OBJECTIVE: To develop a polymerase chain reaction-sequence specific primer (PCR-SSP) method for simultaneous amplification and identification of the KIR genes among Chinese population. METHODS: Peripheral blood samples from 132 healthy donors who had given blood at Shenzhen Blood Center from January 2015 to November 2015 were selected as the study subjects. Based on the polymorphism and single nucleotide polymorphism (SNP) information of high-resolution KIR alleles in the Chinese population and the IPD-KIR database, specific primers were designed to amplify all the 16 KIR genes and the 2DS4-Normal and 2DS4-Deleted subtypes. The specificity of each pair of PCR primers was verified by using samples with known KIR genotypes. During PCR amplification of the KIR gene, co-amplification the fragment of human growth hormone (HGH) gene by multiplex PCR was used as the internal control to prevent false negative results. A total of 132 samples with known KIR genotypes were randomly selected for blind inspection to verify the reliability of the developed method. RESULTS: The designed primers can specifically amplify the corresponding KIR genes, with clear and bright bands for the internal control and KIR genes. The results of detection are fully consistent with the known results. CONCLUSION The KIR PCR-SSP method established in this study can yield accurate results for the identification of the presence of KIR genes.
Humans ; Receptors, KIR/genetics* ; Reproducibility of Results ; Polymorphism, Genetic ; Genotype ; Multiplex Polymerase Chain Reaction

Objective To observe the effectiveness and safety of Viatorr stent applicated in TIPS for treating primary hepatic carcinoma(PHC)complicated with liver cirrhosis and upper gastrointestinal hemorrhage.Methods Date of 20 PHC patients complicated with liver cirrhosis and upper gastrointestinal hemorrhage who underwent TIPS using Viatorr stent were retrospectively analyzed.The technical success rate of TIPS,clinical success rate,complications,recurrence of upper gastrointestinal hemorrhage and the patency of stent shunt 1,3,6 and 12 months after TIPS were recorded.Results TIPS was successfully completed in all 20 cases,and the technical success rate was 100%.The portal venous pressure was(37.58±7.26)mmHg before TIPS and(18.35±6.47)mmHg after TIPS,and the difference was significant(P＜0.05).No active bleeding nor declined hemoglobin was found within 72 h after TIPS,and the clinical success rate was 100%(20/20).No severe complication such as intraperitoneal hemorrhage,hemorrhage caused by hepatic carcinoma rupture nor bile leakage occurred.During the follow-up period,hepatic encephalopathy occurred in 4 cases(4/20,20.00%)but alleviated after symptomatic treatments.Recurrence of upper gastrointestinal hemorrhage was noticed in 2 cases(2/20,10.00%),while 6 patients died(6/20,30.00%).One,3,6 and 12 months after TIPS,the patency of stent shunt was 100%(20/20),100%(20/20),100%(20/20)and 95.00%(19/20),respectively.Conclusion Viatorr stent applicated in TIPS was effective and safe for treating PHC complicated with liver cirrhosis and upper gastrointestinal hemorrhage.

Objective: To determine the effect of a motor-specific neurotrophic factor, glial-derived neurotrophic factor (GDNF) on motor nerve regeneration. Methods: We used a nerve conduit filled with a fibrin-based delivery system that provided controlled release of GDNF during nerve regeneration. The motor branch of the rat femoral nerve was used to assess motor nerve regeneration across a 5-mm gap. Four experimental groups (n=5) were evaluated. These included GDNF with the fibrin-based delivery system (GDNF-DS group), fibrin alone(fibrin group), empty conduit (negative control group), and nerve isograft (positive control group). Nerves were harvested at 5 weeks for analysis by histomorphometry and electron microscopy. Results: At 5 mm distal to the conduit or isografts, the GDNF-DS group was not significantly different from the nerve isograft group in the following histomorphometric measures: total nerve fibers, percentage of neural tissue, and nerve density. The number of nerve fibers (respectively: 1 744±274 , 1 481±288) and the percentage of nerve tissue [(14.2±3.9)%, (11.0±2.2)%] in theisograft group and the GDNF-DS group were significantly higher than that in the fibrin group and the empty conduit group [(respectively: 538±93, 535±96) and the percentage of nerve tissue respectively: (4.3±1.6)%, (3.7±0.9)%]. There were no differences in fiber width among all groups. By electron microscopy, the GDNF-DS and isograft groups also demonstrated more organized nerve architecture than the fibrin alone and empty conduit groups. Conclusions The delivery of GDNF from the fibrin-based delivery system promotes motor nerve regeneration at a level similar to an isograft in the femoral motor nerve model. This study gives insight into the potential beneficial role of GDNF in the treatment of motor nerve injuries.

