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Abstract The direct and selective coupling of benzenes with aliphatic hydrocarbons is a promising strategy for C(sp2)-C(sp3) bond formation using readily available starting materials, yet it remains a significant challenge. In this study, we have developed a simplified photochemical system that incorporates catalytic amounts of iron(III) halides as multifunctional reagents and air as a green oxidant to address this synthetic problem. Under mild conditions, the reaction between a strong C(sp2)-H bond and a robust C(sp3)-H bond has been achieved, affording a broad range of cross-coupling products with high yields and commendable chemo-, site-selectivity. The iron halide acts as a multifunctional reagent that responds to visible light, initiates C-centered radicals, induces single-electron oxidation to carbocations, and participates in a subsequent Friedel-Crafts-type process. The gradual release of radical species and carbocation intermediates appears to be critical for achieving desirable reactivity and selectivity. This eco-friendly, cost-efficient approach offers access to various building blocks from abundant hydrocarbon feedstocks, and demonstrates the potential of iron halides in sustainable synthesis.

Program synthesis means that the computer automatically constructs code that conforms to the specified grammar and user’s given specifications.Programming by examples is a kind of paradigm in program synthesis that takes input and output examples as the specifications.It has high usability and low learning cost.In recent years,it has been successfully implemented in applications such as data wrangling and string transformation,and has great potential for development.There are two main pro-blems to be solved in programming by examples.One is the problem of efficient search in huge program space,and the other is the problem of ambiguity in solutions.To solve the first problem,a method for programming by examples needs to select the appropriate domain-specific language and formulate the search algorithm when specifying the searching strategy.The applied searching algorithm can be classified into a rule-based algorithm and an algorithm based on statistical model.To solve the second problem,a method for programming by examples needs to formulate a sorting strategy.The applied sorting strategy can be classified into a sorting method based on given examples and a sorting method based on user interaction.This paper sorts out related literature on programming by examples in recent years,summarizes the methods and key technical points to solve the above two problems,and finally gives suggestions for future research directions in the field of programming by examples.

Neuroblastoma has obvious heterogeneity. It is one of the few undifferentiated malignant tumors that can spontaneously degenerate into completely benign tumors. However, for its high-risk type, even with various intensive treatment options, the prognosis is still unsatisfactory. At the same time, a large number of research data show that the abnormal amplification and high-level expression of the MYCN gene are positively correlated with the malignant progression, poor prognosis, and mortality of neuroblastoma. In this context, this article explores the role of the N-Myc, MYCN gene expression product on its target genes related to the cell cycle and reveals its regulatory network in promoting tumor proliferation and malignant progression. We hope it can provide ideas and direction for the research and development of drugs targeting N-Myc and its downstream target genes.

Chalcone is a common scaffold found in many biologically active compounds. The chalcone scaffold was also frequently utilized to design novel anticancer agents with potent biological efficacy. Aiming to continue the research of effective chalcone derivatives to treat cancers with potent anticancer activity, fourteen amino chalcone derivatives were designed and synthesized. The antiproliferative activity of amino chalcone derivatives was studied in vitro and 5-Fu as a control group. Some of the compounds showed moderate to good activity against three human cancer cells (MGC-803, HCT-116 and MCF-7 cells) and compound 13e displayed the best antiproliferative activity against MGC-803 cells, HCT-116 cells and MCF-7 cells with IC50 values of 1.52 μM (MGC-803), 1.83 μM (HCT-116) and 2.54 μM (MCF-7), respectively which was more potent than the positive control (5-Fu). Further mechanism studies were explored. The results of cell colony formatting assay suggested compound 10e inhibited the colony formation of MGC-803 cells. DAPI fluorescent staining and flow cytometry assay showed compound 13e induced MGC-803 cells apoptosis. Western blotting experiment indicated compound 13e induced cell apoptosis via the extrinsic/intrinsic apoptosis pathway in MGC-803 cells. Therefore, compound 13e might be a valuable lead compound as antiproliferative agents and amino chalcone derivatives worth further effort to improve amino chalcone derivatives’ potency.

