0.1-0.2 mM) and Ni2+ (1-5 mM). A sustained Ca(2+)-independent current component including the delayed rectifier current (IK) was recorded at potentials positive to -20 mV in the absence of extracellular Na+ and Ca2+ and was partially blocked by tetraethylammonium chloride (TEA, 30mM). Whole-cell currents recorded before and after the projected dawn and normalized to the cell capacitance revealed a circadian modulation of the delayed rectifier current (IK). However, the IA and IK(Ca) currents were not affected by the circadian pacemaker.     

Differentiation to the CCR2+ inflammatory phenotype in vivo is a constitutive, time-limited property of blood monocytes and is independent of local inflammatory mediators

It is proposed that CCR2+ monocytes are specifically recruited to inflammatory sites, whereas CCR2- monocytes are recruited to normal tissue to become resident macrophages. Whether these subsets represent separate lineages, how differential trafficking is regulated and whether monocytes undergo further differentiation is uncertain. Using a mouse model of autoimmune uveoretinitis we examined monocyte trafficking to the inflamed retina in vivo. We show that bone marrow-derived CD11b+ F4/80- monocytes require 24 to 48 h within the circulation and lymphoid system before acquiring the CCR2+ phenotype and trafficking to the inflamed retina is enabled. This phenotype, and the capacity to traffic were lost by 72 h. Monocyte CCR2 expression followed a similar time course in normal mice indicating that differentiation to an inflammatory phenotype is a constitutive, time-limited property, independent of local inflammatory mediators. Phenotypic analysis of adoptively transferred cells indicated that circulating inflammatory monocytes also differentiate into CD11c+ and B220+ dendritic cells and F4/80+ tissue macrophages in vivo. Our data supports the hypothesis of continuous extravasation and progressive differentiation over time of inflammatory monocytes in the circulation rather than replication within the actively inflamed tissue, and supports the concept of myeloid dendritic cell differentiation from trafficking monocytes under physiological conditions in vivo.     

Combinatorial effect of fish oil (Maxepa) and 1α,25-dihydroxyvitamin D3 in the chemoprevention of DMBA-induced mammary carcinogenesis in rats

The present study demonstrates the anti-tumor effects of combined supplementations of dietary fish oil (Maxepa) and 1α,25-dihydroxyvitamin D₃ (vitamin D₃) on 7,12-dimethylbenz(α)anthracene (DMBA)-induced rat mammary carcinogenesis. Female Sprague–Dawley rats at 50 days of age were treated with 7,12-dimethylbenz(α)anthracene (DMBA; 0.5 mg/100 g body weight) by a single tail vein injection in an oil emulsion. Both fish oil (rich in EPA and DHA) and vitamin D₃ were administered orally at a dose of 0.5 ml/day/rat and 0.3 μg/100 μL propylene glycol twice a week respectively and continued to 35 weeks after DMBA administration. Fish oil in combination with vitamin D₃ resulted in a significant reduction in incidence, multiplicity and volume of mammary tumors. These supplementation also inhibited DMBA-induced mammary 7-methylguanine DNA adducts formation, which was measured by HPLC-fluorescence assay (at four sequential time points; ANOVA, F = 42.56, P < 0.0001). Immunohistochemical analysis revealed that the effect of fish oil and vitamin D₃ occurred through suppression of cell proliferation (BrdU-LI: P < 0.0001). Fish oil and vitamin D₃ together also reduced the mRNA expression of iNOS (84%, P < 0.05). In view of their natural availability, non-toxicity and acceptability; combined supplementation of fish oil and vitamin D₃ might be effective for chemoprevention of mammary carcinogenesis.     

Loss of the tumor suppressor Pten promotes proliferation of Drosophila melanogaster cells in vitro and gives rise to continuous cell lines

In vivo analysis of Drosophila melanogaster has enhanced our understanding of many biological processes, notably the mechanisms of heredity and development. While in vivo analysis of mutants has been a strength of the field, analyzing fly cells in culture is valuable for cell biological, biochemical and whole genome approaches in which large numbers of homogeneous cells are required. An efficient genetic method to derive Drosophila cell lines using expression of an oncogenic form of Ras (Ras(V12)) has been developed. Mutations in tumor suppressors, which are known to cause cell hyperproliferation in vivo, could provide another method for generating Drosophila cell lines. Here we screened Drosophila tumor suppressor mutations to test if they promoted cell proliferation in vitro. We generated primary cultures and determined when patches of proliferating cells first emerged. These cells emerged on average at 37 days in wild-type cultures. Using this assay we found that a Pten mutation had a strong effect. Patches of proliferating cells appeared on average at 11 days and the cultures became confluent in about 3 weeks, which is similar to the timeframe for cultures expressing Ras(V12). Three Pten mutant cell lines were generated and these have now been cultured for between 250 and 630 cell doublings suggesting the life of the mutant cells is likely to be indefinite. We conclude that the use of Pten mutants is a powerful means to derive new Drosophila cell lines.