AIM:To characterize the effect of estradiol on proliferation,differentiation and extracellular matrix(ECM) accumulation in stromal cells through regulation of BPH-1 paracrine.METHODS:BPH-1 cells were stimulated with different concentrations of estradiol.Conditioned media(CM) were harvested and their effects on stromal cell cultures were tested.Cell proliferation was determined by MTT assay.mRNA of smoothelin,fibronectin,collagen Ⅳ and transforming growth factor ?1(TGF-?1) were analyzed by real-time RT-PCR.Western blotting was used to determine smooth muscle myosin heavy chain(SMMHC).ELISA and radioimmunoassay were respectively used to measure fibronectin,TGF-?1 and collagen Ⅳ protein expressions.RESULTS:Estrodiol stimulated the expression and secretion of TGF-?1 in BPH-1 cells.The proliferation of stromal cells increased when they were cultured with CM harvested from estrogen treated BPH-1 cells.The mRNA levels of collagen Ⅳ and smoothelin increased in stromal cells treated with CM from BPH-1 cells.The results of radioimmunoassay also showed that the collagen Ⅳ protein level up-regulated in the supernatants and cell extracts of CM-treated stromal cells.A neutralizing antibody to TGF-?1 inhibited the stimulation of collagen Ⅳ and SMMHC by BPH-1 CM.The expression of fibronectin was only marginally changed in stromal cells cultured in the presence of BPH-1 CM.CONCLUSION:The BPH-1 cells increase ECM accumulation and differentiation of stromal cells through TGF-?1.Estradiol stimulate differentiation of stromal cells by induction of TGF-?1 expression.Estradiol stimulate proliferation by influencing the factors secreted from prostatic epithelial cells.

Objective To evaluate the efficacy and feasibility of laparoscopie intervention for congenital obstructive megaureter in children. Methods Eleven children with congenital obstructive megaureter(left in 4,right in 7)underwent laparoseopie ureteroplasty.One had congenital ureter oririce stenosis,9 had been diagnosed as simple congenital ureter orifice stricture,1 had recurrent ureter orifice stricture after open ureterovesical reimplantation.B-ultrasound and IVU showed severe hydronephrosis in 7 cases and moderate in 4. Results The operation was successful in all cases and none had urine leakage.The mean operating time was 103.0±35.3 min(range 70-190 min).The mean blood loss was 18.0±9.5 ml(range 10-40 ml)and the mean postoperative hospital stay was 8.0±1.4 d(range 7-10 days).The double J stent was removed 6 weeks after operation.The patients were followed up for 3-24 months(mean,6 months).Cystography showed no reflux in all cases during follow-up. Conclusion Laparoscopical ureteroplasty could be a minimal invasive,less suffering technique for the treatment of congenital obstructive megaureter in children.

BACKGROUND:Gemcitabine hydrochloride is a water-soluble anticancer drug that induces apoptosis in tumor cells, but it has an excessive release in vivo. OBJECTIVE:To evaluate the effect of poly(lactic acid) (PLA) electrospun fiber membranes carrying gemcitabine hydrochloride on the growth of human osteosarcoma cell lines MG-63. METHODS:PLA electrospun fiber members with (experimental) or without (control) gemcitabine hydrochloride were fabricated and characterized. Two kinds of fiber membranes were immersed in low-glucose DMEM medium, and the supernatants were collected in the two groups at 3, 5, 7 days, respectively. Passage 5 human osteosarcoma cell lines MG-63 were inoculated into 96-well plates containing low-glucose DEME with 15％ fetal bovine serum, and divided into seven groups. Groups 1-3 were cultured in the experimental supernatants of 3, 5, 7 culture days, and groups 4-6 were cultured in the control supernatants of 3, 5, 7 culture days, respectively. The remaining group acted as the negative control with no supernatant. Thereafter, cell counting kit-8 was used to detect cell proliferation, and RT-PCR was used to measure expression of Bcl-2 and Bax at 3 days of culture. RESULTS AND CONCLUSION:(1) No obvious particle was found on the smooth and even surface of the fiber members in the experimental and control groups. There was no significant difference in fiber diameter, contact angle and tensile strength between the two kinds of fiber membranes. (2) The results of cell counting kit-8 showed that compared with the negative control group, the supernatant released from the control group had no effect on the MG-63 proliferation at different time points, while the supernatant released from the experimental group could inhibit the MG-63 proliferation at different time points (P<0.05), and the inhibitory effect became more and more obvious with the prolongation of release time. (3) RT-PCR findings showed that compared with the control group, the supernatant released from the experimental group could increase Bax mRNA expression and decrease Bcl-2 mRNA expression at the same time point. To conclude, the PLA electrospun fiber membranes carrying gemcitabine hydrochloride can sustainably inhibit MG-63 proliferation and promote cell apoptosis.

OBJECTIVETo study the function of the SP3111 protein in fertilization and early embryo development through in vitro fertilization (IVF) experiments following anti-SP111 antibody (Ab2438) blocking.

