BACKGROUND:There are a lot of studies on isolation and culture methods of human placental chorionic-derived mesenchymal stem cells (hpcMSCs), but how to simply and efficiently harvest a large amount of primary MSCs has not been resolved. OBJECTIVE:To optimize the tissue explants method of isolating and culturing hpcMSCsin vitro. METHODS:Human placental chorionic villi were collected from full-term deliveries under aseptic condition and isolated by electric homogenizer. hpcMSCs were prepared by tissue explants method. The fluid and tissue of the primary culture flask and douching normal saline of the initial culture were centrifuged and prepared for secondary culture. RESULTS AND CONCLUSION: It saved time and effort to treat human placental chorionic villi with electric homogenizer, with good effects on tissue dispersion and removal of red blood cells. The average time of cell acquisition in initial culture and secondary culture was (17.73±1.14) and (10.03±1.30) days, respectively. The yields of primary cultured cells in initial culture and secondary culture were (6.97±0.98)×105 and (13.82±1.44)×105per Φ100 mm culture dish, respectively. The adherent cells showed fibroblast-cell-like shape, which were in parallel or circinate arrangement. Highly expressed CD73, CD105 and CD90 could be detected in the third generation of hpcMSCs, but CD34, CD45, CD14, CD19 and HLA-DR were negative. Following induction, alizarin red staining and oil red O staining produced a strong reaction in cells. In a word, the optimized method is a simple and efficient method for obtaining a large amount of primary hpcMSCs.

Objective To establish a cultivating method for obtaining a large number of P0 generation human placental chorionic-derived mesenchymal stem cells (hpcMSCs).Methods The hpcMSCs were isolated from human placental chorion.After primary culturing and culturing for seven days,the culture medium,the non-adherent tissue and the douching normal saline of the primary culture were centrifuged and re-cultured twice.Cell morphology was observed by an inverted microscope.CCK-8 was used to measure the cell growth curve.Flow cytometry was used to detect cell surface markers.Adipogenic and osteogenic differentiation kits were used to assess the cell differentiation potential.Results The obtained hpcMSCs were fibroblast-like adherent cells and (25.54±3.38)×106 cells were obtained per placenta.The total yield of the primary culture,secondary culture and tertiary culture were (11.73±2.09)×106,(11.12±1.42)×106 and (2.69±0.71)×106,respectively,and the incubation time were (12.00±0.64) d,(8.87±0.63) d and (12.33±0.80) d.There was significant differences in incubation time between the secondary culture and the primary culture as well as the tertiary culture (all P＜0.05),and there was no significant difference between the primary culture and the tertiary culture.However,the incubation time of the tertiary culture had an increasing trend (P＞0.05).The yield per culture flask of the primary culture,secondary culture and tertiary culture were (1.12±0.15) × 106,(2.10±0.16)×106 and (1.04±0.16)×106,respectively.There was significant differences in the yield per culture flask between the secondary culture and the primary culture as well as the tertiary culture (all P＜0.05),and there was no significant difference between the primary culture and the tertiary culture.However,the yield per culture flask of the tertiary culture had a decreasing trend (P＞0.05).There was no difference among the three cultures in the growth curve and the expression of surface markers,and the osteogenic and adipogenic differentiation were all positive.Conclusions The P0 generation hpcMSCs isolated from a choriocarcinoma sample can be doubled by the three cultures compared with the primary culture,which can provide plenty stem cell source for the regenerative medicine.

