ABSTRACT
@#Abstract: Objective To screening new compounds that can inhibit the growth and biofilm formation of Staphylococcus aureus. Methods Compounds that can inhibit the growth of Staphylococcus aureus were screened from the FDA approved drug library by 96 well plates. The absorbance value of 600 nm wavelength (OD600) was measured by Microplate Reader to detect the growth of Staphylococcus aureus planktonic cells in the culture supernatant. The minimum inhibitory concentration (MIC) of ozanimod against Staphylococcus aureus clinical isolates were detected by micro broth dilution method. The inhibitory effect of sub-inhibitory concentrations of ozanimod on the biofilm formation of Staphylococcus aureus was detected by crystal violet staining. Results This study found that ozanimod could significantly inhibit the growth of Staphylococcus aureus SA113 (screening reference strain), and the MIC was 25.00 μmol/L. The MIC of ozanimod against 119 clinical isolates of Staphylococcus aureus [65 isolates of methicillin sensitive (MSSA) and 54 isolates of methicillin resistant (MRSA)] was 12.50 or 25.00 μmol/L. The MIC50 and MIC90 of ozanimod against the 119 Staphylococcus aureus isolates all were 25.00 μmol/L. This study found that 6.25, 12.50, 25.00 μmol/L of ozanimod could significantly inhibit the biofilm formation of 2 MSSA and 2 MRSA. The sub-MIC concentration of ozanimod (12.50 μmol/L) could significantly inhibit the biofilm formation of 14 MSSA and 11 MRSA, but had no inhibitory effect on the growth of planktonic cells of these Staphylococcus aureus isolates. Conclusion Ozanimod can inhibit the growth of Staphylococcus aureus, including MRSA, and has good antibacterial activity. The sub-MIC concentration of ozanimod could significantly inhibit the biofilm formation of Staphylococcus aureus.
ABSTRACT
Macrophages play pivotal roles in host defense and immune homeostasis, which have two major functional polarization states, the classically activated M1 and the alternatively activated M2. Interleukin (IL)-17A is an immune modulator able to shape macrophage phenotypes. Wnt/β-catenin is a developmental signaling pathway that plays crucial roles in morphogenesis and tissue homeostasis, which has also been recently demonstrated playing roles in immune regulation. A growing amount of evidence suggests that both Wnt and IL-17A signaling are involved in macrophage polarization. However, their interaction in macrophage polarization remains elusive. The aim of present study was to explore impacts of Wnt/β-catenin on IL-17A-mediated macrophage M1/M2 polarization in murine monocyte/macrophage-like cell line RAW264.7. Results revealed that IL-17A activated Wnt/β-catenin signaling and induced macrophage M1 polarization, but inhibited M2 polarization. In contrast, the activation of Wnt/β-catenin signaling led to the inhibition of M1 macrophage polarization but the promotion of M2 polarization. Importantly, the activation of Wnt/β-catenin also showed abilities to inhibit the IL-17A-induced M1 macrophage polarization while diminishing the IL-17A-inhibited M2 polarization. Molecular analysis further uncovered that the JAK/STAT signaling pathway was involved in the interaction of Wnt/β-catenin and IL-17A in the modulation of macrophage polarization. These results suggested that the Wnt/β-catenin signaling modulated IL-17A-altered macrophage polarization in part by regulating the JAK/STAT signaling pathway. This study thus revealed a novel function of Wnt/β-catenin signaling in regulating IL-17A-altered macrophage polarization.
Subject(s)
Animals , Rats , Interleukin-17 , beta Catenin , Wnt Signaling Pathway , Macrophage Activation , MacrophagesABSTRACT
The iron chelators can be utilized in target cells to improve 5-aminolaevulinic acid (ALA)-based photodynamic therapy (PDT). The purpose of this study is to compare the effect of two kinds of iron chelators,desferrioxamine (DFO) and ethylenediaminetetraacetic acid (EDTA) on the enhancement of ALA-PDT. HaCat cells were cultured in medium containing 2.0 mmol/L of ALA and 0.5 mmol/L of DFO or EDTA. After 3-h incubation in the dark,the concentration of cellular protoporphyrin IX (PpIX) was detected by high performance liquid chromatography (HPLC),and the fluorescence of PpIX was observed at 630 nm emission under confocal laser scanning microscope.For PDT,HaCat cells were irradiated using 632.8 nm laser,and the fractions of apoptotic and necrotic cells were flow cytometricaUy assayed. Related differences in morphology and ultrastructure of HaCat cells were observed using optical microscope or transmission electron microscope. Compared to incubation with ALA alone,the addition of DFO or EDTA increased the concentration of cellular PpIX and the fluorescent density of PpIX,and also increased cell death ratio after PDT. PDT using ALA plus DFO produced the highest cellular PpIX level,greatest cell death ratio and most severe structural damage to the cells. It was concluded that both DFO and EDTA could enhance ALA-based PpIX production and PDT. Compared to the non-specific iron chelator of EDTA,the specific chelator,DFO,showed more potential for the enhancement.
