BACKGROUND:The research of dental stem cells in the fields of regenerative medicine and tissue engineering has been deepening,bringing hope for the repair of tooth-related tissues and the treatment of systemic diseases.However,there is a lack of systematic research and analysis on the biological characteristics of dental stem cells in different age groups. OBJECTIVE:To explore the biological characteristics of the human deciduous tooth and permanent tooth pulp stem cells cultured in umbilical cord blood platelet lysate to provide a reliable basis for human platelet lysates to replace fetal bovine serum. METHODS:The pulp tissues of deciduous teeth,juvenile permanent teeth and adult permanent teeth were taken out and cultured in DMEM/F-12 medium supplemented with 10%fetal bovine serum or different concentrations(5%,10%and 15%)of human platelet lysates.Cell proliferation in the four groups was detected by cytometry.The optimal concentration of human platelet lysates was selected for subsequent experiments.Under the optimal concentration of human platelet lysates,human deciduous tooth and juvenile and adult permanent tooth pulp stem cells were cultured in vitro.The cell growth status was observed under the microscope.The specific antigen on the cell surface was detected by flow cytometry.The cell proliferation ability was tested by the cell counting method and CCK-8 assay.The cell differentiation ability in vitro was observed by a three-line differentiation assay. RESULTS AND CONCLUSION:(1)The cell proliferation rate of the 10%human platelet lysate group was the highest.(2)In all three groups,fusiform fibrous cells grew and expanded from around the tissue block.There was no significant difference between deciduous teeth and juvenile permanent tooth cells,but the adult permanent tooth cells were larger than the deciduous and juvenile permanent tooth cells of the same generation.(3)The results of flow cytometry showed that deciduous teeth,juvenile permanent teeth and adult permanent teeth conformed to the phenotypic characteristics of mesenchymal stem cells.(4)The proliferative capacity of adult permanent dental pulp stem cells was significantly lower than those of deciduous teeth and juvenile permanent dental pulp stem cells(P<0.01).(5)mRNA expressions of osteoblast-related genes alkaline phosphatase and bone morphogenetic protein 2,lipoprotein lipase and peroxisome proliferator-activated receptor γ2,mRNA expressions of chondroblast related gene type II collagen α1 and cartilage oligomeric matrix protein in adult pulp stem cells of permanent teeth were significantly lower than those of deciduous teeth and juvenile permanent teeth pulp stem cells(P<0.01).(6)Compared with adult dental pulp stem cells,human deciduous teeth and juvenile permanent teeth dental pulp stem cells have the stronger proliferative capacity and multidirectional differentiation potential,and are more suitable for clinical research and disease treatment.

Bisecting N-acetylglucosamine(GlcNAc),a GlcNAc linked to the core β-mannose resi-due via a β1,4 linkage,is a special type of N-glycosylation that has been reported to be involved in various biological processes,such as cell adhesion and fetal development.This N-glycan structure is abundant in human trophoblasts,which is postulated to be resistant to natural killer cell-mediated cytotoxicity,enabling a mother to nourish a fetus without rejection.In this study,we hypothesized that the human amniotic membrane,which serves as the last barrier for the fetus,may also express bisected-type glycans.To test this hypothesis,glycomic analysis of the human amniotic membrane was performed,and bisected N-glycans were detected.Furthermore,our pro-teomic data,which have been previously employed to explore human missing proteins,were ana-lyzed and the presence of bisecting GlcNAc-modified peptides was confirmed.A total of 41 glycoproteins with 43 glycopeptides were found to possess a bisecting GlcNAc,and 25 of these gly-coproteins were reported to exhibit this type of modification for the first time.These results provide insights into the potential roles of bisecting GlcNAc modification in the human amniotic membrane,and can be beneficial to functional studies on glycoproteins with bisecting GlcNAc modifications and functional studies on immune suppression in human placenta.

