OBJECTIVETo investigate the role of cyclin D1 and p27(kip1) in the occurrence and development of conventional renal cell carcinoma(CRCC).

METHODSRT-PCR and Western-blot were used to detect mRNA and protein contents of cyclin D1 and p27(kip1) in 25 CRCCs and 10 normal renal tissue distant to tumor. Immunohistochemistry was used to investigate the expression of cyclin D1 and p27(kip1) in pathological tissue sections of 76 CRCCs. The relationship between those index and clinicopathological parameters was analyzed statistically.

RESULTIn CRCC, the expression of cyclin D1 was higher than that of the control group. The higher cyclin D1 content was related to big tumor size (P<0.05); The expression of p27(kip1) was lower than that of the control group, and the lower p27(kip1) was related to higher nuclear grade and TNM stage (P<0.01). The 5-year cumulative survival rate of the p27(kip1) high expression group is longer than that of the low group (P<0.01).

CONCLUSIONThe excessive expression of cyclin D1 and lower expression of p27(kip1) play an important role in the carcinogenesis of CRCC. The lower expression of p27(kip1) may affect the progression of the tumors. The detection of p27(kip1) may be as a reference marker in the prognosis of CRCC.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Blotting, Western ; Carcinoma, Renal Cell ; genetics ; metabolism ; pathology ; Cyclin D1 ; biosynthesis ; genetics ; Cyclin-Dependent Kinase Inhibitor p27 ; biosynthesis ; genetics ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Kidney Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate the role of nitric oxide in the inhibitory effect of somatic afferent input on the pressor response caused by electrical stimulation of the paraventricular nucleus of the hypothalamus (PVN).

METHODSSD rats anesthetized by urethane were used in the study. Bipolar stainless stimulating electrode was inserted into PVN for electrical stimulation. Multi-barreled micropipettes were used for microinjection of L-NAME or normal saline into the lateral ventricle or amygdala. Deep peroneal nerve (DPN) was stimulated with electrical current pulses of 0.4 mA with duration of 0.5 ms at 4 Hz for 5 min. PVN was stimulated by electrical current pulses of 0.3 mA with duration of 0.5 ms at 80 Hz for 10 sec.

RESULTElectrical stimulation of PVN increased mean arterial pressure. Stimulation of DPN significantly inhibited the pressor response induced by stimulation of PVN (P<0.01), with the inhibitory percentage of 43.27%. Microinjection of L-NAME (0.5 mol/L,10 microl) into the lateral ventricle of brain attenuated the inhibitory effect of DPN. The inhibitory percentage decreased from 47.73% to 12.49% (P<0.05). Microinjection of L-NAME (2 mol/L,100 nl) into amygdala reduced the inhibitory effect of DPN. The inhibitory percentage of stimulating DPN on the pressor response decreased from 50.71% to 25.30% (P<0.05).

CONCLUSIONNitric oxide in the brain and amygdala are involved in the inhibitory effect of somatic afferent input on central pressor response.

Afferent Pathways ; drug effects ; physiology ; Amygdala ; drug effects ; physiology ; Animals ; Blood Pressure ; physiology ; Electric Stimulation ; Enzyme Inhibitors ; pharmacology ; NG-Nitroarginine Methyl Ester ; pharmacology ; Nitric Oxide ; antagonists & inhibitors ; physiology ; Nitric Oxide Synthase ; antagonists & inhibitors ; Paraventricular Hypothalamic Nucleus ; drug effects ; physiology ; Peroneal Nerve ; physiology ; Pressoreceptors ; drug effects ; physiology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the effect of PPAR alpha activator fenofibrate on left ventricular hypertrophy and myocardium PPAR alpha (peroxisome proliferator-activated receptor-alpha) expression in spontaneously hypertensive rats (SHR).

METHODSSixteen nine-week-old male spontaneously hypertensive rats were randomly divided into two groups: SHR received fenofibrate 100 mg x kg(-1) x d(-1) by oral gavage once daily for 8 weeks (SHR-F, n=8), and SHR received vechile (0.9 % saline) acted as controls (SHR, n=8). Age-matched Wistar-kyoto rats received vehicle for 8 weeks were served as negative controls (WKY, n=8). Systolic blood pressure was measured at the beginning, 2, 4, and 8 weeks of the experiment. At the end of the experiment, plasma BNP (brain natriuretic peptide)and lipid levels were measured. Left ventricular hypertrophy was accessed by pathological analysis. The expression of PPAR alpha and nuclear factor-kappa B (NF-kappa B p65) were investigated by the method of Western blotting.

