Copy number variations in the human genome,one of the causes of complex diseases and genetic diseases,can lead to genomic disorders.As these diseases are difficult to diagnose,it is significantly meaningful to conduct genetic researches and molecular diagnosis.Chromosomal microarray can be used to detect copy number variations on a genome-wide scale.With the advantage of high throughput and resolution,chromosomal microarray is perceived as an important means of identifying copy number variations in genomic disorders.As technology advancements of chromosomal microarray and accumulations of clinical experiences,chromosomal microarray has played a significant role in etiological diagnosis of multiple malformations,mental retardation and autism.

Objective To strengthen the control of blood quality, ensure the safety of blood for clinical use and effectively contain the occurrence of transfusion transmitted disease. Methods Using the method of random sample survey, appraisal was made once again of the blood for clinical use in the blood stations of 14 cities(regions) of Hunan Province in 1997～2001, blood that had passed reexamination by relevant blood collecting and supplying institutions. The appraisal, conducted in the proportion of 10% of the yearly use of blood, focused on 5 indexes, including HBsAg, Anti HCV, Anti HIV, RPR, and ALT. Results 6 211 blood samples were checked, with the total positive rate being 7.89‰. Among theses, the positive rate of samples sent for testing was 7.78‰, the positive rate of samples collected and tested on the spot was 9.35‰, the positive rate of HBsAg was 2.59‰, the positive rate of Anti HCV was 3.86‰, the positive rate of Anti HIV after primary screening was 0.64‰(ascertained to be negative through Western Blot testing), the rate of ALT≥25 IU was 0.81‰, and the testing of syphilis was all negative. Conclusion Serious problems remain to be solved with regard to the safety of blood for clinical use.

Objective To establish the method of gene mutation screening for HME and investigate the relationship between genotype and clinical phenotype in HME patients. Methods Fifteen cases of HME probands were divided into the following four subgroups: mild (M) and severe ( Ⅰ S, Ⅱ S, Ⅲ S) according to the clinical diagnosis. DNA samples were obtained from the probands and family members. All of the EXT1 and EXT2 gene exons and their boundary sequences were amplified by PCR, and sequenced by directsequencing. Then the relationship between the genotypes and clinical phenotype was analyzed. Results Among the fifteen cases of HME probands, nine harbored EXT1 gene mutation, while the other 6 were positive for EXT2 gene mutation. Moreover, six novel mutations in EXT1 gene, including I8 + 2T ＞ G, c. 1182delG,c. 1108G ＞T(p. E370X) ,c. 335delA,c. 361C ＞T(p. Q121X) and c. 1879_1881delCAC were identified. In 9 patients with EXT1 gene mutation, 2 (22. 2% ) were M-type, 2 (22. 2% ) were Ⅰ S -type, 4 (44. 4% )were Ⅱ S-type,and 1 ( 11.1% ) was ⅢS-type. Whereas, 5 cases (83.3%) were M-type and only one case was Ⅱ S-type( 16. 7% ) in 6 patients with EXT2 gene mutation. Conclusions An accurate and simple gene diagnostic method for HME was established. Six novel EXT1 gene mutations, including I8 + 2T ＞ G,c. 1182delG, c. 1108G ＞T(p. E370X), c. 335delA, c. 361C ＞T(p. Q121X)and c. 1879_1881delCAC were identified as well. The clinical phenotype of the patients with EXT1 gene mutation was more severe compared to those with EXT2 gene mutations.

Objective To evaluate the value of PCR-restriction fragment length polymorphism (PCR-RFLP),real-time PCR and multiplex ligation-dependent probe amplification (MLPA) in the genetic diagnosis of spinal muscular atrophy (SMA) and make laboratory support accessible to clinicians for the molecular diagnosis of SMA.Methods Methodological evaluation.Forty-one suspected SMA cases and 359 control individuals received in Shanghai Children's Medical Centre from March 2013 to June 2014 were detected for the deletion of exon 7 and 8 in the survival motor neuron gene 1 (SMN 1) by PCR-RFLP,realtime PCR and MLPA,respectively.Then the results of the three methods were compared and the benefits and limitations of the three methods were evaluated.Results The result of real-time PCR was in complete agreement with that of MLPA,showing that 29 suspected cases harbored homozygous deletions of SMN1 and 1 case possessed heterozygous deletion.Among the homozygous deletions,27 patients demonstrated absence of exon 7 and 8,and 2 cases demonstrated only the absence of exon 7.Meanwhile,both PCR-RFLP and MLPA analysis showed the same results that only 5 out of 395 control cases carried heterozygous deletion.As for cases without heterozygous deletions,PCR-RFLP demonstrated the same result with real-time PCR and MLPA but it missed all the heterozygous ones.Conclusions PCR-RFLP,the conventional SMA gene diagnosis method,was only capable of detecting homozygous deletion of exon 7 and/or 8 of SMN1,but was not as sensitive as to find out the carriers with heterozygous deletions.MLPA could detect the deletion and quantify the copy numbers of exon 7 and 8 of SMN1,efficiently,while the price was relatively high,which brings challenges for its application in the carrier screening of SMA.Compared with these two methods,realtime PCR with high throughput and low input was a rapid,acourate and economic method for the genetic diagnosis of SMA and carrier screening in large populations.