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This study examined the change of p 16INK4a and PNCA protein expression in myocardium after injection of hIGF-1 gene modified skeletal myoblasts into post-infarction rats. HIGF-1 gene modified skeletal myoblasts (hIGF-1-myoblasts) were injected into hind limb muscles of 18post-infraction rats (experimental group). Primary-myoblasts were injected into 18 post-infraction rats (control group) and 12 non-infarction rats (sham group). Expression of p16INK4a and PCNA protein in myocardiums were separately detected immunocytochemically 1, 2 and 4 weeks after the inuection. The level of hIGF-1 and rIGF-1 protein in serum and myocardium were detected by enzyme-linked immunosorbent assay (ELISA). Compared with the sham group, the percentage of p16INK4a and PCNA positive cells reached a peak after 1 week in the control group and the experimental group (P＜0.01). Moreover, the percentage of p16INK4a-positive cells in the experimental group was lower than in control group whereas the percentage of PCNA-positive cells was lower in the control group than in the experimental group (P＜0.01). The percentage of p16INK4a-positive cells in the experimental group and the percentage of PCNA-positive cells in the control group were close to that in the sham group from the 2nd week (P＞0.05). ELISA analysis disclosed that the myocardium level of rIGF-1 protein increased gradually in the controls and especially in the experimental group (P＜0.01). The serum level of rIGF-1 decreased significantly in post-infraction rats, but these conditions were improved in the experimental group (P＜0.01). The hIGF-1 protein in serum and myocardium were detected from the 1st week to the 4th week in the experimental group. Statistical analysis revealed significant associations of myocardium level of hIGF-1 protein with expression of p16INK4a and PCNA protein (r=-0.323, P＜0.05; r=0.647, P＜0.01). It is concluded that genetically hIGF-1-myoblast provides a means for constant synthesis and release of hIGF-1. It could not only improve the expression of rIGF-1 and PCNA protein in myocardium, but also suppress the expression of p16INK4a protein for 30 days in post-infraction rats. Myoblasts-mediated IGF-1 gene therapy may provide a new alternative for the clinical treatment of heart failure.
ABSTRACT
BACKGROUND & OBJECTIVES: Erythropoietin (EPO), originally identified for its critical hormonal role in promoting erythrocyte survival and differentiation, has shown to a protective effect in myocardial ischaemia-reperfusion (I-R) injury in animal model. However, the precise mechanisms remain unclear. The objective of this study was to determine the roles of nuclear factor-kappa B (NF-kB) and associated cytokines induced by I-R in the cardioprotection by recombinant human erythropoietin (rhEPO). Morphopathological observations were also made on the ultrastructure of myocardial tissue. METHODS: Myocardial I-R rat model was established by 30 min ligation of left descending coronary and 3 h reperfusion. RhEPO or saline solution was intraperitoneally injected 24 h before I-R insult. The infarct sizes were measured by triphenyltetrazolium chloride (TTC)-Evans blue technique and ultrastructural organizations were observed by a transmission electron microscope. Tumour necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and IL-10 concentrations were analyzed by enzyme-linked immunosorbance assays and NF-kB by electrophoretic mobility shift assay. TNF-alpha and IL-6 mRNA expression were studied by the reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: A single bolus injection of 5,000 units/kg of rhEPO 24 h before insult remarkably reduced infarct size and improved ultrastructural organization of I-R myocardium. It greatly suppressed TNF-alpha and IL-6 expression, but enhanced IL-10 production. It modestly activated NF-kB before I-R insult and markedly attenuated subsequent NF-kB activation during sustained I-R. INTERPRETATION & CONCLUSION: The suppression of proinflammatory cytokines expression may act by inhibiting NF-kB activation during I-R, but not by induction of IL-10. This might be one of the molecular mechanisms of rhEPO in cardioprotection. In addition, NF-kB was suggested to play a dual role in cardioprotective effects of rhEPO.