Objective To understand morphological characteristics of sinoatrial node cells(SNCs) by taking purified isolation and culture of neonatal rat's SNCs in vitro and to understand the expression characteristics of surface antigens hyperpolarization activated cyclic nucleotide gated cation channel 4(HCN4) and connexin 45(Cx45) on SNCs.Methods The SNCs were isolated from sinus node tissues removed from neonatal rats by trypsin and purified cultured with differential attachment and 5'-bromodeoxyuridine(BrdU) treatment.Results Three different types of cells were observed among the purified cultured SNCs: spindle,triangular and irregular cells.The spindle cells took up the greatest proportion,had the fastest beats,and had less developed organelle and scarce muscular fibrils.Immunofluorescence staining was positive against HCN4 and Cx45 on the spindle cells.Small amounts of HCN4 was observed on triangular cells.Conclusion ① Isolation by trypsin combined with differential attachment and BrdU treatment is a reliable approach to culturing sinoatrial node cells.② The spindle cells with the fastest beats and positive expressions of HCN4 and Cx45 may be the sinus node pacemaker cells.

AIM To study the boosting effect of Maximin 2 on expression of HLA DR and HLA ABC by cultured bladder cancer cell lines and compare with TNF ??IFN ?. METHODS Cell culture and FCM were used to dectect the effects of Maximin 2,TNF and IFN on the cell growth of three types of bladder cancer cell lines and HLA DR and HLA ABC expression under the IC 50 culture concentration. RESULTS Maximin 2 had show inhibition to cells growth under the small dosage. TNF ??IFN ? also had the effect on restrain all 3 cell lines growth. The expression of HLA DR was not any change in all 3 cell lines after being stimulated by Maximin 2?TNF ? and IFN ?, Maximin 2 and TNF ? can not change HLA ABC expression, IFN ? can up regulate HLA ABC expression. CONCLUSION Maximin 2 show the dose dependent effect on cell growth suppression; The mechanism of antitumor of Maximin 2 seems not relate to the enhanced expression of HLA DR or HLA ABC, TNF ? and IFN ? also not found effect to HLA DR expression. Since IFN ? are able to increase the expression of HLA ABC expression,enhanced recognization and cytotoxity of CTL to bladder cancer cells. It is may be one of the mechanism of antitumor of IFN ?.

Objective To study the effects of Toll-like receptor 4(TLR4) on oxidized low density lipoprotein ( ox-LDL) induced macrophage apoptosis and its possible mechanism .Methods THP-1 derived macrophages were divided into four groups including untreated control group , ox-LDL treated group , ox-LDL+LPS treated group and tunicamycin treated group .MTT assay and flow cytometry analysis were performed to measure cell vitality and cell apoptosis , respectively .Oil red O staining was used to observe the phagocytosis of lipids by macrophages .The persistent and intense endoplasmic reticulum ( ER) stress markers were de-tected by analyzing the expression of glucose-regulated protein 78 ( GRP78 ) and CCAAT/enhancer-binding protein homologous protein ( CHOP) at mRNA and protein levels by q-RT-PCR and Western blot .Small in-terfering RNA ( siRNA) was used to silence the expression of TLR 4 to further elucidate its possible mecha-nism.Results Flow cytomotry and MTT assay showed that the number of apoptotic cells in ox-LDL+LPS treated group were increased more significantly than that in ox-LDL treated group (P<0.01), and cell apop-tosis in both two groups were greater than that in control group (P<0.01).Compared with control group, the expression of GRP78 and CHOP at mRNA and protein levels were up-regulated in ox-LDL+LPS treated group and ox-LDL treated group (P<0.01), and the expression of GRP78 and CHOP in ox-LDL+LPS treated group was significantly higher than that in ox-LDL treated group (P<0.01).Silenced expression of TLR4 al-leviated the endoplasmic reticulum stress (P<0.05).Conclusion Increased expression of CHOP contribu-ted to cell apoptosis .TLR4 might promote ox-LDL induced macrophage apoptosis through accelerating endo-plasmic reticulum stress .

