OBJECTIVETo explore the role of TBX3 gene in the pathogenesis of breast cancer.

METHODSThe total RNA of 51 fresh breast cancer tissues and the corresponding adjacent tissues were extracted and reverse transcribed into cDNA to detect the expression of TBX3 mRNA by real-time PCR. The correlation between TBX3 mRNA expression and the clinicopathologic parameters in relation to breast cancer metastasis was analyzed.

RESULTCompared to that in the adjacent tissues, the expression of TBX3 mRNA was markedly increased in breast cancer tissues. TBX3 mRNA expression was significantly higher in metastatic breast cancer than in non-metastatic tumors.

CONCLUSIONIncreased expression of TBX3 mRNA suggests the involvement of TBX3 in the pathogenesis and metastasis of breast cancer.

Breast Neoplasms ; etiology ; genetics ; pathology ; Female ; Humans ; Neoplasm Metastasis ; genetics ; RNA, Messenger ; genetics ; metabolism ; T-Box Domain Proteins ; genetics ; metabolism

The aim of this paper is to report the synthesis of the mPEG-PCL-g-PEI copolymers as small interfering RNA (siRNA) delivery vector, and exploration of the siRNA delivery potential of mPEG-PCL-g-PEI in vitro. The diblock copolymers mPEG-PCL-OH was prepared through the ring-opening polymerization. Then, the hydroxyl terminal (-OH) of mPEG-PCL-OH was chemically converted into the carboxy (-COOH) and N-hydroxysuccinimide (NHS) in turn to prepare mPEG-PCL-NHS. The branched PEI was reacted with mPEG-PCL-NHS to synthesize the ternary copolymers mPEG-PCL-g-PEI. The structure of mPEG-PCL-g-PEI copolymers was characterized with Fourier transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR) and gel permeation chromatography (GPC). The mPEG-PCL-g-PEI/siRNA nanoparticles were prepared by complex coacervation, and the nanoparticles size and zeta potential were determined, separately. The cytotoxicities of mPEG-PCL-g-PEI/siRNA nanoparticles and PEI/siRNA nanoparticles were compared through cells MTT assays in vitro. The inhibition efficiencies of firefly luciferase gene expression by mPEG-PCL-g-PEI/ siRNA nanoparticle at various N/P ratios were investigated through cell transfection in vitro. The experimental results suggested that the ternary (mPEG5k-PCL(1.2k))1.4-g-PEI(10k) copolymers were successfully synthesized. (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k) could condense siRNA into nanoparticles (50-200 nm) with positive zeta potential. MTT assay results showed that the cytotoxicity of (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k)/siRNA nanoparticles was significantly lower than that of PEI(10k)/siRNA nanoparticles (P < 0.05). The expression of firefly luciferase gene could be significantly down-regulated at a range of N/P ratio from 50 to 150 (P < 0.01), and maximally inhibited at the N/P ratio of 125. The mPEG-PCL-g-PEI polymers could delivery siRNA into cells to inhibit the expression of target gene with very low cytotoxicity, which suggested that mPEG-PCL-g-PEI could serve as a new type of siRNA delivery vector.
Cell Line, Tumor ; Cell Survival ; Drug Carriers ; Genes, Reporter ; Genetic Vectors ; Humans ; Luciferases ; metabolism ; Molecular Weight ; Nanoparticles ; Particle Size ; Polyesters ; chemistry ; Polyethylene Glycols ; chemistry ; Polyethyleneimine ; chemistry ; Polymers ; chemistry ; RNA, Small Interfering ; administration & dosage ; genetics ; metabolism ; Transfection

OBJECTIVETo investigate the effects of Cox7a2 on the LH-induced testosterone production and the involved autophagy regulating signals in TM3 mouse Leydig cells.

METHODSThe Cox7a2-pEYFP-N1 fluorescent protein vector was constructed and transfected into TM3 mouse Leydig cells. The level of testosterone was determined by ELISA, and the effects of Cox7a2 on the expression of the steroidogenic acute regulatory protein (StAR) and the phosphorylation of the autophagy regulatory factor P70S6K were detected by Western blot.

RESULTSLH stimulation increased the StAR protein expression and testosterone production, while Cox7a2 decreased P70S6K phosphorylation, reduced StAR expression and consequently inhibited LH-induced testosterone biosynthesis in the TM3 Leydig cells.

