OBJECTIVEThe expression of C/EBPalpha protein and mRNA during automatically activation process in primary cultures of HSCs were observed in order to explore its possible association with the proliferation and activation of HSCs.

METHODSImmunocytochemistry, Western blot and RT-PCR were used to evaluated the expression of C/EBPalpha protein and mRNA; as well as the expression of alpha-SMA, Desmin, MMP2, type I procollagen (alpha1). The eukaryotic vector harboring the full length cDNA of C/EBPalpha was transfected into activated HSC, then immunocytochemistry was applied to confirm the transfection and evaluate the effect of transfection on the proliferation of HSC by calculating the PCNA-positive cells. The morphological changes of HSC were observed by use of phase-contrast microscope.

RESULTSConstitutive expression of mRNA and protein of C/EBPalpha were detected in primarily cultured HSCs, and the protein was seen in both nuclei and cytoplasm with the latter being dominant. Their expression levels reached highest at day 2 of the culture, then decreased gradually when continually cultured to the day 4, 7, 10, on the other hand, the expression of alpha-SMA, MMP2 and ColI(alpha1) increased steadily. Transient transfection was verified by the fact that much more and stronger C/EBPalpha stain was observed in transfected HSCs than in void-vector transfected cells. In C/EBPalpha gene transfected HSCs, the number of PCNA-positive cells dramatically decreased compared with the void-vector transfected cells 24h after transfection. In addition, the C/EBPalpha gene transfected HSCs died 36 h after transfection, a few surviving cells became longer and thinner in morphology, however the void-vector transfected cells almost all remained alive.

CONCLUSIONSC/EBPalpha was likely involved in the HSCs activation, and over-expressed C/EBPalpha by transfection had inhibitory influence on the proliferation of cultured rat HSCs.

Animals ; CCAAT-Enhancer-Binding Protein-alpha ; genetics ; Cells, Cultured ; Collagen Type I ; genetics ; Liver ; cytology ; metabolism ; Male ; Matrix Metalloproteinase 2 ; genetics ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Transfection

OBJECTIVETo study the effect of RAR-beta transfection plus treatment with the corresponding ligand ATRA on the proliferation and phenotype of platelet-derived growth factor (PDGF)-activated hepatic stellate cells (HSC).

METHODSPDGF-activated hepatic stellate cells of rats were transfected with eukaryotic expression vector pCMV-script-RAR-beta, which was verified by western blot. The proliferation of transfected HSC was assayed by BrdU incorporation as well as MTT methods. Their phenotype (alpha-SMA and desmin) was observed by immunocytochemistry assay with image analysis and RAR-beta protein expression was detected by western blot.

RESULTSTransfection of RAR-beta gene and treatment with ligand ATRA could increase the expression of RAR-beta protein for at least 144h and inhibit the proliferation and the expression of alpha-SMA and desmin in PDGF-activated HSC. Significant statistical differences were perceived comparing with sham-transfected, only-PDGF treated, non-ligand treated and irrelevant ligand-treated HSC.

CONCLUSIONSTransfected with RAR-beta gene as well as using related ligand ATRA could suppress the proliferation and reverse the activation phenotype of activated HSC.

Animals ; Blotting, Western ; Cell Division ; Liver ; cytology ; Phenotype ; Platelet-Derived Growth Factor ; pharmacology ; Rats ; Receptors, Retinoic Acid ; physiology ; Transfection ; Tretinoin ; pharmacology

OBJECTIVETo investigate the safety and efficiency of the disposable circumcision suture device (DCSD) in the surgical treatment of phimosis and redundant prepuce.

METHODSWe randomly assigned 249 outpatients with phimosis or redundant prepuce to be treated with DCSD (n = 129) and by conventional circumcision (CC, n = 120), respectively. Then we compared the safety and efficiency of the two strategies.

RESULTSComparisons between DCSD and CC showed that the operation time was (4.02 +/- 0.69) vs (30.8 +/- 4.05) min, blood loss was (1.07 +/- 1.29) vs (8.72 +/- 2.15) ml, intraoperative pain score was 0.81 +/- 0.81 vs 2.42 +/- 1.15, 24-hour postoperative pain score was 1.84 +/- 1.02 vs 4.99 +/- 1.36, postoperative complication rate was 13. 95% (18/129) vs 9.17% (11/120), wound healing time was (13.99 +/- 9.06) vs (17.48 +/- 3.49) d, satisfaction with the penile appearance was 98.4% (127/129) vs 95% (109/120), and treatment cost was (2215.62 +/- 17.67) vs (576.47 + 15.58) Y RMB. DCSD exhibited obvious superiority over CC for shorter operation time, less blood loss, milder intraoperative pain, sooner wound healing, and better penile appearance, but it also had a higher rate of postoperative complications (P > 0.05) and involved more treatment cost than the latter (P < 0.05).