Objective: To explore the long-term effect of neuroendoscopy followed by radiotherapy on cystic craniopharyngiomas. Methods: Cystic craniopharyngiomas in 9 patients were treated with neuroendoscopic cyst aspiration and fenestration, followed by fractionated stereotactic radiotherapy (FSRT). The neuroendoscopic procedure focused on widening of cyst fenestration and extensive irrigation of the cyst contents. The collimator of FSRT ranged from 2.5 cm to 3.0 cm, and the target volume 1.1-43.8 cm^3, dose per fraction 1.8 Gy, total dose 50.4 Gy. Results: The median follow-up period was 72.9 months. Tumor control was achieved in 8 of 9 patients. Marked tumor volume reduction was obtained with the neuroendoscopic procedure alone at 6 months, 1 year, and 2 years. One recurrent case showed multilobulated cysts, and a second surgery was required 1 year after the treatment. Clinical symptoms such as headache and visual disruption were rapidly alleviated after the neuroendoscopic procedure. No new visual disturbances, endocrinopathy, or hypothalamic dysfunction was observed during follow up. Conclusions Stereotactic radiotherapy for cystic craniopharyngioma after endoscopic fenestration can effectively control the tumor for a long period of time, improve the clinical symptoms and avoid endocrine diseases.

Objective To explore the expression levels of chemokine (C-C motif) ligand 18 (CCL18) in human glioma tissues and its effects on the invasion,migration and proliferation ofglioma cells.Methods (1) Sixty samples were harvested from the glioma which was excised surgically and confirmed pathologically from the patients at the Department of Neurosurgery,Affiliated Hospital to Binzhou Medical College from January 2012 to December 2017.Of the samples,by the WHO grading,26 belonged to grade Ⅱ,18 to grade Ⅲ and 16 to grade Ⅳ.Ten samples of normal brain tissue were harvested as controls from the contemporary patients who underwent intracranial decompression and excision due to brain injury.CCL18 mRNA expression was determined by real-time RT-PCR and CCL18 protein expression in tumor cells by immunochemically histological staining.(2) U251 glioma cells cultured in vitro were incubated with CCL18 serum-free culture media (0 ng/mL,5 ng/mL and 10 ng/mL) for 24 h before they were subjected to Transwell,scarification and CCK-8 assays to measure cellular invasion,migration and proliferation.Results (1) The expression of CCL18 mRNA was significantly increased in the order from normal brain,glioma of grade Ⅱ,glioma of grade Ⅲ to glioma of grade Ⅳ (P＜0.05);the expression of CCL18 protein was significantly increased in the order from glioma of grade Ⅱ,glioma of grade Ⅲ to glioma of grade Ⅳ (P＜0.05).(2) The 24 h Transwell assay for invasion showed that the number of transmembrane cells was significantly increased in the order from 0 ng/mL group to 5 ng/mL group to 10 ng/mL group (43.5±8.3,202.0±18.5 and 279.7±18.6 cells) (P＜0.05).The widths of scratch (pixels) in the scarification assay for migration were 498.4±75.3,381.3±21.4 and 347.7±14.2 at 12 h,and 299.5±15.3,284.6±7.8 and 237.3±20.6 at 24 h,respectively,showing significant differences between the groups of 0 ng/mL,5 ng/mL and 10 ng/mL recombinant CCL18 (P＜0.05).The cell growth in CCK-8 assay for proliferation was 1.000±0.019,1.260±0.094 and 2.070±0.138 fold at 24 h,respectively,also showing significant differences between the groups of 0 ng/mL,5 ng/mL and 10 ng/mL recombinant CCL18 (P＜0.05).Conclusions Expression of CCL18 in glioma is associated with the malignancy of the tumor.As CCL 18 promotes invasion,migration and proliferation of glioma cells,it may be a potential biomarker for detecting and grading human glioma.

OBJECTIVETo explore the serological characteristics and molecular basis for an individual with para-Bombay phenotype.

METHODSBlood type of the proband was determined with routine serological methods. Exons 6 and 7 of the ABO gene and coding regions of the FUT1 and FUT2 genes were amplified by PCR and sequenced.

RESULTSThe para-Bombay phenotype was confirmed to be of Ah-secretion type. The genotype of the individual was determined as A102/O01. Position 328 of the FUT1 gene was mutated from A to G, resulting in replacement of Alanine (Ala) at position 110 by Threonine (Thr).

CONCLUSIONThe G to A mutation of nt328 of the FUT1 gene probably underlies the para-Bombay phenotype in this individual.