RESULTCompared with SHR group, systolic blood pressure was slightly lowered in SHR-F group, but it didn't reach significant level(p>0.05). Fenofibrate administration lowered plasma BNP in SHR-F group (P<0.01). There were not much difference of plasma lipid levels between SHR-F and SHR group. Left ventricular mass index (assessed by left ventricular weight/body weight, g x kg(-1)), transdiameter of cardiomyocyte (TDM), cardiomyocyte area (CA), collagen volume fraction (CVF), and perivascular circumferential area (PVCA) decreased significantly in SHR-F group (P<0.05, P<0.01). The myocardium PPAR alpha expression increased significantly (P<0.01), and NF-kappa B p65 expression decreased significantly (P<0.01) in SHR-F group.

CONCLUSIONPPAR alpha activator fenofibrate can regress left ventricular hypertrophy and increase myocardium PPAR alpha expression in spontaneously hypertensive rats, which is perhaps independent of its lipid-lowering activity.

Animals ; Blood Pressure ; drug effects ; Blotting, Western ; Fenofibrate ; therapeutic use ; Hypertension ; drug therapy ; metabolism ; physiopathology ; Hypertrophy, Left Ventricular ; blood ; drug therapy ; metabolism ; Lipids ; blood ; Male ; Myocardium ; metabolism ; Natriuretic Peptide, Brain ; blood ; PPAR alpha ; biosynthesis ; Random Allocation ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; Time Factors ; Transcription Factor RelA ; biosynthesis

OBJECTIVETo screen metronidazole (MTZ) -resistance associated gene fragments of Helicobacter pylori (H.pylori) by suppression subtractive hybridization(SSH).

METHODSThe suppression subtractive hybridization (SSH) was used to screen the different DNA fragments between MTZ-resistant and -susceptible clinical strains of H.pylori. The resistant strains specific gene fragments were obtained by SSH and identified by dot blotting.

RESULTAmong the 120 subtractive colonies which were randomly chosen, 37 DNA fragments were different (>or=2 times) in DNA-copy numbers between resistant and susceptible strains and 17 of them were significantly different (>or=3 times). These 17 DNA fragments were sequenced subsequently. Ten of them were new sequences and the other 7 were duplicated sequences. These sequences represented respectively: depeptide ABC transporter periplasmic dipeptide-binding protein (dppA), permease protein (dppB), et al.

CONCLUSIONGene fragments specific to MTZ-resistant H. pylori strains were obtained by SSH and these genes may associated with MTZ-resistance of H.pylori.

Anti-Infective Agents ; pharmacology ; Cloning, Molecular ; DNA, Bacterial ; chemistry ; genetics ; Drug Resistance, Bacterial ; genetics ; Helicobacter pylori ; drug effects ; genetics ; Humans ; Metronidazole ; pharmacology ; Nucleic Acid Hybridization ; methods ; Sequence Analysis, DNA

OBJECTIVETo reconstruct the nucleotide sequence of Leptospira interrogans lipL21 gene for increasing the output of prokaryotic expression and to understand the changes on immunogenicity of the expression products before and after reconstruction, and to determine the position of envelope lipoprotein LipL21 on the surface of leptospiral body.

METHODSAccording to the preferred codons of E.coli, the nucleotide sequence of lipL21 gene was designed and synthesized, and then its prokaryotic expression system was constructed. By using SDS-PAGE plus BioRad agarose image analysor, the expression level changes of lipL21 genes before and after reconstruction were measured. A Western blot assay using rabbit anti-TR/Patoc I serum as the first antibody was performed to identify the immunoreactivity of the two target recombinant proteins rLipL21s before and after reconstruction. The changes of cross agglutination titers of antisera against two rLipL21s before and after reconstruction to the different leptospiral serogroups were demonstrated using microscope agglutination test (MAT). Immuno-electronmicroscopy was applied to confirm the location of LipL21s.

RESULTThe expression outputs of original and reconstructed lipL21 genes were 8.5 % and 46.5 % of the total bacterial proteins, respectively. Both the two rLipL21s could take place immune conjugation reaction with TR/Patoc I antiserum. After immunization with each of the two rLipL21s in rabbits, the animals could produce specific antibody. Similar MAT titers with 1:80 - 1:320 of the two antisera against rLipL21s were present. LipL21 was confirmed to locate on the surface of leptospiral envelope.

