Inhibition of bfl-1/A1 by siRNA inhibits mycobacterial growth in THP-1 cells by enhancing phagosomal acidification

Virulent tubercle bacilli inhibit apoptosis to establish a safe environment within the host cells. Here, we report that NF-kappaB dependent antiapoptotic protein bfl-1/A1 plays an important role in this process. Both virulent and avirulent mycobacteria bearing THP-1 cells expressed considerable amount of bfl-1/A1 after 4 h of infection. However, after 48 h of infection, bfl-1/A1 expression was evident only in Mycobacterium tuberculosis H37Rv but not in M. tuberculosis H37Ra infected cells. When parallel experiments were performed with Human monocyte-derived macrophages (MDMs), differential expression of bfl-1/A1 mRNA was observed in case of M. tuberculosis H37Rv and M. tuberculosis H37Ra infection. siRNA mediated inhibition of bfl-1/A1 induced apoptosis in M. tuberculosis H37Rv infected THP-1 and MDMs. Reduction in intracellular mycobacterial growth was observed in bfl-1/A1 siRNA transfected, M. tuberculosis H37Rv infected THP-1 cells. Enhancement of phagosome-lysosome fusion was observed in bfl-1/A1 siRNA treated and M. tuberculosis H37Rv infected THP-1 cells. These results clearly indicated that differential expression of bfl-1/A1 in M. tuberculosis H37Rv and M. tuberculosis H37Ra infected THP-1 cells probably account for the difference in infection outcome.     

A comprehensive network map of IL-17A signaling pathway

Interleukin-17A (IL-17A) is one of the member of IL-17 family consisting of other five members (IL-17B to IL-17F). The Gamma delta (γδ) T cells and T helper 17 (Th17) cells are the major producers of IL-17A. Aberrant signaling by IL-17A has been implicated in the pathogenesis of several autoimmune diseases including idiopathic pulmonary fibrosis, acute lung injury, chronic airway diseases, and cancer. Activation of the IL-17A/IL-17 receptor A (IL-17RA) system regulates phosphoinositide 3-kinase/AKT serine/threonine kinase/mammalian target of rapamycin (PI3K/AKT/mTOR), mitogen-activated protein kinases (MAPKs) and activation of nuclear factor-κB (NF-κB) mediated signaling pathways. The IL-17RA activation orchestrates multiple downstream signaling cascades resulting in the release of pro-inflammatory cytokines such as interleukins (IL)-1β, IL-6, and IL-8, chemokines (C-X-C motif) and promotes neutrophil-mediated immune response. Considering the biomedical importance of IL-17A, we developed a pathway resource of signaling events mediated by IL-17A/IL-17RA in this study. The curation of literature data pertaining to the IL-17A system was performed manually by the NetPath criteria. Using data mined from the published literature, we describe an integrated pathway reaction map of IL-17A/IL-17RA consisting of 114 proteins and 68 reactions. That includes detailed information on IL-17A/IL-17RA mediated signaling events of 9 activation/inhibition events, 17 catalysis events, 3 molecular association events, 68 gene regulation events, 109 protein expression events, and 6 protein translocation events. The IL-17A signaling pathway map data is made freely accessible through the WikiPathways Database (https://www.wikipathways.org/index.php/Pathway: WP5242).Supplementary InformationThe online version contains supplementary material available at 10.1007/s12079-022-00686-y.     

Follistatin Protein Enhances Satellite Cell Counts in Reinnervated Muscle

Background Muscle recovery following peripheral nerve repair is sup-optimal. Follistatin (FST), a potent muscle stimulant, enhances muscle size and satellite cell counts following reinnervation when administered as recombinant FST DNA via viral vectors. Local administration of recombinant FST protein, if effective, would be more clinically translatable but has yet to be investigated following muscle reinnervation. Objective  The aim of this study is to assess the effect of direct delivery of recombinant FST protein on muscle recovery following muscle reinnervation. Materials and Methods  In total, 72 Sprague-Dawley rats underwent temporary (3 or 6 months) denervation or sham denervation. After reinnervation, rats received FST protein (isoform FS-288) or sham treatment via a subcutaneous osmotic pump delivery system. Outcome measures included muscle force, muscle histomorphology, and FST protein quantification. Results  Follistatin treatment resulted in smaller muscles after 3 months denervation ( p  = 0.019) and reduced force after 3 months sham denervation ( p  < 0.001). Conversely, after 6 months of denervation, FST treatment trended toward increased force output ( p  = 0.066). Follistatin increased satellite cell counts after denervation ( p  < 0.001) but reduced satellite cell counts after sham denervation ( p  = 0.037). Conclusion  Follistatin had mixed effects on muscle weight and force. Direct FST protein delivery enhanced satellite cell counts following reinnervation. The positive effect on the satellite cell population is intriguing and warrants further investigation.     

