BACKGROUND:There is still controversy whether human bone marrow mesenchymal stem cells can maintain their biological characteristics,energy metabolism patterns,and multidirectional differentiation potential after long-term expression in vitro.Further comprehensive and systematic research is needed. OBJECTIVE:To evaluate the effects of long-term expansion in vitro on the biological characteristics of mesenchymal stem cells. METHODS:Human bone marrow mesenchymal stem cells cultured to passage 5,10,and 15 in vitro.MTT assay was used to detect cell proliferation ability.Flow cytometry was used to detect cell cycle.The multi-differentiation potential of mesenchymal stem cells was detected by inducing to adipogenic,osteogenic and chondrogenic differentiation.Cell migration and invasion abilities were detected by scratch test and Transwell assay.The mitochondrial oxidative phosphorylation and glycolysis function were analyzed using energy metabolism analyzer.The cell senescence was detected by senescence-associated β-galactosidase staining.The expression levels of p21,p16,and p53 proteins were detected by western blot assay. RESULTS AND CONCLUSION:Bone marrow mesenchymal stem cells at passages 5,10,and 15 grew adherently;the volume of passage 15 mesenchymal stem cells increased and its proliferation ability decreased;the percentage of S-phase cells decreased(P<0.05).With the increase of culture passages,the migration and invasion abilities decreased gradually(P<0.05).There was no significant difference in the differentiation potential,demonstrated by adipogenic,osteogenic and chondrogenesis induction.The ability of oxidative phosphorylation of mitochondria and glycolysis decreased gradually(P<0.05).The number of senescence-associated β-galactosidase-positive cells increased with the increase of passages(P<0.05),and the expression of senescence protein p21,p16,and p53 increased gradually(P<0.05).The results indicated that the biological characterization of mesenchymal stem cells changed after long-term in vitro expansion.Mesenchymal stem cells cultured over 10 passages may have a reduced activity due to increasing senescence.Therefore,bone marrow mesenchymal stem cells cultured less than 10 passages are suitable for clinical research/therapy.

Background:In the clinical setting, the effect of intra-abdominal hypertension on the human body is dependent on time, but its role is not yet clear.Objective:To investigate the effect of the duration of intra-abdominal hypertension (IAH) on the prognosis of critically ill patients.Methods:This prospective cohort study enrolled 256 IAH patients who were admitted to the Surgical ICU of 10 Grade A hospitals in Fujian Province from January 2018 to December 2020. The duration of IAH (DIAH) was obtained after monitoring IAP, and ICU length of stay, duration of mechanical ventilation, duration of continuous renal replacement therapy (CRRT) and average daily energy intake from enteral nutrition during ICU stay were observed and recorded. The correlation was analyzed by Spearman rank correlation. The patients were divided into the survival group and the death group according to their survival state at 60 days after enrollment. Thereafter, clinical characteristics between the two groups were compared. Multivariable logistic regression was used to study and validate the relationship between DIAH and 60-day mortality. The receiver operating characteristics (ROC) curve was established to evaluate the predictive abilities of DIAH on the mortality risk.Results:In critically ill patients, DIAH was positively correlated with duration of mechanical ventilation ( r=0.679, P<0.001), duration of CRRT ( r=0.541, P<0.001) and ICU length of stay ( r=0.794, P<0.001), respectively. In addition, there was a negative correlation between DIAH and average daily energy intake from enteral nutrition ( r=-0.669, P<0.001). After multivariable adjustment, DIAH was an independent risk factor for 60-day mortality in critically patients with IAH ( OR=1.05, 95% CI: 1.01-1.12; P = 0.012), and exhibited a linearity change trend relationship with mortality risk. The ROC curve analysis of DIAH showed that the area under ROC curve (AUC) was 0.825 (95% CI: 0.763~0.886, P<0.01). When the cut-off value was 16.5 days, the sensitivity was 78.4% and the specificity was 75.4%. Conclusions:DIAH is an important risk factor for prognosis in critically ill patients. Early identification and rapid intervention for the etiology of IAH should be performed to shorten DIAH.