Objective:To investigate the risk factors of citrate accumulation in patients with liver failure treated with regional citrate anticoagulated continuous renal replacement therapy (RCA-CRRT).Methods:The clinical data of liver failure patients with RCA-CRRT admitted to department of intensive care unit (ICU) of Nantong Third People's Hospital from January 2017 to June 2020 were retrospectively analyzed. The selected patients were divided into citrate accumulation group and control group according to whether there was citrate accumulation (serum total calcium/free calcium ratio ≥ 2.4) during CRRT. The age, acute physiology and chronic health evaluation Ⅱ (APACHEⅡ), mean arterial pressure (MAP), norepinephrine (NE) dose, blood lactic acid (Lac) concentration, liver function status, citrate dose, filter time and prognosis of the patients were compared between the two groups. Unconditional Logistic regression was used to analyze the risk factors for citrate accumulation.Results:Among 48 patients with RCA-CRRT and liver failure, 20 patients had citrate accumulation (accumulation group), and a total of 96 CRRTs were performed; the remaining 28 patients did not have citrate accumulation (control group), a total of 106 CRRTs were performed. There were no significant differences in age and APACHEⅡ score between the two groups. Compared with the control group, the MAP in the accumulation group was lower [mmHg (1 mmHg = 0.133 kPa): 66.9±13.6 vs. 86.4±8.3, P = 0.032], and the dosage of NE (μg/min: 16.3±8.4 vs. 5.9±2.8, P = 0.015) and lactic acid level (mmol/L: 4.89±1.45 vs. 2.98±0.87, P = 0.004) were higher, the damage of liver function was more serious [total bilirubin (TBil, μmol/L): 220.4±45.2 vs. 163.4±43.8, P = 0.012; Child-Pugh score: 12.0±2.5 vs. 8.8±1.4, P = 0.029; model for end-stage liver disease (MELD) score: 31.30±8.22 vs. 21.78±6.40, P = 0.041], hourly citric acid dosage (mmol/h: 27.4±6.9 vs. 19.3±4.9, P = 0.032) and total citric acid dosage (mmol: 3 393±809 vs. 1 819±502, P = 0.039) were higher. Although there were no significant differences in the length of ICU stay, total length of hospitalization stay and cost of hospitalization between the two groups, the 28-day mortality of the accumulation group was higher than that of the control group (60.0% vs. 28.6%, P = 0.039). Unconditional Logistic regression analysis showed that MAP [odds ratio ( OR) = 2.901, 95% confidence interval (95% CI) was 0.921-19.493, P = 0.019], NE dosage ( OR = 2.098, 95% CI was 1.923-12.342, P = 0.002), Lac level ( OR = 5.201, 95% CI was 3.211-9.433, P = 0.012), Child-Pugh score ( OR = 1.843, 95% CI was 0.437-7.420, P = 0.018), MELD score ( OR = 3.012, 95% CI was 0.384-12.843, P = 0.031), hourly citric acid dosage ( OR = 4.254, 95% CI was 1.734-11.839, P = 0.011) and total citric acid dosage ( OR = 4.109, 95% CI was 1.283-18.343, P = 0.001) were risk factors for citrate accumulation. Conclusion:In patients with tissue hypoperfusion and severe liver function damage, citrate anticoagulation should be avoided or the dosage of citric acid should be reduced, in order to avoid citrate accumulation.

