Objective To explore the pattern and quantify the heat shock protein HSP)70 and HSP70 mRNA in Vero-E6 cells after infection with Hantann virus(HTNV).Methods The expres- sion of HSP70 and change of its mRNA level were detected by immunocytochemical staining,nucleic acid hybridization in situ and RT-PCR.Results In situ hybridization and RT-PCR were used to eval- uate the level of HSP70 mRNA during Hantaan 76-118 infection.HSP70 mRNA increased 0.5 h after infection,reached its peak by 12 h and gradually declined to steady state level by 72 h(vs.sham infec- ted group,P＜0.05).The expression of HSP70 protein induced by Hantaan 76-118 infection was e- valuated by quantitative immunocytochemical staining.HSP70 increased 0.5 h after infection,reached its peak by 12 h and decreased at 72 h after infection(vs.sham infected group,P＜0.05).Conclu- sions HSP70 can be induced directly by HTNV infection at both mRNA and protein levels,It pro- vides a basis for the further study of the pathogenesis,prevention and treatment of hemorrhagic fever with renal syndrome(HFRS).

Objective To investigate symptom characteristics and their their prevalence in terminally ill patients with ovarian carcinoma.Methods A retrospective study was carried out based on clinical data of 98 terminally ill patients with ovarian carcinoma who died in our hospital during January 1995 to December 2004.Fifteen most common symptoms were analyzed with a focus on the followings:symptom incidence,survival time after symptom occurrence,regularity of symptom cluster,and common causes of death.Fifteen symptoms were:pain,cachexia,pleural effusion and ascites,dyspnea,fever,intestinal obstruction,renal failure,bone marrow depression,lung infection,hemorrhage,deep venous thrombosis (DVT),intestinal or pancreatic fistula,mycotic infection,jaundice and emergency conditions.Results (1)The most prevalent symptom was pleural effusion and ascites(63%),followed by pain(60%), cachexia(59%),dyspnea(52%)and intestinal obstruction(49 %).(2)The symptom which lasted longest survival time was mycotic infection(77 days),followed by intestinal or pancreatic fistula(75 days), intestinal obstruction(67 days),pain(60 days)and eachexia(60 days).Symptoms such as bone marrow depression,renal failure,dyspnea and emergency conditions were comparatively critical associated with shorter survival times(14,13,12,7 days,respectively).(3)Terminal symptoms occurred typically in clusters,with 4.9?1.5 symptoms per case.Of 98 cases,84 cases(86%)had 4 or more symptoms,with the median survival time of 63 days from the last day of anti-cancer therapy,and a slow death process.The remaining 14 cases(14%)with 3 or fewer symptoms survived only 25 days,of which 10 cases(71%)died of emergency diseases.The survival time for two groups was significantly different(P

OBJECTIVETo evaluate the effects of paroxetine on protein kinase PKA, PKC and CaMKII activities in different brain regions in a rat model of depression.

METHODSThirty-six adult male SD rats were randomized into 6 groups, including one control group (I) and 5 groups of depression model established by forcing the rats to swim for 4 weeks. The 5 depression groups received no treatment (II) or were treated with paroxetine at a single dose (III), for a week (IV), 2 weeks (V) or 4 weeks (VI). The radioactivity of PKA, PKC and CaMKII in the hippocampus and prefrontal cortex was quantitatively measured using a liquid scintillation counter.

RESULTSIn the rat hippocampus, PKA and CaMKII activities were significantly lower in groups II, III, IV, and V than in groups I and VI (P<0.01 or P<0.05), but comparable between groups VI and I (P>0.05). PKC activity was significantly lower in group II than in group I (P<0.01), but showed no significant difference between the paroxetine-treated groups and group I (P>0.05). In the prefrontal cortex, the activity of PKA in groups I, II, III, and IV was similar (P>0.05), but all significantly lower than that in groups V and VI (P<0.01). PKC activity was significantly higher in groups II and III than that in group I and other paroxetine-treated groups (P<0.01), and similar between groups IV and I (P>0.05); groups V and VI had significantly lower PKC activity than group I (P<0.01). Group I had the highest CaMKII activity among the groups (P<0.01).

CONCLUSIONChronic administration of paroxetine can reverse chronic stress-induced inhibition of PKA, PKC and CaMKII activity in rat hippocampus, while the effects of paroxetine on the protein kinases can be more complex in prefrontal cortex.

