Biomedical subjects

Hsin-Shih Wang

Publications and source records attributed to Hsin-Shih Wang.

18 recordsLinked to original sources

PubMed2006

Evaluation of MMP3 and TIMP1 as candidate genes for high myopia in young Taiwanese men.

PURPOSE: We studied the relationship between high myopia and the MMP3 and TIMP1 genes. DESIGN: Case-control study. METHODS: We enrolled 150 high myopic individuals (< or = -6 diopters) and 262 controls (> or = -1.5 diopters) initially, and another 216 cases and 474 controls were enrolled for genotyping promising polymorphisms for replication. Thirteen polymorphisms were genotyped in the initial data. Logistic regression was used to test for genetic effects. Subset analyses were performed according to the education level and family history. RESULTS: There was no significant association between any polymorphism and high myopia in the initial data. Among the highly educated subjects, the 5A/6A polymorphism at the MMP3 gene suggested a potential relationship with high myopia, and it was genotyped in the follow-up samples. However, this polymorphism failed to show any significant results in the overall subjects. CONCLUSIONS: The two genes may not play a crucial role for high myopia in young Taiwanese men.

Adolescent↗

PubMed2006

Interleukin-18 system messenger RNA and protein expression in human endometrium during the menstrual cycle.

OBJECTIVE: To investigate the IL-18 system, including IL-18, IL-18 receptor (IL-18R), IL-18 binding protein (IL-18BP), and IL-18 precursor (proIL-18), mRNA and protein expression in human endometrium during the menstrual cycle. DESIGN: Controlled study. SETTING: Clinical and academic research setting in a university medical center. PATIENT(S): Human endometrium was obtained from surgical specimens of normal cycling women undergoing hysterectomy for benign reasons. INTERVENTION(S): A total of 24 human endometrium samples were obtained at proliferative and secretory phase from surgical specimens of normal cycling women undergoing hysterectomy. Quantitative competitive polymerase chain reaction (QC PCR) and immunohistochemistry were performed. MAIN OUTCOME MEASURE(S): The differences of IL-18 system mRNA and the ratio of antagonist to agonist in both proliferative and secretory phases of endometrium were analyzed. RESULT(S): Our results showed a complete IL-18 system, including mRNA expression and protein production in both proliferative and secretory phases of the endometrium. The QC PCR demonstrated that both IL-18 and IL-18R decreased and IL-18BP increased mRNA expression in human endometrium in secretory phase compared with proliferative phase. A positive correlation of IL-18 and IL-18R in human endometrium was demonstrated in proliferative phase but not in secretory phase. In addition, there is a significantly higher ratio of IL-18BP to IL-18 mRNA levels in secretory endometrium compared with proliferative phase endometrium (P<.05). CONCLUSION(S): The expression and ratio of IL-18 antagonist to agonist may be responsible for embryo implantation.

Endometrium↗

PubMed2006

Insulin-like growth factor-II (IGF-II), IGF-binding protein-3 (IGFBP-3), and IGFBP-4 in follicular fluid are associated with oocyte maturation and embryo development.

