Peroxisome proliferator-activated receptor δ inhibits Porphyromonas gingivalis lipopolysaccharide-induced activation of matrix metalloproteinase-2 by downregulating NADPH oxidase 4 in human gingival fibroblasts.

We investigated the roles of peroxisome proliferator-activated receptor δ (PPARδ) in Porphyromonas gingivalis-derived lipopolysaccharide (Pg-LPS)-induced activation of matrix metalloproteinase 2 (MMP-2). In human gingival fibroblasts (HGFs), activation of PPARδ by GW501516, a specific ligand of PPARδ, inhibited Pg-LPS-induced activation of MMP-2 and generation of reactive oxygen species (ROS), which was associated with reduced expression of NADPH oxidase 4 (Nox4). These effects were significantly smaller in the presence of small interfering RNA targeting PPARδ or the specific PPARδ inhibitor GSK0660, indicating that PPARδ is involved in these events. In addition, modulation of Nox4 expression by small interfering RNA influenced the effect of PPARδ on MMP-2 activity, suggesting a mechanism in which Nox4-derived ROS modulates MMP-2 activity. Furthermore, c-Jun N-terminal kinase and p38, but not extracellular signal-regulated kinase, mediated PPARδ-dependent inhibition of MMP-2 activity in HGFs treated with Pg-LPS. Concomitantly, PPARδ-mediated inhibition of MMP-2 activity was associated with the restoration of types I and III collagen to levels approaching those in HGFs not treated with Pg-LPS. These results indicate that PPARδ-mediated downregulation of Nox4 modulates cellular redox status, which in turn plays a critical role in extracellular matrix homeostasis through ROS-dependent regulation of MMP-2 activity.

Ligand-activated peroxisome proliferator-activated receptor-δ and -γ inhibit lipopolysaccharide-primed release of high mobility group box 1 through upregulation of SIRT1.

Peroxisome proliferator-activated receptors (PPARs) inhibit lipopolysaccharide (LPS)-primed release of high mobility group box 1 (HMGB1), a late proinflammatory mediator, but the underlying molecular mechanism is not completely understood. In this study, we demonstrated that the inhibition of HMGB1 release by PPAR-δ and -γ is associated with the deacetylase activity of SIRT1. Ligand-activated PPAR-δ and -γ inhibited LPS-primed release of HMGB1, concomitant with elevation in SIRT1 expression and promoter activity. These effects were significantly reduced in the presence of small interfering (si)RNAs against PPAR, indicating that PPAR-δ and -γ are involved in both HMGB1 release and SIRT1 expression. In addition, modulation of SIRT1 expression and activity by siRNA or chemicals correspondingly influenced the effects of PPARs on HMGB1 release, suggesting a mechanism in which SIRT1 modulates HMGB1 release. Furthermore, we showed for the first time that HMGB1 acetylated in response to LPS or p300/CBP-associated factor (PCAF) is an effective substrate for SIRT1, and that deacetylation of HMGB1 is responsible for blockade of HMGB1 release in macrophages. Finally, acetylation of HMGB1 was elevated in mouse embryonic fibroblasts from SIRT1-knockout mice, whereas this increase was completely reversed by ectopic expression of SIRT1. These results indicate that PPAR-mediated upregulation of SIRT1 modulates the status of HMGB1 acetylation, which, in turn, has a critical role in the cellular response to inflammation through deacetylation-mediated regulation of HMGB1 release.

15-Deoxy-Δ(12,14)-prostaglandin J2 prevents oxidative injury by upregulating the expression of aldose reductase in vascular smooth muscle cells.

