Length–weight relationships of six indigenous fish species from the River Cauvery and its estuary,India

The present study estimated length–weight relationships (LWRs) for six indigenous fish species (Barilius gatensis, Salmostoma acinaces, S. boopis, Puntius amphibius, Hemibagrus punctatus and Ambassis miops) based on specimens collected from River Cauvery (including estuary) during July 2017–January 2020. The sampling surveys were carried out in three distinct sampling seasons, viz., the pre-monsoon (March–May), the monsoon (July–October) and the post-monsoon (November–February). Majority of the fish specimens dealt in the study were collected from multi-meshed monofilament gill nets (mesh sizes 18, 30, 45, 60, 90, 110, 120 and 150 mm) operated by local fishers. For those sites situated in the protected areas, sampling was carried out by cast nets with prior permission from the local administration and the collected fishes were released back into river after length–weight measurements. The length measurements were noted as total length (TL) measured to the nearest 0.1 cm by using a digital Vernier caliper. A digital balance was used for weight measurements with an accuracy of 0.01 g. The study recorded a new maximum length of 48 cm for H. punctatus. The LWR data generated from the present study are significant for proper assessment of the stock status and their management, if collected together with other essential biological and physical parameters.     

Comparative modeling of Rab6 proteins: identification of key residues and their interactions with guanine nucleotides

The cytoplasm of a eukaryotic cell consists of a wide variety of membrane bound cell organelles and continuous flow of proteins amongst these organelles is a major challenge and must be stringently maintained in order to continue the correct biochemical functioning inside a cell. The transportation of various proteins amongst these organelles is facilitated by a vast Tubulo-vesicular network mediated by carrier proteins. The Rabs belong to small G proteins super family involved in the regulation and vesicle transport in between the organelles by shuttling between the active GTP and inactive GDP bound states. In this paper we put forth the homology modeling and docking studies of Rab6A proteins (Mus musculus, Gallus gallus and Caenorhabditis elegans) with GTP, GMP-PNP and GDP molecules and a comparative study between these proteins is done to identify key residues out of which serine of the phosphate binding loop (P – loop) and aspartic acid showed prominent interactions with the GTP, GDP and GMP-PNP nucleotides and cogitate that aspartic acid might also help in the stabilization of the switch I region of the Rab proteins besides serine.     

Decreased Expression of DREAM Promotes the Degeneration of Retinal Neurons

The intrinsic mechanisms that promote the degeneration of retinal ganglion cells (RGCs) following the activation of N-Methyl-D-aspartic acid-type glutamate receptors (NMDARs) are unclear. In this study, we have investigated the role of downstream regulatory element antagonist modulator (DREAM) in NMDA-mediated degeneration of the retina. NMDA, phosphate-buffered saline (PBS), and MK801 were injected into the vitreous humor of C57BL/6 mice. At 12, 24, and 48 hours after injection, expression of DREAM in the retina was determined by immunohistochemistry, western blot analysis, and electrophoretic mobility-shift assay (EMSA). Apoptotic death of cells in the retina was determined by terminal deoxynucleotidyl transferace dUTP nick end labeling (TUNEL) assays. Degeneration of RGCs in cross sections and in whole mount retinas was determined by using antibodies against Tuj1 and Brn3a respectively. Degeneration of amacrine cells and bipolar cells was determined by using antibodies against calretinin and protein kinase C (PKC)-alpha respectively. DREAM was expressed constitutively in RGCs, amacrine cells, bipolar cells, as well as in the inner plexiform layer (IPL). NMDA promoted a progressive decrease in DREAM levels in all three cell types over time, and at 48 h after NMDA-treatment very low DREAM levels were evident in the IPL only. DREAM expression in retinal nuclear proteins was decreased progressively after NMDA-treatment, and correlated with its decreased binding to the c-fos-DRE oligonucleotides. A decrease in DREAM expression correlated significantly with apoptotic death of RGCs, amacrine cells and bipolar cells. Treatment of eyes with NMDA antagonist MK801, restored DREAM expression to almost normal levels in the retina, and significantly decreased NMDA-mediated apoptotic death of RGCs, amacrine cells, and bipolar cells. Results presented in this study show for the first time that down-regulation of DREAM promotes the degeneration of RGCs, amacrine cells, and bipolar cells.     

Anthranilate derivatives as TACE inhibitors: Docking based CoMFA and CoMSIA analyses

Anthranilic acid based derivatives (ANTs) have been identified as a novel class of potent tumor necrosis factor-α converting enzyme (TACE) inhibitors. A computational strategy based on molecular docking studies, followed by CoMFA and CoMSIA analyses has been performed to elucidate the atomic details of the TACE/ANT interactions and also to identify the most important features impacting TACE inhibitory activity of ANTs. The CoMSIA model resulted to be slightly more predictive than CoMFA model, and gave conventional r² 0.991, r_cv² 0.793, q² 0.777, SEE 0.050, F-value 655.610, and r_test² 0.871. The 3D-QSAR field contributions and the structural features of the TACE binding site showed a good correlation. These studies will be useful to design new TACE inhibitors with improved potency.     

