Over-expression of the pathway specific positive regulator gene is an effective way to activate silent gene cluster. In the curret study, the SARP family regulatory gene, vasR2, was over-expressed in strain Verrucosispora sp. NS0172 and the cryptic gene cluster responsible for the biosynthesis of pentaketide ansamycin was partially activated. Two tetraketides (1 and 2) and a triketide (3) ansamycins, together with five known compounds (4-8), were isolated and elucidated from strain NS0172OEvasR2. Their NMR data were completely assigned by analysis of their HR-ESI-MS and
Micromonosporaceae/metabolism* ; Multigene Family ; Polyketides/metabolism* ; Rifabutin/metabolism*

OBJECTIVE: To investigate the role of focal adhesion kinase (FAK) in extracellular signal-regulated kinase (ERK) signaling pathway mediated invadsion of trophoblasts. METHODS: We established a human extravillous cytotrophoblasts in vitro invasion model. Different concentrations of herbimycin A(FAK inhibitor)and PD98059 (ERK inhibitor) were given to observe the influence on the growth of trophoblast cells, FAK, ERK phosphorylation, and trophoblast invasion abilities. RESULTS: The expression of phosphorylated FAK in the extravillous cytotrophoblasts (EVCT) was inhibited by herbimycin A in a concentration-dependent manner and expression of phosphorylated ERK1/2 was also partially reduced. PD98059 had no effect on the expression of phosphorylated FAK. Herbimycin A and PD98059 suppressed the in vitro invasion of EVCT to various degrees. CONCLUSION ERK signaling pathway may be the common pathway for many invasive signals,and play a key role in the regulation of trophoblast invasion.
Benzoquinones ; pharmacology ; Cell Division ; physiology ; Cell Movement ; physiology ; Extracellular Signal-Regulated MAP Kinases ; antagonists & inhibitors ; metabolism ; Flavonoids ; pharmacology ; Focal Adhesion Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Humans ; Lactams, Macrocyclic ; pharmacology ; Phosphorylation ; Rifabutin ; analogs & derivatives ; Signal Transduction ; physiology ; Trophoblasts ; cytology ; physiology

OBJECTIVETo investigate the influence of various signal transduction modulators on the splenic T lymphocytes secretion of IL-2 and IL-10 in severely scalded mice, and to explore its mechanism.

METHODSThe mice were inflicted with 18% TBSA full-thickness scald by high-pressure heat vapour, and T lymphocytes were isolated from murine splenocytes through nylon wool column at 12 and 96 post-scald hours (PSH). Then the cells were divided into following groups: i. e. control, scald, scald and modulator [1 ml of 50 micromol/L PKC inhibitor ( H-7) , 30 micromol/L tetradecanoylphorbol-13-acetate (TPA) , 10micromol/L nonreceptor tyrosine protein kinase inhibitor (herbimycin) , 25 microg/ml of mitogen activated protein kinase kinase inhibitor (PD098059) , 100 nmol/L Calcium ionophore ( A23187) were added to the cells, respectively] groups. The scald group was subdivided into S1 (with scald at 12 PSH) and S2 (with scald at 96 PSH) groups. The modulator group was subdivided into modulator, S1 and modulator( the modulators were added into cells at 12 PSH) , and S2 and modulator( the modulators were added to cells at 96 PSH) groups. The influence of modulators to T lymphocyte secretion of IL-2 and IL-10 were observed.

RESULTSAfter the addition of H-7, the IL-2 and IL-10 levels in each group were obviously lower than that in controls( P <0. 05 or 0.01) , and that in S1 and H7 group, S2 and H7 group were obviously lower than that in scald group at corresponding time-points( P <0.01). The levels of IL-10, and especially IL-2 were elevated by TPA, but they were markedly lower than that in control group after PD098059 pretreatment. The secretion of IL-2 and IL-10 was significantly suppressed by herbimycin in S1 and herbimycin, and S2 and herbimycin groups, but those in Sl and A21387[ (2 417+/-39) pg/ml, (2 793+/-25)pg/ml] , S2 and A21387 [ (921+/-50) pg/ml, (2 633+/-35)pg/ml] groups were evidently higher than those in S1[ (1 542+/-40)pg/ml, (2 390+/-15)pg/ml] , S2 [(328+/-19)pg/ml, (1 618+/-21)pg/ml,( P <0.05 or <0.01)]groups.

CONCLUSIONPKC, calcium, MAPKK and TPK play critical roles in the dysfunction of splenic T lymphocyte secretion of IL-2 and IL-10 in severely scalded mice, among which TPK and PKC are mainly targeted to IL-2 secretion, and MAPKK is targeted to IL-10 secretion. TPA and A23187 can markedly rectify the disturbance of IL-2/IL-10 secretion ratio by increasing the IL-2 secretion after scald.

Animals ; Benzoquinones ; pharmacology ; Burns ; metabolism ; Calcimycin ; pharmacology ; Calcium ; metabolism ; Cells, Cultured ; Female ; Flavonoids ; pharmacology ; Interleukin-10 ; metabolism ; Interleukin-2 ; metabolism ; Lactams, Macrocyclic ; pharmacology ; Lymphocyte Activation ; Male ; Mice ; Mice, Inbred Strains ; Mitogen-Activated Protein Kinase Kinases ; antagonists & inhibitors ; Protein Kinase C ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Rifabutin ; analogs & derivatives ; Signal Transduction ; Spleen ; cytology ; T-Lymphocytes ; drug effects ; metabolism ; Tetradecanoylphorbol Acetate ; pharmacology

OBJECTIVETo investigate the role of adhesion between fibronectin and fibroblasts in wound healing as well as tyrosine phosphorylation proteins in procollagen mRNA expression.

METHODSThe level of proalpha1 (I) mRNA and tyrosine phosphorylation protein were detected employing the techniques of RT-PCR and immunoblotting. After inhibition of tyrosine kinases, herbimycin A was added to the medium to block the pathway of tyrosine phosphorylation, the changes of procollagen mRNA and tyrosine phosphorylation proteins were further investigated.

RESULTSThe adhesion between fibroblasts and fibronectin in wound healing not only induced the production of 98kd and 65kd tyrosine phosphorylation protein, but also enhanced obviously the expression of procollagen alpha1 (I) mRNA. When the pathway of tyrosine phosphorylation was blocked, the level of procollagen alpha1 (I) mRNA lowered remarkably, accompanied by the decrease of 98kd, 65kd tyrosine phosphorylation proteins.

CONCLUSIONThe adhesion between fibronectin and fibroblasts plays an important role in expression increase of procollagen mRNA during wound healing, in the process of which tyrosine phosphorylation is a key step.

Adolescent ; Adult ; Benzoquinones ; Blotting, Western ; Child ; Female ; Fibroblasts ; metabolism ; Gene Expression ; Humans ; Lactams, Macrocyclic ; Male ; Phosphoproteins ; metabolism ; physiology ; Phosphorylation ; drug effects ; Procollagen ; genetics ; Quinones ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Rifabutin ; analogs & derivatives ; Skin ; drug effects ; metabolism ; pathology ; Tyrosine ; metabolism ; Wound Healing ; genetics ; physiology

OBJECTIVETo explore correlation of seven apoptosis-related proteins (Hsp90a, p53, MDM2, Bcl-2, Bax, Cytochrome C, and Cleaved caspase3) with clinical outcomes of ALK+ anaplastic large-cell lymphoma (ALCL).

METHODSUsing immunohistochemistry and immunofluorescence double staining methods, the expressions of these seven apoptosis-associated proteins were studied to clarify their relationship with clinical outcomes of 36 ALK+ and 25 ALK-systemic ALCL patients enrolled between 1996 and 2006. The relationship of these apoptosis-regulating proteins with NPM-ALK status was also evaluated with the tyrosine inhibitor herbimycin A (HA) in vitro by immunocytochemistry, Western blotting and flow cytometric assays.

RESULTSThe presence of Hsp90α-, MDM2-, Bax-, Cytochrome C, and Cleaved caspase3-positive tumor cells was found significantly different in ALK+ and ALK-ALCLs, which was correlated with highly favorable clinical outcome. The Bcl-2- and p53-positive tumor cells were found in groups of patients with unfavorable prognosis. Inhibition of NPM-ALK by HA could reactivate the p53 protein and subsequent apoptosis-related proteins and therefore induced apoptosis in ALK+ ALCL cells.

CONCLUSIONOur results suggest that these seven proteins might be involved in apoptosis regulation and associated with clinical outcome of ALK+ systemic ALCLs. We also reveal a dynamic chain relation that NPM-ALK regulates p53 expression and subsequent apoptosis cascade in ALK+ ALCLs.

Adolescent ; Adult ; Aged ; Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Benzoquinones ; pharmacology ; Biomarkers, Tumor ; metabolism ; Blotting, Western ; Cell Culture Techniques ; Cell Survival ; drug effects ; Child ; Child, Preschool ; Disease-Free Survival ; Enzyme Inhibitors ; pharmacology ; Female ; Flow Cytometry ; Humans ; Immunohistochemistry ; Kaplan-Meier Estimate ; Lactams, Macrocyclic ; pharmacology ; Lymphoma, Large-Cell, Anaplastic ; enzymology ; metabolism ; pathology ; Male ; Microscopy, Fluorescence ; Middle Aged ; Neoplasm Staging ; Prognosis ; Protein-Tyrosine Kinases ; metabolism ; Receptor Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Retrospective Studies ; Rifabutin ; analogs & derivatives ; Young Adult