Various hydrogels have been studied for nucleus pulposus regeneration. However, they failed to overcome the changes in the acidic environment during intervertebral disc degeneration. Therefore, a new functionalized peptide RAD/SA1 was designed by conjugating Sa12b, an inhibitor of acid-sensing ion channels, onto the C-terminus of RADA16-I. Then, the material characteristics and biocompatibility of RAD/SA1, and the bioactivities and mechanisms of degenerated human nucleus pulposus mesenchymal stem cells (hNPMSCs) were evaluated. Atomic force microscopy (AFM) and scanning electron microscopy (SEM) confirmed that RAD/SA1 self-assembling into three-dimensional (3D) nanofiber hydrogel scaffolds under acidic conditions. Analysis of the hNPMSCs cultured in the 3D scaffolds revealed that both RADA16-I and RAD/SA1 exhibited reliable attachment and extremely low cytotoxicity, which were verified by SEM and cytotoxicity assays, respectively. The results also showed that RAD/SA1 increased the proliferation of hNPMSCs compared to that in culture plates and pure RADA16-I. Quantitative reverse transcription polymerase chain reaction, enzyme-linked immunosorbent assay, and western blotting demonstrated that the expression of collagen I was downregulated, while collagen II, aggrecan, and SOX-9 were upregulated. Furthermore, Ca2+ concentration measurement and western blotting showed that RAD/SA1 inhibited the expression of p-ERK through Ca2+-dependent p-ERK signaling pathways. Therefore, the functional self-assembling peptide nanofiber hydrogel designed with the short motif of Sa12b could be used as an excellent scaffold for nucleus pulposus tissue engineering. Moreover, RAD/SA1 exhibits great potential applications in the regeneration of mildly degenerated nucleus pulposus.

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Salmonella enterica subspecies enterica serovar abortus equi (S. Abortus equi) is the most common cause of abortion in mares. It has recently been found to cause abortion in donkeys more frequently in China. A novel virulent bacteriophage vB_SabS_Sds2 (hereafter designated as Sds2) was isolated from the feces of donkeys using a S. Abortus equi strain as a host. Phage Sds2 had an isometric polyhedral head and an uncontracted long tail, belonging to the Tequintavirus, Markadamsvirinae, Demerecviridae, Caudovirales. The genome of phage Sds2 was 114,770 bp, with a GC content of 40.26%. The genome contained 160 open reading frames (ORFs), and no ORFs were associated with pathogenicity, drug resistance, or lysogenization by sequence analysis. Both genome annotation and phylogenetic analysis indicated that phage Sds2 was highly similar to T5-like bacteriophages. Phage Sds2 could lyse 100% (30/30) of S. Abortus equi strains, 25.3% (24/95) of other serotypes of Salmonella strains, and 27.6% (8/29) of Escherichia coli strains using the double-layer agar plate method. The in vitro test showed that phage Sds2 had high bactericidal activity against S. Abortus equi at a wide range of MOIs. The in vivo test indicated that phage Sds2 had an inhibitory effect on abortion in mice challenged with S. Abortus equi. In general, phage Sds2 is a novel lytic phage with a wide host range and has the potential to prevent abortion caused by S. Abortus equi.

Sa12b is a wasp peptide that can inhibit acid-sensitive ion channels (ASICs). The biological effects of nucleus pulposus mesenchymal stem cells (NP-MSCs) have not been investigated. Therefore, this study investigated the effect of Sa12b on the biological activity of NP-MSCs through ASICs in the acidic environment of intervertebral disc degeneration (IVDD). In this study, NP-MSCs were isolated from the nucleus pulposus (NP) in patients who underwent lumbar disc herniation surgery, identified by flow cytometry and tertiary differentiation, and cultured in vitro in an acidic environment model of IVDD with a pH of 6.2. Proliferation, and apoptosis were observed after different Sa12b concentrations were added to P2 generation NP-MSCs. The Ca2+ influx was detected using flow cytometry and laser confocal scanning microscopy, and qPCR was used to detect the relative expression of stem cell–associated genes (Oct4, Nanog, Jag1, and Notch1), the relative expression of extracellular matrix (ECM)–associated genes (collagen II, aggrecan, and SOX-9), and the relative expression of genes encoding ASICs (ASIC1, ASIC2, ASIC3, and ASIC4). Western blotting was used to detect the protein expression of collagen II and aggrecan in different treatment groups. Cells isolated and cultured from normal NP were spindle-shaped and adherent, and they exhibited expansion in vitro. Flow cytometry results showed that the cells exhibited high expression of CD73 (98.1%), CD90 (97.5%), and CD105 (98.3%) and low expression of HLA-DR (0.93%), CD34 (2.63%), and CD45 (0.33%). The cells differentiated into osteoblasts, adipocytes, and chondrocytes. According to the International Society for Cellular Therapy criteria, the isolated and cultured cells were NP-MSCs. With an increase in Sa12b concentration, the cell proliferation rate of NP-MSCs increased, and the apoptosis rate decreased significantly, reaching the optimal level when the concentration of Sa12b was 8 μg/μl. When the Sa12b concentration was 8 μg/μl and contained the ASIC non-specific inhibitor amiloride, the Ca2+ influx was the lowest, followed by that when the Sa12b concentration was 8 μg/μl. The Ca2+ influx was the highest in the untreated control group. qPCR results showed that as the concentration of Sa12b increased, the relative expression of Oct4, Nanog, Jag1, Notch1, collagen II, aggrecan, and SOX-9 increased, while that of ASIC1, ASIC2, ASIC3, and ASIC4 decreased. The difference was statistically significant (p < 0.05). In conclusion, Sa12b can improve the biological activity of NP-MSCs in severely acidic environments of the intervertebral disc by reducing Ca2+ influx via AISC inhibition and, probably, the Notch signaling pathway. This study provides a new approach for the biological treatment of IVDD. Inhibition of AISCs by Sa12b may delay IVDD and improve low back pain.

2021) Verbascoside attenuates experimental varicocele-induced damage to testes and sperm levels through up-regulation of the hypothalamus-pituitary-gonadal (HPG) axis,