Objective:To explore the value of radiographic assessment of lung edema (RALE) score in evaluating the severity and prognosis of patients with acute respiratory distress syndrome (ARDS).Methods:A retrospective study was conducted. Patients with ARDS admitted to the department of intensive care unit (ICU) of Affiliated Nantong Third Hospital of Nantong University from January 2016 to November 2020 were enrolled. Clinical data of those patients were collected, and two senior radiologists who did not know the outcome of the patients independently scored each chest radiograph, the mean value of which was taken as the RALE score. The patients were divided into death group and survival group according to the 28-day prognosis. The differences of the basic data, PaO 2/FiO 2, sequential organ failure assessment (SOFA) score, acute physiology and chronic health evaluation Ⅱ (APACHEⅡ) score and RALE score between groups were analyzed. ARDS patients were classified according to the Berlin standard and RALE scores were compared between groups. Then, the correlations between RALE score and PaO 2/FiO 2, SOFA score, APACHEⅡ score were analyzed. The prognostic capacity of RALE score for 28-day prognosis of ARDS patients were analyzed by Kaplan-Meier survival curve. Results:Of the 98 ARDS patients, 62 were included in the final analysis, 39 patients survived and 23 patients died. The 28-day mortality was 37.1%. Compared with the survival group, patients in the death group were older (years old: 72.83±12.21 vs. 64.44±14.68), had lower PaO 2/FiO 2 [mmHg (1 mmHg = 0.133 kPa): 122.66±48.32 vs. 150.26±50.40], and higher SOFA score and greater difference of RALE score between the third day and the first day after admission (D3-D1 RALE score) (SOFA score: 11.26±3.91 vs. 9.04±3.72, D3-D1 RALE score: 1.35±6.42 vs. -2.74±7.35), with statistically significant differences (all P < 0.05). However, there were no significant differences in gender, cause of ARDS, APACHEⅡ score, and RALE scores on the first and the third day of admission (D1 RALE, D3 RALE) between the two groups. Among the 62 patients, there were 11 mild cases (17.7%), 36 moderate cases (58.1%), and 15 severe cases (24.2%). The D1 RALE score of patients with mild and moderate ARDS were lower than those of patients with severe ARDS (19.09±3.65, 22.58±6.79 vs. 27.07±5.23, both P < 0.05). Correlation analysis showed that D1 RALE score was negatively correlated with PaO 2/FiO 2 ( r = -0.385, P = 0.002), and positively correlated with SOFA score and APACHEⅡ score ( r1 = 0.433, r2 = 0.442, both P < 0.001). Kaplan-Meier survival curve analysis showed that the 28-day survival rate of ARDS patients in D3-D1 RALE score ≥ -1 group was significantly higher than that in D3-D1 RALE score < -1 group (73.08% vs. 55.56%; log-rank test: χ 2 = 3.979, P = 0.046). Conclusions:The RALE score is a simple and reliable non-invasive evaluation index, which can be used to evaluate the severity of ARDS patients. The difference of RALE score in early stage is helpful to identify ARDS patients with poor prognosis.

OBJECTIVE: To analyze the current status of diagnosis and management of acute appendicitis (AA) in China. METHODS: Questionnaire survey was used to retrospectively collect data of hospitalized patients with AA from 43 medical centers nationwide in 2017 (Sort by number of cases provided: Jinling Hospital of Medical School of Nanjing University, The First Affiliated Hospital of Xinjiang Medical University, Lu'an People's Hospital, Tengzhou Central People's Hospital, Dalian Central Hospital, The Affiliated Hospital of Xuzhou Medical University, Dongying People's Hospital, Jinjiang Hospital of Traditional Chinese Medicine, Huangshan Shoukang Hospital, Xuyi People's Hospital, Nanjing Jiangbei People's Hospital, Lanzhou 940th Hospital of PLA, Heze Municipal Hospital, The First College of Clinical Medical Science of China Three Gorges University, Affiliated Jiujiang Hospital of Nanchang University, The Second People's Hospital of Hefei, Affiliated Central Hospital of Shandong Zaozhuang Mining Group, The Third People's Hospital of Kunshan City, Xuzhou First People's Hospital, The 81st Group Army Hospital of PLA, Linyi Central Hospital, The General Hospital of Huainan Eastern Hospital Group, The 908th Hospital of PLA, Liyang People's Hospital, The 901th Hospital of Joint Logistic Support Force, The Third Affiliated Hospital of Chongqing Medical University, The Fourth Hospital of Jilin University, Harbin Acheng District People's Hospital, The First Affiliated Hospital of Zhengzhou University, Nanjing Luhe People's Hospital, Taixing Municipal People's Hospital, Baotou Central Hospital, The Affiliated Hospital of Nantong University, Linyi People's Hospital, The 72st Group Army Hospital of PLA, Zaozhuang Municipal