CONCLUSIONCox7a2 inhibits testosterone production by decreasing the StAR protein expression, which might be at least in part related with the activation of autophagy in TM3 mouse Leydig cells.

Animals ; Autophagy ; Cells, Cultured ; Electron Transport Complex IV ; genetics ; Leydig Cells ; metabolism ; Luteinizing Hormone ; pharmacology ; Male ; Mice ; Phosphoproteins ; metabolism ; Phosphorylation ; Ribosomal Protein S6 Kinases, 70-kDa ; metabolism ; Testosterone ; biosynthesis

BACKGROUNDY-27632 is a specific inhibitor of Rho-associated coiled kinase (ROCK) and has been shown to promote the survival and induce the differentiation of a variety of cells types. However, the effects of Y-27632 on adult human adipose tissue-derived stem cells (ADSCs) are unclear. This study aimed to investigate the effects of Y-27632 on the neuronal-like differentiation of ADSCs.

METHODSADSCs were isolated from women undergoing plastic surgery and cultured. ADSCs were treated with different doses of Y-27632 and observed morphological changes under microscope. The expression of nestin, neuron specific enolase (NSE) and microtubule-associated protein-2 (MAP-2) in ADSCs treated with Y-27632 was detected by immunocytochemistry and Western blotting analysis.

RESULTSY-27632 had the potency to induce neuronal-like differentiation in ADSCs in a dose-dependent manner. Moreover, the differentiation induced by Y-27632 was recovered upon drug withdraw. ADSCs treated with Y-27632 expressed neuronal markers such as NSE, MAP-2 and nestin while untreated ADSCs did not express these markers.

CONCLUSIONSelective ROCK inhibitor Y-27632 could potentiate the neuronal-like differentiation of ADSCs, suggesting that Y-27632 could be utilized to induce the differentiation of ADSCs to neurons and facilitate the clinical application of ADSCs in tissue engineering.

Adipose Tissue ; cytology ; Adult ; Amides ; pharmacology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Female ; Humans ; Neurons ; cytology ; Pyridines ; pharmacology ; Stem Cells ; cytology ; drug effects

BACKGROUNDRh blood group system is the most complex and immunogenetic blood group system. Prevalent RHD alleles varied in different populations. The purpose of this study is to determine the molecular basis of weak D and DEL phenotype in Anhui Chinese Han population.

METHODSThe D antigen was determined with IgM monoclonal anti-D conformed to the guidelines for donor testing in China. Weak D samples were identified by an indirect antiglobulin test. DEL phenotype was determined by adsorption and elution test. All the RHD 10 exons were screened by PCR with sequence-specific priming or sequenced for the first-time donors who typed weak D, DEL or D negative by serologic test.

RESULTSOf all the 30 799 blood donors, 155 blood samples were found D negative with IgM anti-D; 34 blood samples were found D positive by indirect antiglobulin test or absorption elution test. RHD alleles were identified by nucleotide sequencing. Total 4 RHD alleles were found including two new. One hundred and twenty of 155 (77.4%) of the serologically D negative samples lacked the RHD gene. One D negative was RHD(615del2). Thirty-two of 155 (20.6%) carried RHD(K409K) among them one carrying 1227G > A and 845G > A. Two of 155 (1.3%) was weak D type 15.

CONCLUSIONSIn this study at the molecular level, all DEL phenotype is RHD(K409K); weak D type 15 is the prevalent weak D allele in Anhui Chinese Han population. Additionally, an improved more efficient method was adopted to amplify all the RHD exons in one PCR program. Our study added to the understanding of molecular mechanisms underlying D antigen expression in Anhui Han population and provided useful information for adopting suitable genotyping strategies in routine use.