ObjectiveTo establish an effective method for F9 gene mutation detection by using high resolution melting ( HRM ) curve analysis.Methods Peripheral blood samples of 55 hemophilia B (HB) patients were collected from Shanghai Ruijin Hospital during January 2005 to June 2010.Genomic DNA was extracted from the peripheral blood.PCR amplification combined with sequencing was used to identify the F9 gene mutations in 40 patients.HRM assay was established on the 21 DNA samples with known mutations in exonl to exon7 of F9 gene.Mutation scanning of exonl to exon7 by HRM combined with direct sequencing of exon8 was used in the molecular diagnosis of 15 HB patients with unknown F9 gene mutations.ResultsF9 gene mutation was detected in each of the 40 HB patients by direct sequencing.By HRM,the different melting curve patterns were identified in 19 out of 21 cases.The detection rate was about 90％.Through mutation scanning of exonl to exon7 by HRM combined with direct sequencing of exon8,F9 gene mutations were detected in all the 15 HB patients with unknown F9 gene mutations.Thirty-four F9 gene mutations had been identified in the 55 HB patients.ConclusionsA new strategy of HB genetic diagnosis,scanning mutations of exonl to exon7 combined with DNA sequencing of exon8 of F9 gene,is established in this study.The new strategy is efficient and reliable.

Objective To establish a method of gene mutation detection for congenital adrenal hyperplasia (CAH) by using sequencing, single nucleotide polymorphisms (SNP) analysis and T-A cloning. Methods The blood samples of 33 patients with 21-hydroxylase deficiency (21-OHD) , 2 patients with 17α-hydroxylase deficiency (17-OHD) , the parents of all the patients and 105 healthy children were collected. Genomic DNA were extracted form the blood samples. To detect the gene mutation of CYP21A2,highly specific primers for CYP21A2 gene were designed according to the sequence differences between CYP21A2 gene and its pseudogene. The whole CYP21A2 gene was amplified and sequenced. SNP analysis and TA cloning of PCR products were also carried out. The molecular diagnosis of 17-OHD was based on the amplification and sequencing of CYP17A1 gene. Results The corresponding gene mutations was determined in all the patients based on the method established in this study. Thirteen mutations of CYP21A2 gene were identified in 33 patients with 21-OHD. The 3 most frequent mutation of CYP21A2 gene were IVS2-13A/C >G, p. I172N and chimeric mutation, which accounted for 32% (21/66) ,27% (18/66) and 15% (10/66) respectively. Ninety-one persent mutations of CYP21A2 gene resulted from pseudogene conversion. In 2 patients with 17-OHD, homozygous mutations of CYP17A1 gene, IVS4-6A > G and p. 487_489del were identified separately. All the gene mutations detected in the patients were inherited from their parents. No mutation of CYP21A2 gene or CYP17A1 gene was found in the healthy children. Conclusion A method of gene mutation detection for CAH has been established. It will be beneficial to clinical diagnosis of CAH.

