is a pathogen that causes a variety of infectious diseases such as pneumonia, endocarditis, and septic shock. Methicillin-resistant
(MRSA) evades virtually all available treatments, creating the need ...for an alternative control strategy. Although we previously demonstrated the inhibitory effect of sodium propionate (NaP) on MRSA, the regulatory mechanism of this effect remains unclear. In this study, we investigated the regulatory mechanism responsible for the inhibitory effect of NaP on MRSA using RNA-Seq analysis. Total RNAs were isolated from non-treated and 50 mM NaP-treated S. aureus USA300 for 3 h and transcriptional profiling was conducted by RNA-Seq analysis. A total of 171 differentially expressed genes (DEGs) with log
fold change ≥2 and
< 0.05 was identified in the NaP treatment group compared with the control group. Among the 171 genes, 131 were up-regulated and 40 were down-regulated. Upon gene ontology (GO) annotation analysis, total 26 specific GO terms in "Biological process," "Molecular function," and "Cellular component" were identified in MRSA treated with NaP for 3 h. "Purine metabolism"; "riboflavin metabolism"; and "glycine, serine, and threonine metabolism" were identified as major altered metabolic pathways among the eight significantly enriched KEGG pathways in MRSA treated with NaP. Furthermore, the MRSA strains deficient in
,
,
, or
, which were the up-regulated DEGs in the metabolic pathways, were more susceptible to NaP than wild-type MRSA. Collectively, these results demonstrate that NaP attenuates MRSA growth by altering its metabolic pathways, suggesting that NaP can be used as a potential bacteriostatic agent for prevention of MRSA infection.
E. faecalis lipoteichoic acid induces chemokine expression via TLR2/CD14/MyD88 and PAFR/JAK/STAT1 signaling pathways, without induction of IFN‐β in murine macrophages.
Enterococcus faecalis is one of ...the most common opportunistic pathogens responsible for nosocomial infections, and its LTA is known as an important virulence factor causing inflammatory responses. As chemokines play a key role in inflammatory diseases by triggering leukocyte infiltration into the infection site, we purified EfLTA and investigated its effect on the expression of chemokines, IP‐10, MIP‐1α, and MCP‐1, in murine macrophages. EfLTA induced the expression of these chemokines at the mRNA and protein levels. TLR2, CD14, and MyD88 were involved in the EfLTA‐induced chemokine expression, as the expression was reduced remarkably in macrophages derived from TLR2‐, CD14‐, or MyD88‐deficient mice. EfLTA induced phosphorylation of MAPKs and enhanced the DNA‐binding activity of NF‐κB, AP‐1, and NF‐IL6 transcription factors. The induction of IP‐10 required ERK, JNK, p38 MAPK, PKC, PTK, PI3K, and ROS. We noticed that all of these signaling molecules, except p38 MAPK and ROS, were indispensable for the induction of MCP‐1 and MIP‐1α. Interestingly, the EfLTA‐induced chemokine expression was mediated through PAFR/JAK/STAT1 signaling pathways without IFN‐β involvement, which is different from LPS‐induced chemokine expression requiring IFN‐β/JAK/STAT1 signaling pathways. Furthermore, the culture supernatant of EfLTA‐treated RAW 264.7 cells promoted the platelet aggregation, and exogenous PAF induced the chemokine expression in macrophages derived from WT and TLR2‐deficient mice. These results suggest that EfLTA induces the expression of chemokines via signaling pathways requiring TLR2 and PAFR, which is distinct from that of LPS‐induced chemokine expression.
Periodontitis is a chronic inflammatory disease of the gum caused by infection with multispecies oral bacteria. Since the periodontopathic bacteria, Porphyromonas gingivalis together with ...Enterococcus faecalis are frequently detected in patients with a severe form of periodontitis, interactions between their virulence factors might play an important role in progression of the disease. P. gingivalis and E. faecalis possess lipopolysaccharide (Pg.LPS) and lipoteichoic acid (Ef.LTA), respectively, as the major virulence factors inducing inflammatory responses. However, the combinatorial effect of these virulence factors on chemokine expression was poorly understood. Here, we examined the interaction between Ef.LTA and Pg.LPS on IL‐8 induction in human periodontal ligament (PDL) cells. Pg.LPS, but not Ef.LTA, induced IL‐8 expression at both mRNA and protein levels, which was suppressed in the presence of Ef.LTA. Although Ef.LTA and Pg.LPS could stimulate Toll‐like receptor 2 (TLR2), Ef.LTA did not interfere with Pg.LPS induced‐TLR2 activation. However, Ef.LTA decreased Pg.LPS‐induced phosphorylation of ERK, JNK, and p38 kinase. Furthermore, Ef.LTA suppressed Pg.LPS‐induced IL‐8 promoter activity as well as AP‐1, NF‐IL6 and NF‐κB transcription factors, which are indispensable for IL‐8 expression. Interestingly, Ef.LTA enhanced only IL‐1 receptor‐associated kinase‐M (IRAK‐M) expression among the tested negative regulators of TLR intracellular signaling cascades in the presence of Pg.LPS. In addition, silencing IRAK‐M restored the decreased IL‐8 expression by Ef.LTA in the presence of Pg.LPS. Collectively, these results suggest that Ef.LTA inhibits Pg.LPS‐induced IL‐8 expression in human PDL cells via inducing the expression of a negative regulator of TLR signaling cascades, IRAK‐M.
