Objective To construct 6 kinds of human soluble proliferation-inducing ligand (sAPRIL) cDNA mutants. Methods Six pair primers according to the cDNA sequence of human sAPRIL for different mutants were designed as follows: HEL Th epitope was added at C or N end, the last or first 45 bp DNA was deleted and HEL Th epitope was added, the last or first 90 bp DNA was deleted and HEL Th epitope was added. Different enzyme site was designed at 5′ end of 12 primers, BamH Ⅰ or SalⅠ or HandⅢ. At the same time, 2 pairs of HEL Th epitope sequence were designed and synthesized, and different incision enzyme sticky end sequence was added at the 5′ or 3′ end of each epitope DNA. Mutant sequences were amplified by 6 pairs primers with sAPRIL cloning plasmid as template. The PCR fragment was digested and then ligated with the HEL Th epitope DNA that had been reannealed. The ligation fragments of the head were ligated with digested pQE80 vector fragment again, and were transformed into DH5? competent cell. Recombinant plasmids were identified by digestion and sequencing. Results Six sAPRIL cDNA mutant DNA sequences were obtained by PCR, and the expression plasmids of sAPRIL cDNA mutants including HEL Th epitope sequence were constructed successfully. The result of sequencing proved that mutations generated as designed fully. Conclusion Six sAPRIL cDNA mutant expression plasmids were constructed successfully.

Objective To construct, express, purify and screen immunosuppressive molecule against human soluble APRIL (a proliferation-inducing ligand). Methods The cDNA of soluble APRIL (sAPRIL) was mutated and TEL Th epitope was added. Mutants was expressed by pQE-80L/DH5? system, identified by SDS-PAGE and Western blotting, purified by Ni-NTA resin. Their activity on stimulating the proliferation of Raji cell was detected. Results Four sAPRIL mutants were constructed and expressed as follows: HEL Th epitope was added at C end (Ⅰ); HEL Th epitope was added at N end (Ⅱ); the last 45-bp DNA was deleted at C end and HEL Th epitope was added (Ⅲ); the first 45-bp DNA was deleted at N end and HEL Th epitope was added (Ⅳ). Specific protein bands according to mutant Ⅰ-Ⅳ were detected by SDS-PAGE and Western blotting. Mutant protein was purified by Ni-NTA successfully. Mutants Ⅰand Ⅱ promoted cell proliferation remarkably, and mutant Ⅲand Ⅳnot. Conclusion Four sAPRIL mutants was constructed and expressed successfully. Immunosuppressive molecule against sAPRIL that can not promote cell proliferation was screened out, and it laid a foundation for further study on their immunosuppressive function.

To compare the differences between Hengshanhuangqi (HH) and Chuanhuangqi (CH) at molecular level, 1H NMR based plant metabolomics approach was used to reveal the chemical difference between HH and CH. Then, the contents of astragaloside IV and calycosin-7-O-beta-D-glucoside, the marker compounds specified in China Pharmacopoeia, were determined. In addition, the ITS2 fragments of HH and CH were sequenced. Twenty-three metabolites were identified in the 1H NMR spectrum, and the principal component analysis showed CH and HH could be separated clearly. HH contained more aspartic acid, GABA, citric acid, astragaloside IV and calycosin-7-O-beta-D-glucoside, while CH contained more threonine, alanine, acetic acid, choline, arginine, fructose and sucrose. And the astragaloside IV is almost undetectable in CH. In addition, the ITS2 fragment sequences of HH and CH were different at eight bases. Thus, the HH and CH showed significant differences chemically and genetically.