ABO Blood-Group System ; genetics ; Adult ; Alleles ; Base Sequence ; Exons ; Female ; Genotype ; Humans ; Molecular Sequence Data ; Mutation ; Point Mutation

Objective To investigate the expression of inositol requiting enzyme1 α (IRE1 α) and tumor necrosis factor receptor-associated factor 2 (TRAF2) and its significance through establishing models of intestinal ischemia reperfusion injury (IIRI) in rats.Methods According to the random number table,50 male SD rats were randomly divided into 2 groups:sham operation group (n =10) and ischemia reperfusion (I/R) group (n =40).Sham group animals underwent laparotomy.I/R group rats were subjected to occlusion of the superior mesenteric artery for 30 min;then the blood flow was restored.I/R group animals were divided into 4 subgroups:2 h,6 h,12 h,24 h according to the time of reperfusion.Eight rats were examined based on the number of live rats in each subgroup.The HE staining pathological changes in intestinal samples were observed by the light microscope.The small intestinal epithelial cell apoptosis index (AI) was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL).The expression levels of intestinal tissues tumor necrosis factor α (TNF-α) and plasma intestinal fatty acid-binding protein (Ⅰ-FABP) were detected by ELISA tests.Situ end labeling method was used to detect intestinal cell AI.Western blot was applied to investigate the expression of endoplasmic reticulum stress(ERS) proteins IRE1α,phosphorylation IREIα (p-IRE1 α) and TRAF2 in all group rats intestinal tissues.Results (1)The pathological changes showed that the intestinal injury of I/R groups was more severe than that of sham group,especially at 6 h.(2) Compared with sham group,the expression levels of TNF-α [sham group (16.41 ± 4.44)ng/ L,2 h group:(79.71 ± 8.20) ng/L,6 h group:(131.70 ± 11.59) ng/L,12 h group:(94.23 ±7.66) ng/L,24 h group:(69.78 ± 9.58) ng/L],AI[sham group:(3.93 ±0.77)％,2 h group:(16.24 ± 1.97)％,6 h group:(42.19 ±2.40)％,12 h group:(37.79 ± 2.34)％,24 h goup:(10.38 ±1.46)％] and plasma Ⅰ-FABP [sham group:(0.65 ±0.10) × 103 ng/L,2 h group:(1.47 ±0.10) ×103 ng/L,6 h group:(2.36 ±0.17) ×103 ng/L,12 h group:(37.79 ±2.34) ×103 ng/L,24 group:(l.41 ±0.09) × 103 ng/L] were higher (F =231.462,149.032,162.491,all P ＜ 0.01).(3) The expression of TRAF-2 protein and p-IRE1 α/IRE1 α could be up-regulated after IIRI (F =40.473,59.59,P ＜ 0.01).The expression of these proteins was up-regulated 2 h after reperfusion,peaking at 6-12 h reperfusion,and then decreased at 24 h,and the variation tendencies of all groups were the same.Conclusions IIRI could induce ERS,activate IRE1 α and up-regulate TRAF2.IRE1α/TRAF2 mediating ERS might be involved in regulating the cell inflammation,apoptosis and increasing intestinal permeability after IIRI.

Objective To provide reference for protection evaluation of soldiers working in a complex electromagnetic environment by investigating their knowledge on electromagnetic radiation (EMR) protection via means of a questionnaire . Methods Ninety-eight soldiers working in complex electromagnetic environments were selected by random sampling .Ques-tionnaires were designed ,involving the hazard of and protection against EMR .Then the results of the survey were analyzed . Results Ninety-four questionnaires were collected .Results showed that the soldiers had some knowledge of the difference between EMR and ionizing radiation , and the hazard of and protection against EMR , but professional training was needed . In addition, their knowledge of the hazard of and protection against EMR could be improved through education .Conclusion The knowledge about EMR is insufficient among soldiers and needs to be improved .

Objective To determine the level of five trace elements(Fe 2+ ?Cu 2+ ?Zn 2+ ?Ca 2+ ?Mg 2+ )in the spleen and changes of T lymphocyte and its subtype variations in peripheral blood from the rats infected with Toxoplasma gondii . Methods Twenty rats were randomly and equally divided into two groups: control group and experiment group. Each rat in the experiment group received an ip injection of 2 ml normal saline containing 1.5?10 6 tachyzoites of T. gondii . On the 64th day after injection of T.gondii , the changes in T lymphocytes (TL) and their subgroups, the helper T lymphocytes (Th) and the suppressor T lymphocytes(Ts) in the peripheral blood of the rats with T.gondii were determined by the assay of the lymphocytes labeled with intercellular acid ? naphthyl acetate esterase. All the rats were killed and the atomic absorption method were used for detecting the level of trace elements in the spleen tissue. Results The number of TL and Th in experiment group was significantly lower than that of control ( P

Objective To study the significance of DNA of Toxoplasma gondii in peripheral blood. Methods DNA of T.gondii in peripheral blood of 50 infected rats was detected by polymerase chain reaction. A pair of primers was designed, according to the sequence P30 gene specific to T.gondii, to amplify DNA from T.gondii by PCR. Results The primers amplified DNA specifically from T.gondii and could not amplify DNA from humans, uninfected rat and mouse and from Trichomonas vaginalis and Entamoeba histolytica. DNA of two Toxoplasma parasites was detected by 35 cycles of amplification, indicating a fair sensitivity of the PCR system. Conclusion PCR may have a value for early diagnosis of T.gondii infection in rat.