Reconstitution of rapeseed high molecular weight protein from isolated subunits

The high molecular weight protein was isolated from rapeseed and characterised. Six subunits were isolated in SDS (0.01%) solution on polyacrylamide-gel electrophoresis and by gel filtration on Sephadex G-100. Reassociation by removing SDS by co-dialysis, against 10 mM sodium phosphate buffer (pH 7.9) was done and the yield was about 90%. The reconstituted protein was indistinguishable from the intact protein in all respects. The subunits isolated from the native protein and the reconstituted protein were found to have identical molecular weights and N-terminal amino acids. No disulphide bonds were observed in the subunit association. Amino acid analysis of the proteins and the six subunits was performed and the number of each amino acid residue calculated.     

Characterization and Cytotoxic Evaluation of Bacteriocins Possessing Antibiofilm Activity Produced by Lactobacillus plantarum SJ33

International Journal of Peptide Research and Therapeutics - Biofilm forming pathogens are among the major causes of hospital-acquired infections and are not much affected by antibiotic treatment....     

Noninvasive Method of DNA Isolation From Fecal Epithelial Tissue of Dairy Animals

A novel noninvasive genomic DNA isolation protocol from fecal tissue, by the proteinase K digestion and guanidine hydrochloride extraction method, was assessed for the genotyping of cattle and buffalo. The epithelial tissues present on the surface of the feces were used as source for isolation of genomic DNA. The DNA isolated from fecal tissue was found to be similar as those obtained from other body tissues such as skin, brain, liver, kidney, and muscle. The quality of DNA was checked by agarose gel electrophoresis and polymerase chain reaction (PCR). We successfully amplified a 320 bp MHC class II DRB gene and a 125 bp mt-DNA D-loop region from isolated genomic DNA of cattle. Thus, the DNA isolated using this method was suitable for common molecular biology methods, such as restriction enzyme digestion and genotyping of dairy animals through PCR.     

Influence of plant growth regulators on micropropagation and in vitro flowering of Trichodesma indicum (Linn) R. Br

An efficient protocol for micropropagation and in vitro flowering of Trichodesma indicum (Linn) R. Br. was developed using shoot tip explants. The physiological role of cytokinin and its combination with auxins on micropropagation and in vitro flowering was investigated. The highest number of shoots (9.94 ± 0.10) and the maximum average shoot length (5.56 ± 0.35 cm) were recorded on Murashige and Skoog (MS) medium supplemented with benzylaminopurine (BAP) (4.44 μM) and naphthaleneacetic acid (NAA) (2.69 μM). The effect of sucrose concentration on in vitro floral development was studied in plantlets cultured on MS medium supplemented with gibberellic acid (GA₃) and BAP. The highest percentage of flowering (93.2%) was obtained on MS medium supplemented with GA₃ (1.44 μM), BAP (1.33 μM) and sucrose (30 g l^?1). Root formation from the adventitious shoots was easily achieved on MS medium containing indole-3-butyric acid (IBA) (2.46 μM). The regenerated plantlets showed 86% survival rate and were phenotypically normal. The described method can be successfully employed for large-scale multiplication and in vitro flowering of T. indicum.     

Anticariogenic and phytochemical evaluation of Eucalyptus globules Labill.

In the present study, in vitro anticariogenic potential of ethyl acetate, hexane and methanol and aqueous extracts of plant leaves of Eucalyptus globules Labill. were evaluated by using four cariogenic bacteria, Lactobacillus acidophilus, Lactobacillus casei, Staphylococcus aureus and Streptococcus mutans. Agar well diffusion method and minimum inhibitory concentration (MIC) were used for this purpose. The ethyl acetate extracted fraction of plant leaves showed good inhibitory effects against all selected bacteria. In Eucalyptus globules, hexane and ethyl acetate extracts found highly effective against, Lactobacillus acidophilus with MIC value of 0.031 and 0.062 mg/mL, respectively. Qualitative phytochemical investigation of above extracts showed the presence of alkaloids, phenolic compounds, steroids, cardiac glycosides and terpenes. Based on the MIC value and bioautography, ethyl acetate of plant leaf was selected for further study. Further investigation on the structure elucidation of the bioactive compound using IR, GC-MS and NMR techniques revealed the presence of alpha-farnesene, a sesquiterpene. Eucalyptus globules plant leaf extracts have great potential as anticariogenic agents that may be useful in the treatment of oral disease.