Objective To differential the biologic characteristics and multiple differentiation potential of mesenchymal stem cells in nucleus pulposus in (NP-MSC) scoliosis patient and patient with degenerative interverthral disc.Methods The human nucleus pulposus-mesenchymal stem cells were isolated and cultured with enzyme digestion from 2 patients of scoliosis and 2 patient with degenerative intervertbral disc separately.Cellular proliferation was detected with MTT assay and trypan blue.The immunophenotype expression of NP-MSC was detected by flow cytometry in scoliosis and degenerative group.The multiple differentiation ability of cells was assessed respectively using alizarin red dye,Oil red O dye and immunohistochemical staining.Results The primary cell morphology of scoliosis NP-MSC was the shape of spindle,while the degenerative NP-MSC was inhomogeneous.However,both of them became spindle shape after passages.The scoliosis NP-MSC was stronger than degenerative one in metabolic activity and proliferation.The percentage of positive antigen expression of CD44,CD105 and CD29 was 97％-100％ in scoliosis NP-MSC group,but 88.7％-97％ in the degenerative group.The expression of mature cell marker CD24 was negative in both groups.Furthermore,the MSC i.isolated from both groups differentiated along the osteogenic,chondrogenic but not adipogenic lineages.Conclusion The scoliosis and degenerative NP contains mesenchymal stem cells.Moreover the scoliosis NP-MSC had stronger ability of cell metabolic activity and proliferation.These cells have multiple differentiation potential with the exception of their adipogenic differentiation ability.

Objective:To analyze the molecular characteristics of Echovirus 11 (Echo11) strains isolated in Xiangyang, Hubei Province from 2016 to 2017 based on the sequences of VP1 gene.Methods:Rectal and throat swab specimens were collected from children with hand, foot and mouth disease (HFMD) in Xiangyang from 2016 to 2017. Echo11 strains were detected by real-time reverse transcriptase PCR (RT-PCR) and isolated after cultured in human rhabdosarcoma (RD) cells. The VP1 regions of Echo11 strains isolated from RD cells and the whole genomes of three representative Echo11 strains were amplified by conventional RT-PCR and the sequences were analyzed. DNAStar7.0 (MegAlign) and MEGA6.0 (Data) were used to analyze the homology and mutation sites in nucleotide and amino acid sequences. Neighbor-joining method was used to construct phylogenetic trees. Recombination analysis was performed with SimPlot software (BootScanning).Results:A total of 11 Echo11 strains were isolated from 3 494 HFMD cases, accounting for 0.31%. They were highly homologous in the VP1 gene. These strains shared 98.4%-100.0% homology in nucleotide sequences and 98.3%-100.0% homology in amino acid sequences. The homology between the 11 Echo11 strains and the prototype strain (Echo11/Gregory, X80059) was 73.9%-74.8% in nucleotide sequences and 87.7%-88.7% in amino acid sequences. All of the Echo11 strains circulating in Xiangyang were classified into lineage D, having a similarity to the strains circulating in some regions of mainland China since 2013. In multiple regions of the genome, the Echo11 strains isolated in Xiangyang were highly similar to the Henan Echo1 strains in 2010 and the Hubei Echo6 strains in 2015, suggesting there was recombination within the genome of Echo11 strains in Xiangyang.Conclusions:The Echo11 strains circulating in Xiangyang from 2016 to 2017 belonged to lineage D and were recombinant strains.

Objective To investigate the clinical significance of plasma methylated Septin 9 (SEPT9)gene test for colorectal cancer(CRC).Methods Clinical data of this retrospective study were obtained from Huashan Hospital of Fudan University(2016-2017).The subjects were divided into three groups,84 patients in CRC group,50 patients with adenoma in precancerous group,and 20 cases as healthy controls.A fluorescent PCR assay was used to analyze SEPT 9 methylation in DNA extracted from plasma. Chi square test was used for statistical analysis.Results The positive incidence of SEPT9 gene methylation in plasma was 63.1%(53/84)in CRC group,significantly higher than 10%(5/50)in precancerous group (χ2=35.993, P<0.001), and undetectable in healthy group.The sensitivity of the methylated SEPT9 gene test was 63.1%(53/84), and the sensitivity of a joint detection combined with carcinoembryonic antigen(CEA)was 75%(63/84).The receiver operating characteristic curve(ROC)showed that methylated SEPT9 gene test had 0.828 in the area under the curve(AUC),higher than 0.795 in the AUC of CEA test.In CRC patients,51.4%(19/37)in the stage Ⅰ-Ⅱand 72.3%(34/47)in the stage Ⅲ-Ⅳ were positive for methylated SEPT9 gene test(χ2=3.917, P<0.05).There were no significant differences in gender,age and primary tumor site.Conclusion The SEPT9 gene methylation in plasma is helpful for early screening for CRC,and is associated with CRC progression.