OBJECTIVE: The objective of this study was to investigate the association between follicular fluid (FF) levels of insulin-like growth factors (IGFs), IGF binding proteins (IGFBPs), and pregnancy-associated plasma protein-A (PAPP-A), which is a protease for IGFBP-4, and the quality of subsequent embryo development from in vitro fertilized oocytes aspirated from the same follicle. DESIGN: Prospective study. SETTING: University infertility clinic and academic research laboratory. PATIENT(S): One hundred sixty-two infertile women undergoing controlled ovarian hyperstimulation for IVF and embryo transfer were recruited in a university hospital. INTERVENTION(S): During oocyte retrieval, samples of 225 FFs and matched mature oocytes were collected and studied. MAIN OUTCOME MEASURE(S): Concentrations of FF IGF-I, IGF-II, IGFBP-1, IGFBP-3, IGFBP-4, and PAPP-A were determined using ELISA. Progesterone secretion by cultured granulosa cells (GC) was measured by RIA. RESULT(S): Levels of IGF-II, IGFBP-3, and IGFBP-4 in FF on the day of oocyte retrieval were significantly correlated with embryo scores on day 3 (72 hours after oocyte retrieval). Levels of IGF-II, IGFBP-3, and IGFBP-4 in FF from follicles in which oocytes developed into day 2 embryos (48 hours after oocyte retrieval; 20 >or= embryo score >or= 6) after fertilization were significantly higher than those from follicles in which oocytes were unable to be fertilized and were arrested in embryo development on day 2, whereas the levels of PAPP-A were significantly lower in the former than the latter group. Using multiple regression analysis, we found that high levels of IGFBP-3 and IGFBP-4 combined with low levels of PAPP-A in FF were significantly correlated with successful fertilization and early development into day 2 embryos. In contrast, high FF IGFBP-1 and IGFBP-4 in combination with low FF IGF-I were significantly correlated with a later (day 2-day 3) embryo development. A significant stimulation of P secretion in cultured GCs by the combination of recombinant IGF-II, IGFBP-3, and IGFBP-4 further strengthened these proteins' functional roles in promoting late follicular development. CONCLUSION(S): High IGF-II, IGFBP-3, IGFBP-4, and low PAPP-A levels in FF at the time of oocyte retrieval suggest better oocyte maturation and early embryo development (within 48 hours after oocyte retrieval), whereas high IGFBP-1, IGFBP-4, and low FF IGF-I levels may favor later embryo development (between 48 and 72 hours after oocyte retrieval).

Cells, Cultured↗

PubMed2005

Touchdown thermocycling program enables a robust single nucleotide polymorphism typing method based on allele-specific real-time polymerase chain reaction.

Different methods have been developed for single nucleotide polymorphism (SNP) typing during recent years. Allele-specific polymerase chain reaction (ASPCR) is a cost-saving method that scores SNPs by difference of the PCR efficiency of allele-specific primers. However, ASPCR for SNP typing is notoriously confounded for its locus-specific unpredictability and the laborious gel electrophoresis. In the current study, we investigated the real-time kinetics of ASPCR and found that a simple touchdown thermocycling protocol improved its specificity significantly. Combined with real-time PCR, we developed a homogeneous genotyping method and scored more than 1000 genotypes, including all transition and transversion SNPs. A clear genotyping result was identified and validated the robustness of the method. Optimization of reactions and intrinsic modification of allele-specific primers, a laborious process but one that is repeatedly reported to be inevitable for successful ASPCR, was proved to be unnecessary with our method. Accuracy was confirmed with mass spectrometry. These characters enabled real-time ASPCR with the touchdown thermocycling protocol being very competitive among various SNP typing methods for large-scale genetic studies.

Alleles↗

PubMed2005

The genome sequence of Salmonella enterica serovar Choleraesuis, a highly invasive and resistant zoonotic pathogen.

Salmonella enterica serovar Choleraesuis (S. Choleraesuis), a highly invasive serovar among non-typhoidal Salmonella, usually causes sepsis or extra-intestinal focal infections in humans. S. Choleraesuis infections have now become particularly difficult to treat because of the emergence of resistance to multiple antimicrobial agents. The 4.7 Mb genome sequence of a multidrug-resistant S. Choleraesuis strain SC-B67 was determined. Genome wide comparison of three sequenced Salmonella genomes revealed that more deletion events occurred in S. Choleraesuis SC-B67 and S.Typhi CT18 relative to S. Typhimurium LT2. S. Choleraesuis has 151 pseudogenes, which, among the three Salmonella genomes, include the highest percentage of pseudogenes arising from the genes involved in bacterial chemotaxis signal-transduction pathways. Mutations in these genes may increase smooth swimming of the bacteria, potentially allowing more effective interactions with and invasion of host cells to occur. A key regulatory gene of TetR/AcrR family, acrR, was inactivated through the introduction of an internal stop codon resulting in overexpression of AcrAB that appears to be associated with ciprofloxacin resistance. While lateral gene transfer providing basic functions to allow niche expansion in the host and environment is maintained during the evolution of different serovars of Salmonella, genes providing little overall selective benefit may be lost rapidly. Our findings suggest that the formation of pseudogenes may provide a simple evolutionary pathway that complements gene acquisition to enhance virulence and antimicrobial resistance in S. Choleraesuis.

Bacterial Proteins↗

PubMed2005

Linear allele-specific long-range amplification: a novel method of long-range molecular haplotyping.

