Objective To explore the method for early diagnosis of masked polycythemia vera (mPV) by comparing and analyzing the features of mPV and polycythemia vera (PV). Methods A total of 200 newly diagnosed male patients were collected based on diagnostic criteria, including 100 cases of PV and 100 cases of mPV. Erythropoietin (EPO), neutrophil alkaline phosphatase (NAP) score, bone marrow biopsy (BMB), and JAK2 V617F mutation were evaluated in all cases. After 6 months, hemoglobin (Hb) and JAK2 V617F mutation load were detected in patients without special treatment in two groups during follow-up. Results EPO, NAP score, BMB hematopoietic volume and the number of megakaryocyte had no statistical differences between PV group and mPV group [(3.4 ± 0.7) U/ml vs. (3.2±0.6) U/ml, (276±20) score vs. (278±21) score, (78±10) % vs. (76±9) %, (53±6) vs. (51±5), respectively], while JAK2 V617F mutation load in PV group was higher than that in mPV group[(89.2±9.4) % vs. (78.1±8.6) %, P<0.05]. In mPV patients without special treatment, Hb ≥185 g/L was found in 37 patients after 6 months, and the level of Hb and JAK2 V617F mutation load in these 37 patients reached (194±8) g/L and (90.7±9.1) %, respectively. Conclusions There is no significant difference in EPO, NAP score and BMB between PV and mPV, but the JAK2 V617F mutation load is different. In mPV patients without special treatment, Hb level can reach the typical PV diagnostic criteria after 6 months, meanwhile, JAK2 V617F mutation load is also increased.

Animals ; Glutamine ; pharmacology ; Kidney ; drug effects ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Malondialdehyde ; metabolism ; Rats ; Rats, Sprague-Dawley ; Serum ; metabolism ; Stress, Physiological ; Swimming ; physiology

Asthma ; diagnosis ; Child ; Humans ; Perception

OBJECTIVETo establish a one-step real-time quantitative RT-PCR assay for detecting the expression of WT1 mRNA, which allows detection of the minimal residue disease and prognostic prediction in leukemic patients.

METHODSWT1 gene fragment was amplified from the RNAs extracted from K562 cells using one-step RT-PCR. The quantitative standard were constructed by pMD 18-T vector cloning, and a Taqman-MGB fluorescent probe and a pair of primers were used to establish the one-step real-time fluorescence quantitative RT-PCR assay for WT1 gene detection. The sensitivity, repeatability and stability of this assay were evaluated and verified.

RESULTSThe sensitivity of this assay reached the 10(-4) level. The standard template of 1.0 x 10(6)-1.0 x 10(2) copies/ml were amplified by the one-step real-time fluorescence quantitative RT-PCR assay , and the Ct value was strongly correlated (r=0.998) to the logarithm of the initial template concentration. The repetition Ct value and both the inter-tube and inter-batch coefficients of variation (CV%) were less than 8%.

CONCLUSIONThe one-step real-time fluorescence quantitative RT-PCR assay has good sensitivity, repeatability and specificity, and the one-step completion of the reverse transcription and PCR processes may reduce the operational complexities and the possibility of contamination.

Humans ; K562 Cells ; Leukemia ; metabolism ; RNA, Messenger ; analysis ; metabolism ; Reproducibility of Results ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Sensitivity and Specificity ; WT1 Proteins ; analysis ; metabolism