Objective:To construct a three-dimensional imaging model of the pancreatic head based on the embryological fusion plane, and to provide morphological parameters of the pancreatic head for future developments of basic and clinical researches on the pancreas.Methods:Histologic cross-sections of the pancreatic head with its adjacent structures were made from healthy cadavers. Immunohistochemical analysis of pancreatic polypeptide antibody was then performed to verify the existence and location of the embryological fusion plane reported previously. The histologically positioning method of the embryological fusion plane was then applied to the corresponding sections on computed tomography (CT). Based on the results of the above work, volunteers from the Affiliated Changzhou No. 2 People's Hospital of Nanjing Medical University were then used as research objects. A three-dimensional visualization reconstruction software was used to perform CT image-based structures to include the abdominal pancreas, dorsal pancreas head, and embryo fusion surface. Three-dimensional reconstruction of the pancreatic head, and morphological measurements of the relevant structures of the pancreatic head were then made.Results:Immunohistochemical analysis verified the existence and the position of the embryological fusion plane. The histologically positioning method was then successfully applied to the CT sections. The three-dimensional imaging model of the pancreatic head containing the embryological fusion plane, dorsal segment and ventral segment of head were built based on CT images. A total of 35 volunteers were included, including 19 males and 16 females, aged (48.26±8.26) years, and with a BMI of (22.29±1.78) kg/m 2. The morphological results showed that the volume of the pancreatic head, dorsal pancreas and abdominal pancreas were (32.80±8.15) cm 3, (22.21±6.94) cm 3, (10.59±3.87) cm 3, and the area of the embryo fusion surface was (12.46±3.20) cm 2. All volunteers were then grouped according to gender. Statistical analysis showed that there were no significant differences in the total pancreatic head volume, dorsal pancreatic head volume, abdominal pancreatic volume, and embryo fusion area among the groups ( P>0.05). Conclusions:It was feasible and practical to build a three-dimensional imaging model of the pancreatic head based on the embryological fusion plane by using a 3D computer system. This model and its morphological parameters could provide a new tool for research on pancreas.

Objective To investigate the quality of life, sexual quality of life and its influencing factors in cervical cancer patients undergoing radiotherapy. Methods Pre-menopausal patients of 87 cases were diagnosed with cervical cancer treated with radiotherapy at Peking Union Medical College Hospital from December 2013 to December 2018 were recruited in this study. The functional assessment of cancer therapy-cervix ( FACT-Cx V4. 0) and the female sexual function index ( FSFI) were evaluated. Demographic and treatment data were collected. The influencing factors were analyzed by multiple linear regression analysis. Results The total score of quality of life was 114( 100-126) , 19( 16-21) for physiology, 21( 18-24) for social/family, 19( 17-21 ) for emotion, 19 ( 14-22 ) for function and 39 ( 34-43 ) for additional attention, respectively. The end time of radiotherapy was the influencing factor of the total score ( P= 0.034 ) , physiology ( P=0) and function ( P=0.008) . Family was the influencing factor of social family dimension ( P=0.010) . The total score of FSFI was 8.0( 5.0-20.5) , 1.8( 1.2-3.0) for sexual desire, 1.5( 0-2.7) for sexual arousal, 0.3( 0-3.6) for vaginal lubrication, 0( 0-3.6) for orgasm, 3.2( 2-4) for satisfaction and 0( 0-3.6) for sexual pain, respectively. The total score was less than 27 in 97%( 84/87) of the patients and had sexual dysfunction. Age was the influencing factor of sexual arousal ( P=0.005) and vaginal lubrication ( P=0.039) . Occupation was the influencing factor of sexual pain ( P=0.004) . Conclusions The quality of life of premenopausal patients with cervical cancer is affected by multiple factors. Radiotherapy-related complications should be actively treated to improve the quality of life and sexual life of patients.