Objective: To investigate the intervention effect of Buyang Huanwu Decoction on endothelial progenitor cell function (EPCs), and to explore the therapeutic mechanism of Buyang Huanwu Decoction. Methods: This research take 54 rats were divided into blank group, the control group, model group, Chinese medicine, western medicine group and combine traditional Chinese and western medicine group, tonifying Yang also five decoction and atorvastatin calcium for acute myocardial infarction (AMI) rats group intervention, by using the method of density gradient centrifugation separation training each rat endothelial progenitor cells, using tetramethyl azo salt trace enzyme reaction colorimetry, used in the determination of adhesion ability and improved Boyden chamber, the method of analysis and comparison between groups of endothelial progenitor cells proliferation, adhesion and migration. Results: In 7 days, 14 days and 4 weeks, acute myocardial infarction model of rat peripheral blood EPCs count, proliferation, migration and adhesion ability were compared with control group were significantly decline, tonifying Yang also five decoction group, western medicine control group and combine traditional Chinese and western medicine group of peripheral blood EPCs count, proliferation, migration and adhesion ability compared with model group were significantly increased (P < 0.05), and combine traditional Chinese and western medicine group is the traditional Chinese medicine and western medicine group on the improvement of the function of EPCs effect more significantly (P < 0.05). Conclusion: Statins combined with Buyang Huanwu Decoction can better improve the endothelial function of AMI rats than traditional Chinese medicine or western medicine. Further clinical studies on statins may better improve the endothelial function of AMI patients, and provide a new research entry point for improving the long-term prognosis of AMI patients. [ABSTRACT FROM AUTHOR]

Reductive coupling of o-substituted carbonyl compounds and m-substituted carbonyl compounds by the direct transfer of electron to carbonyl group respectively gave 1-(4-(1-hydroxy-1-phenylethyl/methyl)phenyl)ethanones/methanones and 2,3-bis(3-substitutedphenyl)butane/ethane-2,3-diols. 4-Methoxyacetophenone surprisingly gave 4-methoxybenzoic acid as oxidation product. Even acetophenone conjugated with alkyne group afforded interesting reductive addition product. Finally, imine also furnished reductively coupled diamino compound. Probable mechanisms for the formation of products is proposed. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is a polygenic disease and the fourth leading cause of cancer-associated mortality worldwide; however, the tumorigenesis of pancreatic cancer remains poorly understood. Research at a molecular level, which includes the exploration of biomarkers for early diagnosis and specific targets for therapy, may effectively aid in the diagnosis of pancreatic cancer in its early stages and in the development of targeted molecular-biological approaches for treatment, thus improving prognosis. By conducting expression profiling in para-carcinoma, carcinoma and relapse of human pancreatic tissues, 319 genes or transcripts with differential expression levels >3-fold between these tissue types were identified. Further analysis with Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes demonstrated that the translation, nucleus assembly processes and molecular functions associated with vitamin B6 and pyridoxal phosphate binding in pancreatic carcinoma were abnormal. Pancreatic cancer was additionally identified to be closely associated with certain autoimmune diseases, including type I diabetes mellitus and systemic lupus erythematosus. [ABSTRACT FROM AUTHOR]

In this study, we aimed to determine the effects of hepatitis B virus (HBV) infection on sperm quality and the outcome of assisted reproductive technology (ART). A total of 916 men (457 HBV-positive and 459 HBV-negative) seeking fertility assistance from January 2008 to December 2009 at the Women's Hospital in the School of Medicine at Zhejiang University were analysed for semen parameters. Couples in which the men were hepatitis B surface antigen (HBsAg)-seropositive were categorized as HBV-positive and included 587 in vitro fertilisation (IVF) and 325 intracytoplasmic sperm injection (ICSI) cycles from January 2004 to December 2009; negative controls were matched for female age, date of ova retrieval, ART approach used (IVF or ICSI) and randomized in a ratio of 1:1 according to the ART treatment cycles (587 for IVF and 325 for ICSI). HBV-infected men exhibited lower semen volume, lower total sperm count as well as poor sperm motility and morphology (P<0.05) when compared to control individuals. Rates of two-pronuclear (2PN) fertilisation, high-grade embryo acquisition, implantation and clinical pregnancy were also lower among HBV-positive patients compared to those of HBV-negative patients after ICSI and embryo transfer (P<0.05); IVF outcomes were similar between the two groups (P>0.05). Logistic regression analysis showed that HBV infection independently contributed to increased rates of asthenozoospermia and oligozoospermia/azoospermia (P<0.05) as well as decreased rates of implantation and clinical pregnancy in ICSI cycles (P<0.05). Our results suggest that HBV infection in men is associated with poor sperm quality and worse ICSI and embryo transfer outcomes but does not affect the outcome of IVF and embryo transfer. [ABSTRACT FROM AUTHOR]