BACKGROUND:The homing ability of mesenchymal stem cells after transplantation can decrease along with culture and passage in vitro. And the decline of homing abilities can further influence the implantation of mesenchymal stem cells in the target tissue, thus seriously affecting the repair effect.
OBJECTIVE:To investigate the effect and its related mechanisms by which in vitro culture and passage affect the homing ability of mesenchymal stem cells.
METHODS:Mesenchymal stem cells were isolated from the bone marrow using Ficol density gradient centrifugation, and then purified using adhesion method. The mesenchymal stem cells were cultured into the seventh generations with the normal cultural condition, and the morphological features of the 3rd, 5th and 7th generations of mesenchymal stem cells were observed. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide chromatometry was used to detect the growth feature of the 3rd, 5th and 7th generations of mesenchymal stem cells, and the growth curve was drawn. Real-time PCR was used to detect the expression of CXCR4, CXCR6, CXCL12, CD44 in the 3rd, 5th and 7th generations of mesenchymal stem cells, 2-△△Ct was calculated to get the relative value of each target gene, and the differences in expression of CXCR4, CXCR6, CXCL12, CD44 between different generations were compared.
RESULTS AND CONCLUSION:Mononuclear cells could be obtained from the bone marrow by using Ficol density gradient centrifugation. High-purity mesenchymal stem cells could be obtained using adherent method. The ability of in vitro growing was strong, but fol owing the passage, the cellmorphology became wider and shorter. And the proliferation rate, the overal proliferated multiple and the expression of homing related factors decreased fol owing the passage. The expression of CXCR4, CXCR6, CXCL12 and CD44 of mesenchymal stem cells decreased fol owing the passage. The homing ability of mesenchymal stem cells was decreased fol owing the passage, and may be relevant with the lower expression of CXCR4, CXCR6, CXCL12 and CD44 in cultured mesenchymal stem cells.

OBJECTIVETo compare the clinical effects of circumcision and the foreskin-deglove plus shaft-fix (FDSF) procedure in the treatment of phimosis or redundant prepuce in obese adult males (body mass index [BMI] ≥ 28 kg/m²).

METHODSForty-four obese adult men with phimosis or redundant prepuce underwent circumcision (n = 24) or FDSF (n = 20) according to their own wishes. The patients in the circumcision and FDSF groups were aged (26.38 ± 4.24) and (26.90 ± 3.14) years, with BMIs of (27.77 ± 0.77) and (28.07 ± 2.28) kg/m² and penis lengths of (3.51 ± 0.46) and (3.50 ± 0.59) cm, respectively. The operations were performed under local anesthesia with lidocaine plus ropivacaine mesylate.

RESULTSThe operation time of circumcision was (28.04 ± 2.65) min and that of FDSF was (45.45 ± 3.49) min. At 6 months after surgery, normal penile erection was found in all the patients, the penis length was significantly longer in the FDSF than in the circumcision group ([5.01 ± 0.73] vs [3.70 ± 0.47] cm) , and the rate of satisfaction with penile appearance was markedly higher in the former than in the latter group (3.25 ± 0.71 vs 2.83 ± 0.56).

CONCLUSIONThe foreskin-deglove plus shaft-fix procedure under local anesthesia with lidocaine and ropivacaine mesylate may achieve desirable penile erection and appearance in the treatment of phimosis or redundant prepuce in obese adult patients.

Adult ; Amides ; Anesthetics, Local ; Body Mass Index ; Circumcision, Male ; methods ; Foreskin ; abnormalities ; surgery ; Humans ; Lidocaine ; Male ; Mesylates ; Obesity ; complications ; Operative Time ; Penile Erection ; Penis ; abnormalities ; Phimosis ; surgery

OBJECTIVETo investigate the effects of small interfering RNA (siRNA) targeting YAP on the proliferation and apoptosis of human periodontal ligament stem cells (hPDLSCs).

METHODSSynthesized sequences of siRNA were transfected into hPDLSCs by Lipofectamine™ 2000. The expression of YAP was identified by using real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. Proliferation activity was detected by using cell counting kit-8 (CCK-8). Changes in the cell cycle and apoptosis rate were detected by using flow cytometry. Results were analyzed by using SPSS 19.0, and P < 0.05 was considered statistically significant.

RESULTSExpression of YAP mRNA and protein were significantly downregulated after 48 h of transfection (P < 0.001). No obvious difference was found in the expression levels of YAP protein between 48 and 72 h, thus indicating that siRNA could inhibit the expression of YAP persistently and effectively. Proliferation activity was inhibited, and apoptosis rate was increased. Cell cycle was changed as the proportion of G₁and S phases increased (P < 0.01) and G₂ phase decreased (P < 0.05).

CONCLUSIONKnocking down YAP gene by siRNA could inhibit proliferation activity, induce apoptosis, and change the cell cycle of hPDLSCs. Thus, YAP could regulate the proliferation and apoptosis of hPDLSCs.