Objective: To investigate the application value of caesarean section scar or bikini line incisional approach in laparoscopic sleeve gastrectomy. Methods: The retrospective cohort study was conducted. The clinical data of 162 patients with obesity and metabolic diseases who were admitted to the First Affiliated Hospital of Jinan University between March 2018 and April 2019 were collected. There were 51 males and 111 females, aged (35±8)years, with a range from 12 to 47 years. Of 162 patients, 72 undergoing laparoscopic sleeve gastrectomy via caesarean section scar or bikini line incisional approach and 90 undergoing laparoscopic sleeve gastrectomy via traditional incisional approach were respectively allocated into concealed incision group and traditional incision group. Observation indicators: (1) surgical situations; (2) intraoperative situations; (3) postoperative situations; (4) follow-up. Follow-up using outpatient examination, telephone interview, and WeChat was performed to detect the postoperative complications at 1, 3, 6 months and 1, 2, 5 years postoperatively up to May 2019. Measurement data with normal distribution were represented as Mean±SD, and the t test was used for comparison between groups. Count data were represented as absolute numbers, and were analyzed by the chi-square test or fisher exact probability. Ordinal data were analyzed by the Wilcoxon rank sum test. Results: (1) Surgical situations: patients in the concealed incision group and traditional incision group underwent successfully laparoscopic sleeve gastrectomy, without conversion to open surgery, reoperation, or perioperative death. (2) Intraoperative situations: the operation time and volume of intraoperative blood loss of the concealed incision group were (102±17)minutes and (11±4)mL, respectively, versus (105±19)minutes and (11±4)mL of the traditional incision group (t=-1.232, -0.676, P>0.05). There were 6 cases and 21 cases with additional surgical ports during operation in the concealed incision group and traditional incision group, respectively, with a significant difference between the two groups (χ²=6.280, P<0.05). (3) Postoperative situations: in the concealed incision group, 56 patients were very satisfied with scar appearance and 16 patients were satisfied with scar appearance. In the traditional incision group, 42 patients were very satisfied with scar appearance and 48 patients were satisfied with scar appearance. There was a significant difference in the overall satisfaction with the incision scar between the two groups (Z=-4.012, P<0.05). Duration of postoperative hospital stay and total hospital expenses of the concealed incision group were (4.9±0.9)days and (64 408±5 123)yuan, versus (5.2±1.5)days and (64 953±3 477)yuan of the traditional incision group (t=-1.788, -0.804, P>0.05). There were 19 and 14 patients with gastroesophageal reflux in the concealed incision group and traditional incision group, respectively, with no significant difference between the two groups (χ²=2.894, P>0.05). There was no postoperative complication such as infection, hemorrhage or anastomotic leakage in either group. (4) Follow-up: 32 of 162 patients were lost to follow-up, including 10 in the concealed incision group and 22 in the traditional incision group; other 130 patients were followed up for 1-14 months, with a median follow-up time of 7 months. During the follow-up, 1 patient in the traditional incision group was hospitalized again at 3 months after surgery due to upper gastrointestinal bleeding, and was cured after symptomatic supportive treatment. One patient in the concealed incision group was admitted to the local hospital for functional obstruction at 1 month after surgery and was discharged after symptomatic treatment. Other patients had no readmission due to postoperative complications. Conclusion Laparoscopic sleeve gastrectomy via caesarean section scar or bikini line incision is safe and feasible, with good cosmetic effects.