Haplotypes have been repeatedly shown to be more powerful than collections of single-locus markers in gene-mapping studies. Various haplotyping methods including statistical estimation are employed but molecular haplotyping, the acquisition of information directly on physical DNA sequences, has been in demand for its accuracy and independence from family pedigrees. We investigated the allelic specificity of long-range PCR, which was successful for long-range haplotyping in recent reports, and found problems of initial mispriming and crossover amplification significantly confounded its application. Based on these observations, we designed a novel method based on linear amplification of a hemizygous DNA segment with a single phosphorothioate-modified oligonucleotide. Our results revealed, with a single nucleotide polymorphism as the discriminative marker, downstream haplotypes of 14-15 kb DNA segment could be confidently scored. With two rounds of the method and five single nucleotide polymorphisms, molecular haplotypes of 29.3 kb spanning the HCR and CDSN genes, two genes associated with the susceptibility of psoriasis, of 11 members, belonging to a CEPH family, were revealed. Clear Mendelian segregation of 35 highly heterozygous SNPs confirmed the accuracy of the method. Problems of low specificity associated with long-range PCR were not observed. The simplicity, along with long-sequence accessibility and feasibility of a single nucleotide difference as the discriminative marker indicated our method holds promise for future gene-mapping studies.

Adenomatous Polyposis Coli↗

PubMed2005

Rapid detection of fetal aneuploidy using proteomics approaches on amniotic fluid supernatant.

OBJECTIVE: Conventional chromosomal studies or fluorescent in situ hybridization takes days to diagnose fetal aneuploidies during amniocentesis. Here, we evaluated the value of mass spectrometry-based clinical proteomics analysis on amniotic fluid supernatant (AFS) as a rapid detection of fetal aneuploidies. METHODS: Proteomics profiles generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) after fractionating samples with functionalized magnetic beads were used for differentiating 60 normal karyotypic from 20 aneuploid AFS. After the discriminating models were generated using genetic algorithm, we evaluated the clinical efficacy of the models in detecting aneuploidies in two batches (each n=30) of AFS prior to the release of chromosomal diagnoses. RESULTS: Within hours, the two-step proteomics analysis of AFS with the C18 model, followed by the weak cation exchange model, was able to detect aneuploid AFS at 3.3% disease prevalence rate with 100% sensitivity, 72 to 96% specificity, 11 to 50% positive predictive value, and 100% negative predictive value. CONCLUSION: Clinical proteomics analysis of AFS using magnetic beads-based sample preparation and MALDI-TOF-MS can be used as a rapid detection for fetal aneuploidies. With perfect sensitivity and negative predictive value of the two-step proteomics method, it may be used for rapid detection of aneuploid AFS immediately after amniocentesis. Further large-scale examinations are apparently needed to verify the clinical value of this rapid detection.

Amniotic Fluid↗

PubMed2004

Evaluation of meiotic spindles in thawed oocytes after vitrification using polarized light microscopy.

OBJECTIVE: To investigate the efficacy of the PolScope on imaging the spindle morphology in oocytes at the metaphase II stage before vitrification and after thawing. DESIGN: In vitro study. SETTING: University infertility clinic and academic research laboratory. INTERVENTION(S): Oocytes at the metaphase II stage that were obtained from superovulating mice were vitrified and then thawed. MAIN OUTCOME MEASURE(S): Morphological features of the spindle in oocytes were evaluated by both the PolScope and immunofluorescent staining. RESULT(S): Using the PolScope, the morphological features of the spindle of intact thawed oocytes were undetected by 3 hours of thawing in only 25% of cases. Most of the spindle images were recognized during the first hour of observation. Additionally, the statistical analysis of agreement of spindle morphology by both the PolScope and fluorescent staining showed a weighted Kappa value of 0.70, indicating good agreement. Oocytes with good spindle morphology verified by the PolScope before vitrification had a higher survival rate of intact oocytes after thawing compared with those with poor or undetected spindle images. CONCLUSION(S): The morphological features of the spindle in oocytes evaluated by the PolScope before freezing and after thawing are significantly correlated with those assessed by immunofluorescent staining after fixation. With the assistance of the PolScope, thawed oocytes with good spindle morphology can be verified and selected for further manipulation without fixation and staining.

