Epidemiology and clinical features of Skin and Soft Tissue Infections Caused by PVL-Positive and PVL-Negative Methicillin-Resistant <i>Staphylococcus aureus</i> Isolates in inpatients in China: a single-center retrospective 7-year study.

Previous studies have mainly focused on outpatient cases of skin and soft tissue infections (SSTIs), with limited attention to inpatient occurrences. Thus, we aimed to compare the clinical parameters of inpatients with SSTIs, performed genomic characterization, and determined the subtypes of Panton-Valentine leucocidin (PVL) bacteriophages of methicillin-resistant Staphylococcus aureus (MRSA) strains isolated from these patients. We found that PVL-positive patients had shorter hospital stays (mean, 9 vs. 24 days; p lukSF-PV gene, revealing that major clusters were associated with specific STs, suggesting independent acquisition of PVL by different strain types and indicating that significant diversity was observed even within PVL-positive strains detected in the same facility. Overall, our study provides comprehensive insights into the clinical, genetic, and phage-related aspects of MRSA-induced SSTIs in hospitalized patients and contributes to a more profound understanding of the epidemiology and evolution of these pathogens in the Chinese population.</p

The bioactive sphingolipid S1P induces secretion of IL-8 from ASM cells, but this can be repressed by the corticosteroid dexamethasone in a concentration-dependent manner.

<p>(A) Growth-arrested ASM cells were treated with vehicle or S1P (1 μM) for 24 h and IL-8 secretion measured by ELISA. Statistical analysis was performed using the Student's unpaired <i>t</i> test, where § denotes a significant effect of S1P (<i>P</i><0.05). (B) To demonstrate corticosteroid-mediated repression of S1P-induced IL-8 secretion we then pretreated growth-arrested ASM cells for 1 h with vehicle or dexamethasone (0.0001-1M), before stimulation for 24 h with S1P (1 μM). IL-8 protein was measured by ELISA and results expressed as % S1P-induced IL-8 secretion at 24 h. Statistical analysis was performed using one-way ANOVA then Fisher's post-hoc multiple comparison test, where * denotes significant repression by dexamethasone (<i>P</i><0.05). Data are mean+SEM values from n = 12 primary ASM cell lines.</p

S1P induces IL-8 secretion via a MSK1 dependent pathway and MSK1 knock-down by siRNA attenuates S1P-induced neutrophil chemotaxis.

<p>ASM cells were transiently transfected using nucleofection with scrambled control or MSK1 siRNA, growth-arrested, then treated for 24 h with vehicle or S1P (1 μM). (A) To confirm that MSK1 siRNA reduces protein levels of MSK1, cells were lysed and immunoblotted for MSK1, using α-tubulin as the loading control. To measure the effect of MSK1 knock-down on IL-8 inducibility and neutrophil chemotaxis, supernatants were removed and (B) IL-8 protein measured by ELISA and (C) neutrophil chemotaxis assessed using microchemotaxis chambers. Results are expressed as: (A) representative Western blots; (B) % S1P-induced IL-8 secretion in cells transfected with scrambled control; or (C) % S1P-induced neutrophil chemotaxis in the corresponding conditioned media (data are mean+SEM values from n = 6 primary ASM cell lines). Statistical analysis was performed using the Student's unpaired <i>t</i> test where * indicates that knocking down MSK-1 significantly attenuates S1P-induced effects (<i>P</i><0.05).</p

Time course of S1P-induced IL-8 mRNA expression and protein secretion and its repression by dexamethasone.

<p>Growth-arrested ASM cells were pretreated for 1 h with vehicle or 100 nM dexamethasone, followed by treatment with vehicle or S1P (1 μM) for 0, 1, 2, 4, 8, and 24 h. (A) IL-8 mRNA expression was quantified by real-time RT-PCR and results expressed as fold increase compared to vehicle-treated cells at 0 h. (B) IL-8 protein secretion was measured by ELISA and results expressed as % S1P-induced IL-8 secretion at 24 h. Statistical analysis was performed using two-way ANOVA then Bonferroni's post-test where * indicates significant repression by dexamethasone, compared to vehicle-treated cells at the same time point (<i>P</i><0.05). Data are mean+SEM values from n = 7 primary ASM cell lines.</p

IL-8 induced by S1P causes human neutrophil chemotaxis <i>in vitro</i> and this can be repressed by corticosteroids or by blocking the p38 MAPK- or ERK-mediated pathways.