Objective:To explore the serum levels of copper and zinc and the application value of the ratio in assessing disease activity in patients with inflammatory bowel disease (IBD).Methods:From March 2019 to April 2020, 200 patients with IBD hospitalized at the Department of Gastroenterology of the First Affiliated Hospital of Anhui Medical University were selected by prospective random direct sampling method, including 100 patients with Crohn′s disease (CD) and 100 patients with ulcerative colitis (UC). The Crohn′s disease activity index (CDAI) and the modified Mayo score were used to evaluate the disease activity of CD patients and UC patients. In the same period 100 healthy individuals in the routine physical examination were selected as healthy control group. The serum levels of copper and zinc of the healthy control group, the CD group and the UC group were determined by atomic absorption spectrometry. The levels and the ratio of serum copper to zinc of three groups were compared. The ratio of serum copper to zinc of CD patients and UC patients with different disease activity were compared. The correlation between the ratio of serum copper to zinc and IBD activity indexes were analyzed, which included fecal calprotectin (FC), C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), CDAI and Mayo score. Receiver operating characteristic curve was drawn to analyze the value of the ratio of serum copper to zinc, CRP and ESR in predicting disease activity of patients with IBD. Independent sample t test, least significant difference- t test and Pearson correlation analysis were performed for statistical analysis. Results:The serum copper levels and the ratio of serum copper to zinc of the CD group and the UC group were both higher than that of the healthy control group, however the serum zinc levels were lower than that of the healthy control group ( (32.27±7.69) and (29.80±9.68) mol/L vs. (20.16±6.67) mol/L; 2.81±1.57 and 2.29±1.09 vs. 0.68±0.36; (14.64±7.11) and (15.65±8.17) mol/L vs. (34.29±16.40) mol/L), and the differences were statistically significant ( t=2.81, 5.87, 1.47, 7.21, 1.73 and 2.56, all P<0.05). Among CD patients, the the ratio of serum copper to zinc of patients at remission stage (29 cases), mild activity stage (23 cases), moderate activity stage (30 cases) and severe activity stage (18 cases) was 2.61±1.43, 2.75±1.35, 3.15±2.37 and 4.17±1.77, respectively, and the ratios of serum copper to zinc of patients at mild activity stage, moderate activity stage and severe activity stage were all higher than that of patients at the remission stage, and the differences were statistically significant ( t=3.41, 7.92 and 5.84, all P<0.05). There were statistically significant differences in the ratios of serum copper to zinc between patients at mild activity stage and moderate activity stage, severe activity stage, and between patients at moderate activity stage and severe activity stage ( t=5.82, 6.23 and 3.45, all P<0.05). Among UC patients, the ratio of serum copper to zinc of patients at remission stage (10 cases), mild activity stage (30 cases), moderate activity stage (45 cases) and severe activity stage (15 cases) was 1.52±0.44, 1.74±0.58, 2.38±0.83 and 3.80±1.19, respectively, the ratio of serum copper to zinc of patients at moderate activity stage was higher than that of patients at remission stage and mild activity stage, and the ratio of serum copper to zinc of patients at severe activity stage was higher than those of patients at remission stage, mild activity stage and moderate activity stage, and the differences were statistically significant ( t=7.92, 5.83, 3.21, 9.54 and 2.83, all P<0.05). There was no statistically significant difference in serum copper to zinc ratio between patients at remission and at mild activity stage ( P>0.05). The ratio of serum copper to zinc of CD patients was positively correlated with FC and CRP ( r=0.697 and 0.586, P=0.014 and 0.001), however was not correlated with ESR or CDAI score (both P>0.05). The ratio of serum copper to zinc of UC patients was positively correlated with FC, ESR and Mayo score ( r=0.488, 0.452 and 0.331, P=0.001, P<0.01 and P=0.041), however was not correlated with CRP ( P>0.05). The cut-off value of the ratio of serum copper to zinc, CRP and ESR for the diagnosis of CD activity was 1.76, 8 mg/L and 20 mm/1 h, respectively. Among them, ESR was the most effective in the diagnosis of CD activity with an area under the curve (AUC) value of 0.830, and to the sensitivity and specificity was 69.0% and 86.2%, respectively. The cut-off value of the ratio of serum copper to zinc, CRP and ESR for the diagnosis of UC activity was 1.63, 8 mg/L and 20 mm/1 h, respectively; among which the the ratio of serum copper to zinc had the highest efficacy in the diagnosis of UC activity, with an AUC value of 0.862, sensitivity and specificity of 73.0% and 90.9%, respectively. Conclusion:The the ratio of serum copper to zinc is correlated with the disease activity of IBD, which may become a new auxiliary indicator for the evaluation of disease activity.