CONCLUSIONLipL21 is a superficial antigen of Leptospira interrogans. The expression output of the reconstructed lipL21 gene is remarkably increased. The expression rLipL21 maintains fine antigenicity and immunoreactivity and its antibody still shows an extensive cross immunoagglutination activity. The high expression of the reconstructed lipL21 gene will offer a favorable condition to use its product for further developing a novel universal vaccine as well as detection kit of leptospirosis.

Amino Acid Sequence ; Animals ; Antigens, Bacterial ; genetics ; immunology ; metabolism ; Bacterial Outer Membrane Proteins ; genetics ; immunology ; metabolism ; Bacterial Vaccines ; immunology ; Base Sequence ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Immune Sera ; immunology ; Leptospira interrogans ; genetics ; immunology ; ultrastructure ; Lipoproteins ; genetics ; immunology ; metabolism ; Microscopy, Immunoelectron ; Molecular Sequence Data ; Rabbits ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Sequence Analysis, DNA ; Vaccines, DNA ; immunology

OBJECTIVETo investigate the effects of focused ultrasound (FUS) treatment on cervical microenvironment in infertility women with chronic cervicitis.

METHODSA total of 10 women treated in Infertility Clinic of Obstetrics and Gynecology Hospital of College of Medicine of Zhejiang University were assessed. The women aged from 25 to 34 with normal menstrual cycles had moderate or severe cervical erosion. Ultrasound showed they had normal ovulation. The sperms of their husbands had normal motility and number. The FUS they received had a focal depth range of 4 - 6 mm, a resonant frequency of 9 - 11 MHz, and an output power of 3.5 W. The follow-up time was three months. The changes of cervical mucus and the results of postcoital test were evaluated. The clinical effects on cervical erosion and complications were also evaluated.

RESULTIn 6 cases of severe cervical erosion, 2 were cured, 2 were improved significantly (more than 50 % reduction of erosion area) and 2 were improved (less than 50 % reduction of erosion area). In 4 cases of moderate cervical erosion, 2 were cured, 2 were improved significantly. After the treatment, the median of cervical mucus score during LH peak were increased from 11 to 13 (Wilcoxon test, P=0.014). Postcoital test showed that the median of highly motile sperm (grade III and grade II) of 5 high-power field were increased from 22 to 52 (Wilcoxon test, P=0.015). Mild side effects without medical intervention included vaginal fluid in all 10 women, vaginal spotting in 2 women. No serious side effects were observed.

CONCLUSIONThe focused ultrasound therapy can help to improve the cervical microenvironment for infertility women with moderate or severe cervical erosion without serious side effects.

Adult ; Chronic Disease ; Female ; Humans ; Infertility, Female ; therapy ; Male ; Treatment Outcome ; Ultrasonic Therapy ; methods ; Uterine Cervical Erosion ; therapy ; Uterine Cervicitis ; therapy

OBJECTIVETo investigate aquaporin 9 (AQP9) mRNA and protein expression in antrum follicle and luteinizing granulosa cells of polycystic ovarian syndrome (PCOS) ovary, and its relation to follicular fluid steroids hormone levels during IVF cycles.

METHODSAQP9 mRNA expression on luteinizing granulosa cells in IVF cycles was detected by RT-PCR. AQP9 protein expression in antrum follicles of PCOS ovary and luteinizing granulosa cells was measured by immunohistochemistry. The concentrations of estradiol (E2), progesterone (P) and testerone (T) in follicular fluid were measured by radioimmunoassay (RIA).

RESULTThe expression of AQP9 mRNA in luteinizing granulosa cells during IVF cycles was positive by RT-PCR. No significant differences in AQP9 mRNA levels in granulosa cells between PCOS and control group were found during IVF cycles. The expression level of AQP9 mRNA in large follicles was higher than that in small follicles, but not significantly. The immunoreactivity for AQP9 was localized in membrane and cytoplast of granulosa cells in antrum follicles from PCOS ovary and luteinizing granulosa cells during IVF cycles. Multiple regression analysis showed that AQP9 mRNA levels on granulosa cells were not correlated with E2, P and T levels in follicular fluid during IVF cycles.

CONCLUSIONAQP9 may play an important role in the follicle development and antrum formation through water transport and AQP9 may be involved in the mechanism of follicle development in PCOS.

Adult ; Aquaporins ; biosynthesis ; genetics ; Embryo Transfer ; Female ; Fertilization in Vitro ; Follicular Fluid ; metabolism ; Granulosa Cells ; metabolism ; Humans ; Immunohistochemistry ; Infertility, Female ; etiology ; genetics ; metabolism ; Polycystic Ovary Syndrome ; complications ; genetics ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate the influence of different cycles, ovarian follicle size and IVM culture media on the number of retrieved immature oocytes, maturation rate, fertilization rate, embryo quality and implantation rate, pregnancy rate, delivery rate, survival and development of frozen-thawed embryos from IVM.