Animals↗

PubMed2004

Feasibility of human telomerase reverse transcriptase mRNA expression in individual blastomeres as an indicator of early embryo development.

PURPOSE: The study was undertaken to test whether human telomerase reverse transcriptase (hTERT) transcripts in an individual blastomere could be used as an indicator for embryo development. METHODS: Group A consisted of day 3 normal cleaving embryos at 4- to 8-cell stage, which were surplus and not allocated for uterine transfer. Group B consisted of arrested or fragmented embryos at the same stage, which were considered to be compromised. After blastomere dissociation, RNA purification, reverse transcription, and hTERT PCR amplification, the amplified product was separated by 3% gel electrophoresis. RESULTS: Eighty-six (90.5%) of the 95 intact blastomeres in group A and 78 (70.9%) of the 110 blastomeres in group B demonstrated hTERT mRNA expression. The difference was statistically significant (P < 0.05, chi-square). Eight (38.1%) of the 21 embryos in group A and 22 (62.9%) of the 35 embryos in group B had at least one blastomere that did not express hTERT mRNA under this procedure. The difference was not significant (P > 0.05, chi-square). CONCLUSIONS: General hTERT mRNA transcripts can be detected in most of the individual blastomeres but cannot be used as an indicator for early embryo development. Further investigations are necessary to elucidate its clinical application.

Blastomeres↗

PubMed2003

Detection of the subendometrial vascularization flow index by three-dimensional ultrasound may be useful for predicting the pregnancy rate for patients undergoing in vitro fertilization-embryo transfer.

OBJECTIVE: To obtain quantitative data on endometrial volume and subendometrial blood flow by three-dimensional ultrasound clue to endometrial receptivity. DESIGN: A prospective, nonrandomized clinical study. SETTING: One tertiary center for assisted reproduction. PATIENT(S): Fifty-four patients aged <38 years with normal basal serum FSH level experiencing their first IVF cycle, whose uteri were morphologically normal as confirmed by 3-dimensional (3-D) ultrasound, were studied. INTERVENTION(S): Ultrasound evaluation was performed for all patients with 3-D facility by a single operator on the day of hCG administration. MAIN OUTCOME MEASURE(S): Ultrasonographic parameters and conception rates. RESULT(S): Subendometrial vascularization flow index (VFI) in predicting the pregnancy rate of IVF was superior to that using vascularization index, flow index, or endometrial volume in the receiver operating characteristics curve analysis. The best prediction rate was achieved by a VFI cutoff value of >0.24. CONCLUSION(S): The detection of subendometrial blood flow by 3-D power Doppler ultrasound may be a useful ultrasound parameter in the prediction of pregnancy rate of IVF, especially subendometrial VFI.

Adult↗

PubMed2003

Detection and treatment of mycoplasma contamination in cultured cells.

BACKGROUND: Mycoplasmas, the smallest and simplest prokaryotes that reside in endosomes of mammalian cells, are widespread contaminants found in cell cultures. About 30% of all cell cultures, varying from 15 to 80%, are reportedly contaminated with mycoplasmas. Here, we present our experience in successfully detecting and treating mycoplasmal infection in various cell lines. METHODS: The nested polymerase chain reaction (PCR) detection and microscopic examination, including phase-contrast, fluorescent, as well as differential interference contrast, were used for detecting potential mycoplasma contamination of cell lines used in our laboratory. As soon as mycoplasma was identified, antibiotic treatment was initiated. RESULTS: Mycoplasmal contamination was detected in six of 15 cell lines using the nested PCR amplification of mycoplasma DNA, which was further demonstrated using 4, 6-Diamidino-2-phenylindole (DAPI) staining and fluorescent microscopy. Alternate treatment with two antibiotics, macrolide (tiamulin) and tetracycline (minocycline), effectively eliminated mycoplasma, which was validated by both PCR and microscopic studies. CONCLUSIONS: The nested PCR using genomic DNA extracted from cultured cells as templates is a rapid and sensitive method for detecting mycoplasma contamination. Treatment with combined antibiotics can completely eradicate mycoplasmal infection from cultured cells. For the ease of use, PCR and/or DAPI staining appear suitable for detecting potential mycoplasmal contamination in laboratories that rely heavily on the cell culture system.