METHODSSperm samples collected from male mice were divided into an experimental, a blank control and a negative control group before IVF. The sperm of the experimental group was incubated with Ab2438 for 1 h followed by IVF and observed for the rates of fertilization and embryo fragmentation at 2, 4, 6, 8 and 22 h. Then the fertilized eggs were incubated with Ab2438, and the rates of fertilization embryo fragmentation were observed at 22 h.

RESULTSAfter the sperm was incubated with Ab2438, the incidences of embryo fragmentation were 5.26, 8.77, 23.25, 43.42 and 59.21% at 2, 4, 6, 8 and 22 h, respectively, with significant differences from the control groups (P < 0.01). After 22 h Ab2438 incubation of the fertilized eggs, the rates of normal and fragmented embryos of the experimental group were 23.64 and 63.64%, respectively, significantly different from those of the control groups (P < 0.01).

CONCLUSIONAnti-SP3111 antibodies remarkably affected fertilization and early embryo development in mice. The SP3111 protein may be a signal molecule and plays a role in fertilization and early embryo development together with other proteins. Further studies on the function of the SP3111 protein in reproduction may offer a new insight into the molecular mechanism of infertility.

Animals ; Antibodies ; immunology ; Embryonic Development ; Female ; Fertilization in Vitro ; Male ; Membrane Proteins ; metabolism ; Mice ; Mice, Inbred ICR ; Spermatozoa ; immunology

OBJECTIVETo detect cadmium in environmental and food samples by graphite furnace atomic absorption spectroscopy (GFAAS) and inductively coupled plasma atomic emission spectroscopy (ICPAES).

METHODSAn indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) was developed based on a cadmium-specific monoclonal antibody. IC-ELISA for cadmium in environmental and food samples was evaluated.

RESULTSIC-ELISA showed an IC50 of 45.6 microg/L with a detection limit of 1.95 microg/L for cadmium, and showed a mean recovery ranging 97.67%-107.08%. The coefficient of variations for intra- and interassay was 3.41%-6.61% and 4.70%-9.21%, respectively. The correlation coefficient between IC-ELISA and GFAAS was 0.998.

CONCLUSIONIC-ELISA can detect and quantify cadmium residue in environmental or food samples.

Animals ; Antibodies, Monoclonal ; Cadmium ; chemistry ; Environmental Pollutants ; chemistry ; Food Contamination ; analysis ; Immunoassay ; methods ; Mice ; Mice, Inbred BALB C ; Reproducibility of Results ; Sensitivity and Specificity

Animals ; Cell Adhesion Molecules ; metabolism ; Liver Cirrhosis, Experimental ; drug therapy ; metabolism ; Male ; Rats ; Rats, Wistar ; Thalidomide ; pharmacology ; therapeutic use

OBJECTIVETo determine the effects of Chinese herbal monomers such as baicalin, berberine, and matrine on the androgen receptor (AR) mRNA expression in SZ95 sebocytes in vitro and to explore the possible mechanism of using traditional Chinese medicines to treat acne.

METHODSSZ95 sebocytes were cultured and then treated with berberine, baicalin, matrine, and 13-cis-retinoic acid for 24 hours. Reverse transcription polymerase chain reaction was applied to detect the changes of AR.

RESULTAR mRNA was downregulated by 13-cis-retinoic acid of 1 x 10(-5) mol/L and 1 x 10(-6) mol/L, and by baicalin of 1 x 10(-4) mol/L (P < 0.05).

CONCLUSION13-cis-retinoic acid and baicalin may exert antiandrogenitic action by inhibiting AR mRNA expression in human sebocytes.

Androgen Antagonists ; pharmacology ; Cell Line ; Down-Regulation ; Drugs, Chinese Herbal ; pharmacology ; Flavonoids ; pharmacology ; Humans ; RNA, Messenger ; biosynthesis ; Receptors, Androgen ; biosynthesis ; genetics ; Skin ; cytology

The human basic Krüppel-like factor (hBKLF) is a newly cloned human transcription factor from the cDNA library of fetal liver. It belongs to the Krüppel-like transcription factor family. Previous expression study showed that it is a hematopoietic related factor. This study was aimed to investigate the effect of hBKLF on cell proliferation, differentiation and hemoglobin synthesis by using K562 cell line as model. The sense and antisense expression plasmids of hBKLF were constructed, and transfected into K562 cells by lipofectamine. After G418 selection for 4 weeks, the cell line with stable expression of the gene was obtained. Then the hBKLF expression level, proliferation ability, colony formation and hemoglobin production were detected by RT-PCR and Western blot, MTT method, methyl cellulose semisolid culture method and benzidine test respectively. The morphologic change of cell was observed with inverted microscope. The results showed that the sense plasmid could increase hBKLF level and antisense plasmid could decrease hBKLF expression. When hBKLF level was down-regulated, K562 cells could proliferate more quickly and synthesize more hemoglobin. But there were no differences in colony formation ability and no apparent morphologic change. It is concluded that hBKLF can inhibit hematopoietic cell proliferation and hemoglobin synthesis. It is suggested that hBKLF plays an important role in the proliferation and differentiation of hematopoietic cells.
Animals ; COS Cells ; Cell Differentiation ; physiology ; Cell Proliferation ; drug effects ; Cell Transformation, Neoplastic ; drug effects ; Cercopithecus aethiops ; Hemoglobins ; biosynthesis ; Humans ; K562 Cells ; Kruppel-Like Transcription Factors ; biosynthesis ; genetics ; pharmacology ; Transcription Factors ; biosynthesis ; genetics ; Transfection

OBJECTIVETo investigate the inhibitory effects of CaMKIIN on acute myeloid leukemia cell line HL-60 to explore a novel therapeutic target of leukemia.