We compared the social adjustment among Chinese children born after intracytoplasmic sperm injection (ICSI) vs those after in vitro fertilisation (IVF) and identify factors related to the adjustment. The social adaptation of 86 ICSI and 165 IVF conceived children of 4-6 years of age was assessed using the Infants-Junior Middle School Students' Social-Life Abilities Scale. There was no significant difference between the ICSI and IVF-conceived groups on the item of communication, self-dependence, locomotion, work skills, socialisation, self-management and total scores. Compared with routine IVF, ICSI does no harm to the social adaptation of children conceived through this technology. [ABSTRACT FROM AUTHOR]

Objective: To assess the effects of autologous mature follicular fluid (FF) on the in vitro maturation (IVM) outcome of human cumulus-removed oocytes from routine stimulated ovaries, particularly on the IVM co-culture outcome with autologous cumulus cells. Design: Prospective comparison study. Setting: A provincial reproductive medicine and research center. Patient(s): A total of 196 germinal vesicle (GV) and 192 metaphase I (MI) stage oocytes were recruited from 151 infertile women by ovarian stimulation for routine intracytoplasmic sperm injection (ICSI) program. Intervention(s): Two novel IVM systems assisted with either autologous mature FF alone or both autologous mature FF and autologous cumulus cells were developed to mature in vitro MI and GV stage oocytes, respectively. A standard IVM system was used as the control. Main Outcome Measure(s): Cumulative IVM rate of oocytes and the subsequent fertilization, cleavage, and preimplantation embryonic development. Result(s): By adding autologous mature FF to the medium of the IVM system, the nuclear maturation of human cumulus-removed oocytes was significantly promoted in both speed and rate, but not the corresponding fertilization, cleavage, and preimplantation embryonic development. Conclusion(s): Autologous mature FF favors the IVM of human cumulus-removed oocytes, including the IVM co-culture with autologous cumulus cells, mainly in nuclear, but not cytoplasmic, maturation. [Copyright &y& Elsevier]

Objective: To analyze the aberrant der(X) chromosome using conventional and molecular cytogenetic approaches in a fetus of second trimester and to discuss its clinical effect., Methods: Conventional cytogenetic procedures (GTG and CBG banding) were performed on cultured amniotic fluid cells. Three-color fluorescence in situ hybridization (FISH) consisting of X chromosome enumeration probes(CEPX), CEPY and Tel Xp/Yp was further performed to study the aberrant der(X) chromosome., Results: Der(X) was a rare X/Y translocation. The final karyotypes of the fetus was designated as: 46,X,der(X)t(X;Y)(p22.3;q11.2). ish der(X)t(X;Y)(p22.3;q11.2)(X/Ypter-, DXZ1+, DYZ1+)mat., Conclusion: The combination of FISH and conventional cytogenetic techniques is a powerful tool to determine derivative chromosome and to offer an accurate genetic counseling. Identification of Xp; Yq rearrangement can help estimate the risk of fetus abnormalities and give a more precise prognosis.

Objective: To analyze the numerical aberration rate of X, Y and chromosome 18 in sperms from an oligozoospermic male with mosaic trisomy 18 and to perform preimplantation genetic diagnosis (PGD) for the couple., Methods: G-banding and fluorescence in situ hybridization (FISH) were performed on metaphase chromosome. Sperm was analyzed in three-color FISH with a probe mixture containing CEP18, CEPY and Tel Xq/Yq. A healthy man with normal semen parameters was used as control., Results: Significant difference in the rates of disomy for chromosome 18 (0.63% vs. 0.16%) and the gonosomes (0.945% vs. 0.35%) and diploidy (0.87% vs. 0.31%) was found in the spermatozoa between the patient and the control. After four embryos were biopsied in one PGD cycle, two embryos with XY1818 and XX1818 were selected for implanting and clinical pregnancy was ongoing., Conclusion: Sperm-FISH allows further understanding of aneuploidy rate and accurate genetic counseling. FISHPGD was effective for patient with mosaic trisomy 18.

