Cysteiniphilum litorale gen. nov., sp. nov., isolated from coastal seawater.

A taxonomic study was performed on strain SYSU D3-2T, isolated from coastal seawater near the estuary of Pearl River in southern China. The strain was observed to be Gram-reaction-negative, non-motile and non-spore-forming. Cells were found to be of coccobacilli shape. Chemotaxonomic analysis of the plasma membrane revealed ubiquinone-8 as the respiratory quinone, diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, an unidentified aminolipid, an unidentified aminophospholipid and an unidentified phospholipid as the polar lipids, and anteiso-C15 : 0, C18 : 0 and anteiso-C17 : 0 as the major fatty acids (>10 % of total fatty acids). Comparison of 16S rRNA gene sequences showed that strain SYSU D3-2T shared maximum similarities with Caedibacter taeniospiralis 51T (92.3 %) and Fangia hongkongensis UST040201-002T (90.6 %), while sharing 85.8-90.0 % similarity with species of the genera Allofrancisella and Francisella. Phylogenetic dendrograms based on the 16S rRNA gene sequences showed that the strain clustered within the family Francisellaceae, but formed a separate lineage closely linked to Caedibactertaeniospiralis 51T and F. hongkongensis UST040201-002T. Based on the findings of the polyphasic taxonomic study, strain SYSU D3-2T is proposed to be recognized as a representative of a novel species of a new genus within the order Thiotrichales, with the name Cysteiniphilum litorale gen. nov., sp. nov. The type strain of the type species is SYSU D3-2T (=NBRC 112441T=DSM 101832T=KCTC 52386T=CGMCC 1.15758T).

[Characteristics and expression of Mip5, a novel gene associated with myocardial ischemia/reperfusion in rats].

OBJECTIVE: To determine the characteristics of a novel gene Mip5 (GenBank accession number AY553870) and its expression under physiological and pathological conditions. METHODS: The characteristics of Mip5 were analyzed by bioinformatic programs including BLAST, spidey, psort, ClustalW and so on. RT-PCR was performed to detect Mip5 expression. RESULTS: Bioinformatic analysis showed that Mip5 gene lied in the 13th chromosome and contained 8 exons and 7 introns, its open reading frame contained 909 bp and its protein production was 302 amino acid residues including 6 kelth domains. Under normal conditions, MIP5 expressed abundantly in the heart, brain and kidney, but its expression could not be detected in the liver and muscle. Expression of Mip5 gene was increased significantly after ischemia-reperfusion compared with the sham groups, and reached its peak at 3 h and recovered at 12 h after the reperfusion. Conclusion Mip5 gene is a novel gene containing a putative open reading frame of 302 amino acids residues and may play an important role in rat cardiomyocytes suffering ischemia processing.

[Effect of nucleolin down-regulation on the proliferation and apoptosis in C2C12 cells].

OBJECTIVE: To clarify the effect of nucleolin on the proliferation and apoptosis in C2C12 cells. METHODS: After inhibiting the expression of nucleolin using antisense oligonucleotides, the cellular proliferation was determined by MTT, and the apoptosis was detected by flow cytometry (FCM) assays and DNA ladder assays. RESULTS: After being transfected with antisense oligonucleotides for 24 hours, Western blotting showed that the expression of nucleolin was repressed significantly. In cells treated with antisense oligonucleotides, the cellular proliferation was obviously inhibited; the apoptotic cell increased significantly; and the "DNA ladder" was clearly observed. But the sense and random oligonucleotides had no effect on the cellular proliferation and apoptosis. CONCLUSION: The down-regulation of nucleolin can inhibit the cellular proliferation and initiate the apoptosis in C2C12cells.

[Effect of heat shock response on the cleavage of nucleolin induced by oxidative stress].

OBJECTIVE: To observe the cleavage of nucleolin (C23) during apoptosis induced by oxidative stress and to clarify the effect of heat shock response (HSR) on the cleavage of nucleolin and its possible molecular mechanism. METHODS: We added 0.5 mmol/L peroxide hydrogen (H2O2 ) into cultured cells to mimic oxidative stress. Apoptosis and cleavage of C23 were detected using caspase-3 colorimetric assay and Western blotting respectively. HSR was performed to observe the effect of HSR on cleavage of C23 induced by oxidative stress, and over-expressions of HSP70 and HSP25 were detected by Western blotting. RESULTS: Activity of caspase-3 increased significantly after 2 hours of 0.5 mmol/L H2O2 treatment, and reached the peak after 12 hours. The cleavage of C23 appeared 30 minutes to 1 hour after the treatment of H2O2 as indicated by a cleaved fragmentation of 80 kD, which was significantly inhibited by HSR. Moreover, HSR could induce HSP70 and HSP25 over-expressions. CONCLUSION: Oxidative stress can induce the activation of caspase-3, cleavage of C23, and apoptosis. HSR can significantly inhibit the cleavage of C23 induced by oxidative stress, which is related to the over-expressions of HSP70, HSP25, and other stress proteins.

Immortalization of embryonic fibroblasts in heat shock transcription factor 1 knockout mouse / 中南大学学报(医学版)

OBJECTIVE@#To establish immortalized embryonic fibroblast lines in heat shock transcription factor 1 (HSF1) HSF1-/- and HSF1+/+ mice and to provide experimental models to study the function of HSF1.@*METHODS@#A mammalian expression vector (pSV3neo) containing the SV40 large T antigen was used to transfect the HSF1-/- and HSF1+/+ mouse embryonic fibroblast using Lipofectamine 2000. Colonies were screened by G418 and expanded to immortalized cell lines. PCR was used to detect the integration of the large T antigen with genome in the mouse embryonic fibroblast. Expression of SV40 large T antigen gene in expanded cells was identified by RT-PCR. HSP70 expression was examined by Western blot in the embryonic fibroblast lines.@*RESULTS@#The stable growth and serial propagation were observed in the HSF1-/- and HSF1+/+ cell lines for six months. The mRNA of SV40 T antigen gene expressed in the two cell lines. HSP70 expression could not be induced in the heat-treated HSF1-/- mouse embryo fibroblasts.@*CONCLUSION@#The immortalized cells of HSF1+/+ and HSF1-/- mouse embryo fibroblasts are successfully established.