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Cui Y.,Collaborative Center for Research in Human Reproduction | Cui Y.,Peking Union Medical College | Liu L.,Tangshan College | Xia H.,Collaborative Center for Research in Human Reproduction | And 9 more authors.
Frontiers in Bioscience - Elite | Year: 2011

It is well-known that the majority of malformations found in the human population is based on complex gene-environment interactions. As an industrial chemical sodium thiosulfate (STS) is used heavily in many industries. Nevertheless, there is little known about the effects of STS on embryo development. In the present study, we have investigated the effects of STS on cardiac development in rat cardiomyocyte H9C2 cell line and chick embryos. As determined by MTT assays, the proliferation of H9C2 cells was inhibited by STS in a dose-dependent manner. Fertilized eggs injected via the yolk sac with STS at Hamburger-Hamilton (HH) stages 6, 9 and 12 showed significantly increased cardiotoxicity at HH stage 18, including cardiomyocyte apoptosis and animal mortality. Western blot analysis showed that STS significantly affected the expression of the apoptosis-related genes bcl-2, bax, and caspase-3 in a dose-dependent manner in the H9C2 cell line and in chick embryos. Dysregulation of apoptosis was correlated with embryonic heart malformations. Thus, STS may be a potent cardiac teratogen during embryo development.


Gao X.,Peking Union Medical College | Gao X.,Collaborative Center for Research in Human Reproduction | Cheng J.,Capital Medical University | Lu C.,Peking Union Medical College | And 12 more authors.
Current Eye Research | Year: 2010

Purpose: To identify the genetic defect in a four-generation Chinese family with autosomal dominant congenital nuclear cataract. Methods: Family history data were recorded. Clinical and ophthalmologic examinations were performed on family members. All the members were genotyped with microsatellite markers at loci associated with cataracts. Linkage analysis was performed after genotyping. Candidate genes were screened for mutation using direct sequencing. Results: Linkage analysis was obtained at markers D1S1653 (LOD score [Z]1.50, recombination fraction [θ]0.0) and D1S498 (LOD score Z0.90, recombination fraction [θ]0.0), which encompasses the connexin 50 gene (GJA8). Sequencing the coding regions of GJA8 revealed a novel, heterozygous c.773C>T transition that resulted in the substitution of a highly conserved serine by phenylalanine at codon 258 (S258F). Bioinformatics analysis showed that the mutation altered the hydrophobicity and secondary structure of the protein. This mutation co-segregated with the disease phenotype in all affected individuals and was not found in the unaffected family members or in 100 normal unrelated individuals. Conclusions: This study has identified a novel missense mutation located in the carboxyl terminus of GJA8 (S258F) associated with autosomal dominant nuclear cataract. © 2010 Informa Healthcare USA, Inc.


An L.-S.,National Research Institute for Family Planning | An L.-S.,Peking Union Medical College | An L.-S.,Collaborative Center for Research in Human Reproduction | Yuan X.-H.,Shaanxi Provincial Peoples Hospital | And 12 more authors.
Steroids | Year: 2012

Granulosa cells proliferate, differentiate, and undergo apoptosis throughout follicular development. Previous studies have demonstrated that stimulation of progesterone production is accompanied by caspase-3 activation. Moreover, we previously reported that arsenic enhanced caspase-3 activity coupled with progesterone production. Inhibition of caspase-3 activity can significantly inhibit progesterone production induced by arsenic or follicle-stimulating hormone (FSH). Here, we report that serum starvation induces caspase-3 activation coupled with augmentation of progesterone production. Serum starvation also increased the levels of cytochrome P450 cholesterol side chain cleavage enzyme (P450scc) and steroidogenic acute regulatory (StAR) protein, both of which may contribute to progesterone synthesis in preovulatory granulosa cells. Inhibition of caspase-3 activity resulted in a decrease in progesterone production. Deactivation of caspase-3 activity by caspase-3 specific inhibitor also resulted in decreases in P450scc and StAR expression, which may partly contribute to the observed decrease in progesterone production. Our study demonstrates for the first time that progesterone production in preovulatory granulosa cells is required for caspase-3 activation in a serum starvation model. Inhibition of caspase-3 activity can result in decreased expression of the steroidogenic proteins P450scc and StAR. Our work provides further details on the relationship between caspase-3 activation and steroidogenesis and indicates that caspase-3 plays a critical role in progesterone production by granulosa cells. © 2012 Elsevier Inc. All rights reserved.


Li F.-F.,National Research Institute for Family Planning | Li F.-F.,Peking Union Medical College | Yang M.,Capital Medical University | Ma X.,National Research Institute for Family Planning | And 6 more authors.
Current Eye Research | Year: 2010

Purpose: We sought to identify the genetic defect in a four-generation Chinese family with autosomal dominant congenital nuclear cataracts, examine the clinical features in detail and demonstrate the functional analysis of a candidate gene in the family. Methods: Family history data were recorded. Clinical and ophthalmological examinations were performed on affected and unaffected family members. All the members were genotyped with microsatellite markers at loci considered to be associated with cataracts. Two-point LOD scores were calculated using the LINKAGE program package after genotyping. A mutation was detected by dilff521229rect sequencing and verified by denaturing high-performance liquid chromatography (DHPLC). Wild-type and mutant proteins were analyzed with online softwares. Results: All affected members of this family had nuclear cataracts. Genetic analysis revealed a heterozygous previously described Arg116Cys mutation in the CRYAA gene in all of the affected members of the family but not in unaffected or 100 normal, unrelated individuals. Data generated with online software revealed that the different amino acid side chain, impact the aa116 interaction with other amino acids, thereby affecting the proteins secondary structure. Conclusions: This study identified a mutation in the CRYAA gene causing autosomal dominant nuclear cataracts and some patients show nystagmus or small blepharophimosis clinical features. These results provide evidence that CRYAA is a pathogenic gene for congenital cataracts, congenital cataracts are a clinically and genetically heterogeneous lens condition; at the same time, demonstrates a possible mechanism of action for the mutant gene. © 2010 Informa Healthcare USA, Inc.


Ma Y.,Peking Union Medical College | Ma Y.,National Research Institute for Family Planning | Wu M.,Peking Union Medical College | Wu M.,National Research Institute for Family Planning | And 18 more authors.
Food and Chemical Toxicology | Year: 2012

Selenium (Se) is an essential micronutrient, but also a potential toxin, which may be absorbed in excess. Relatively little is known about selenium embryotoxicity in zebrafish. In this study, we evaluated the effect of selenite exposure in zebrafish embryos. Selenite treatment decreased survival and resulted in abnormal development in a dose- and time-dependent manner. We observed irregular growth of neurons in selenite treated embryos, characterized by the absence of neurons in the brain, trunk and tail. Selenite exposure also induced defects in heart function, such as bradycardia and cardiac dysplasia with irregular and smaller chamber shape. In addition, selenite exposure caused ectopic cell proliferation, apoptosis, and a change in the pattern of DNA methylation. Our results suggested that supplementation with folic acid (FA) ameliorated the cardiac and neural defects in selenite-treated embryos. In conclusion, we demonstrated that selenite exposure caused cardiac and neural defects in zebrafish embryos and that folic acid protected against this embryotoxicity. It will give insight into the risk assessment and prevention of Se-mediated embryotoxicity. © 2012 Elsevier Ltd.