Objective: To perform genetic analysis of a complex chromosome rearrangement (CCR) 46,XY, t(3;11)(q27; q13), ins(11;3)(q13;p26p13) in an azoospermic man., Methods: Peripheral blood lymphocytes we re obtained for karyotyping, and metaphases were studied by multicolor fluorescence in situ hybridization procedure, Y chromosomal microdeletions in the azoospermia factor (AZF) region were analyzed with multiplex polymerase chain reaction., Results: The case was a complex chromosomal translocation between chromosomes 3 and 11 with four breakpoints, and accompanied with a band of chromosome 3 inserting into chromosome 11. No Y-chromosome microdeletions were identified at 6 STS sequences of the AZF loci., Conclusion: CCR can have a significant impact on male fertility. Molecular cytogenetic techniques may contribute to improving and personalizing reproductive counseling.

Objective: To analyze the sex chromosome meiotic segregation in inv(Y) patients by fluorescence in situ hybridization (FISH)., Methods: Conventional cytogenetic procedures (GTG and CBG banding) and FISH were performed on metaphase chromosome. Three-color FISH was performed on sperm samples using a probe mixture containing CEPX, Tel Xp/Yp and Tel Xq/Yq to investigate the sex chromosome segregation of five inv(Y) (p11.1q11.2) carriers. A healthy man with normal semen parameters was used as control., Results: There was no statistical difference in the abnormal sex chromosome number and recombination frequencies in each spermatozoon from the patient in comparison with that in the control., Conclusion: There was no apparent sex chromosome abnormality in the sperm of the inv(Y) (p11.1q11.2) carriers. Sperm-FISH allows further understanding of the sex chromosome segregation pattern and an accurate genetic counseling.

Objectives: To observe the genetic characteristics of chromosomes and the rates of implantation and pregnancy in couples of translocation carriers who undergo preimplantation genetic diagnosis (PGD) and to evaluate the significance of PGD in the treatment of translocation carriers., Methods: Fluorescence in situ hybridization (FISH) was performed to analyze the embryos of 12 carriers of reciprocal translocation and 22 carriers of Robertsonian translocation. The results of diagnosis and the implantation and pregnancy rates were analyzed., Results: A total of 253 embryos from 36 couples were retrieved and FISH was applied for the examination. The characteristics of chromosomes were diagnosed in 225 embryos and the rate of successful PGD was 88.9%. Fifty-eight embryos were found to have normal chromosome or balanced translocation and were transferred into the uterus. The rate of implantation was 36% (5/14) and 14% (6/44) and the rate of pregnancy was 4/9 and 26% (5/19) for carriers of Robertsonian translocation and reciprocal translocation, respectively., Conclusions: The FISH-based PGD is effective in the diagnosis of Robertsonian translocation and reciprocal translocation of embryos. It provides the possibility of a high rate of implantation and pregnancy, and avoids recurrent abortion and unwilling termination of pregnancy.

Objective: To 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., Methods: The 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., Result: When 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., Conclusion: When 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.

Objective: To 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., Methods: AQP9 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)., Result: The 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., Conclusion: AQP9 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.

Background: The impact of different ovarian stimulation (OS) protocols on endometrial receptivity remains controversial. In this study, the effects of different OS on the expression of endometrial integrin beta3 subunit and leukaemia-inhibitory factor (LIF) during the implantation window and the implantation rate in mice were investigated., Methods: Three OS protocols were used, involving either pregnant mare's serum gonadotrophin (PMSG) alone, PMSG plus GnRH agonist or PMSG plus GnRH antagonist. Uterus samples were collected at 48 h after OS or ovulation and were detected with immunohistochemistry, Western blot and RT-PCR analyses. Normal embryos at gestation day 4 were transferred into the uteri of mice in the control and OS groups., Results: All OS groups showed a significant decrease in the expression of both the endometrial integrin beta3 subunit and LIF during the implantation window and the implantation rate. Among the three OS groups, GnRH agonist-treated mice showed a higher endometrial integrin beta3 subunit and LIF expression and a higher implantation rate. No significant difference was found in the measured indices between the GnRH antagonist and PMSG groups., Conclusions: OS may inhibit the expression of endometrial integrin beta3 subunit and LIF and impair endometrial receptivity in mice. OS with GnRH agonist, but not GnRH antagonist, may partially restore the endometrial physiological secretion and improve uterine receptivity.