METHODSThe oocytes were obtained by follicular aspiration from 19 women undergoing oocyte retrieval for in vitro maturation due to the possible risk of ovarian hyperstimulation in IVF-ET program. One patient was in natural cycle, four patients were in ovulation induction cycles with gonadotropine and fourteen patients is controlled ovarian stimulated cycles. All the oocytes retrieved from follicles with 10.0 - 13.5 mm in maximumdiameter were allowed to culture in medium M-199 (TCM 199) or HTF supplemented with other substance.

RESULTWhen there were nonuniform diameters of follicles and the diameter of largest oocyte exceeded 12 mm, the retrieval rate of oocytes, fertilization rate, and the number of high-quality embryos decreased. The high-quality embryos formation rate was higher for the oocytes cultured in TCM 199 medium than in HTF medium (P<0.01). After being frozen-thawed, the IVM embryos could achieve the same outcome when compared with the conventional IVF treatment. In addition, the offspring were healthy.

CONCLUSIONWhen the nonuniform diameters of follicles and the diameter of largest oocyte exceeds 12 mm,the retrieval rate of oocytes, fertilization rate, and the number of high-quality embryos decreased. TCM199-based medium is better to improve the developmental potential and implantation rate of embryos derived from in vitro matured oocytes. After being frozen-thawed, the IVM embryos could achieve the same outcome when compared with the conventional IVF treatment. In addition, the offspring are healthy.

Adult ; Cryopreservation ; methods ; Embryo Transfer ; Embryo, Mammalian ; cytology ; physiology ; Female ; Fertilization in Vitro ; methods ; Humans ; Infertility, Female ; therapy ; Oocytes ; cytology ; physiology ; Pregnancy ; Pregnancy Outcome ; Pregnancy Rate

OBJECTIVETo evaluate the levels of bone morphogenetic protein-15 (BMP-15) in human follicular fluid (FF) and its association with response to ovarian stimulation.

METHODSWestern blotting was performed to determine the levels of BMP-15 in FF obtained from follicle aspirates in 70 patients undergoing IVF treatment. According to the response to ovarian stimulation the patients were divided into poor responder group and normal responder group.

RESULTBMP-15 levels in FF of poor responders were significantly higher than those in normal responders (1.01 +/- 0.34 vs 0.77 +/- 0.24, P<0.01).

CONCLUSIONIncreased levels of BMP-15 in FF may be associated with poor response to ovarian stimulation.

Adult ; Blotting, Western ; Bone Morphogenetic Protein 15 ; Female ; Follicle Stimulating Hormone ; administration & dosage ; Follicular Fluid ; drug effects ; metabolism ; Gonadotropin-Releasing Hormone ; administration & dosage ; analogs & derivatives ; Growth Differentiation Factor 9 ; Humans ; Infertility, Female ; metabolism ; Intercellular Signaling Peptides and Proteins ; biosynthesis ; Ovary ; drug effects ; metabolism ; Ovulation Induction

OBJECTIVETo investigate the expression of aquaporin 1 (AQP1) in endometrium of patients with menorrhagia.

METHODSRT-PCR and immunohistochemistry were carried out in twenty women with normal menstrual cycle to confirm the expression of AQP1 in endometrium and locate it. Then 51 women with menorrhagia and 40 women with normal menstrual cycle were included in the study. RT-PCR and immunohistochemistry were used to examine the expression of AQP1.

RESULTAQP1 mRNA was expressed in the human endometrium throughout menstruation cycle, which was mainly located in the endothelia of the capillaries and small blood vessels. Quantification of the immunostaining revealed higher density during secretary phase than that in proliferative phase (P<0.01). The staining intensity and density of AQP1-positive microvessel decreased significantly in simple hyperplasia group (P<0.01) and then gradually increased in complex hyperplasia and atypical hyperplasia groups (P<0.001).

CONCLUSIONDecreased expression of AQP1 may lead to disturbed endometrial vascular remodeling and may be involved in the occurrence of menorrhagia.

Adult ; Aquaporin 1 ; biosynthesis ; genetics ; Endometrium ; blood supply ; metabolism ; Female ; Gene Expression ; Humans ; Immunohistochemistry ; Menorrhagia ; genetics ; metabolism ; pathology ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction