To find a rapid and accurate genotyping method for specific non-syndromic hearing loss (NSHL)-causing gene mutations for disease diagnosis in different ethnic populations.Methods We performed a novel multiplex primer extension (PE) reaction in combination with denaturing high-performance liquid chromatography (DHPLC) to simultaneously detect and genotype the 6 most common mutations in 180 patients with NSHL (GJB2-235delC,GJB2-299delAT,PDS-A2168G,PDS IVS7-2A ＞ G,mtDNA-A1555G,and mtDNA-C1494T) in Chinese population.This method involved the amplification of the target sequence,followed by a purification step,a multiplex PE reaction,and DHPLC analysis performed on the Transgenomic Wave DNA fragment analysis system under fully-denaturing conditions.Results In a blind analysis,this technique successfully and accurately genotyped 100％ of the samples simultaneously characterized by direct sequencing.Conclusion Combination of PE and DHPLC is simple,rapid,accurate,and cost-effective for genotyping common disease-causing mutations,including substitutions,insertions,and deletions in NSHL,and may be successfully used in other genetic diseases.

Objective The aim of this study was to investigate the differences of the cell ultrastucture of normal mouse hatched blastocysts and their dormant ones cultured in vitro after freezing-thawing, and to explore whether the dor-mant embryos have a better anti-freezing shock property than the normal hatched mouse embryos .Methods By transmis-sion electron microscopy , the ultrastructure of these two types of mouse embryos was observed and analyzed .Results By comparative analysis of their ultrastructure , the results showed that the dormant embryos before freezing are being austerity and with lower energy metabolism at a ‘ground state ’ .After freezing-thawing and culture , their cellular structure seemed to be similar to that of the normal embryos cultured in vitro before freezing.However, after freezing-thawing and culture, the number of mitochondria decreased , the nuclei were loose , and their heterochromatin also increased .Conclusions From the ultrastructural observation , compared with the normal mouse hatched embryos , the cellular state of dormant mouse em-bryos after freezing-thawing is more favorable for material storage and energy metabolism , thus, indicating that they have a better anti-freezing property than normal hatched embryos .

Objective To study the cellular senescence and molecular mechanism of olaparib in MCF-7 breast cancer cells.Methods The effects of olaparib on the proliferation and migration of MCF-7 cells were detected dynamically by real-time cell analysis(RTCA)technology.The effects of olaparib on the Senescence was detected by using the senescence-associated β-galactosidase(SA-β-gal).Quantitative polymerase chain reaction was used to analyze the effects of olaparib on the expression levels of genes encoding the senescence-associated factors p16,p21,C/EBP homologous protein,interleukin(IL)-6,IL-8,plasminogen activator inhibitor 1,phosphatase and tensin homolog deleted on chromosome 10,p27,retinoblastoma gene,Ki67,and E2F1.The effects of olaparib on the expression levels of the senescence-associated proteins p21,γH2AX,pRB,cyclin D1,insulin-like growth factor binding protein 3,and Ki67 were analyzed by Western Blot.Results Olaparib inhibited the proliferation and migration and induced the senescence of MCF-7 cells.Long-term(96 h)treatment with olaparib significantly up-regulated the gene expression levels of p16,p21,p27,C/EBP homologous protein,IL-6,IL-8,plasminogen activator inhibitor 1,phosphatase and tensin homolog deleted on chromosome 10,and retinoblastoma protein(P＜0.01)and significantly down-regulated the gene expression levels of Ki67 and E2F1(P＜0.01)in MCF-7 cells.Olaparib significantly increased protein expression levels of p21,γH2AX,and insulin-like growth factor binding protein 3 in MCF-7 cells(P＜0.01,P＜0.01,P＜0.05)and significantly decreased cyclin D1,pRB,and Ki67 levels(P＜0.05,P＜0.01,P＜0.05).Conclusions Olaparib can inhibit proliferation and migration and induce senescence in MCF-7 breast cancer cells.

Objective To establish the instability model of goat cervical vertebrae, and test biomechanical stability of the novel arc cervical titanium mesh with endplate ring. Methods The anatomical data from cervical vertebrae of adult goats were measured, so as to select a new type of arch cervical titanium mesh with endplate ring which was suitable for goat cervical vertebrae. A total of 24 goats with preserved articular capsule, ligaments and intervertebral disc were randomly divided into 4 groups. Group A (n=6, normal group) didn’t receive any special treatment, while Group B (n=6, model group) received partial resection of C4 vertebrae as well as upper and lower intervertebral disc. On the basis of models in Group B, Group C (n=6, experimental group) was installed with the novel arch cervical titanium mesh and fixed by plate and screw, and Group D (n=6,control group) was installed with traditional straight titanium mesh and fixed by plate and screw. The ranges of motion (ROMs) for 4 groups of specimens during flexion, extension, left/right lateral bending, left/right rotation under 2.0 N·m load were measured, and their three-dimensional (3D) motion stability was tested. The displacement of Group C and Group D under 200 N compression force was measured, the stiffness was calculated, and the anti settlement ability of the whole specimen was tested. Results The ROMs of Group A in all directions were smaller than those of Group B,the ROMs of Group A were larger than those of Group C, and the ROMs of Group C during flexion were smaller than those of Group D (P<0.05). The stiffness of Group C was higher than that of Group D (P<0.05).The compression displacement of Group C was smaller than that of Group D(P<0.05). Conclusions Compared with the straight titanium mesh, the novel arc titanium mesh is more consistent with the physiological curvature of cervical verebrae, and has better stability than the traditional titanium mesh during the most frequent flexion activities of cervical verebrae. Moreover, compression displacement of the novel arc titanium mesh under short-term stress is smaller than that of the straight titanium mesh, and its postoperative anti-settlement is better than that of the straight titanium mesh, which is worthy of further experiment and clinical promotion.