Hospital, People's Hospital of Dayu County, Taixing City Hospital of Traditional Chinese Medicine, Suzhou Municipal Hospital, Beijing Guang'anmen Hospital, Langxi County Hospital of Traditional Chinese Medicine, Nanyang Central Hospital, The Affiliated People's Hospital of Inner Mongolia Medical University).The diagnosis and management of AA were analyzed through unified summary. Different centers collected and summarized their data in 2017 and sent back the questionnaires for summary. RESULTS: A total of 8 766 AA patients were enrolled from 43 medical centers, including 4 711 males (53.7%) with median age of 39 years and 958 (10.9%) patients over 65 years old. Of 8 776 patients, 5 677 cases (64.6%) received one or more imaging examinations, and the other 3 099 (35.4%) did not receive any imaging examination. A total of 1 858 (21.2%) cases received medical treatment, mainly a combination of nitroimidazoles (1 107 cases, 59.8%) doublet regimen, followed by a single-agent regimen of non-nitroimidazoles (451 cases, 24.4%), a nitroimidazole-free doublet regimen (134 cases, 7.2%), a triple regimen of combined nitroimidazoles (116 cases, 6.3%), nitroimidazole alone (39 cases, 2.1%) and nitroimidazole-free triple regimen (3 cases, 0.2%). Of the 6 908 patients (78.8%) who underwent surgery, 4 319 (62.5%) underwent laparoscopic appendectomy and 2589 (37.5%) underwent open surgery. Ratio of laparotomy was higher in those patients under 16 years old (392 cases) or over 65 years old (258 cases) [15.1%(392/2 589) and 10.0%(258/2 589), respectively, compared with 8.5%(367/4 316) and 8.0%(347/4 316) in the same age group for laparoscopic surgery, χ²=91.415, P<0.001; χ²=15.915,P<0.001]. Patients with complicated appendicitis had higher ratio of undergoing open surgery as compared to those undergoing laparoscopic surgery [26.7%(692/2 589) vs. 15.6%(672/4 316), χ²=125.726, P<0.001].The cure rates of laparoscopic and open surgery were 100.0% and 99.8%(2 585/2 589) respectively without significant difference (P=0.206). Postoperative complication rates were 4.5%(121/2 589) and 4.7%(196/4 316) respectively, and the difference was not statistically significant (χ²=0.065, P=0.799). The incidence of surgical site infection was lower (0.6% vs. 1.7%, χ²=17.315, P<0.001), and hospital stay was shorter [6(4-7) days vs. 6(5-8) days, U=4 384 348.0, P<0.001] in the laparoscopic surgery group, while hospitalization cost was higher (median 12 527 yuan vs. 9 342 yuan, U=2 586 809.0, P<0.001). CONCLUSIONS The diagnosis of acute appendicitis is still clinically based, supplemented by imaging examination. Appendectomy is still the most effective treatment at present. Laparoscopic appendectomy has become the main treatment strategy, but anti-infective drugs are also very effective.
Acute Disease ; Adolescent ; Adult ; Aged ; Anti-Bacterial Agents ; therapeutic use ; Appendectomy ; Appendicitis ; diagnosis ; therapy ; China ; Female ; Health Care Surveys ; Humans ; Laparoscopy ; Male ; Middle Aged ; Retrospective Studies ; Treatment Outcome ; Young Adult

Objective To investigate the effects of angiotensin Ⅱ (Ang Ⅱ) on the expression of C-terminal Src kinase (Csk) in Ang Ⅱ-infused rat model and cultured podocytes,and to explore the role of Csk in Ang Ⅱ-induced cytoskeletal rearrangement of podocytes.Methods Twenty-four Wista rats were randomly subjected to normal saline infusion,or Ang Ⅱ infusion at 400 ng · kg1 · min-1 (via subcutaneous osmotic minipumps) for 2 or 4 weeks.Renal histomorphology was evaluated through electron microscopy.The expression of glomerular Csk was analyzed by immunofluorescence and Western blotting.In vitro,conditionally immortalized mouse podocytes were cultured and treated with Ang Ⅱ doses ranging from 10-9 mol/L to 10-5 mol/L and for different hours.The expression of podocytes Csk was assessed by Western blotting.After transfection to podocytes with Csk siRNA,FITC-conjugated phalloidin was used to stain F-actin,to investigate the role of Csk in Ang Ⅱ-induced or cytochalasin D-induced cytoskeletal rearrangement.Results (1) Examination of Ang Ⅱ infusion rats glomerular and podocyte ultrastructure by electron microscopy revealed foot process effacement and fusion; (2) In Ang Ⅱ infusion rats,the expression of glomerular Csk was increased (P ＜ 0.05); (3) In vitro,Ang Ⅱ-stimuli up-regulated the expression of Csk (P ＜ 0.05),and the effects of Ang Ⅱ were on dose-dependent and time-dependent manner; (4) Ang Ⅱ-induced disruption of F-actin was alleviated by Csk siRNA transfection in cultured podocytes; furthermore,cytochalasin D depolymerized the F-actin cytoskeleton,while Csk siRNA stabilized the actin filaments.Conclusion The enhanced expression of Csk may be involved in Ang II-induced podocytes cytoskeletal rearrangement and foot process fusion.