To investigate the expression patterns of histone acetyltransferase (GCN5) and histone deacetylase 1 (HDAC1) in preimplantation mouse embryos and the effects of in-vitro cultures on their expressions, immunocytochemistry was used to detect the expressions of GCN5 and HDAC1 in mouse embryos at the stages of two-cell, four-cell, eight-cell, morula and blastocyst in both in-vivo and invitro groups. In in-vivo group, the obvious expressions of GCN5 were observed in the cytoplasm of all kinds of mouse embryos but blastocysts. Meanwhile, HDAC1 was highly expressed in the nuclei of four and eight-cell embryos and morula but mainly seen in the cytoplasm of two-cell embryos. In blastocysts, the HDAC1 fluorescence was limited to the nuclei of trophoblast cells. In in-vitro group, there were no obvious GCN5 expressions in all kinds of embryos and the expression of HDAC1 was significantly reduced although its expression pattern was similar to that of in-vivo group. Those results showed that in -vitro culture environments could inhibit GCN5 expression and decrease the expression of HDAC1 in mouse preimplantation embryos, which might affect the correct embryonic gene expression.

Objective: To investigate protein and mRNA expression of aquaporin-1 (AQP1) in epithelial ovarian tumors and its clinic significance., Methods: The protein and mRNA expressions of AQP1 were measured by immunohistochemical technique, western blot and RT-PCR in 65 cases of epithelial ovarian tumors and 13 cases of normal ovary tissue., Results: AQP1 located in microvascular and small vessel epithelial cells. The protein and mRNA expressions of AQP1 in ovarian cancer (0.39 +/- 0.12, 0.93 +/- 0.51, respectively) and ovarian borderline tumors (0.43 +/- 0.21, 0.95 +/- 0.34, respectively) were significantly higher than that of ovarian benign tumors (0.27 +/- 0.13, 0.51 +/- 0.41, respectively; P < 0.05) and normal ovary tissue (0.24 +/- 0.13, 0.34 +/- 0.29, respectively; P < 0.05). Of all ovarian cancers, expression of AQP1 in cases with ascites more than 1000 ml (0.46 +/- 0.13, 1.25 +/- 0.57, respectively) was higher than that of ascites less than 1 approximately 499 ml (0.35 +/- 0.11, 0.75 +/- 0.45, respectively; P < 0.05)., Conclusion: Over-expression of AQP1 plays an important role in development of epithelial ovarian tumors, and may be related with formation of ascites of ovarian carcinoma.

Objective: To investigate the effects of mouse preimplantation embryos on the expressions of DNA methyltransferase 1(Dnmt1) of mouse oviduct epithelial cells., Methods: The histological location of Dnmt1 protein was detected by immunohistochemical staining and the expression levels of Dnmt1 mRNA and protein in mouse oviduct were assayed by real-time reverse transcription-PCR(RT-PCR) and Western blotting in both pregnant and pseudopregnant mice at the 2-cell, 4-cell and 8-cell stages., Results: The expressions of Dnmt1 protein were mainly located in the epithelial cells of mouse oviduct. It was found that during all three stages, the expression levels of Dnmt1 mRNA in the epithelial cells of the pregnant mice were significantly lower than those in the pseudopregnant mice (P< 0.05), and the level of Dnmt1 protein expression in the pregnant mice was significantly decreased as compared with that in pseudopregnant mice at the 4-cell stage., Conclusion: Expressions of both Dnmt1 mRNA and protein in the epithelial cells of mouse oviduct could be regulated by mouse preimplantation embryos, which might play an important role in the expression changes of some genes in oviduct epithelial cells during the preimplantation period.