Objective:To investigate the effect of nurse-led stress inoculation training on fear of self-injecting and self-testing and self-management behaviors in elderly type 2 diabetic patients and provide reference for diabetes nursing care.Methods:A total of 110 elderly type 2 diabetic patients of Department of Endocrinology of Hainan People′s Hospital from January 2018 to January 2020 were divided into experimental group and control group according to odd and even numbers, with 55 patients in each group. The control group received routine nursing care, while the experimental group implemented nurse-led stress inoculation training for 4 weeks. The intervention effect was assessed by Diabetes Fear of Injecting and Self-testing Ouestionnaire (D-FISQ) and Diabetes self-management behaviors among older (DSMB-O), respectively.Results:In the study, one patient in the experimental group fell off, and finally included 54 cases in the experimental group and 55 cases in the control group. After intervention, the fear of self-injecting scores, fear of self-testing scores, and total D-FISQ scores were 13.15 ± 3.02, 15.67 ± 3.59 and 28.81 ± 5.08 in the experimental group, significantly lower than those in the control group (15.25 ± 3.18, 17.56 ± 3.92 and 32.82 ± 4.89), the difference was statistically significant ( t=3.55, 2.63, 4.19, P<0.05). Active exercises, current medication, blood glucose monitoring, dealing with problem, active response, reducing risks scores and total DSMB-O scores were 2.39 ± 0.49, 2.39 ± 0.49, 2.20 ± 0.81, 4.41 ± 0.92, 4.70 ± 1.13, 5.06 ± 0.79 and 25.28 ± 2.57 in the experimental group, significantly higher than those in the control group (3.95 ± 0.85, 2.11 ± 0.85, 1.51 ± 0.50, 3.95 ± 0.78, 4.13 ± 1.43, 4.38 ± 1.16 and 22.09 ± 2.24), the difference was statistically significant ( t values were 2.10-6.90, P<0.05). Conclusions:Nurse-led stress inoculation training can effectively alleviate fear of self-injecting and self-testing and promote self-management behaviors of elderly patients with type 2 diabetes.

Objective:To establish a classification model for rapid identification of hypervirulent subtype ST17 clones of Group B Streptococcus (GBS) using matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS).Methods:In a retrospective study, 235 strains of GBS strains were selected from multiple centers in China during 2015-2018. For model generation,45 strains of ST17 and 50 strains of non-ST17 (20 ST19, 15 ST12 and 15 ST10 strains) were enrolled as the modeling group. The remaining 90 main ST strains (40 ST17, 16 ST10, 17 ST12 and 17 ST19) were served as validation group. 50 GBS strains classified as other minor ST subtypes were regarded as taxonomic groups. MS spectra were collected by Bruker mass spectrometry, and then loaded for model generation and verification, and screening of differential peptide peaks by genetic algorithm (GA) and model verification on ClinProTools 3.0 software.Results:The recognition rate for ST17-GA model were 99.4% with cross validation value of 96.9%. Among the ten differential peptide peaks for the classification model, the weights of both two main peptide peaks m/z 2 956 and m/z 5 912 were greater than 1, while the weights of the all other eight peptide peaks were less than 0.5. Model validation showed only one of the ST17 was misjudged as non-ST17 strain, resulting in diagnostic accuracy of 98.9%, sensitivity of 97.5% and specificity of 100%, positive predictive value of 100% and negative predictive value of 98.0%, respectively. For other sporadic STs, 42.0% (21/50) of them were misdiagnosed as ST17 subtype.Conclusion:A MALDI-TOF MS classification model for hypervirulent subtype of ST17 GBS strains has been successfully established with good diagnostic efficacy.

Objective: To evaluate diagnostic performance of Todd-Hewitt (T-H) broth culture method, direct culture method, liquid chromogenic culture method, and loop-mediated isothermal amplification (LAMP) method for screening group B streptococcus (GBS) during late pregnancy. Methods: In the retrospective study, the rectal vaginal secretions samples were collected from pregnant women at 35 to 37 weeks at the obstetrics clinic of Guangzhou Women and Children′s Medical Center affiliated to Guangzhou Medical University during October 2016 to April 2018. For the purposes of clinical evaluation, T-H broth culture was used as the standard reference method, and double-blind trials were used to evaluate diagnostic performance of direct culture method, liquid chromogenic culture method, and LAMP method for screening group B streptococcus during late pregnancy in three research stages. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), coincidence rate and Yoden index for each method were calculated. Also, the level of agreement between each method and T-H broth was assessed using the kappa (k) coefficient. Results: A total of 969 specimens were detected by the T-H enrichment culture method, and 90 were positive (9.3%). The sensitivities from high to low were LAMP method [100% (25/25)], direct culture method [81.5% (22/27), 95%CI:65.8%-97.1%], and liquid color culture method [71.1% (27/38), 95%CI:55.9%-86.2%]. Specificities were direct culture method [100% (282/282)], liquid color culture method [98.1% (455/464), 95%CI:96.8%-99.3%], and LAMP method [94.0% (125/133), 95%CI: 89.9%-98.1%]. The coincidence rates were direct culture method [98.4% (22+282)/309], liquid color culture method [96.0% (27+455)/502], and LAMP method [94.9% (25+125)/158]. The Kappa values of the direct culture method (0.889), LAMP method (0.832) and the enrichment culture method were all ≥0.75, and that of the liquid color culture method was 0.708. The false negative rate of direct culture method was 18.5% (5/27), and no false negative case by LAMP method, but its false positive rate was 6.0% (8/133). The false negative rate and false positive rate of liquid color culture method were 28.9% (11/38) and 1.9% (9/464), respectively. Conclusions Of the three screening methods compared in this study, only the LAMP method has the advantages in sensitivity, specificity, and coincidence rate compared with T-H enriched culture method, while the others have a certain degree of false negatives rate. The clinical laboratory can introduce these methods based on laboratory facilities and staffing, or refer to the European and American guidelines and combine the recommended antenatal GBS screening method with intrapartum nucleic acid amplification tests to best meet the clinical demands.