Objective To analyze the correlation between the tubular gastric width and the anti gastroesophageal reflux after esophageal cancer operation,and to provide reference for the choice of surgical methods in treatment of esophageal cancer.Methods Selected 60 patients who received radical surgery for esophageal carcinoma combined with gastric tube reconstruction surgery in our hospital from January 2015 to October 2015,and divided them into two groups according to the way of stomach tube anastomosis (cervical anastomosis,thoracic anastomosis) and different width of gastric tube (greater than or equal to or less than 3 cm).Namely:cervical anastomosis + greater than or equal to 3 cm group(14 cases),cervical anastomosis + less than 3 cm group(15 cases),thoracic anastomosis + greater than or equal to 3 cm group(15 cases) and thoracic anastomosis + less than 3 cm group(16 cases).All patients recieved esophageal pH monitoring for 3 days continuously from the 11 th day after operation.The monitoring indicators include:number of reflux,accumulation time of pH ＜ 4,whether there were clinical symptoms (heartburn,chest pain,pharyngeal foreign body sensation,cough,asthma,etc.) after surgery,and the frequency and time of these clinical symptoms appeared.All the patients were given endoscopic examination at the 14th days postoperatively.Observed the esophageal mucosa of patients and conducted histopathological grading of gastric mucosal inflammation.And then made a correlation analysis of gastric tube width and esophageal mucosal inflammation grade among all the patients with reflux symptoms.Results The cumulative time and number of reflux,incidence rate of clinical symptoms,and pH values less than 4 were significantly different(P ＜ 0.05).The cumulative time and number of reflux,incidence rate of clinical symptoms,and pH values less than 4 in the cervical anastomosis + less than 3 cm group were significantly lower than that in the other 3 groups(P ＜ 0.05),with statistical significance between different groups of endoscopic esophageal mucosa inflammation grade difference (P ＜ 0.05).Esophageal mucosal inflammation grading in patients of the cervical anastomosis + less than 3 cm group was the lightest.It showed a linear correlation between the gastric tube width and esophageal mucosal inflammation grading in patients with reflux symptoms.Conclusion Postoperative gastroesophageal reflux is closely related to stomach esophagus width after resection of esophageal carcinoma with tubular stomach reconstruction of stomach esophagus,because it is unable to control gastric tube width to the appropriate range.And it should be strengthened in patients with reflux related indicators for monitoring,so as to take measures to prevent gastroesophageal reflux as soon as possible to improve the prognosis of patients with quality.

Objective To explore the clinical results of total knee arthroplasty (TKA) in the treatment of knees with bony ankylosis in extension.Methods From June 2000 to June 2007,10 patients had knees with bony ankylosis in extension were treated with TKA,including 4 males and 6 females,with an average age of 49 years (range,29 to 63 years).The primary diseases were as follows:ankylosing spondylitis in 3 cases,rheumatoid arthritis in 2 cases,hemophilic arthritis in 1 care,traumatic arthritis in 3 cases and pyogenic arthritis in 1 case.The range of motion (ROM) was 0° in all knees,and the average HSS (the Hospital for Special Surgery) score before operation was 32.5±10.26.Two patients underwent bilateral TKA,and 8 underwent unilateral TKA.Results All patients were followed up for 3 to 10 years (average,5.3 years).The average HSS score was improved to 87.75±6.45 at the last follow-up,and there was significant difference compared with that before operation (t=18.668,P=0.000).The average ROM was improved to 97.08°±11.57°at the last follow-up,and there was remarkable difference compared with that before operation (t=29.063,P=0.000).The postoperative complications included cutaneous necrosis in 2 cases,deep venous thrombosis of lower limb in 1 case,periprosthetic fracture in 1 case,and deep prosthetic infection in 1 case who underwent revised TKA later.The postopertative X-ray showed no looseness of the prostheses.Conclusion TKA through medial parapatellar approach,with dissection of the rectus femoris,secondary osteotomy and soft tissue balance intraoperatively is effective in the treatment of knees with bony ankylosis in extension,which can correct the ankylosed knee deformitis successfully.Combining with the correct rehabilitation exercise,it is possible to improve significantly the function of knee and life quality in these patients.

Objective The aim of present study was to detect methylation rate of CpG unit of brain derived neurotrophic factor (BDNF) promoter and to study the epigenetic mechanism of autism spectrum disorders (ASD).Methods Total of 12 ASD patients and 12 healthy controls were recruited.The methylation rate of CpG unit in BDNF promoter Ⅰ and Ⅳ were detected using Sequenom MassArray method.The methylation model,correlationship,evolutionary relationship of CpG units in BDNF promoter Ⅰ and Ⅳ were detected and compared between ASD patients and healthy controls.Results The methylation rate was identified in 17 and 8 CpG units in BDNF promoter][and BDNF promoter Ⅳ.A close correlation distance was detected in BDNF promoter Ⅰ CpG units 4,7,10,35,and BDNF promoter Ⅳ CpG units 11.12,14.BDNF promoter][CpG units 4,7,10,35,and BDNF promoter Ⅳ CpG units 11.12,14 could be clustered.ASD patients had a significant lower methylation rate in BDNF promoter Ⅰ CpG unit 5.6 and Ⅳ CpG units 3 and 15 compare with healthy controls (P＜0.05).Conclusions The DNA methylation rate in BDNF pronoter Ⅰ CpG unit 5.6 and Ⅳ CpG units 3 and 15 may be used as potential biomarkers of ASD.