Streptococcus gordonii is a commensal Gram‐positive bacterium that acts as an opportunistic pathogen that can cause apical periodontitis, endocarditis, and pneumonia. Biofilm formation of bacteria is ...important for the initiation and progression of such diseases. Although lipoproteins play key roles in physiological functions, the role of lipoproteins of S. gordonii in its biofilm formation has not been clearly understood. In this study, we investigated the role of lipoproteins of S. gordonii in the bacterial biofilm formation using its lipoprotein‐deficient strain (Δlgt). The S. gordonii Δlgt exhibited increased biofilm formation on the human dentin slices or on the polystyrene surfaces compared to the wild‐type strain, while its growth rate did not differ from that of the wild‐type. In addition, the S. gordonii Δlgt strain exhibited the enhanced LuxS mRNA expression and AI‐2 production, which is known to be a positive regulator of biofilm formation, compared to the wild‐type. Concordantly, the augmented biofilm formation of S. gordonii Δlgt was attenuated by an AI‐2 inhibitor, D‐ribose. In addition, lipoproteins from purified S. gordonii inhibited the biofilm formation of S. gordonii wild‐type and Δlgt. Taken together, these results suggest that lipoprotein‐deficient S. gordonii form biofilms more effectively than the wild‐type strain, which might be related to the AI‐2 quorum‐sensing system.
, a Gram-positive bacterium, is a commensal bacterium that is commonly found in the skin, oral cavity, and intestine. It is also known as an opportunistic pathogen that can cause local or systemic ...diseases, such as apical periodontitis and infective endocarditis.
, an early colonizer, easily attaches to host tissues, including tooth surfaces and heart valves, forming biofilms.
penetrates into root canals and blood streams, subsequently interacting with various host immune and non-immune cells. The cell wall components of
which include lipoteichoic acids, lipoproteins, serine-rich repeat adhesins, peptidoglycans, and cell wall proteins, are recognizable by individual host receptors. They are involved in virulence and immunoregulatory processes causing host inflammatory responses. Therefore,
cell wall components act as virulence factors that often progressively develop diseases through overwhelming host responses. This review provides an overview of
and how its cell wall components could contribute to the pathogenesis and development of therapeutic strategies.
Osteoporosis and bone disorders related to the metabolic syndrome are often associated with adipokines secreted by adipocytes in bone. Adiponectin, a type of adipokine, is a regulator of immune ...responses and metabolic processes, but its role in bone biology remains uncertain. We investigated the role of adiponectin in bone metabolism using adiponectin-deficient mice
and
. Adiponectin-deficient mice exhibited reduced bone mass and increased adiposity. Adiponectin-deficient calvarial cells were prone to differentiate into adipocytes rather than osteoblasts. Although bone marrow macrophages (BMMs) from adiponectin-deficient mice had low osteoclastogenic potential as osteoclast precursors with increasing interferon regulatory factor 5 expression, under co-culture conditions of calvarial cells and BMMs, the enhanced receptor activator of nuclear factor κB ligand/osteoprotegerin (RANKL/OPG) ratio of adiponectin-deficient mesenchymal progenitor cells facilitated osteoclast differentiation. In addition, increased RANKL/OPG ratio was observed in the bone marrow extracellular fluid of adiponectin-deficient mice compared to that of wild-type mice. Notably, recombinant adiponectin treatment enhanced RANKL-induced osteoclast differentiation from BMMs but up-regulated OPG production in recombinant adiponectin-exposed calvarial cells, which inhibited osteoclast differentiation. Taken together, these results suggest that adiponectin plays an inhibitory role in bone metabolism through cross talk between precursor cells of both osteoclasts and osteoblasts by regulating RANKL/OPG ratio in the bone marrow microenvironment.
Initiation and progression of oral infectious diseases are associated with streptococcal species. Bacterial infection induces inflammatory responses together with reactive oxygen species (ROS), often ...causing cell death and tissue damage in the host. In the present study, we investigated the effects of oral streptococci on cytotoxicity and ROS production in human periodontal ligament (PDL) cells.