Alleles ; China ; Exons ; genetics ; Humans ; Immunoglobulin M ; blood ; Multiplex Polymerase Chain Reaction ; Phenotype ; Polymerase Chain Reaction ; Rh-Hr Blood-Group System ; genetics

To study the effect of cytochrome C on HL-60 cells in vitro and the mechanism of expression changes of relevant apoptotic genes, the inhibition rate of cytochrome C on HL-60 cells was detected by MTT, the morphology of HL-60 cells was observed by light microscopy and fluorescence microscopy, the changes of apoptosis rate and cell cycle were assayed by flow cytometry (FCM), DNA ladder was investigated on electrophoresis, the expression changes of bax and bcl-2 mRNA were examined by RT-PCR, when HL-60 cells were treated with different concentrations of cytochrome C for 24 hours. The results showed that the inhibition rate increased with increase of the cytochrome C concentration within 0 - 150 mg/L; when treated with 0 - 37.5 mg/L cytochtome C for 24 hours, the percentage of apoptotic HL-60 cells increased with the dose increasing, and the typical apoptotic cells and the apoptotic DNA ladder were observed. At the same time, within this range of concentration, the expression of bcl-2 mRNA decreased gradually and the expression of bax increased gradually. When the cytochrome C concentration was higher than 37.5 mg/L, the percentage of apoptotic HL-60 cells not increased, but decreased, while the cells necrosed. The above metioned results suggested that at certain range of concentration of cytochrome C, apoptosis or necrosis can be induced by cytochrome C, and cell cycle arrests at G(1) phase in HL-60 cells, the percentage of apoptotic cells and the changes of expression of bax and bcl-2 depend on the dose of cytochrome C. The mechanism that cytochtome C induced apoptosis in HL-60 cells may be related to the activation of bax and inhibition of bcl-2.
Apoptosis ; drug effects ; Cytochromes c ; pharmacology ; Dose-Response Relationship, Drug ; Flow Cytometry ; Gene Expression Regulation, Leukemic ; drug effects ; HL-60 Cells ; Humans ; Microscopy, Fluorescence ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; bcl-2-Associated X Protein ; genetics

The purpose was to study the responses of AML cell treated with cytochrome C and to explore the influence of cytochrome C on apoptosis of AML cell induced by daunorudicine (DNR). The differentiation of AML cell was detected by Wright-Giemsa staining and NBT test, the apoptosis of AML cell was assayed by flow cytometry and fluorescence microscopy. The results showed as follows: (1) different concentrations of cytochrome C could induce different effects on AML cells. Concentration of cytochrome C for differentiation was 10 microl/ml, for apoptosis was 20 microl/ml, and for necrosis was 40 microl/ml. (2) the apoptosis of AML cells decreased with the administration of cytochrome C in 10.0 microg/ml before treating AML cells with DNR (P < 0.01), but no change was shown with the administration of cytochrome C in 20.0 microg/ml (P > 0.05). (3) in reverse sequence, administrating of cytochrome C in 10 microl/ml and 20 microl/ml after treating AML cells with DNR, two different concentrations of cytochrome C could increase the apoptosis of AML cells (P < 0.01). It is suggested that cytochrome C may probably affect the apoptosis of AML cells induced by DNR.
Antibiotics, Antineoplastic ; pharmacology ; Apoptosis ; drug effects ; Cytochromes c ; pharmacology ; Daunorubicin ; pharmacology ; Dose-Response Relationship, Drug ; Drug Synergism ; Flow Cytometry ; Humans ; Leukemia, Myeloid, Acute ; pathology ; Microscopy, Fluorescence ; Tumor Cells, Cultured

OBJECTIVETo evaluate the effect of cluster of differentiation 14 (CD-14) and Toll like receptors (TLR) on the expression of interleukin-6 (IL-6) mRNA induced by Porphyromonas endodontalis (Pe) lipopolysaccharides (LPS).

METHODSMC3T3-E1 cells were treated with 10 mg/L Pe-LPS for different hours, and the cells uninvolved by anything as the blank group. The expression of IL-6 was detected by reverse transcription polymerse chain reaction (RT-PCR) and enzyme-liked immunosorbent assay (ELISA). The expression of CD-14, TLR-2 and TLR-4 mRNA was observed at different time point (0 - 24 h) by RT-PCR. The protein of CD-14, TLR-2 and TLR-4 was analyzed with a flow cytometer. MC3T3-E1 cells were pretreated with anti-CD-14, anti-TLR-2 and anti-TLR-4 antibody for 1 h, and then cells were stimulated with 10 mg/L Pe-LPS for 6 h. The expression of IL-6 mRNA was examined by RT-PCR. Statistical analysis was performed using one-way ANOVA Dunnett-t test with SPSS 11.0 software package.