Anti-Bacterial Agents↗

PubMed2003

Polycystic ovary syndrome (PCOS), insulin resistance and insulin-like growth factors (IGfs)/IGF-binding proteins (IGFBPs).

Polycystic ovary syndrome (PCOS) is the most frequent androgen disorder of ovarian function. Hyperinsulinemia with insulin resistance is believed to be a key link in the enigmatic generation of the symptoms of PCOS such as anovulatory infertility and hyperandrogenism. Regression of these symptoms may be achieved by reducing the hyperinsulinemia. A growing body of evidence suggests that PCOS patients with hyperinsulinemia have a higher risk to develop diabetes mellitus, hypertension and cardiovascular disease as compared to age-matched women. Although oral contraceptives, progestins, antiandrogens, and ovulation induction agents remain standard therapies, weight loss should also be vigorously encouraged to ameliorate the metabolic consequences of PCOS. In addition, insulin-sensitizing agents are now being shown to be useful alone or combined with standard therapies to alleviate hyperinsulinemia in PCOS. Finally and most importantly, early identification of patients at risk and prompt initiation of therapies, followed by long-term surveillance and management, may promote the patient's long-term health.

Androgen Antagonists↗

PubMed2002

Resumption of meiosis-I tissue to enucleated preovulatory oocytes: a preliminary report.

PURPOSE: Ovarian tissue banking may be the best strategy to preserve female fertility. But optimal method to obtain viable mature oocytes remains challenging. In order to bypass the long in vitro oocyte growth period, we developed this study to test whether reconstruction of thawed primordial oocytes with enucleated preovulatory germinal vesicle (GV) oocytes could induce dictyate nuclei to undergo chromosomal condensation and meiotic maturation. METHODS: Isolated primordial oocytes from thawed mouse ovarian tissue were reconstructed with enucleated GV oocytes. After electrofusion and in vitro maturation, the reconstituted oocytes were assessed for first polar body extrusion, cytoskeleton configuration, and chromosome abnormalities. RESULTS: Primordial oocytes from thawed ovarian tissue showed a high survival rate. Following transfer and electrofusion, they could be fused with enucleated GV oocytes (35.6%, 36/101) and extruded a first polar body (52.8%,19/36). These mature oocytes showed a normal spindle configuration and chromosome number. CONCLUSIONS: We successfully established a mouse cell model to prove that omitting the whole growth and maturation period by transfer of primordial oocytes to developmentally older enucleated oocytes would bypass the long growth period required to the preovulatory stage. Polar body extrusion could also ensue after in vitro growth. This study provided an alternative approach for future investigations in oocyte maturation.

Animals↗

PubMed2002

The concentration of inhibin B in follicular fluid: relation to oocyte maturation and embryo development.

BACKGROUND: The objective of the present study was to investigate the correlation between inhibin B and estradiol levels in follicular fluid (FF) with the quality of subsequent embryo development from in-vitro fertilized oocytes aspirated from the same follicle. METHODS: A total of 156 infertile women undergoing controlled ovarian stimulation for IVF and embryo transfer was recruited to the present study. Prospectively, 233 FF samples and matched mature oocytes were studied. Concentrations of inhibin B and estradiol were determined by enzyme-linked immunosorbent assay (ELISA) and immunofluorometric assay (IFMA) respectively. RESULTS: Inhibin B levels in FF were significantly correlated with embryo scores on days 2 and 3 (48 and 72 h after oocyte retrieval). In contrast, both inhibin B and estradiol levels in FF were inversely related to age. Furthermore, FF inhibin B levels were inversely associated with serum FSH levels on day 3 of the menstrual cycle, which was believed to reflect the ovarian reserve. CONCLUSION: Inhibin B in FF may serve as an effective marker of follicular development and a useful predictor of quality of embryo. In addition, quality of oocyte is age-related and declines as age increases.

Adult↗

PubMed2002

Human chorionic gonadotropin-induced ovarian hyperstimulation syndrome is associated with up-regulation of vascular endothelial growth factor.