Enhanced Bioavailability of Buspirone From Reservoir-Based Transdermal Therapeutic System,Optimization of Formulation Employing Box–Behnken Statistical Design

The purpose of the present study was to develop and optimize reservoir-based transdermal therapeutic system (TTS) for buspirone (BUSP), a low bioavailable drug. A three-factor, three-level Box–Behnken design was employed to optimize the TTS. Hydroxypropyl methylcellulose, d-limonene and propylene glycol were varied as independent variables; cumulative amount permeated across rat abdominal skin in 24 h, flux and lag time were selected as dependent variables. Mathematical equations and response surface plots were used to relate the dependent and independent variables. The statistical validity of polynomials was established, and optimized formulation factors were selected by feasibility and grid search. Validation of the optimization study with seven confirmatory runs indicated high degree of prognostic ability of response surface methodology. BUSP-OPT (optimized formulation) showed a flux 104.6 μg cm⁻² h⁻¹, which could meet target flux. The bioavailability studies in rabbits showed that about 2.65 times improvement (p < 0.05) in bioavailability, after transdermal administration of BUSP-OPT compared to oral solution. The ex vivo–in vivo correlation was found to have biphasic pattern and followed type A correlation. Reservoir-based TTS for BUSP was developed and optimized using Box–Behnken statistical design and could provide an effective treatment in the management of anxiety.     

A single-nucleotide polymorphism in tumor necrosis factor-α (− 308 G/A) as a biomarker in chronic pancreatitis

Objective

Chronic pancreatitis is a gradual, long-term inflammation of the pancreas that results in alteration of its normal structure and function. The study aims to investigate the role of − 308 (G/A) polymorphism of TNF-α gene in chronic pancreatitis.

Material and methods

A total of 200 subjects were included in this case–control study. A total of 100 in patients admitted in the Gastroenterology Unit of Gandhi Hospital and Osmania General Hospital, Hyderabad were included in the present study. An equal number of healthy control subjects were randomly selected for the study. The genotyping of TNF-α gene was carried out by tetra-primer ARMS PCR followed by gel electrophoresis. The TNF-α levels were assayed by enzyme-linked immunosorbent assay.

Results

A significant variation with respect to the genotypic and allelic distribution in the disease group when compared to control subjects [OR = 2.001 (1.33–3.005), p < 0.0001**] was observed. Subjects homozygous for the A allele had higher TNF-α levels compared to G allele.

Conclusion

The present study revealed a significant association of the TNF-α gene promoter polymorphism with chronic pancreatitis. Thus, TNF-α genotype can be considered as one of the biological markers in the etiology of chronic pancreatitis.     

The conserved ubiquitin-like protein Hub1 plays a critical role in splicing in human cells

Different from canonical ubiquitin-like proteins, Hub1 does not form covalent conjugates with substrates but binds proteins noncovalently. In Socchoromyces cerevisioe, Hub1 associates with spUceosomes and mediates alternative splicing of SRCI, without affecting pre-mRNA splicing generaity. Human Hub1 is highty similar to its yeast homotog, but its cellular function remains largely unexplored. Here, we show that human Hub1 binds to the spliceosomal protein Snu66 as in yeast; however, unlike its 5. cerevisioe homolos, human Hub1 is essential for viability. Prolonged in vivo depletion of human Hub1 leads to various cellular defects, including splicing speckle abnormalities, partial nuclear retention of mRNAs, mitotic catastrophe, and consequently cell death by apoptosis. Early consequences of Hub1 depletion are severe splicing defects, however, only for specific splice sites leading to exon skipping and intron retention. Thus, the ubiquitin-iike protein Hub1 is not a canonlcal spliceosomal factor needed generally for splicing, but rather a modulator of spliceosome performance and facilitator of alternative splicing.     

A Novel Cell Fixation Method that Greatly Enhances Protein Identification in Microproteomic Studies Using Laser Capture Microdissection and Mass Spectrometry

Microproteomic studies have improved our knowledge of cell biology. Yet, with mass spectrometry (MS) analysis, accuracy can be lost for protein identification and quantification when using heterogeneous samples. Laser capture microdissection (LCM) allows for the enrichment of specific subsets of cells to study their proteome; however, sample fixation is necessary. Unfortunately, fixation hampers MS results due to protein cross‐linking. The aim of this study was to identify both a fixation protocol and an extraction method that returns the best yield of proteins for downstream MS analysis, while preserving cellular structures. We compared glutaraldehyde (GLU), a common fixative to preserve cells, to dithiobispropionimidate (DTBP), a cleavable cross‐linker. Our DTBP fixation/extraction protocol greatly increased the protein recovery. In fact, while 1000 GLU fixed cells returned only 159 unique protein hits, from 1464 unique peptides of 1994 unique collected spectra, 1000 DTBP fixed cells resulted in 567 unique collected protein hits, from 7542 unique peptides, of 10,401 unique collected spectra. That is, a 3.57‐fold increase in protein hits, 5.15‐fold increase in unique peptides, and a 5.22‐fold increase in unique collected spectra. Overall, the novel protocol introduced here allows for a very efficient protein recovery and good sample quality for MS after sample collection using LCM.     

A novel templates of piperazinyl-1,2-dihydroquinoline-3-carboxylates: Synthesis,anti-microbial evaluation and molecular docking studies

A series of piperazinyl-1,2-dihydroquinoline carboxylates were synthesized by the reaction of ethyl 4-chloro-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylates with various piperazines and their structures were confirmed by ¹H NMR, ¹³C NMR, IR and mass spectral analysis. All the synthesized compounds were screened for their in vitro antimicrobial activities. Further, the in silico molecular docking studies of the active compounds was performed to explore the binding interactions between piperazinyl-1,2-dihydroquinoline carboxylate derivatives and the active site of the Staphylococcus aureus (CrtM) dehydrosqualene synthase (PDB ID: 2ZCQ). The docking studies revealed that the synthesized derivatives showed high binding energies and strong H-bond interactions with the dehydrosqualene synthase validating the observed antimicrobial activity data. Based on antimicrobial activity and docking studies, the compounds 9b and 10c were identified as promising antimicrobial lead molecules. This study might provide insights to identify new drug candidates that target the S. aureus virulence factor, dehydrosqualene synthase.