RESULTSThe IL-6 mRNA and proteins increased significantly after treatment with Pe-LPS. When MC3T3-E1 cells treated by Pe-LPS for 6 h, the expression of proteins soared from (11.696 ± 0.672) ng/L to (36.534 ± 0.574) ng/L (P < 0.01); In the control group, the CD-14 and TLR-4 mRNA are ambly-expression, and the ratios of CD-14 and TLR-4 positive cells were (39.038 ± 3.131)% and (11.438 ± 0.385)% respectively in MC3T3-E1. After treatment by Pe-LPS, the expression of CD-14 and TLR-4 mRNA increased significantly, and the ratios of CD-14 and TLR-4 positive cells markedly increased to (62.407 ± 1.800)% and (21.367 ± 2.271)%. TLR-2 expression did not change apparently after Pe-LPS treatment. The expression of IL-6 mRNA was partly inhibited by anti-CD-14 or anti-TLR-4 antibody, but not by TLR-2.

CONCLUSIONSPe-LPS can induce the expression of IL-6 in osteoblast MC3T3-E1 through CD-14 and TLR-4, but not TLR-2.

3T3 Cells ; Animals ; Antibodies ; immunology ; Interleukin-6 ; genetics ; metabolism ; Lipopolysaccharide Receptors ; genetics ; metabolism ; Lipopolysaccharides ; isolation & purification ; pharmacology ; Mice ; Porphyromonas endodontalis ; RNA, Messenger ; metabolism ; Toll-Like Receptor 2 ; genetics ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism

OBJECTIVE: To evaluate the therapeutic effects of intraarticular injection of ligustrazine on knee osteoarthritis (OA). METHODS: Seventy-one cases of knee osteoarthritis (82 knees) were randomly divided into ligustrazine (LI) group and sodium hyaluronate (SH) group. The patients were intraarticularly injected ligustrazine or sodium hyaluronate once a week for 5 consecutive weeks, and were followed-up for 3 months. Lequesneos protocol for the evaluation of OA severity and activity was used. The therapeutic effects and changes of Lequesneos index were observed after the treatment. RESULTS: There was significant decrease in Lequesneos index in SH group after the treatment (P<0.01), but not in LI group (P>0.05). Three weeks later, there was significant decrease in Lequesneos index in both groups after the treatment (P<0.01), with no significant difference between SH and LI group (P>0.05). After the 5-week treatment, the efficacy rate of the LI group was 82.1%, and that of the SH group was 87.2%. No serious toxic or side effect was observed during the treatment and the follow-up. CONCLUSION Intraarticular injection of ligustrazine has a therapeutic effect on knee OA. No adverse effect is observed, but it needs long time to take effect.
Anti-Inflammatory Agents, Non-Steroidal ; administration & dosage ; Female ; Follow-Up Studies ; Humans ; Injections, Intra-Articular ; Male ; Middle Aged ; Osteoarthritis, Knee ; drug therapy ; Pyrazines ; administration & dosage

Objective To explore the clinical outcomes of patients with hypertensive intracerebral hemorrhage in the thalamus received stereotactic combined with microsurgical treatment,and find the best treated method. Methods Forty-three patients with hypertensive intracerebral hemorrhage in the thalamus,admitted to our hospital from October 2009 to March 2011,were selected in our study; these patients were divided into stereotactic combined with microsurgical treatment group (observation group,n=22) and direct surgery group (control group,n=21).Preoperative hematoma,and hematoma at 1 week and 1 month after the treatments under CT were compared; clinical curative effect at 1 month after the treatments and activities of daily living (ADL) 3 months after the treatments were compared. Results The hematoma in the observation group 1 week and 1 month after the treatments was statistically different as compared with that in the control group (P＜0.05).Clinical curative effect in the observation group (average rank=18.022) was better than that in the control group (26.166) with significant differences (P＜0.05); the total effective rate in the observation group was 77.3％ and that in the control group was 52.4％ with significant differences (P＜0.05).ADL scale indicated that the ability of the observation group (average rank=18.363) were better than that in the control group (25.809) with significant differences (P＜0.05).The postoperative complication rate in the observation group (22.7％) was lower than that in the control group (42.8％) with significant differences (P＜0.05). Conclusion Stereotactic combined with microsurgical treatment in patients with hypertensive intracerebral hemorrhage in the thalamus is safe and effective; this treatment can improve the postoperative living quality and enjoy few complications,which is worth in the clinical application.