Objective: To investigate the role of interleukin-17 (IL-17) in the pathogenesis of endometriosis., Methods: Enzyme-linked immunosorbent assay (ELISA) was used to detect the concentrations of IL-17 in peritoneal fluid of 36 patients with different stage endometriosis and 26 patients without endometriosis., Results: The concentrations of IL-17 in peritoneal fluid of the patients with and without endometriosis were (5.7 +/- 1.9) ng/L and (5.3 +/- 1.4) ng/L, respectively, without significant difference between the two groups (P > 0.05). According to staging criteria of r-AFS, the concentrations of peritoneal IL-17 in the patients with stage I-II endometriosis (6.4 +/- 1.7) ng/L were significantly higher than those in the patients with stage III-IV endometriosis (5.1 +/- 1.8) ng/L and in the patients without endometriosis (P < 0.05). There was no difference with regard to peritoneal IL-17 concentrations between proliferative and secretory phases in the patients with or without endometriosis (P > 0.05). The levels of peritoneal IL-17 were significantly higher in the endometriosis patients with infertility (6.4 +/- 1.8) ng/L than in the endometriosis patients without infertility (5.1 +/- 1.8) ng/L (P < 0.05)., Conclusion: IL-17 may play an important role in the pathogenesis of early endometriosis and pathophysiology of endometriosis-associated infertility.

Objective: To investigate insulin receptor (INSR) genotype exon 17 frequencies in women with polycystic ovary syndrome (PCOS) and to elucidate its role in the pathogenesis of PCOS., Methods: The study involved 33 women with PCOS and 28 healthy control women who were genotyped for polymorphism of INSR gene exon 17 by single strand conformation polymorphism (SSCP) analysis. Body mass index (BMI), insulin sensitive index (ISI), the expression of INSR beta subunit, and serum concentration of luteinizing hormone (LH), total testosterone between the genotypes were compared., Results: (1) The T-to-C mutation was observed in the INSR gene exon 17 (1008 bp). The frequency of the C/C genotype was significantly higher in patients (39%) than in the controls (11%) (P < 0.05). There was no significant difference in the distribution of genotypes between obese PCOS and non-obese PCOS, and between PCOS with insulin resistance (IR) and PCOS without IR. (2) In comparison of mutation genotype groups with wild genotype (T/T), ISI was significantly decreased (C/C genotype, P < 0.01; C/T genotype, P < 0.05), and no significant difference was observed in the other indices., Conclusions: There is polymorphism in INSR gene exon 17 in patients with PCOS. This variant leads to increased risk of IR in women with PCOS. It does not influence the expression of INSR beta subunit.

Objective: To explore the relationship between the patients' high follicle-stimulating hormone (FSH) azoospermia and microdeletions in Y chromosome., Methods: Eleven sequence tagged sites (STSs) in Yq were detected by PCR in 16 male patients' high FSH azoospermia., Results: Microdeletions were observed in 6 of 16 male patients and the deletion rate was 37.5%(6/16). Five types of microdeletions were detected: AZFc(SY152), AZFc (SY152+SY254)+AZFd (SY153), AZFc (SY152+SY254+SY255)+AZFd (SY153), AZFc (SY152+SY158+SY255)+AZFd (SY153),and AZFb (SY130)+AZFc (SY158+SY254+SY255)+AZFd (SY153) respectively., Conclusion: Microdeletion of Y chromosome was one of the important reasons of the patients' high FSH azoospermia. Before the application of assisted-reproductive technology (ART) to the patients, it is necessary to detect the microdeletions, especially AZFc and AZFd.

Objective: To perform preimplantation genetic diagnosis (PGD) of chromosome abnormality using fluorescence in-situ hybridization (FISH)., Methods: Ten couples were presented for preimplantation genetic diagnosis. They had a total of 10 oocyte pick-up cycles. The collected oocytes were inseminated by intracytoplasmic sperm injection. PGD was carried out using cleavage-stage (day 3) embryo biopsy, fluorescence in-situ hybridization, and day 4 embryo transfer., Results: Ten oocyte pick-up cycles yielded 158 oocytes. Among the 94 embryos obtained, 54 embryos were biopsied and FISH analyses were performed for 51 blastomeres. Twenty-four embryos were transferred on the fourth day. There were 4 clinical pregnancies: 3 infants have been born, and 1 couple had ectopic pregnancy., Conclusion: PGD is a valuable method to prevent the high risk of spontaneous miscarriages and conceiving chromosomally unbalanced offspring.