<p>Chemotaxis of human neutrophils toward conditioned media from growth-arrested ASM cells pretreated with vehicle, dexamethasone (100 nM), SB203580 (1 μM) or PD98059 (10 μM), followed by treatment with vehicle or S1P (1 μM) for 24 h, was measured using microchemotaxis chambers. Results are expressed as cells per high-power (x200) field. Statistical analysis was performed using the Student's unpaired <i>t</i> test where § denotes a significant effect of S1P on neutrophil chemotaxis, while * denotes significant repression (<i>P</i><0.05). Data are mean+SEM values using conditioned media from n = 3 primary ASM cell lines.</p

S1P does not activate NF-κB in ASM cells.

<p>ASM cells transfected with a NF-κB reporter vector, pNF-κB-Luc, were growth-arrested, then treated with vehicle or TNFα (10 ng/ml), in the absence or presence of S1P (1 μM) for 6 h. Cells were then harvested and luciferase and β-galactosidase activities assessed. Data represent normalized luciferase activity, relative to vehicle-treated cells (expressed as fold difference). Statistical analysis was performed using the Student's unpaired <i>t</i> test (where § denotes significant effect of TNFα on NF-κB activity (<i>P</i><0.05)). Data are mean+SEM values from n = 6 primary ASM cell lines.</p

S1P-induced IL-8 protein secretion is repressed by inhibitors of the p38 MAPK- and ERK-mediated pathways.

<p>Growth-arrested ASM cells were pretreated for 30 min with vehicle, 1 μM SB203580, or 10 μM PD98059 to inhibit p38 MAPK and ERK, respectively. Cells were then stimulated with 1 μM S1P for 24 h. IL-8 protein was measured by ELISA and results expressed as % S1P-induced IL-8 secretion at 24 h. Statistical analysis was performed using the Student's unpaired <i>t</i> test, where * denotes significant inhibition (<i>P</i><0.05). Data are mean+SEM values from n = 10 primary ASM cell lines.</p

FBLN-1 from a soluble source is incorporated into the ECM.

<p>Cycloheximide treated and untreated human ASM cells were stimulated with or without 10 ng/ml TGF-β₁ in quiescing medium or soluble FBLN-1 containing medium for 72 hours (n = 7). The incorporation of FBLN-1 into the matrix was detected by ECM ELISA and data were expressed as absorbance at 450 nm–570 nm. Data were expressed as mean ± SEM and analysed by one-way ANOVA with Bonferroni’s multiple comparison test, *P<0.05, ***P<0.001. V: vehicle; CHX: cycloheximide; S: soluble FBLN-1 containing medium; T: TGF-β₁.</p

TGF-β₁ decreased soluble FBLN-1 from human ASM cells.

<p>Soluble FBLN-1 released from human ASM cells was detected by western blot, following 72 hours stimulation with 10 ng/ml TGF-β₁ (panel A). Data from COPD (black bar, n = 9) group and non-COPD (grey bar, n = 9) group were expressed as fold change relative to control (panel B). Data were expressed as mean ± SEM and analysed by two-way ANOVA with Bonferroni post tests, ***P<0.001, compared with control.</p

Characteristics of volunteers.

<p>T, transplant; R, resection; B, bronchoscopy; M, male; F, female; Y, yes; N, no; N/A, not available; FEV₁%, forced expiratory volume in 1 second of predicted %; FVC%, forced vital capacity of predicted %; Ca, carcinoma; SCCa, small cell carcinoma; NSCCa, non-small cell carcinoma; NSCLC: non-small cell lung carcinoma; IPF: idiopathic pulmonary fibrosis.</p