The omega-6 fatty acid derivative 15-Deoxy-Δ(12,14)-prostaglandin J2 (15d-PGJ2) is believed to play a role in cellular protection against oxidative stress in diverse cell systems. However, the cellular mechanisms by which protection is afforded by 15d-PGJ2 are not fully elucidated in vascular smooth muscle cells (VSMCs). In this study, we report the finding that 15d-PGJ2 elicited a time and concentration- dependent increase in aldose reductase (AR) expression. This induction was independent of the activation of peroxisome proliferator- activated receptor γ. Inhibition of phosphatidylinositol 3-kinase (PI3K) significantly suppressed the increase in expression and promoter activity of AR induced by 15d-PGJ2. Luciferase reporter assays demonstrated that 15d-PGJ2 targets the multiple stress response regions comprising the antioxidant response element in the promoter of the AR gene. 15d-PGJ2-mediated induction of AR promoter activity was potentiated in the presence of nuclear factor-erythroid 2-related factor 2 (Nrf2), but not in cells expressing dominant negative Nrf2. Cells treated with 15d-PGJ2 were resistant to oxidant-induced apoptotic cell death by inhibiting production of reactive oxygen species. These effects were significantly attenuated in the presence of an AR inhibitor or small interfering RNA against AR, indicating that AR plays a protective role against oxidative injury. Taken together, these findings demonstrate that activation of PI3K by 15d-PGJ2 increases the expression of AR through Nrf2, and increased AR activity may function as an important cellular response against oxidative injury.

Production of a monoclonal anti-myostatin antibody and the effects of in ovo administration of the antibody on posthatch broiler growth and muscle mass.

Myostatin, a member of the transforming growth factor-beta (TGF-beta) superfamily, is a potent negative regulator of skeletal muscle growth. The objective of this study was to produce a monoclonal anti-myostatin antibody and to examine the effects of in ovo administration of the antibody on posthatch broiler growth and muscle mass. The mature form of myostatin was expressed in Escherichia coli and used as an immunogen in producing a monoclonal antibody against myostatin. One hybridoma clone (mAb-c134) that showed the strongest affinity to the immunogen in Western blot analysis was used in producing a large quantity of monoclonal anti-myostatin antibody. In Western blot analysis, this antibody showed a strong binding affinity to commercially available mature myostatin and demonstrated a certain level of cross-reactivity with recombinant human BMP2 but not with recombinant human TGF-beta3 or porcine TGF-beta1. Competitive ELISA demonstrated binding of the antibody to the native form of mature myostatin in solution. To examine the effects of in ovo administration of the mAb-c134 antibody, eggs were injected once with 40 microg of mAb-c134 in 50 mL of PBS either into the albumen or yolk on d 3 of incubation. Controls received no injection. After hatching, chicks were raised for 35 d. Broilers from eggs that had the antibody injected into the yolk had significantly heavier body (4.2%) and muscle (5.5%) mass than the controls in both male and female birds. In contrast, no significant effects on body and muscle mass were observed when the mAb-c134 antibody was injected into the albumen. The results of this study suggest that immunoneutralization of myostatin during embryonic development is a potential means to improve growth potential of broilers.

Phorbol ester up-regulates aldose reductase expression in A549 cells: a potential role for aldose reductase in cell cycle modulation.

Over-expression of aldose reductase (AR) has been observed in many cancer cells. To clarify the role of AR in tumor cells, we investigated the pathways mediating expression of the AR gene induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), a potent tumor promoter. In A549 human lung adenocarcinoma cells, TPA elicited a dose- and time-dependent increase in AR mRNA level with an elevated enzyme activity. The TPA-induced increase in mRNA level and promoter activity of the AR gene was significantly attenuated in the presence of an inhibitor of protein kinase C, tyrosine kinase, or nuclear factor kappaB (NF-kappaB). TPA augmented the NF-kappaB-dependent gene transcription, indicating the involvement of NF-kappaB in this regulation. Accumulation of TPA-treated cells in S phase was almost completely abolished in the presence of ethyl 1-benzyl-3-hydroxy-2(5H)-oxopyrrole-4-carboxylate, an AR inhibitor. Taken together, TPA augmented the promoter activity of the AR gene via the activation of protein kinase and NF-kappaB. The inhibition of AR may assist in the chemotherapy of malignant tumors by suppressing the rapid growth of cancer cells.

In vitro and In vivo activities of LB 10827, a new oral cephalosporin, against respiratory pathogens.