Objective To investigate the expression of Toll-like receptors 2 (TLR2) mRNA,p38 mitogen activated protein kinase(p38 MAPK) mRNA and cytokine in peripheral blood of children with measles and to study the effect and possible mechanism for TLR2-p38 MAPK signal pathway defects on immune suppression in the children with measles during acute phase.Methods Thirty children with measles hospitalized in the department of infectious diseases from June 2012 to July 2013 were enrolled into the measles group,and 30 healthy children were chosen as the healthy control group.The mRNA expressions of TLR and p38 MAPK in peripheral blood mononuclear cells (PBMC) were detected by reverse transcription-polymerase chain reaction (RT-PCR).The protein levels of interferon-γ (IFN-γ),tumor necrosis factor-β (TNF-β),interleukin (IL)-12,IL-6 and IL-10 in plasma were measured by using enzyme linked immunosorbent assay (ELISA),and flow cytometry (FCM)was applied to detect the percentage of lymphocyte subpopulation.The serum IgG,IgA and IgM levels were detected by velocity scatter turbidimetry.Results (1) The expressions of TLR2 mRNA and p38 MAPK mRNA in the measles group were both significantly lower than those in the healthy control group (all P ＜ 0.05).(2) Compared with the healthy control group,the protein levels of IFN-γ,TNF-β and IL-12 in the plasma of the measles group decreased significantly (all P ＜ 0.05),and the levels of IL-6 and IL-10 increased significantly(all P ＜ 0.05).(3)Compared with the healthy control group,the percentage of CD3 +,CD4 +,CD4 +/CD8 + ratio and the level of IgG and IgA in the measles group decreased significantly(all P ＜ 0.05),and the percentage of CD19 + increased significantly(P ＜ 0.05),but there was no any significant change in the percentage of CD8 + and the level of IgM (all P ＞ 0.05).Conclusions The mRNA expressions of TLR2 and p38 MAPK are low in PBMC in the measles children during acute phase.There are different degrees of suppression of cell immunity,humoral immune and cytokines disorder in children with measles.Defects of TLR2-p38 MAPK signal pathway may cause the formation of measles immune suppression.

Objective To investigate expression of 4-1BB and lymphocyte subsets in peripheral blood of children with acute infectious lymphocytosis. Methods Flow cytometry (FCM) was applied to detect the expression of 4-1BB and lymphocyte subsets in peripheral blood of 15 cases of acute infectious lymphocytosis and 20 cases of acute upper respiratory infection, and 20 healthy children. Results The expression of 4-1BB and CD3+, CD4+and CD8+lymphocytes were higher in acute infectious lymphocytosis group than those in acute upper respiratory infection group and healthy control group (P<0.05). There was no sig-niifcant difference of CD19+CD23+lymphocytes among three groups (P>0.05). A positive correlation was found between 4-1BB expression and CD3+ lymphocytes expression in acute infectious lymphocytosis group (r=0.73, P<0.05). Conclusions The abnormal expression of 4-1BB may play a pathological role in the development of acute infectious lymphocytosis. T cells in chil-dren with acute infectious lymphocytosis may not function to activate B cells.

Objective:To study the clinical effect and safety of ubenimex capsules and SOX chemotherapy on the advanced gastric cancer.Methods:90 patients with advanced gastric cancer who were treated in our hospital from September 2013 to September 2015 were selected and randomly divided into the observation group (n=45) and the control group (n=45).The patients in the control group were treated with SOX chemotherapy,while the patients in the observation group were treated with ubenimex capsules on the basis of control group.Then the serum levels of MMP-2 and MMP-9,the immune functions,the clinical efficacy,the adverse reactions and survival rate of two groups were observed and compared before and after the treatment.Results:After treatment,the CD4+,CD4+/CD8+ in the observation group were higher than those of the control group (P＜0.05);The levels of MMP2 and MMP-9 in the observation group were lower than those of the control group (P＜0.05);The total effective rate of the observation group was higher than that of the control group [68.89％(31/45) vs 48.89％(22/45)] (P＜0.05);The incidence of thrombocytopenia,leukopenia,nausea and vomiting and abnormal liver functions in the observation group was lower than that of the control group (P＜0.05);The survival rate of the observation group was higher than that of the control group at 6 months and 12 months [93.33％ (42/45) vs 77.78％ (35/45),82.22％ (37/45) vs 57.78％ (26/45)](P＜0.05).Conclusion:Compared with SOX chemotherapy alone,ubenimex capsules and SOX chemotherapy could effectively improve the immune function,enhance the long-term survival rate with high safety of patients with advanced gastric cancer.

Objective To explore the effects of empowerment education model on glycemic control and self-management behavior of community patients with diabetes. Method A total of 120 diabetics from 2 different communities were set as the observation group and control group. The control group was treated by routine medications and health education and the observation group was intervened with empowerment education model. After intervention for 6 months, the patients′blood glucose index and the diabetes self-management behavior were assessed and compared by scale of the diabetes self-care activities (SDSCA) between the two groups. Result By the end of the study, the glycemic index and SDSCA score in the former group were both significantly decreased than those in the control group (P<0.05), indicating the patients in the experiment group had better glycemic control and self-management behaviors compared with those in the control group. Conclusions Empowerment education model can arouse the internal motivation of patients to change their behaviors from passive to active acceptance in the health education. Empowerment education model can enhance diabetes patients′self-management conduct and the glycemic level.