Ovarian hyperstimulation syndrome (OHSS), a life-threatening complication occurring in stimulated ovarian cycles, arises from treatment with gonadotropin for induction of follicular maturation in infertile women. Clinical characteristics of OHSS include ascites and pleural effusion induced by increased vascular permeability, where vascular endothelial growth factor (VEGF) was suspected to be the culprit. To test whether the effects of human CG (hCG) on the pathogenesis of OHSS were mediated through the VEGF produced by luteinized granulosa cells, we measured estradiol, VEGF, IGF-II levels in serum, and follicular fluid and analyzed their mRNA expression in luteinized granulosa cells obtained from 101 women (58 with OHSS and 43 controls) who underwent in vitro fertilization and embryo transfer. This study presents the first evidence that hCG up-regulated VEGF expression of granulosa cells in the OHSS, not the control groups, and that follicular VEGF worked through an autocrine mechanism using its kinase insert domain-containing receptor, not the fms-like tyrosine kinase receptor. We calculated total follicular production of VEGF, by multiplying follicular concentrations by follicular volumes, and verified that an increase in total follicular production of VEGF accounted for elevated serum levels of VEGF, which was associated with the development of OHSS. These findings demonstrate that through up-regulation of VEGF, hCG plays a significant role in the pathogenesis of OHSS.

Adult↗

PubMed2002

Use of the lambda sign in fetal reduction of dizygotic triplets after intracytoplasmic sperm injection.

With the increased use of artificial reproductive technologies, there are ever greater numbers of multifetal pregnancies. The increased incidences of monozygotic twins and triplet gestations can be attributed to several factors. It is important to differentiate the chorionicity in the management of multifetal pregnancies because monochorionic gestations lead to an increased risk of prematurity, twin-to-twin transfusion, morbidity, and mortality. In a dizygotic triplet pregnancy, increased risks of obstetric complications result from the monozygotic twins, such as twin-to-twin transfusion, as well as from the presence of triplets. Fetal reduction can lower these risks. During the first trimester, a "lambda sign" on ultrasound can differentiate between monozygotic twins and a separate fetus. In this paper, we present selective fetal reduction of 1 of the monozygotic twins in order to reduce the risks resulting from either monozygotic twins or from triplets. By sacrificing 1 monozygotic twin, we believe the quality of life of the remaining babies in this case were improved.

Adult↗

PubMed2002

Laser-assisted hatching of embryos in women of advanced age after in vitro fertilization: a preliminary report.

BACKGROUND: For infertile women aged over 37 years, failure of the zona pellucida to rupture is believed to be associated with a decreased implantation rate in in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). It has been suggested that assisted hatching of embryos will increase the pregnancy rate with IVF by means of mechanically facilitating the hatching process. METHODS: One hundred twenty cycles of IVF/ICSI in females aged over 37 years were analyzed. Women included in this study were allocated into 2 groups. In group I, embryos were cultured and transferred without laser-assisted hatching (LAH), whereas embryos of group II were examined and treated with LAH just before being transferred. Laser manipulations were performed using a 1.48-microm (infrared) diode laser (Fertilase). The laser was aimed at the zona to create openings of about 20 microm in diameter. RESULTS: The mean ages of women in groups I and II were 38.8 +/- 1.8 and 39.5 +/- 1.6 years, respectively (p = 0.17). The number of retrieved oocytes, endometrial thickness, and number and quality of transferred embryos did not significantly differ between the 2 groups. Rates of implantation (7.3% and 6.7%, respectively, p = 0.89), pregnancy (16.3% and 17.5%, respectively, p = 0.86), and early pregnancy loss did not differ between the 2 groups. CONCLUSION: Our data failed to demonstrate any benefit of LAH in improving implantation or pregnancy rates in women of advanced age, suggesting that factors other than laser drilling of the zona should be considered.

Adult↗

PubMedDate not supplied

The role of metformin in the treatment of polycystic ovary syndrome (PCOS).

Hyperinsulinemia is believed to be a key link in the enigmatic generation of the symptoms of polycystic ovarian syndrome (PCOS), which include anovulatory infertility and the skin stigmata induced by hyperandrogenism. Regression of these symptoms may be achieved by reducing the hyperinsulinemia. Metformin, an insulin-sensitizing agent, has been proven to be of clinical usefulness both in the short-term aiding of infertility treatments and, potentially, in the prevention of the long-term sequelae for patients with PCOS.

Androgen Antagonists↗