The in vitro antibacterial activities of LB 10827, a new oral cephalosporin, against common respiratory tract pathogens were compared with those of six beta-lactams (cefdinir, cefuroxime, cefprozil, penicillin G, amoxicillin-clavulanate, and ampicillin), two quinolones (trovafloxacin and ciprofloxacin), and one macrolide (clarithromycin). The MIC of LB 10827 at which 90% of the penicillin-resistant strains of Streptococcus pneumoniae tested were inhibited was 0.5 microg/ml, and the drug was 4- to 32-fold more active than the compared beta-lactams. The potent activity of LB 10827 against Haemophilus influenzae and Moraxella catarrhalis was retained, and the presence of beta-lactamase in both strains had little effect on the in vitro activity of the compound. Time-kill studies revealed that LB 10827 had bactericidal activity against these respiratory pathogens. This agent reduced original counts of all pathogens tested by >/=3 log(10) CFU/ml at the MIC, and the regrowth was completely prevented for 12 h. The potent in vitro antibacterial activity of LB 10827 against respiratory pathogens has been proved in both mouse pneumonia and neutropenic rat models. These results strongly suggest that this agent has potential for the treatment of respiratory tract infections.

Synthesis and antibacterial activities of novel C(3)-aminopyrimidinyl substituted cephalosporins including against respiratory tract pathogens.

The variety of cephalosporins 1 and 2 which possessed C(3)-aminopyrimidinyl substituents were prepared and evaluated for their antibacterial activities. They exhibited excellent in vitro activities especially against respiratory tract pathogens such as penicillin resistant Streptococcus pneumonia. Moraxella catarrhalis and Haemophilus influenza.

In vitro and in vivo activities of LB10522, a new catecholic cephalosporin.

In vitro activity of LB10522 was compared with those of cefpirome, ceftazidime, ceftriaxone, and cefoperazone against clinical isolates. Against gram-positive bacteria, LB10522 was most active among the compounds tested. It was fourfold more active than cefpirome against methicillin-susceptible Staphylococcus aureus and Enterococcus faecalis. LB10522 was highly effective against most members of the family Enterobacteriaceae tested. Ninety percent of isolates of Escherichia coli, Klebsiella oxytoca, Proteus vulgaris, Proteus mirabilis, and Salmonella spp. were inhibited at a concentration of < or = 0.5 micrograms/ml. These activities were comparable to those of cefpirome. Against Pseudomonas aeruginosa, LB10522 with a MIC at which 90% of the isolates are inhibited of 2 micrograms/ml was 16- and 32-fold more active than ceftazidime and ceftazidime against systemic infections caused by Staphylococcus aureus giorgio, Streptococcus pneumoniae III, Pseudomonas aeruginosa 1912E, Escherichia coli 851E, Proteus mirabilis 1315E, Serratia marcescens 1826E, and Acinetobacter calcoaceticus Ac-54. LB10522 was very resistant to hydrolysis by various beta-lactamases such as TEM-3, TEM-7, SHV-1, FEC-1, and P-99. LB10522 did not induce beta-lactamase in Enterobacter cloacae 1194E, although most of the reference cephalosporins acted as inducers of beta-lactamase in this strain. Time-kill study showed that LB10522, at concentrations of two or four times the MIC, had a rapid bactericidal activity against Staphylococcus aureus 6538p, Escherichia coli 851E, and Pseudomonas aeruginosa 1912E.

In vitro and in vivo evaluations of LB20304, a new fluoronaphthyridone.

In vitro activity of LB20304 against 1,231 clinical isolates was evaluated and compared with those of ciprofloxacin, sparfloxacin, lomefloxacin, and ofloxacin. LB20304 demonstrated the most potent activity against gram-positive bacteria. It was 32- to 64-fold more active than ciprofloxacin against methicillin-susceptible Staphylococcus aureus, methicillin-resistant S. aureus, methicillin-resistant Staphylococcus epidermidis, and Streptococcus pneumoniae (penicillin G resistant). LB20304 was also highly active against most members of the family Enterobacteriaceae. Its activity was more potent than those of sparfloxacin, ofloxacin, and lomefloxacin and comparable to that of ciprofloxacin. The protective activities of LB20304 against systemic infections caused by gram-positive bacteria in mice were superior to those of ciprofloxacin and sparfloxacin. Against infections by gram-negative bacteria, LB20304 was slightly less active than ciprofloxacin.