Activation of Autophagic Flux against Xenoestrogen Bisphenol-A-induced Hippocampal Neurodegeneration via AMP kinase (AMPK)/Mammalian Target of Rapamycin (mTOR) Pathways

The human health hazards related to persisting use of bisphenol-A (BPA) are well documented. BPA-induced neurotoxicity occurs with the generation of oxidative stress, neurodegeneration, and cognitive dysfunctions. However, the cellular and molecular mechanism(s) of the effects of BPA on autophagy and association with oxidative stress and apoptosis are still elusive. We observed that BPA exposure during the early postnatal period enhanced the expression and the levels of autophagy genes/proteins. BPA treatment in the presence of bafilomycin A₁ increased the levels of LC3-II and SQSTM1 and also potentiated GFP-LC3 puncta index in GFP-LC3-transfected hippocampal neural stem cell-derived neurons. BPA-induced generation of reactive oxygen species and apoptosis were mitigated by a pharmacological activator of autophagy (rapamycin). Pharmacological (wortmannin and bafilomycin A₁) and genetic (beclin siRNA) inhibition of autophagy aggravated BPA neurotoxicity. Activation of autophagy against BPA resulted in intracellular energy sensor AMP kinase (AMPK) activation, increased phosphorylation of raptor and acetyl-CoA carboxylase, and decreased phosphorylation of ULK1 (Ser-757), and silencing of AMPK exacerbated BPA neurotoxicity. Conversely, BPA exposure down-regulated the mammalian target of rapamycin (mTOR) pathway by phosphorylation of raptor as a transient cell''s compensatory mechanism to preserve cellular energy pool. Moreover, silencing of mTOR enhanced autophagy, which further alleviated BPA-induced reactive oxygen species generation and apoptosis. BPA-mediated neurotoxicity also resulted in mitochondrial loss, bioenergetic deficits, and increased PARKIN mitochondrial translocation, suggesting enhanced mitophagy. These results suggest implication of autophagy against BPA-mediated neurodegeneration through involvement of AMPK and mTOR pathways. Hence, autophagy, which arbitrates cell survival and demise during stress conditions, requires further assessment to be established as a biomarker of xenoestrogen exposure.     

Genome-wide analysis of signatures of selection in populations of African honey bees (Apis mellifera) using new web-based tools

Background

With the development of inexpensive, high-throughput sequencing technologies, it has become feasible to examine questions related to population genetics and molecular evolution of non-model species in their ecological contexts on a genome-wide scale. Here, we employed a newly developed suite of integrated, web-based programs to examine population dynamics and signatures of selection across the genome using several well-established tests, including F_ST, pN/pS, and McDonald-Kreitman. We applied these techniques to study populations of honey bees (Apis mellifera) in East Africa. In Kenya, there are several described A. mellifera subspecies, which are thought to be localized to distinct ecological regions.

Results

We performed whole genome sequencing of 11 worker honey bees from apiaries distributed throughout Kenya and identified 3.6 million putative single-nucleotide polymorphisms. The dense coverage allowed us to apply several computational procedures to study population structure and the evolutionary relationships among the populations, and to detect signs of adaptive evolution across the genome. While there is considerable gene flow among the sampled populations, there are clear distinctions between populations from the northern desert region and those from the temperate, savannah region. We identified several genes showing population genetic patterns consistent with positive selection within African bee populations, and between these populations and European A. mellifera or Asian Apis florea.

Conclusions

These results lay the groundwork for future studies of adaptive ecological evolution in honey bees, and demonstrate the use of new, freely available web-based tools and workflows (http://usegalaxy.org/r/kenyanbee) that can be applied to any model system with genomic information.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1712-0) contains supplementary material, which is available to authorized users.     

Prophylactic role of D-Saccharic acid-1,4-lactone in tertiary butyl hydroperoxide induced cytotoxicity and cell death of murine hepatocytes via mitochondria-dependent pathways

D-Saccharic acid 1,4-lactone (DSL) is a derivative of D-glucaric acid. It is a beta-glucuronidase inhibitor and possesses anticarcinogenic, detoxifying, and antioxidant properties. In the present study, the protective effects of DSL were investigated against tertiary butyl hydroperoxide (TBHP) induced cytotoxicity and cell death in vitro using murine hepatocytes. Exposure of TBHP caused a reduction in cell viability, enhanced the membrane leakage, and disturbed the intracellular antioxidant machineries in murine hepatocytes. Investigating the signaling mechanism of TBHP-induced cellular pathophysiology and protective action of DSL, we found that TBHP exposure disrupted mitochondrial membrane potential, facilitated cytochrome c release in the cytosol, and led to apoptotic cell death via mitochondria-dependent pathways. DSL counteracted these changes and maintained normalcy in hepatocytes. Combining, results suggest that DSL possesses the ability to ameliorate TBHP-induced oxidative insult, cytotoxicity, and apoptotic cell death probably due to its antioxidant activity and functioning via mitochondria-dependent pathways.     

Cellulase and xylanase activity in relation to biofilm formation by two intertidal filamentous fungi in a novel polymethylmethacrylate conico-cylindrical flask

A polymethylmethacrylate (PMMA) conico-cylindrical flask (CCF) with an inner arrangement consisting of eight equidistantly spaced rectangular strips mounted radially on a circular disk to provide additional surface area for fungal attachment was employed for production of cellulase by Chaetomium crispatum and xylanase by Gliocladium viride. The design allowed comparison of production between CCFs with hydrophobic surface (PMMA-CCF), hydrophilic glass surface (GS-CCF) and 500-ml Erlenmeyer flask (EF). Compared with the EF, endo-β-1,4-glucanase and FPase (filter paper degradation) activities increased from 0.044 to 0.156 and from 0.008 to 0.021 IU/ml, respectively, in the PMMA-CCF, while growth of C. crispatum was higher by at most 1.38-fold compared with the other vessels. Xylanase production in the EF was at most 5.08-fold higher and growth of G. viride was at most 1.52-fold higher compared with the other vessels. Temporal pattern of biofilm development based on two-channel fluorescence detection of extracellular polymeric substances (EPSs) and whole cells in a confocal laser scanning microscope demonstrated increase by 100% in biovolume, 25% in thickness and 62.5% both in substratum coverage and total spreading of C. crispatum biofilm in PMMA-CCF over 6 days. Biovolume of G. viride biofilm in GS-CCF increased by 150% over 4 days while that in PMMA-CCF enhanced by 200% over 2 days. Biofilm thickness in PMMA-CCF was 44% higher compared with GS-CCF and increased by 175% over 2 days. Substratum coverage was 38% higher in GS-CCF compared with PMMA-CCF. Thus, reactor surface area and property, shear forces and biofilm formation influenced enzyme production.     

Synthesis and evaluation of anti-HIV activity of isatin beta-thiosemicarbazone derivatives

On the basis of pharmacophoric modelling studies of existing NNRTIs, a series of isatin beta-thiosemicarbazone derivatives was synthesized and evaluated for their anti-HIV activity in HTLV-III(B) strain in the CEM cell line. Three compounds showed significant anti-HIV activity, whereupon compound 6 was found to be the most active compound with an EC(50) value of 2.62 microM and a selectivity index of 17.41, while not being cytotoxic to the cell line at a CC(50) value of 44.90 microM. Other tested compounds exhibited marked activity below their toxicity threshold.