Influence of Frequent Corticosteroid Local Injections on the Expression of Genes and Proteins Related to Fatty Infiltration, Muscle Atrophy, Inflammation, and Fibrosis in Patients With Chronic Rotator Cuff Tears: A Pilot Study.

Background: The effect of local corticosteroid (CS) injections on rotator cuff muscles remains poorly defined, despite the significance of muscle quality as a crucial prognostic factor for patients with rotator cuff tears (RCTs). Purpose: To compare alterations in gene and protein expression patterns in the rotator cuff muscles of patients with RCTs who received frequent joint CS injections with alterations in those without a history of CS injections. Study Design: Controlled laboratory study. Methods: A total of 24 rotator cuff muscle samples with medium-sized tears from 12 patients with a frequent joint CS injection history (steroid group; 7 men and 5 women who had received ≥5 injections with at least 1 within the previous 3 months; mean age, 63.0 ± 7.2 years) and 12 age- and sex-matched control patients without a history of CS injections (no-steroid group) were acquired. Alterations in the expression of genes and proteins associated with adipogenesis, myogenesis, inflammation, and muscle fibrosis were compared between the groups using quantitative reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemistry. Statistical analysis included comparison of group means using the Mann-Whitney U test, chi-square test, or Fisher exact test and logistic regression for multivariate analysis. Results: In the steroid group, the mRNA expression levels of adipogenic CCAAT/enhancer-binding protein alpha (C/EBPα; P = .008) and muscle atrophy-related genes (atrogin; P = .019) were significantly higher, and those of myogenic differentiation 1 (MyoD; P = .035), inflammatory interleukin 6 (IL-6; P = .035), and high mobility group box 1 (P = .003) were significantly lower compared with the no-steroid group. In addition, MyoD (P = .041) and IL-6 (P = .026) expression were decreased in the steroid versus no-steroid group. Immunohistochemistry revealed increased expression of C/EBPα and atrogin and decreased expression of MyoD and IL-6 in the steroid versus no-steroid group. Conclusion: Patients with RCTs and a history of frequent CS injections exhibited an upregulation of adipogenic and muscle atrophy-related genes and proteins within the rotator cuff muscles and a downregulation in the expression of myogenic and inflammatory genes and proteins in the same muscles. Clinical Relevance: These altered gene and protein expressions by frequent local CS injections may cause poor outcomes in patients with RCTs.

Multivalent electrostatic pi-cation interaction between synaptophysin and synapsin is responsible for the coacervation.

We recently showed that synaptophysin (Syph) and synapsin (Syn) can induce liquid-liquid phase separation (LLPS) to cluster small synaptic-like microvesicles in living cells which are highly reminiscent of SV cluster. However, as there is no physical interaction between them, the underlying mechanism for their coacervation remains unknown. Here, we showed that the coacervation between Syph and Syn is primarily governed by multivalent pi-cation electrostatic interactions among tyrosine residues of Syph C-terminal (Ct) and positively charged Syn. We found that Syph Ct is intrinsically disordered and it alone can form liquid droplets by interactions among themselves at high concentration in a crowding environment in vitro or when assisted by additional interactions by tagging with light-sensitive CRY2PHR or subunits of a multimeric protein in living cells. Syph Ct contains 10 repeated sequences, 9 of them start with tyrosine, and mutating 9 tyrosine to serine (9YS) completely abolished the phase separating property of Syph Ct, indicating tyrosine-mediated pi-interactions are critical. We further found that 9YS mutation failed to coacervate with Syn, and since 9YS retains Syph's negative charge, the results indicate that pi-cation interactions rather than simple charge interactions are responsible for their coacervation. In addition to revealing the underlying mechanism of Syph and Syn coacervation, our results also raise the possibility that physiological regulation of pi-cation interactions between Syph and Syn during synaptic activity may contribute to the dynamics of synaptic vesicle clustering.

DA-9601, a standardized extract of Artemisia asiatica, blocks TNF-alpha-induced IL-8 and CCL20 production by inhibiting p38 kinase and NF-kappaB pathways in human gastric epithelial cells.

AIM: To investigate whether, or how, DA-9601, which is a new gastroprotective agent, inhibits TNF-alpha-induced inflammatory signals in gastric epithelial AGS cells. METHODS: Cell viability was determined by MTT assay. IL-8 and CCL20 promoter activities were determined by a luciferase reporter gene assay. NF-kappaB-dependent transcriptional activity was determined by I-kappaB alpha degradation, NF-kappaB p65 nuclear translocation and a luciferase activity assay. IL-8 and CCL20 gene expression and protein secretion were determined by RT-PCR and an enzyme-linked immunosorbent assay (ELISA). Total and phosphorylated forms of mitogen-activated protein kinases (MAPKs) were determined by Western blot. RESULTS: Treatment of AGS cells with DA-9601 reduced TNF-alpha-induced IL-8 and CCL20 promoter activities, as well as their gene expression and protein release. TNF-alpha also induced NF-kappaB-dependent transcriptional activity in AGS cells. In contrast, in cells treated with DA-9601, TNF-alpha-induced NF-kappaB activity was significantly blocked. Although all three MAP kinase family members were phosphorylated in response to TNF-alpha, a selective inhibitor of p38 kinase SB203580 only could inhibit both NF-kappaB-dependent transcriptional activity and IL-8 and CCL20 production, suggesting a potential link between p38 kinase and NF-kappaB-dependent pathways in AGS cells. Interestingly, DA-9601 also selectively inhibited p38 kinase phosphorylation induced by TNF-alpha. CONCLUSION: DA-9601 blocked TNF-alpha-mediated inflammatory signals by potentially modulating the p38 kinase pathway and/or a signal leading to NF-kappaB-dependent pathways in gastric epithelial cells.