Objective To raise awareness of the late-onset meningitis caused by group B streptococcus (GBS) which was homogenous to the maternal colonization.Methods The clinical data of late-onset GBS meningitis in neonates twins whose pathogens were homogenous to their maternal colonization were collected from Department of Neonatology,Guangzhou Women and Children's Medical Center.The general conditions,clinical symptoms,laboratory tests and drug treatment of the twins and their mother were retrospectively analyzed,and the GBS homology during inpatient care was tested.And the progress of the twins' condition was investigated by telephone follow-up.Results The mother had two pregnancies without prenatal GBS screening or intrapartum antimicrobial intervention for GBS,everything was normal during pregnancy and delivery.Twins were born through cesarean section.The elder sister was discharged with Linezolid taken orally after 167 days in hospital without convulsions,shaking or other discomfort.The elder sister was followed up for every 2 weeks,and in the last time of follow-up,cerebrospinal fluid white blood cell counts were 45 × 106/L,protein level was 1.52 g/L and Linezolid was withdrawn.The younger brother was discharged after 58 days in hospital with follow-up for every 2 weeks,and in the last time of follow-up,cerebrospinal fluid white blood cell counts were 30 × 106/L,protein level was 0.66 g/L.During the hospitalization and follow-up without convulsions and irritation,and the cranial magnetic resonance imaging of the twin brother was normal.Test results showed that the GBS bacteria strain for twins and their mother were all serotype Ⅲ.The possibility of the GBS homology was more than 90％.Conclusions The toxicity of serotype Ⅲ GBS strain was strong.More proactive precautions should be considered to apply for the mother whose first birth already had GBS infection.Early identification and intervention of infection risk factors would help optimize the anti-infection treatment program and reduce nerve system damage and other adverse outcomes caused by invasive GBS infection.

Objective To investigate the clinical characteristics,antibiotic susceptibilities and serotypes of Group B Streptococcus(GBS)isolated from neonatal meningitis to provide references for the prevention and treatment of neonatal GBS meningitis. Methods From June 2013 to June 2016,we surveyed the GBS strains iso-lated from purulent meningitis of < 90 days infants from Guangzhou Women and Children′s Medical Center. The GBS isolates were identified and the minimum inhibitory concentration of the antibiotics was determined by Vitek 2 Compact automatic bacterial identification system.GBS serotyping was performed using Strep-B-Latex?rapid latex agglutination test kit. Results A total of 46 cases of neonatal GBS meningitis,15 cases of early-onset infection and 31 cases of late-onset infection were diagnosed. 78.3% of GBS meningitis with varying degrees of complica-tions.Among 41 survivors with 3~24 months follow-up,50% of the early-onset and 44.8% of the late-onset GBS meningitis with varying degrees of neurological sequelae.Four capsular types were identified among the 46 isolates, serotype-Ⅲwas the most prevalent(73.9%),followed by Ib(19.6%),V(4.3%)and Ia(2.2%).All the isolates were susceptible to penicillins,cephalosporins,linezolid and vancomycin. Conclusion The highly pathogenic serotype-Ⅲ was the predominant serotype among neonatal GBS meningitis in Guangzhou,Therefore,it is neces-sary to strengthen the epidemiological surveillance of GBS invasive infection and the effective implementation of pre-ventive measures.