METHODSHuman CaMK II N gene expression vector pcDNA3.1/hCaMKIIN or empty vector pcDNA3.1/myc-His (-) B was transfected into HL-60 cells by Lipofectamine 2000. Human CaMK II N proteins of transfected cells were detected by Western blot. Cell proliferation affected by human CaMKIIN was determined by MTT. Colony-forming assay was performed by soft agar growth system. The cells transfected with CaMKIIN were stained with Hoechst 33342 to detect the apoptotic proportion under fluorescence microscopy. Cell cycle was analyzed by flow cytometry.

RESULTSHuman CaMKIIN was stably transfected into HL-60 cells, and overexpression of human CaMKIIN inhibited the proliferation of HL-60/CaMKIIN cells compared to HL-60/mock cells and HL-60 cells [(0.44 ± 0.03) vs (0.94 ± 0.05) vs (0.94 ± 0.04), P<0.01]. The colony formation of HL-60/CaMKIIN was also markedly smaller[(21.00 ± 3.05)/500] than that of mock-transfected [(111.00±4.58)/500]] and control cells [(119.00±6.09)/500] (P<0.01). After 72 hrs-culture, the apoptotic proportion in cells transfected with CaMK II N was obviously higher than of cells transfected with mock DNA or control [(22.49 ± 2.15)% vs (7.17 ± 0.72)% vs (6.40 ± 0.55)%, P<0.01]. Up to (82.97 ± 2.90)% human CaMKIIN/HL-60 cells were arrested at G0/G1 phase, which was more than mock-transfected [(40.53 ± 2.38)%] and control cells [(41.63 ± 2.27)%] (P<0.05). Human CaMKIIN could down-regulate expression of Bcl-2 in transfected cells.

CONCLUSIONCaMK IIN up-regulation could inhibit proliferation and induce apoptosis of human acute myeloid leukemia cell HL-60.

Apoptosis ; Cell Proliferation ; Genetic Vectors ; HL-60 Cells ; Humans ; Proteins ; genetics ; metabolism ; Transfection ; Up-Regulation

BACKGROUNDModern research has provided new insights into the biological mechanisms of noise-induced hearing loss, and a number of studies showed the appearance of increased reactive oxygen species (ROS) and reactive nitrogen species (RNS) during and after noise exposure. This study was designed to investigate the noise exposure induced nitrotyrosine change and the mechanism of outer hair cells death in guinea pig cochlea.

METHODThirty guinea pigs were used in this study. The experimental animals were either exposed for 4 hours per day to broadband noise at 122 dB SPL (A-weighted) for 2 consecutive days or perfused cochleae with 5 mg/ml of the SIN1 solutions, an exogenous NO and superoxide donor, for 30 minutes. Then the cochleae of the animals were dissected. Propidium iodide (PI), a DNA intercalating fluorescent probe, was used to trace morphological changes in OHC nuclei. The distribution of nitrotyrosine (NT) in the organ of Corti and the cochlear lateral wall tissue from the guinea pigs were examined using fluorescence immunohistochemistry method. Whole mounts of organ of Corti were prepared. Morphological and fluorescent changes were examined under a confocal microscope.

RESULTSEither after noise exposure or after SIN1 perfusion, outer hair cells (OHCs) death with characteristics of both apoptotic and necrotic degradation appeared. Nitrotyrosine immunolabeling could be observed in the OHCs from the control animals. After noise exposure, NT immunostaining became much greater than the control animals in OHCs. The apoptotic OHC has significant increase of nitrotyrosine in and around the nucleus following noise exposure. In the normal later wall of cochleae, relatively weak nitrotyrosine immunolabeling could be observed. After noise exposure, nitrotyrosine immunoactivity became stronger in stria vascularis.

CONCLUSIONNoise exposure induced increase of nitrotyrosine production is associated with OHCs death suggesting reactive nitrogen species participation in the cochlear pathophysiology of noise-induced hearing loss.

Animals ; Cell Death ; Cochlea ; chemistry ; pathology ; Female ; Guinea Pigs ; Hair Cells, Auditory, Outer ; pathology ; Immunohistochemistry ; Male ; Noise ; adverse effects ; Organ of Corti ; chemistry ; pathology ; Tyrosine ; analogs & derivatives ; analysis