Animals ; Brain ; drug effects ; enzymology ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; metabolism ; Cyclic AMP-Dependent Protein Kinases ; metabolism ; Depression ; enzymology ; Disease Models, Animal ; Hippocampus ; drug effects ; enzymology ; Male ; Paroxetine ; pharmacology ; Protein Kinase C ; metabolism ; Random Allocation ; Rats

Idiopathic pulmonary fibrosis (IPF) is characterized by myofibroblast foci in lung parenchyma.Myofibroblasts are thought to originate from epithelial-to-mesenchymal transition (EMT).Wnt1 and lithium chloride (LiCl) induce EMT in alveolar epithelial cells (AECs),but the mechanisms are unclear.AECs were treated with Wnt1 and LiCl,respectively;morphological change and molecular changes of EMT,including E-cadherin,fibronectin,and vimentin,were observed.SB203580 was administrated to test the role of p38 MAPK signaling in EMT.Then AECs were treated with siRNAs targeting p38 MAPK to further test the effects of p38 MAPK,and the role was further confirmed by re-expression of p38 MAPK.At last β-catenin siRNA was used to test the role of β-catenin in the EMT process and relationship of β-catenin and p38 MAPK was concluded.Exposure of AECs to Wnt1 and LiCl resulted in upregulation of vimentin and fibronectin with subsequent downregulation of E-cadherin.Wnt1 and LiCl stimulated the p38 MAPK signaling pathways.Perturbing the p38 MAPK pathway either by SB203580 or through p38 MAPK siRNA blocked EMT and inhibited fibronetin synthesis,which were reversed by transfection of p38 MAPK expression plasmid.β-catenin siRNA attenuated the EMT process and decreased p38 MAPK phosphorylation,indicating that β-catenin is involved in the EMT-related changes through regulation of p38 MAPK phosphorylation.These findings suggest that p38 MAPK participates in the pathogenesis of EMT through Wnt pathway and that p38 MAPK may be a novel target for IPF therapy.

To study the effect of Siwu decoction on the function and expression of P-glycoprotein (P-gp) in Caco-2 cells. The Real-time quantitative poly-merase chain reaction (Q-PCR) was used to analyze the mRNA expression of MDR1 gene in Caco-2 cells. Flow cytometer was used to study the effect of Siwu decoction on the uptake of Rhodamine 123 in Caco-2 cells, in order to evaluate the efflux function of P-gp. Western blotting method was used to detect the effect of Siwu decoction on the P-gp protein expression of Caco-2 cells. Compared with the blank control group, after Caco-2 incubation with Siwu decoction at concentrations of 3.3, 5.0, 10.0 g x L(-1) for 24, 48, 72 h, the mRNA expression of MDR1 was up-regulated, suggesting the effect of Siwu decoction in inducing the expression of MDR1. After the administration with Siwu decoction in Caco-2 cells for 48 h, the uptake of Rhodamine 123 in Caco-2 cells decreased by respectively 16.6%, 22.1% (P < 0.05) and 45.4% (P < 0.01), indicating that the long-term administration of Siwu decoction can enhance the P-gp efflux function of Caco-2 cells. After the incubation of Caco-2 cells with Siwu decoction for 48 h, the P-gp protein expression on Caco-2 cell emebranes, demonstrating the effect of Siwu decoction in inducing the protein expression of P-gp.
ATP Binding Cassette Transporter, Sub-Family B ; genetics ; metabolism ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Caco-2 Cells ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Up-Regulation ; drug effects

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Female ; Glasgow Coma Scale ; Hematoma ; diagnosis ; surgery ; Humans ; Infant ; Intracranial Hemorrhage, Traumatic ; diagnosis ; surgery ; Male ; Middle Aged ; Prognosis

Spiramycin (SP) belongs to the 16-member macrolide antibiotics. It contains three components,namely SP I, SP II and SP III, which differ structurally in the acylation moieties on the C3 of the lactone. The SP I component contains a hydroxyl group at C3. SP II, and SP III are formed by further acetylation or propionylation of the C3 of SP I, by the same 3-O-acyltransferase (3-O-AT) . The study focused on simplifying spiramycin components. Theoretically, disruption/deletion of the 3-O-AT gene will reduce/stop the acylation of SP I to SP II and SP III. In this study, degenerated primers were designed according to the conserved regions of 3-O-acyltransferase, MdmB and AcyA in the medicamycin and carbomycin producers of S. mycarofaciens and S. thermotolerans, respectively, and an 878bp DNA fragment was amplified from the spiramycin-producer of S. spiramyceticus F21. Blast analysis of the 878bp DNA fragment suggested that it encoded the 3-O-acyltransferase (3-0-AT, sspA) gene for spiramycin biosynthesis. The flanking regions of this 878bp DNA fragment were then amplified by single-oligonucleotide-nested PCR, and a total of 4.3 kb DNA was obtained (3457nt among the 4.3kb fragment was sequenced, and deposited in GenBank DQ642742),covering the whole putative 3-O-acyltransferase gene, sspA. The sspA was then deleted from the S. spiramyceticus F21 genome by double cross-over homologous recombination, mediated by temperature-sensitive plasmid pKC1139. A comparison was done of the components of spiramycins produced by the sspA-deleted mutant strain with that of the parent strain by HPLC analysis, which showed that sspA-deleted mutant produced SP I (72%), SP II (18%), and SP III (9.6%), whereas parent strain produced SP I (7.8%), SP II (67%), and SP III (25%), respectively, demonstrating the role of ssp A in the acylation of SP I into SP II and SP III. The ssp A-deleted mutant strain obtained in this study may be used for the production of SP I, or may serve as a good starter for the construction of spiramycin derivatives.
Acyltransferases ; genetics ; Aminoglycosides ; biosynthesis ; Gene Deletion ; Genes, Bacterial ; genetics ; Genetic Engineering ; methods ; Streptomyces ; enzymology ; genetics

AIMTo investigate human epidermal growth factor receptor type 2 (HER2) protein expression and gene amplification in Chinese metastatic prostate cancer patients and their potential value as prognostic factors.