ObjectiveTo evaluate the effects of angiotensin Ⅱ (Ang Ⅱ )infusion on renal c-Abl expression in vivo,and on podocyte c-Abl expression change in cultured mouse podocytes.Methods Twenty four male Sprague-Dawley rats (Group C,D,E and F) were assigned to receive Ang Ⅱ(400 ng· kg-1 min-1) by osmotic minipump and of which 12 rats (Group D and F) were assigned to receive telmisartan (3 mg·kg-1·d-1),six rats received normal saline(Group B),and six rats were used as normal control(Group A).Animals were sacrificed at day 14 (Group C and D),day 28 (Group E and F) respectively.Conditionally immortalized mouse podocytes were used in vitro.Podocytes were studied 2 weeks after thermoswitching from 33℃ to 37℃.Cells were fetal bovine serum(FBS) starved for at least 12 hours prior to stimulation.The cultured podocytes were treated withAngⅡdosesranging from10 -9 mol/L to10 -6 mol/L andfor differenthours.Expression of renal and podocytes c-Abl was examined by immunofluorescence staining,real-time PCR and Western blotting.Results(1) Distribution of c-Abl expression was mainly in the cytoplasm and nuclear of the podocytes in vivo and in vitro. (2) Expressions of c-Abl mRNA and protein wereincreasedinAng Ⅱ-infusedratpodocytesandAng Ⅱ-inducedculturedmouse podocytes(P＜0.05),and the effects of Ang Ⅱ were dose-dependent and time-dependent in vitro.Conclusion There are c-Abl mRNA and protein expression in podocytes,and c-Abl may play a critical role in the pathogenesis of Ang Ⅱ -induced podocyte injury.

Objective To evaluate the effect of angiotensin Ⅱ (Ang Ⅱ ) on the change of nephrin phosphorylation both in Ang Ⅱ-infused rat model and cultured podocytes.Methods Thirty Wistar rats were subcutaneously embedded with osmotic minipumps and randomly divided into 3 groups according to receiving either Ang Ⅱ at a dose of 400 ng· kg-1· min-1 or Ang Ⅱ +telmisartan at a dose of 3 mg·kg-1 ·d-1,or normal saline as a control group.Blood pressure and 24-hour urinary albumin were measured at 0 d,7 d,14 d,21 d and 28 d of the experiment.Renal histomorphology was evaluated through electron microscopy.The concentrations of Ang Ⅱ both in blood plasma and kidney were detected by radioimmunoassay.In vitro,cultured murine podocytes were exposed to Ang Ⅱ (10-6 mol/L) pretreated with or without losartan (10-5 mol/L) for different time periods.Nephrin and its phosphorylation expression were analyzed by Western blotting.The distribution of F-actin was presented by FITC-phallodin labeling.The change in phenomenon of F-actin was evaluated by cortical F-actin score index (CFS).Results (1)Ang Ⅱ-infused rats exhibited increased Ang Ⅱ concentration,significant hypertension and marked albuminuria.(2)In Ang Ⅱ-infusion group,nephrin expression was decreased (P＜0.05).Ang Ⅱ-receiving rats displayed diminished phosphorylation of nephrin.(3)In vitro,the phosphorylation of nephrin was significantly reduced after Ang Ⅱ stimulation for 3-6 hours (P＜0.05).(4)Ang Ⅱ stimulatation resulted in irregularly arrangement of F-actin followed by the redistribution of F-actin to podocyte periphery and formation of F-actin ring,in which the CFS obviously increased compared to control (P＜0.05).Conclusions Phosphorylation of nephrin is important for the survival status of podocytes.Ang Ⅱ-induced nephrin dephosphorylation may be an important molecular mechanism for Ang Ⅱ-induced podocyte cytoskeleton rearrangement and foot process effacement.