Objective:To analyze the implementation effects of a medical laboratory talent training program based on local colleges and universities' applied talent-oriented cultivation principal as well as students' interests and industry needs.Methods:Based on the design principals of clarifying the professional orientation, meeting the national standard, reconstructing the curriculum system, introducing the spirit of innovation and entrepreneurship, and multi-dimensional collaborative education, as well as the reverse design path of the outcome-based education concept, we have built a medical laboratory applied talent training system focusing on humanity education, solid foundation, broad employment, and good competency and in accordance with the "three complete education" strategy, along with measures including creating an applied teaching atmosphere, developing an applied curriculum teaching model, providing vocational guidance and improving vocational identity, and promoting education via evaluation. The system was applied to the training and practice of students of grades 2021 and 2022 majoring in medical laboratory technology. SPSS20.0 software was used for statistical analysis.Results:With the concept of application-oriented talent training and the "four-in-one" practical teaching model, students' skills were improved, and the training path was broadened. Compared with those trained with the original program (grades 2019-2020), the graduates trained with the new program (grades 2021-2022) showed a significantly decreased employment rate in medical laboratory jobs in medical institutions from 71.25% to 42.86% ( χ2=12.36, P<0.001), a significantly increased employment rate in in-vitro diagnostics industry from 3.75% to 17.14% ( χ2=7.44, P<0.05), and a significantly increased rate of applying for postgraduate education from 17.05% to 32.86% ( χ2=4.74, P<0.05). Conclusions:The medical laboratory talent training program based on the talent training principal of local colleges and universities combined with students' interests and industry needs can help improve the quality of talent training and broaden the employment path of graduates.

Myostatin, a member of the transforming growth factor beta (TGF-beta) family, is a negative regulator for muscle growth. Loss of the function of this gene is associated with the phenotype described as "double muscling", an extreme form of muscle development characterized by a large increase in muscle mass. Two replacement vectors, pA2T-Mstn4.0 and pA2T-Mstn3.2, were constructed, linearized, and transfected into the bovine fetal fibroblasts through electroporation. 170 drug-resistant cell colonies were obtained in cell culture medium containing 600 microg/mL G418 and 50 nmol/L GCV. Targeted homologous integration occurred in colony No. 58 as identified by PCR, and the targeted colony was further confirmed by sequencing and Southern blotting. This suggested that one allele of myostatin was successfully mutagenized in bovine fetal fibroblasts.
Animals ; Base Sequence ; Cattle ; China ; Electroporation ; Fetus ; Fibroblasts ; cytology ; metabolism ; Gene Targeting ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; veterinary ; Myostatin ; genetics

Objective To compare the difference of cutpoint and clinical significance of HbA1C for the diagnosis of abnormal glucose metabolism in two population groups with different ages.Methods According to oral glucose tolerance test(OGTT),the cutpoint and clinical significance of HbA1C for the diagnosis of type 2 diabetes and impaired glucose regulation(IGR)were investigated in the two population groups.Results The mean HbA1C of 1 064 young subjects in an academy and 1 671 aged subjects in a community were 5.31% ±0.41% and 5.79% ±0.71%,respectively.The cutpoints of HbA1C for diagnosis of diabetes were 5.7%(specificity 86.7%,sensitivity 66.7%)and 5.9%(specificity 73.8%,sensitivity 80.1%)in the two population groups,and 5.6% for diagnosis of IGR (specificity 82.8%,sensitivity 55.8%)and 5.7%(specificity 60.9%,sensitivity 64.3%),respectively.87.8%,78.7%,and 38.5% were diagnosed diabetes by current OGTT criteria at HbA1C levels of ≥5.7%,≥5.9%,and≥6.5%,IGR being 61.6%,39.6%,and 4.1%,and normal glucose tolerance being 24.4%,10.0%,and 0.4%.Conclusion The cutpoints of HbA1C for diagnosis of diabetes and IGR are different in populations with different ages and HbA1C levels.As one of diagnostic criteria for diabetes,HbA1C 6.5% with relatively higher specificity and lower sensitivity must be combined with fasting blood glucose,random blood glucose,and OGTT.

Objective To evaluate the change of clinicopathological features and prognosis of papillary thyroid cancer over a 15-year period.Methods The clinicopathological features and outcomes of papillary thyroid cancer patients were analyzed in three groups according to the time of diagnosis:group Ⅰ (1997-2001),group Ⅱ (2002-2006),and group Ⅲ (2007-2011).Results As time advanced,the average age of papillary thyroid cancer patients increased,tumor stage,like size,extrathyroid invasion and lymph node metastasis decreased dramatically (P ＜ 0.01).The percentage of multifocality and bilaterality increased.The long-term follow up data (median follow up time was 6.6 years),indicated that the 15-year over all survival was 97.8％ and the 15-year disease-free survival was 90.2％.Tumor ≥3 cm,bilaterality,extrathyroid invasion,lymph node metastasis and AJCC stage were correlated with tumor recurrence.By multivariate COX-regression analysis only lymph node metastasis and bilaterality were independent risk factors.Conclusion The clinicopathological features of papillary thyroid cancer changed over 15 years,with the percentage of early-staged patients increased.Lymph node metastasis and bilaterality are two risk factors for tumor recurrence.