CONCLUSIONThe disposable circumcision suture device affords ideal clinical effects and therefore deserves clinical popularization.

Circumcision, Male ; instrumentation ; Disposable Equipment ; Follow-Up Studies ; Humans ; Male ; Phimosis ; surgery ; Surgical Staplers ; Treatment Outcome

To study the detection of weak D and Del from samples initially screened RhD(-), RhD phenotype was initially screened by routine serological test, out of which weak D phenotype was detected by indirect antiglobulin test (IAT) and Del phenotype was detected by chloroform-trichloroethylene absorption-elution test. The results showed that 56 samples were RhD(-) confirmed by routine serology test, which were screened out of 26 200 donors, among them 5 samples were typed as weak D by IAT and 9 cases samples were typed as Del by absorption-elution test. In conclusion, the samples which typed as RhD(-) by routine serological test must be identified by IAT and chloroform-trchloroethylene absorption test is order to detect weak D and Del phenotype. It is important for clinical transfusion safely.
Blood Donors ; Blood Grouping and Crossmatching ; methods ; standards ; Epitopes ; immunology ; Erythrocytes ; immunology ; Humans ; Rh-Hr Blood-Group System ; blood ; immunology

This study was aimed to isolate and culture rhesus mesenchymal stem cells (MSC), and to analyze its phenotype and biological characteristics. Bone marrow was aspirated from femur of rhesus, mononuclear cells were extracted, the nonadherent cells were removed after 24 hours, adherent cells were subcultured when they grew 80% confluence. After the fifth passage, the cells were used for examination. The phenotypes of MSC were analyzed by flow cytometry, differentiated cells were identified by relevant specific staining. Cytokines' mRNA expressed by MSC were detected by RT-PCR. The results showed that rhesus MSC gave rise to a population of adherent cells characterized by the presence of a predominant cell type with a typical fibroblast-like morphology. During the log phase of growth, MSC proliferated to a two-fold population at 31 - 40 hours. MSC could be ex vivo expanded by successive cycles of trypsinization, seeding, and culture. The phenotypes expressed on rhesus MSC were Flk-1, CD29, CD105, CD166 positive and CD34, CD45, CD33 nearly negative. In various induction differentiation conditions, rhesus MSC could differentiate into the osteoblast and lipocyte. IL-6, TGF-beta were highly expressed, TNF-alpha was lowly expressed and IL-2, Fas-L, IFN-gamma were not detected on rhesus MSC using RT-PCR method. It is concluded that rhesus MSC can be successfully isolated and culture-expanded and the biological characteristics of rhesus MSC are similar to those of human MSC.
Adipocytes ; cytology ; immunology ; metabolism ; Alkaline Phosphatase ; metabolism ; Animals ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Cytokines ; genetics ; Female ; Flow Cytometry ; Gene Expression ; Immunohistochemistry ; Immunophenotyping ; Macaca mulatta ; Male ; Mesenchymal Stromal Cells ; cytology ; immunology ; metabolism ; Osteoblasts ; cytology ; immunology ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVEIn order to discuss the chemical foundation of hematopoietic effect of Siwu Tang, three fractions of different polarities (C1, C2 and C3) were prepared from Siwu Tang and the characteristics of these fractions' constituents were investigated.

METHODFraction C1, C2 and C3 of Siwu Tang and corresponding fractions of Siwu Tang's four ingredient drugs were analyzed and compared, synthetically using the three methods of high-performance thin layer chromatography (HPTLC), high-performance liquid chromatography (HPLC) and direct infusion electrospray ionization mass spectrometry (ESI-MS).

RESULTFraction C1 of Siwu Tang contained various types of compounds, including ferulic acid, paeoniflorin and supposedly ligustilide, etc. Saccharide content in fraction C1 was very little. The major constituents in fraction C2 of Siwu Tang were paeoniflorin, monosaccaride and disaccharide. The major constituents in fraction C3 of Siwu Tang were monosaccaride and disaccharide.

CONCLUSIONWith synthetical chromatographic and direct infusion ESI-MS methods, abundant information on composition of fractions of traditional Chinese medicine formulas can be obtained. The results gained with different methods can be compared with each other and corroborate each other, so that the obtained information can be more comprehensive and more definite than that gained with single method. The results of this study are important as references for the discussion of the chemical foundation of hematopoietic effect of Siwu Tang.