METHODSImmunohistochemistry (IHC) was performed to investigate HER2 protein expression in prostate biopsy specimens from 104 Chinese metastatic prostate cancer patients. After 3-11 months of hormonal therapy, 12 patients underwent transurethral resection of the prostate (TURP). HER2 protein expression of TURP specimens was compared with that of the original biopsy specimens. Of these, 10 biopsy and 4 TURP specimens with HER2 IHC staining scores >or=2+ were investigated for HER2 gene amplification status by fluorescent in situ hybridization (FISH).

RESULTSOf the 104 prostate biopsy specimens, HER2 protein expression was 0, 1+, 2+ and 3+ in 49 (47.1%), 45 (43.3%), 8 (7.7%) and 2 (1.9%) cases, respectively. There was a significant association between HER2 expression and Gleason score (P = 0.026). HER2 protein expression of prostate cancer tissues increased in 33.3% of patients after hormonal therapy. None of the 14 specimens with HER2 IHC scores >or= 2+ showed HER2 gene amplification. Patients with HER2 scores >or= 2+ had a significantly higher chance of dying from prostate cancer than those with HER2 scores of 0 (P = 0.004) and 1+ (P = 0.034). Multivariate Cox regression analysis showed that HER2 protein expression intensity was an independent predictor of cancer-related death (P = 0.039).

CONCLUSIONAn HER2 IHC score >or= 2+ should be defined as HER2 protein overexpression in prostate cancer. Overexpression of HER2 protein in cancer tissue might suggest an increased risk of dying from prostate cancer. HER2 protein expression increases in some individual patients after hormonal therapy.

Aged ; Antineoplastic Agents, Hormonal ; therapeutic use ; Asian Continental Ancestry Group ; genetics ; statistics & numerical data ; Biopsy ; China ; epidemiology ; Gene Expression Regulation, Neoplastic ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Kaplan-Meier Estimate ; Male ; Middle Aged ; Prognosis ; Proportional Hazards Models ; Prostatic Neoplasms ; drug therapy ; genetics ; mortality ; secondary ; Receptor, ErbB-2 ; genetics ; metabolism ; Risk Factors

OBJECTIVETo study the effects of Siwu decoction on protein expression of blood deficiency mice induced by cyclophosphamide (CIX) and discuss the possible molecular mechanism on blood enriching function of Siwu decoction.

METHODBlood deficiency mice were established by injecting ip with 250 mg x kg(-1) CTX. Proteomic technologies were applied to identify the different protein.

RESULTSiwu decoction could restore the changes of 12 up-regulated and 3 down-regulated proteins in bone marrow of blood deficiency mice induced by cyclosphosphamide.

CONCLUSIONSiwu decoction could effect expression of proteins which functions including apoptosis, proliferation and differentiation of the haematopoietic stem/progenitor cell. The regulation in the molecular level might be the mechanism of stimulating hematopoiesis in bone marrow fo siwu decocetion.

Actin Depolymerizing Factors ; metabolism ; Anemia ; chemically induced ; metabolism ; pathology ; Animals ; Annexin A1 ; metabolism ; Apoptosis ; drug effects ; Carbonic Anhydrases ; metabolism ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cyclophosphamide ; Drug Combinations ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Fatigue ; chemically induced ; metabolism ; pathology ; Female ; Hematopoietic Stem Cells ; metabolism ; pathology ; Medicine, Chinese Traditional ; Mice ; Mice, Inbred C57BL ; Peroxidases ; metabolism ; Peroxiredoxins ; Plants, Medicinal ; chemistry ; Proteome ; metabolism ; Proteomics ; methods

AIMTo explore the culture method of rat hepatocyte between two collagen gel layers on a sandwich configuration and observe the function and morphological characteristics of the hepatocyte, which will be used in evaluation the effect of traditional Chinese Medicine on cytochrome P450.

METHODSRat hepatocyte were isolated by two-step in situ collagenase perfusion method; the hepatocyte were seeded in dishes coated with type I rat tail collagen, culture medium was added and changed daily after gelation. The morphological characteristics of the hepatocyte were observed and biochemical index were tested. The drug effect on the expression of CYP3A was determined by reverse transcriptase-polymerase chain reaction (RT-PCR).

RESULTSRat hepatocytes were successfully attached in gel. BUN and Alb excretion from cell could be tested in the culture period, however, the release of LDH content were lower than the culture system without collagen gel. The typical cellular morphological characteristics of cultured hepatocytes could be observed. PCN increased CYP3A mRNA expression in dose-dependent manner, while the expression of GAPDH wasn't affected.

CONCLUSIONRat hepatocyte sandwich culture can maintenance the cell function and activity, which simulate the environment that more closely in vivo, especially the activity of drug metabolism enzymes.

Animals ; Cell Culture Techniques ; methods ; Cells, Cultured ; Culture Media ; Cytochrome P-450 Enzyme System ; metabolism ; Hepatocytes ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley