Induction and stimulation of 92-kDa gelatinase/type IV collagenase production in osteosarcoma and fibrosarcoma cell lines by tumor necrosis factor alpha

Production of a 92-kDa gelatinase/type IV collagenase and tissue inhibitor of metalloproteinases (TIMP) was investigated with human sarcoma cell lines. Among the cytokines and growth factors examined, only human recombinant tumor necrosis factor alpha (TNF alpha) induced and stimulated the proteinase with concomitant increase in TIMP expression, but matrix metalloproteinase 2 (72-kDa gelatinase/type IV collagenase) expression was unchanged. These data suggest that gene expression of the two metalloproteinases is regulated in a different fashion and TNF alpha may be important to allow cancer cells to be more invasive and metastatic.     

Evaluation of chemical pretreatment for enzymatic solubilization of rice straw

Summary Rice straw was treated with NaOH, peracetic acid (PA), and sodium chlorite (NaClO₂). Quantitative changes in the composition of the treated straw, crystallinity of the treated straw and extracted cellulose, and susceptibility of the treated straw to Trichoderma reesei cellulase were studied. The alkali treatment resulted in a remarkable decrease in hemicellulose as well as lignin. Consequently, the recovery of residual straw after NaOH treatment was lowest among the three chemical reagents evaluated. The treatment with PA or NaCIO₂ resulted in a slight loss in hemicellulose and cellulose in the straw. The three chemical treatments caused little or no breakdown of the crystalline structure of cellulose in the straw. The treated straw was solubilized with the culture filtrate of T. reesei. The degree of enzymatic solubilization relative to the amount of residual straw was 69% after treatment with 0.25 N NaOH, 42% after treatment with 20% PA, and 50% after treatments with NaClO₂ (twice). The degree of enzymatic solubilization relative to the amount of the untreated straw, however, was 30% after treatment with 0.25 N NaOH, 32% after treatment with 20% PA, and 37% after treatments with NaClO₂ (twice).     

Initiation of DNA replication in Bacillus subtilis. IV. The effect of an intercalating dye, ethidium bromide

Summary The effects of an intercalating dye, ethidium bromide (EtBr), on the initiation of chromosome replication in Bacillus subtilis were studied. Spores of a thymine requiring mutant acquired the ability to initiate one round of replication in the absence of RNA and protein synthesis (initiation potential) during germination in a thymine starved medium. When EtBr was added after the initiation potential was fully established, initiation of replication was completely inhibited. This inhibition was reversible, and initiation was resumed when the drug was removed. The recovery of initiation occurred in the absence of protein synthesis but did require RNA synthesis and an active dna gene product.During germination both a DNA-protein complex and a DNA-membrane complex were formed at the replication origin in parallel with the establishment of initiation potential. EtBr destroyed both of these complexes at the concentration which inhibited initiation.The first round of replication of a plasmid DNA, pSL103, during spore germination was also prevented by EtBr. However a higher concentration was required to inhibit plasmid replication. It was found that the plasmid formed two complexes identical to the S- and M-complex of the chromosome origin. Compared to the chromosome complexes the plasmid complexes were less sensitive to EtBr. The loss of sensitivity was equivalent to that for the initiation of the plasmid compared to the chromosome. These results indicate that the target of EtBr is the DNA in the S- and M-complexes whose conformation is essential for the initiation of chromosome and plasmid replication.III of this series is Murakami et al. 1976     

Stimulatory effect of basic fibroblast growth factor on induction of heat shock protein 27 in osteoblasts: role of protein kinase C

In a previous study we showed that basic fibroblast growth factor (bFGF) stimulates activation of protein kinase C through phosphoinositide hydrolysis by phospholipase C and phosphatidylcholine hydrolysis by phospholipase D in osteoblast-like MC3T3-E1 cells. In the present study, we investigated whether bFGF stimulates the induction of heat shock protein (HSP) 27, a low-molecular-weight HSP, and HSP70, a high-molecular-weight HSP, in MC3T3-E1 cells and the mechanism behind the induction. bFGF increased the level of HSP27 while having little effect on HSP70 level. bFGF stimulated the accumulation of HSP27 dose-dependently in the range between 1 and 30 ng/ml. bFGF induced an increase in the level of the mRNA for HSP27. The bFGF-stimulated accumulation of HSP27 was reduced by inhibitors of protein kinase C. The bFGF-induced HSP27 accumulation was reduced in protein kinase C-downregulated MC3T3-E1 cells. U-73122, an inhibitor of phospholipase C, and propranolol, a phosphatidic acid phosphohydrolase inhibitor, suppressed the bFGF-stimulated HSP27 accumulation. These results strongly suggest that bFGF stimulates HSP27 induction through protein kinase C activation in osteoblasts.     