Angelica sinensis ; chemistry ; Benzoates ; analysis ; Bridged-Ring Compounds ; analysis ; Chromatography, High Pressure Liquid ; Chromatography, Thin Layer ; Coumaric Acids ; analysis ; Disaccharides ; analysis ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Glucosides ; analysis ; Ligusticum ; chemistry ; Monosaccharides ; analysis ; Monoterpenes ; Paeonia ; chemistry ; Plants, Medicinal ; chemistry ; Rehmannia ; chemistry ; Spectrometry, Mass, Electrospray Ionization

OBJECTIVETo detect the levels of nitric oxide synthetase (NOS) and carbon monoxide (CO) in the penile corpus cavernous of adult male Wistar rats with high homocysteine (Hhcy) and to explore the relationship of NOS and CO levels with erectile dysfunction.

METHODSTwenty Wistar rats were equally and randomly divided into a control and an Hhcy group and fed on normal diet and normal diet with 3.0% methionine respectively. Four weeks later, the levels of NOS and CO in the penile corpus cavernous were detected by ultraviolet spectrophotometry and that of serum homocysteine by the cycle enzyme method.

RESULTSCompared with the control group, the levels of NOS and CO in the penile corpus cavernous were significantly lower in the Hhcy group, (6.45 +/- 1.12) nmol/(g x min) vs (10.77 +/- 0.60) nmol/(g x min) and (10.60 +/- 0.92) micromol/L vs (13.36 +/- 0.44) micromol/L, while that of homocysteine was significantly higher, (22.32 +/- 1.65) micromol/L) vs (4.90 +/- 1.73) micromol/L.

CONCLUSIONFour-week diet with methionine can cause Hhcy and significantly decreased levels of NOS and CO in the penile corpus cavernous in Wistar rats. Hhcy is an independent risk factor of erectile dysfunction.

Animals ; Carbon Monoxide ; metabolism ; Homocysteine ; blood ; Hyperhomocysteinemia ; blood ; physiopathology ; Male ; Nitric Oxide Synthase ; metabolism ; Penis ; metabolism ; Rats ; Rats, Wistar ; Spectrophotometry, Ultraviolet

OBJECTIVETo study the distribution of genomovars and microevolution of Yersinia pestis in the Qinghai-Tibet Plateau.

METHODSPrimer pairs targeting the twenty-two different regions(DFRs) were designed for detecting the presence or deletion of each DFR in 297 strains isolated from the Qinghai-Tibet Plateau.

RESULTS9 genomovars, i. e. Genomovar 1, 5, 6, 7, 8, 10, 11, new type and Ype-ancestor were identified in the Marmota himalayana plague focus of the Qinghai-Tibet Plateau. Among these genomovars, genomovar 5,8 and 10 were dominant types. The total rate of the three genomovars was 80.6% (204/253) and the genomovars in different regions were different. All of 44 strains of Y. pestis in the Microtus fuscus plague focus of the Qinghai-Tibet Plateau belonged to genomovar 14.

CONCLUSIONThe distribution of genomovars of Y. pestis in the Qinghai-Tibet plateau had remarkable characteristics geographically. Based on the distribution of genomovars of Y. pestis, the routes of transmission and microevolution of Y. pestis were proposed.

Biological Evolution ; China ; Geography ; Humans ; Plague ; transmission ; Yersinia pestis ; genetics

The growth of three plague phages from Qinghai Plateau in two Yersinia pestis strains(plague vaccine strains EV76 and 614F)and four non-Yersinia pestis strains(Yersinia pseudotuberculosis PTB3,PTB5,Escherichia coli V517,and Yersinia enterocolitica 52302-2)were detected through a micromethod based on the OmniLogTM microbial identification system and by the drop method,to provide a scientific basis for future ecological studies and classification based on the host range.For plague vaccine strains EV76 and 614F,successful phage infection and subsequent phage growth were observed in the host bacte-rium.Diminished bacterial growth and respiration and a concomitant decrease in color were observed with the OmniLogTM mi-crobial identification system at 33 ℃ for 48 h.Yersinia pseudotuberculosis PTB5 was sensitive to Yersinia pestis phage 476,but Yersinia pseudotuberculosis PST5 was insensitive to phage 087 and 072204.Three strains of non-Yersinia pestis(Yersinia pseudotuberculosis PTB3,Escherichia coli V517,and Yersinia enterocolitica 52302-2)were insensitive to Yersinia pestis pha-ges 087,072204,and 476 showed similar growth curves.The growth of phages 476 and 087,as determined with the drop method,in two Yersinia pestis strains(plague vaccine strains EV76 and 614F)and four non-Yersinia pestis strains(Yersinia pseudotuberculosis PTB3,Escherichia coli V517,and Yersin-ia enterocolitica 52302-2)showed the same results at 37 ℃,on the basis of comparisons with the OmniLogTM microbial i-dentification system;in contrast,phages 072204 did not show plaques on solid medium at 37 ℃ with plague vaccine strains EV76 and 614F.Determination based on the OmniLogTM detection system can be used as an alternative to the traditional determination of the host range,thus providing favorable application val-ue for determining the interaction between the phage and host bacteria.