Group B streptococcus ( GBS) is the most common cause of neonatal invasive infections in western countries , leading to severe pneumonia , sepsis and meningitis with high mortality and morbidity . Neonatal GBS infections are mainly transmitted from mother to baby .Since the end of 1990 s, prevention strategies of antenatal GBS screening and intrapartum antibiotic prophylaxis ( IAP) have been implemented by European and American area , decreasing the incidence of neonatal GBS disease significantly .The harm of GBS to neonates has been recognized and valued in recent years in China , but the authorized prevention measures are still lacking.Efficacy of IAP-based prevention depends on the accuracy of GBS screening results during pregnancy .Here, the GBS prevention guideline or consensus recommended by European and American CDC, including the screening strategies , detection methods, sample collection, storage and transportation were interpreted .

ObjectiveTo explore the clinical application of preoperative biliary drainage in the treatment of patients with malignant obstructive jaundice (MOJ) with acute cholangitis (AC). MethodsA retrospective study was performed on the clinical data of 74 patients with MOJ and AC who were treated with preoperative biliary drainage in our hospital from January 2010 to December 2014. In those patients, 29 patients treated with percutaneous transhepatic biliary drainage (PTCD) were assigned to PTCD group, and 35 patients treated with endoscopic retrograde biliary drainage (ERBD) were assigned to ERBD group. The levels of total bilirubin (TBil), direct bilirubin (DBil), and alanine aminotransferase (ALT) before and after treatment, total hospitalization cost, average duration of hospitalization, and postoperative complications were compared between the two groups. The advantages and disadvantages were compared between the two preoperative biliary drainage approaches. Between-group comparison of continuous data was made by t test, and between-group comparison of categorical data was made by χ2 test. ResultsIn both groups, the levels of TBil, DBil, and ALT were significantly reduced after treatment (all P＜0.05). The ERBD group had significantly larger decreases in the levels of the three biochemical indices than the PTCD group (all P＜0.05). The ERBD group had significantly shorter average duration of hospitalization and significantly lower total hospitalization cost than the PTCD group (t=3.172, P＜0.05; t=2.562, P＜0.05). The incidence of acute pancreatitis in the ERBD group was significantly higher than that in the PTCD group (14.28% vs 3.45%, P＜0.05); however, the incidence rates of biliary tract bleeding, biliary tract infection, and resection or puncture site infection were significantly lower in the ERBD group than in the PTCD group (all P＜0.05). ConclusionPreoperative biliary drainage can substantially control AC symptoms and improve liver function. Compared with PTCD, ERBD achieves shorter duration of hospitalization, lower total hospitalization cost, and lower incidence rates of complications after treatment, which can be taken as the first choice for the treatment of MOJ with AC.

ObjectiveTo investigate the association between CD4+/CD8+ ratio in peripheral blood and patient prognosis after hepatectomy for liver cancer. MethodsThe clinical data of 150 patients who received partial hepatectomy for liver cancer in the 180 Hospital of PLA from October 2008 to November 2011 were analyzed retrospectively. CD4+/CD8+ ratio in peripheral venous blood was measured before surgery, and the patients were divided into low-CD4+/CD8+ group (CD4+/CD8+ ratio ≤1, 52 patients) and high-CD4+/CD8+ group (CD4+/CD8+ ratio ＞1, 98 patients). Clinical indices were compared between the two groups, and outpatient follow-up and telephone follow-up were applied to record survival rate and tumor recurrence. The chi-square test was applied for comparison between the two groups, and Kaplan-Meier method (log-rank test) was applied for survival analysis. Univariate and multivariate logistic regression analyses were performed for clinical factors to determine the related risk factors for recurrence after hepatectomy for liver cancer. ResultsThe low-CD4+/CD8+ group had significantly lower 1-, 3-, and 5-year survival rates than the high-CD4+/CD8+ group (χ2=36.473, 41983, and 55.214, respectively; all P＜0.001), and the 5-year survival rate differed significantly between the two groups (χ2=81.471; P＜005); the low-CD4+/CD8+ group had significantly higher 1-, 3-, and 5-year tumor recurrence rates than the high-CD4+/CD8+ group (χ2=44.041, 68.234, and 55.157, respectively; all P＜0.05). Univariate analysis showed that CD4+/CD8+ ratio, tumor diameter, existence of satellite lesions, hepatitis B virus infection, depth of tumor invasion, microvascular invasion, lymph node metastasis, and degree of tumor differentiation were high risk factors for recurrence after resection of liver cancer. Multivariate analysis showed that CD4+/CD8+ ratio, tumor diameter, degree of tumor differentiation, lymph node metastasis, and microvascular invasion were independent risk factors for recurrence after resection of liver cancer. ConclusionThe patients with a CD4+/CD8+ ratio of ≤1 before resection of liver cancer have poor prognosis and high recurrence rates, and CD4+/CD8+ ratio has a certain predictive value for prognosis after resection of liver cancer.