Endothelin-1 induces vascular endothelial growth factor synthesis in osteoblasts: involvement of p38 mitogen-activated protein kinase

We previously reported that endothelin-1 (ET-1) stimulates p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the effect of ET-1 on the synthesis of vascular endothelial growth factor (VEGF) in these cells. ET-1 significantly stimulated VEGF secretion time-dependently 18 hours after the stimulation. The stimulatory effect was dose-dependent in the range between 0.1 nM and 0.1 micro;M. BQ123, an antagonist of endothelin(A) (ET(A)) receptor, inhibited the ET-1-induced VEGF secretion. The ET-1-induced VEGF secretion was suppressed by SB203580 and PD169316, inhibitors of p38 MAP kinase, but not PD98059, an inhibitor of the upstream kinase that activates p44/p42 MAP kinase. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a protein kinase C (PKC)-activating phorbol ester, stimulated VEGF secretion. Calphostin C, a specific PKC inhibitor, suppressed the VEGF secretion by ET-1. TPA-induced VEGF secretion was suppressed by SB203580. Taken together, our results strongly suggest that ET-1 stimulates VEGF synthesis via ET(A) receptor in osteoblasts and that p38 MAP kinase is involved at a point downstream from PKC in the VEGF synthesis.     

Expression of Six Proteins Causes Reprogramming of Porcine Fibroblasts Into Induced Pluripotent Stem Cells With Both Active X Chromosomes

In this study, we created porcine‐induced pluripotent stem (iPS) cells with the expression of six reprogramming factors (Oct3/4, Klf4, Sox2, c‐Myc, Lin28, and Nanog). The resulting cells showed growth dependent on LIF (leukemia inhibitory factor) and expression of multiple stem cell markers. Furthermore, the iPS cells caused teratoma formation with three layers of differentiation and had both active X chromosomes (XaXa). Our iPS cells satisfied the both of important characteristics of stem cells: teratoma formation and activation of both X chromosomes. Injection of these iPS cells into morula stage embryos showed that these cells participate in the early stage of porcine embryogenesis. Furthermore, the RNA‐Seq analysis detected that expression levels of endogenous pluripotent related genes, NANOG, SOX2, ZFP42, OCT3/4, ESRRB, and ERAS were much higher in iPS with six factors than that with four reprogramming factors. We can conclude that the expression of six reprogramming factors enables the creation of porcine iPS cells, which is partially close to naive iPS state. J. Cell. Biochem. 118: 537–553, 2017. © 2016 Wiley Periodicals, Inc.     

Stromal Palladin Expression Is an Independent Prognostic Factor in Pancreatic Ductal Adenocarcinoma

It has been clear that cancer-associated fibroblasts (CAFs) in the tumor microenvironment play an important role in pancreatic ductal adenocarcinoma (PDAC) progression. However, how CAFs relate to the patients’ prognosis and the effects of chemoradiation therapy (CRT) has not been fully investigated. Tissue microarrays (TMAs) representing 167 resected PDACs without preoperative treatment were used for immunohistochemical studies (IHC) of palladin, α-smooth muscle actin (SMA), and podoplanin. Correlations between the expression levels of these markers and clinicopathological findings were analyzed statistically. Whole sections of surgical specimens from PDACs with and without preoperative CRT, designated as the chemotherapy-first group (CF, n = 19) and the surgery-first group (SF, n = 21), respectively, were also analyzed by IHC. In TMAs, the disease-specific survival rate (DSS) at 5 years for all 167 cases was 23.1%. Seventy cases (41.9%) were positive for palladin and had significantly lower DSS (p = 0.0430). α-SMA and podoplanin were positive in 167 cases (100%) and 131 cases (78.4%), respectively, and they were not significantly associated with DSS. On multivariable analysis, palladin expression was an independent poor prognostic factor (p = 0.0243, risk ratio 1.60). In the whole section study, palladin positivity was significantly lower (p = 0.0037) in the CF group (5/19) with a significantly better DSS (p = 0.0144) than in the SF group (16/22), suggesting that stromal palladin expression is a surrogate indicator of the treatment effect after chemoradiation therapy.