Adaptor Proteins, Signal Transducing ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Down-Regulation ; Humans ; Periodontal Ligament ; drug effects ; Phosphoproteins ; RNA, Messenger ; RNA, Small Interfering ; pharmacology ; Stem Cells ; drug effects ; Transfection

OBJECTIVETo test the different contrctile responses of extracellular nucleotides, such as ATP, UTP and nucleotide uridine adenosine tetraphosphate (Up4A) in gastric longitudinal muscle (LM) and circular muscle (CM). Examined the effect of P2X and P2Y receptor antagonists (in this study, we used IP5I and suramin) and cyclooxygenase inhibitor (indomethacin) on Up4A induced contractile responses in LM and CM.

METHODSThe rats were sacrificed and the stomachs were opened to gain LM and CM. Using organ bath system to assess contrctile responses of smooth muscle.

RESULTSUp4A could induce contractile responses in both CM and LM, which were similar with ATP and UTP. IP5 did not attenuate Up4A could induce contractions in both LM and CM, but suramin and indomethacin significantly inhibited Up4A contraction in CM, but not in LM.

CONCLUSIONOur results suggest that extracellular nucleosides and their inhibitors induce different responses between LM and CM.

Adenosine Triphosphate ; pharmacology ; Animals ; Dinucleoside Phosphates ; pharmacology ; Indomethacin ; Muscle Contraction ; Muscle, Smooth ; physiology ; Nucleotides ; pharmacology ; Rats ; Suramin ; Uridine Triphosphate ; pharmacology

A two-step method for the purification of blocking-type anti-human CD80 monoclonal antibody 4E5 from mouse ascites was developed using anion exchange and gel filtration in combination. The ascites was first purified by anion exchange after centrifugation and filtration. The experimental parameters of sample loading and elution were optimized. The optimized loading condition was pH 8.0,50 mmol/L Tris-HCl and satisfactory results were obtained using a 0~0.5mol/L NaCl step elution. The fraction containing the protein of interest was directly loaded on gel filtration column and eluted using a 20 mmol/L phosphate buffer at pH 7.2. The purity of the obtained monoclonal antibody was up to 95% with a recovery of 61%. The purity of mAb could efficiently inhibit the growth of Daudi cells. The amplification of the method was also studied using a Bio-Scale Q5 column and the result was satisfied.

Objective To explore the value of monitoring techniques of the external branch of superior laryngeal nerve (EBSLN) in thyroid surgery and to study its protective effects on vocal function after thyroid surgery.Methods We retrospectively analyzed the clinical data of 139 patients who underwent primary surgery of papillary thyroid cancer with neurological monitoring from Jun.2017 to Mar.2018 in the General Surgery Department of PLA General Hospital.The tumors of 31 cases were located in the upper pole of the gland and elsewhere in 108 cases.The patients' vocal function was assessed at one week and one month after surgery.The rate of EBSLN identified visually and by the intraoperative nerve monitoring (IONM) were counted.EBSLN recognition efficiency and prevalence of EBSLN damage during the operation of tumor in upper pole of thyroid and in other location were compared.Results In the 139 patients,there were 218 upper poles(218 EBSLN) treated intraoperatively,of which 145 were recognized visually (126(57.8％) confirmed by IONM,and 203(93.1％) were identified by IONM,OR=8.27(x2=59.345,P=0.00).The percentage of EBLSN located in the upper pole accurately identified by the naked eye was 20/46(43.5％) while by IONM was 43/46(93.4％).The percentage of EBSLN at the other position accurately identified visually was 106/172(61.6％),and by IONM was 160/172(93.0％).The number of visually identified cases in different locations showed significantly differences according to the chi-square test (x2=4.901,P=0.027),and no significant difference by IONM identification according to chi-square test (x2=0.012,P=0.914).Five patients had a low voice at one week postoperatively and low voice and vocalization change were not observed after one month.Conclusions IONM can effectively increase the proportion of intraoperative EBSLN identification to ensure the safety of surgery.The difficulty of visual identification of EBSLN during the surgery of tumor in upper pole is greater than that in other locations.IONM can provide more evidences for nerve protection and reduce the risk of injury.