Guo Y.,National Research Institute for Family Planning | Guo Y.,Collaborative Center for Research in Human Reproduction | Li F.,Shenyang Pharmaceutical University | Cui F.,Henan University | And 6 more authors.
Frontiers in Bioscience - Elite | Year: 2012

Fragile X syndrome (FXS), which is the most frequently inherited mental retardation after Down syndrome, is caused by the absence of the fragile X mental retardation protein (FMRP) encoded by the fragile X mental retardation 1 (FMR1) gene. Patients with FXS can be identified by antibody tests that detect the absence of FMRP caused by loss-of-function mutations including the prevalent CGG repeat amplification in lymphocytes. Although the expression of recombinant FMRP in prokaryotic and eukaryotic expression systems has been achieved in different laboratories, the solubilization and purification of this protein is time consuming, varies with each protocol, and often results in low yield. In this study, glutathione S-transferase FMRP fusion protein (GST-FMRP) was expressed in and purified from Escherichia coli BL21(DE3) pLysS cells transformed with pGEX-6P-1 fusion expression vector containing the FMR1 cDNA. The recombinant GST-FMRP was purified on a glutathione sepharose 4B affinity column and detected using SDS-PAGE followed by western blotting with anti-FMRP antibody. This highly purified and soluble GST-FMRP protein can be very beneficial for generating anti-FMRP antibodies and developing FXS diagnosis kits.


Su D.,National Research Institute for Family Planning | Guo Y.,Capital Medical University | Li Q.,National Research Institute for Family Planning | Guan L.,National Research Institute for Family Planning | And 3 more authors.
Molecular Vision | Year: 2012

Purpose: To identify the genetic defect in a three-generation Chinese family with congenital cataracts. Methods: The phenotype of a three-generation Chinese family with congenital cataracts was recruited. Detailed family history and clinical data of the family were recorded. Candidate gene sequencing was performed to screen out the disease-causing mutation. Bioinformatics analysis was performed to predict the function of the mutant gene. Results: The phenotype of the family was identified as Y-suture cataract by using slit-lamp photography. Direct sequencing revealed a c.161G>C transversion in exon 1 of crystallin, alpha A (CRYAA). This mutation cosegregated with all affected individuals in the family and was not found in unaffected family members or in the 100 unrelated controls. Bioinformatics analysis indicated that the 54th amino acid position was highly conserved and the mutation R54P caused an increase in local hydrophobicity around the substitution site. Conclusions: This study identified a novel disease-causing mutation c.161G>C (p.R54P) in CRYAA in a Chinese family with autosomal dominant Y-suture cataracts. This is the first report relating a G→C mutation in CRYAA leading to congenital Y-suture cataract. © 2012 Molecular Vision.


Yang Z.,Capital Medical University | Su D.,National Research Institute for Family Planning | Li Q.,National Research Institute for Family Planning | Yang F.,Capital Medical University | And 4 more authors.
Molecular Vision | Year: 2012

Purpose: The purpose of this study was to identify the disease-causing mutation and the molecular phenotype that are responsible for the presence of an autosomal dominant congenital nuclear cataract disease in a Chinese family. Methods: The family history and clinical data were recorded. The patients were given a physical examination and their blood samples were collected for DNA extraction. Direct sequencing was used to detect the mutation. Transcription analysis of the mutant crystallin, beta A1 (CRYBA1/A3) gene was performed to verify whether the defective mutation had influenced the splice of the mature mRNA. Results: The phenotype of the congenital cataract in the family was identified as a nuclear cataract type, by using slitlamp photography. Direct sequencing revealed a novel mutation IVS3+2 T→G in CRYBA1/A3. This mutation cosegregated with all affected individuals in the family, but was not found in unaffected family members nor in the 100 unrelated controls. Transcription analysis of the mutant CRYBA1/A3 gene indicated that this mutation had influenced the splice of the mature mRNA. Conclusions: Our study identified a novel splice site mutation in CRYBA1/A3. This mutation was responsible for aberrant splicing of the mature mRNA and had caused the congenital nuclear cataracts in the family. This is the first report relating an IVS3+2 T→G mutation of CRYBA1/A3 to congenital cataracts. © 2012 Molecular Vision.

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