Streptococcus gordonii
showed cell cytotoxicity in a dose- and time-dependent manner. The cytotoxicity might be due to apoptosis since
S. gordonii
increased annexin V-positive cells, and the cytotoxicity was reduced by an apoptosis inhibitor, Z-VAD-FMK. Other oral streptococci such as
Streptococcus mitis
,
Streptococcus sanguinis
, and
Streptococcus sobrinus
also induced apoptosis, whereas
Streptococcus mutans
did not. All streptococci tested except
S. mutans
triggered ROS production in human PDL cells. Interestingly, however, streptococci-induced apoptosis appears to be ROS-independent, as the cell death induced by
S. gordonii
was not recovered by the ROS inhibitor, resveratrol or
n
-acetylcysteine. Instead, hydrogen peroxide (H
2
O
2
) appears to be important for the cytotoxic effects of streptococci since most oral streptococci except
S. mutans
generated H
2
O
2
, and the cytotoxicity was dramatically reduced by catalase. Furthermore, streptococcal lipoproteins are involved in cytotoxicity, as we observed that cytotoxicity induced by the lipoprotein-deficient
S. gordonii
mutant was less potent than that by the wild-type and was attenuated by anti-TLR2-neutralizing antibody. Indeed, lipoproteins purified from
S. gordonii
alone were sufficient to induce cytotoxicity. Notably,
S. gordonii
lipoproteins did not induce H
2
O
2
or ROS but cooperatively induced cell death when co-treated with H
2
O
2
. Taken together, these results suggest that most oral streptococci except
S. mutans
efficiently induce damage to human PDL cells by inducing apoptotic cell death with bacterial H
2
O
2
and lipoproteins, which might contribute to the progression of oral infectious diseases such as apical periodontitis.
Streptococcus gordonii is commonly found in the periapical endodontic lesions of patients with apical periodontitis, a condition characterized by inflammation and periapical bone loss. Since bone ...metabolism is controlled by osteoclastic bone resorption and osteoblastic bone formation, we investigated the effects of S. gordonii on the differentiation and function of osteoclasts and osteoblasts. For the determination of bone resorption activity in vivo, collagen sheets soaked with heat-killed S. gordonii were implanted on mouse calvaria, and the calvarial bones were scanned by micro-computed tomography. Mouse bone marrow-derived macrophages (BMMs) were stimulated with M-CSF and RANKL for 2 days and then differentiated into osteoclasts in the presence or absence of heat-killed S. gordonii. Tartrate-resistant acid phosphatase staining was performed to determine osteoclast differentiation. Primary osteoblast precursors were differentiated into osteoblasts with ascorbic acid and β-glycerophosphate in the presence or absence of heat-killed S. gordonii. Alkaline phosphatase staining and alizarin red S staining were conducted to determine osteoblast differentiation. Western blotting was performed to examine the expression of transcription factors including c-Fos, NFATc1, and Runx2. Heat-killed S. gordonii induced bone destruction in a mouse calvarial implantation model. The differentiation of RANKL-primed BMMs into osteoclasts was enhanced in the presence of heat-killed S. gordonii. Heat-killed S. gordonii increased the expression of c-Fos and NFATc1, which are essential transcription factors for osteoclast differentiation. On the other hand, heat-killed S. gordonii inhibited osteoblast differentiation and reduced the expression of Runx2, an essential transcription factor for osteoblast differentiation. S. gordonii exerts bone resorptive activity by increasing osteoclast differentiation and reducing osteoblast differentiation, which may be involved in periapical bone resorption.
•An endodontic pathogen S. gordonii induces bone resorption in a mouse calvarial implantation model.•S. gordonii increases osteoclast differentiation through up-regulation of c-Fos and NFATc1.•S. gordonii inhibits osteoblast differentiation by downregulating Runx2.
Runx2 is a key transcription factor regulating osteoblast differentiation and skeletal morphogenesis, and FGF2 is one of the most important regulators of skeletal development. The importance of the ...ERK mitogen-activated protein (MAP) kinase pathway in cranial suture development was demonstrated by the findings that the inhibition of FGF/FGF receptor (FGFR) signaling by a MEK blocker prevents the premature suture closure caused by an Fgfr2 mutation in mice. We previously demonstrated that ERK activation does not affect Runx2 gene expression but that it stimulates Runx2 transcriptional activity. However, the molecular mechanism underlying Runx2 activation by FGF/FGFR or ERK was still unclear. In this study, we found that FGF2 treatment increased the protein level of exogenously overexpressed Runx2 and that this increase is reversed by ERK inhibitors. In contrast, overexpression of constitutively active MEK strongly increased the Runx2 protein level, which paralleled an increase in Runx2 acetylation. As Runx2 protein phosphorylation mediated by ERK directly correlates with Runx2 protein stabilization, acetylation, and ubiquitination, we undertook to identify the ERK-dependent phosphorylation sites in Runx2. Analysis of two C-terminal Runx2 deletion constructs showed that the middle third of the protein is responsible for ERK-induced stabilization and activation. An in silico analysis of highly conserved ERK targets indicated that there are three relevant serine residues in this domain. Site-directed mutagenesis implicated Ser-301 in for ERK-mediated Runx2 stabilization and acetylation. In conclusion, the FGF2-induced ERK MAP kinase strongly increased the Runx2 protein level through an increase in acetylation and a decrease in ubiquitination, and these processes require the phosphorylation of Runx2 Ser-301 residue.
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