ObjectiveTo evaluate the effects of Ang Ⅱ on apoptosis of podocytes and explore the signaling pathwayof nephrin in preventingAng Ⅱ-inducedpodocyte apoptosis.MethodsDifferentiated mouse podocytes were exposed to Ang Ⅱ at different concentrations for 18 h or at 10-8 mol/L for variable incubation times.Undifferentiated mouse pedocytes were transfected using lipofectamine 2000 with the pcDNA3.1-mNPHS1 plasmid and stably transfected cell lines were generated with G418 selection.In separated experiments,untransfected mouse podocytes (MPC) and stably transfected podocytes with pcDNA3.1-neo and PcDNA3.1-mNPHS1 were exposed toAng Ⅱ(10-8 mol/L) or LY294002(a selective Akt inhibitor,50 μmol/L) for indicated times.Apoptosis was evaluated by flow cytometry.The expression of nephrin was assessed by quantitative real-time PCR,immunofluorescence and Western blotting.The phosphorylation level of Akt was determined by Westem blotting.Results(1) AngⅡ promoted podocyte apoptosis in a dose-and time-dependentmanner. PretreatmentwithlosartansignificantlypreventedAngⅡ -induced apoptosis. (2) Nephfin mRNA and protein were obviously decreased in podocytes exposed to 10-8 mol/L Ang Ⅱ for at least 12 h than those in vehicle-treated cells (P＜0.05).(3) Ang Ⅱ exposure for more than 15 min inhibited the phosphorylation of AKT in MPC,which was dramatically reversed by pcDNA3.1-mNPHS1 transfection,but not by pcDNA3.1-neo transfection. (4) Podocyte apoptosis was promoted byLY294002. Conversely,Ang Ⅱ-induced podocyteapoptosis was significantly alleviated by pcDNA3.1-mNPHS1 transfection.ConclusionAng Ⅱinduces mouse podocyte apoptosis which is suppressed by overexpression of nephrin through PI3K-Akt signaling pathway.

Objective To evaluate the effect of aldosterone (Ald) on glomerular mesangial cells apoptosis and to explore the possible mechanisms.Methods Twenty-four Sprngue-Dawley rats were subcutaneously embedded with osmotic mini-pumps and randomly divided into 3 groups.Aldosterone (1.5 μg/h) was administrated subcutaneouly by osmotic mini-pumps in Ald group,eplerenone (Epl,100 mg·kg-1·d-1) and Ald (1.5 μg/h) was given to Epl group.And normal saline was used in control group (Con group).Systolic blood pressure and urinary albumin excretion rate (UAER) were detected on day 0,7,14,21,28.Blood and kidney samples were harvested on day 28.Plasma creatinine,potassium and aldosterone were measured.Renal paraffin sections were stained by PAS and the morphological changes were evaluated by light microscopy.Apoptosis index of mesangial cells were detected by TUNEL assay.The glomerular mesangial cells (MCs) were cultured in a DMEM-F12 media.MCs apoptosis was evaluated by staining cells with Annexin V and propidium iodide (PI) using flow cytometer.Expression of Bcl-2 and Bax mRNA was examined by RT-PCR.The protein level of Bad or phospho-Bad was measured by Western blotting.Results Ald-infused rats developed hyperaldosteronemia and hypokalemia.Rats in Ald group exhibited significant hypertension and marked albuminuria.Ald group rats showed increased number of TUNEL-positive mesangial cells when compared with control rats (P＜0.05).Aldosterone induced mesangial cells apoptosis in a time-dependent manner.Expression of Bcl-2 mRNA was decreased but Bax mRNA was increased in aldosterone treated MCs compared to that in Con group (P＜0.05).Aldosterone promoted dephosphorylation of cytosolic phospho-Bad compared with vehicle treated cells (P＜ 0.05).However,eplerenone attenuated these effects of aldosterone.Conclusion Aldosterone directly promotes mesangial cells apoptosis,and eplerenone can attenuate this effect of aldosterone.Dephosphorylation of cytosolic phospho-Bad may be the key role in the progression of mesangial cells apoptosis induced by aldosterone.