Aim: The human cytochrome P-450 2C18(CYP2C18) has been characterized. However, the protein has not been purified from liver and very little is known regarding the specific substrate of CYP2C18. In order to study its enzymatic activity for drug metabolism, the CYP2C18 cDNA was cloned and a stable CHL cell line expressing recombinant CYP2C18 was established., Methods: The human CYP2C18 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR) from total RNAs extracted from human liver and cloned into pGEM-T vector. The cDNA segment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9. A transgenic cell line was established by transfecting the recombinant plasmid of pREP9-CYP2C18 to Chinese hamster lung (CHL) cell. The enzyme activity of CYP2C18 catalyzing oxidation of tolbutamide to hydroxytolbutamide in postmitochondrial supernant(S9) fraction of the cell was determined by high performance liquid chromatography(HPLC)., Results: The amino acid sequence predicted from the cloned cDNA segment was identical to that of reported by Romkes et al (GenBank accession number: M61856, J05326). The S9 fraction of the established cell line metabolizes tolbutamide to hydroxytolbutamide. Tolbutamide hydroxylase activity was found to be 0.509+/-0.052 micromol x min(-1) x g(-1) S9 protein or 8.82+/-0.90 mol x min(-1) x mol(-1) CYP, but was undetectable in parental CHL cell. In addition, we have identified a CYP2C18 cDNA clone with exon 5 missing., Conclusion: The cDNA of human CYP2C18 was successfully cloned and a cell line, CHL-CYP2C18, efficiently expressing the protein of CYP2C18, was established. A spliced variant of CYP2C18 with exon 5 missing was identified in the cloning process.

OBJECTIVE: To establish a technology of preimplantation genetic diagnosis. METHODS: Intracytoplasm sperm injection and blastomere biopsy were performed on two women at the advanced age with the fallopian tube obstruction. Normal embryos were selected for embryo transfer after fluorescence in-situ hybridziation in biopsied blastomere. RESULTS: The levels of serum HCG were increased 20 days after embryo transfer and ultrasonography in 16 gestation weeks showed the fetal growth and structure are normal. CONCLUSION: Two successful clinical pregnancies achieved after preimplantation genetic diagnosis.

Aim: Using bacterial, yeast, or mammalian cell expressing a human drug metabolism enzyme would seem good way to study drug metabolism-related problems. Human cytochrome P-450 2C9(CYP2C9) is a polymorphic enzyme responsible for the metabolism of a large number of clinically important drugs. It ranks among the most important drug metabolizing enzymes in humans. In order to provide a sufficient amount of the enzyme for drug metabolic research, the CYP2C9 cDNA was cloned and expressed stably in CHL cells., Methods: After extraction of total RNA from human liver tissue, the human CYP2C9 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR), and cloned into cloning vector pGEM-T. The cDNA fragment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9. A transgenic cell line was established by transfecting the recombinant vector of pREP9-CYP2C9 into CHL cells. The enzyme activity of CYP2C9 catalyzing oxidation of tolbutamide to hydroxy tolbutamide in S9 fraction of the cell was determined by high performance liquid chromatography(HPLC)., Results: The amino acid sequence predicted from the cDNA segment was identical to that of CYP2C9*1, the wild type CYP2C9. However, there were two base differences, i.e. 21T>C, 1146C>T, but the encoding amino acid sequence was the same, L7, P382. The S9 fraction of the established cell line metabolizes tolbutamide to hydroxy tolbutamide; tolbutamide hydroxylase activity was found to be 0.465 +/- 0.109 micromol.min(-1).g(-1) S9 protein or 8.62 +/- 2.02mol.min(-1).mol(-1) CYP, but was undetectable in parental CHL cell., Conclusion: The cDNA of human CYP2C9 was successfully cloned and a cell line of CHL- CYP2C9, efficiently expressing the protein of CYP2C9, was established.