Objective:To explore the correlation of serum adiponectin (APN) , D-dimer (D-D) and neutrophil-to-lymphocyte ratio (NLR) levels with disease severity and prognosis in patients with diabetic foot ulcer infection.Methods:92 patients with diabetic foot ulcer infection in the Nantong Third Hospital Affiliated to Nantong University from Feb. 2020 to Feb. 2021 were selected, and they were divided into mild ( n=30) , moderate ( n=44) and severe ( n=18) patients according to the severity of the disease. The serum APN, D-D and NLR levels in patients with different severity were compared, the relationship between serum APN, D-D and NLR levels and disease severity in patients with diabetic foot ulcer infection were analyzed. Patients were followed up for 1 year, and the prognosis of the patients was counted. Factors affecting serum APN, D-D and NLR levels in patients with diabetic foot ulcer infection were analyzed, and the receiver operating curve (ROC) was used to analyze the value of serum APN, D-D and NLR levels in predicting poor prognosis of patients. Results:There were significant differences in serum APN, D-D and NLR levels in patients with different severity ( P<0.05) . APN level in severe patients was 5.35±0.98, in moderate patients was 7.64±1.25, both lower than that of the mild patients 9.19±1.73 ( P<0.05) . Serum APN level in severe patients was lower than that in moderate patients ( P<0.05) . Serum D-D and NLR levels were 3.49±0.72 and 2.86±0.58 in severe patients, respectively; and they were 3.02±0.63 and 2.24±0.46 in moderate patients, higher than that of mild patients 2.43±0.51; 1.71±0.33 ( P<0.05) . The levels of serum D-D and NLR in severe patients were higher than those in moderate patients ( P<0.05) . Spearman correlation analysis showed that the severity of the disease was negatively correlated with serum APN levels ( r=-0.414, P<0.001) , and positively correlated with serum D-D and NLR levels in patients with diabetic foot ulcer infection ( r=0.387, P<0.001; r=0.461, P<0.001) . Univariate analysis showed that the proportion of severe disease, serum fasting blood glucose, glycosylated hemoglobin, fibrinogen, D-D and NLR levels in patients with poor prognosis were higher than those in patients with good prognosis ( P<0.05) , and the APN level in patients with poor prognosis was lower than that in patients with good prognosis ( P<0.05) . Logistic multivariate regression analysis showed that severe disease, serum glycosylated hemoglobin, APN, D-D, and NLR levels were independent risk factors for poor prognosis in patients with diabetic foot ulcer infection ( P<0.05) . ROC analysis showed that the optimal cut-off points of serum APN, D-D and NLR levels for predicting poor prognosis of patients were 5.73 mg/L, 3.06 mg/L, 2.12, the sensitivity was 78.57%, 82.14%, 85.71%, the specificity was 76.56%, 67.19%, 73.44%, the area under the curve (AUC) was 0.793, 0.784, 0.818, the specificity and AUC of the three were 98.44 %, 0.918, respectively. Conclusions:Serum APN, D-D and NLR levels are related to the severity of the disease in patients with diabetic foot ulcer infection. Clinical detection of serum APN, D-D and NLR levels can be used as sensitive indicators to predict poor prognosis.

Objective:To investigate the correlation between serum complement C1q/tumor necrosis factor associated protein 3 (CTRP3) and carotid atherosclerosis in patients with type 2 diabetes mellitus (T2DM) and nonalcoholic fatty liver disease (NAFLD).Methods:From January 2018 to December 2019, 111 T2DM patients hospitalized in the Endocrinology Department of Nantong Third People ′s Hospital Affiliated to Nantong University, and 30 healthy physical examiners in the physical examination center of Nantong Third People 's Hospital Affiliated to Nantong University in the same period were selected. Thirty cases of healthy physical examination were the control group, 111 cases of T2DM were divided into 52 cases of T2DM group and 59 cases of T2DM+NAFLD group according to whether they were combined with NAFLD. The cross-sectional study method was used to collect the relevant clinical data of three groups. The comparison data between multiple groups conformed to the normal distribution and the variance was uniform. One way ANOVA was used. SNK- q test was used for pairwise comparison, χ2 test for qualitative data comparison. The correlation between carotid intima-media thickness (IMT) and influencing factors was analyzed by partial correlation analysis, and the influencing factors of carotid IMT were analyzed by multi factor linear regression. Results:In the control group, T2DM group and T2DM+NAFLD group, body mass index (BMI) (23.65±2.81), (25.52±3.12), (24.90±2.94) kg/m 2,systolic blood pressure (119.43±15.81), (130.63±10.20), (139.37±14.11) mmHg, diastolic blood pressure (72.93±9.74), (73.40±9.44), (77.97±10.00) mmHg, and fasting blood glucose (5.12±0.77), (9.78±1.37), (9.24±1.46) mmol/L,glycosylated hemoglobin (HbA1c) (4.87±1.43)%, (7.99±1.10)%, (8.56±1.29)%,homeostasis model assessment of insulin resistance (HOMA-IR)(1.56±0.37),(2.80±1.00), (3.47±0.94), high density lipoprotein cholesterol (HDL-C) (1.52±0.34),(1.23±0.31), (1.22±0.31) mmol/L,low density lipoprotein cholesterol (LDL-C) (2.41±0.53), (2.73±0.61), (2.93±0.59) mmol/L, CTRP3 (292.93±68.54), (241.69±61.01), (150.80±56.67) μg/L, the difference between groups were statistically significant ( F=3.712,23.023,4.074,134.285,90.818,47.105,10.139,7.941,60.035,all P<0.05). Pairwise comparison shows that the systolic blood pressure, diastolic blood pressure, HbA1c and HOMA-IR in T2DM+NAFLD group were higher than those in control group and T2DM group,and CTRP3 was lower than those in control group and T2DM group, the difference was statistically significant (all P<0.05). BMI, fasting blood glucose, HbA1c, HOMA-IR, HDL-C and LDL-C in T2DM group were higher than those in the control group, CTRP3 was lower than that in the control group (all P<0.05). In the control group, T2DM group and T2DM+NAFLD group, IMT were (0.75±0.13), (1.11±0.17) and (1.25±0.15) cm; Crouse scores were (1.28±0.97), (3.22±1.42) and (4.54±1.22); the plaque detection rates 16.7%(5/30), 65.4%(34/52) and 78.0%(46/59), and there were significant differences between the two groups ( F=105.941,67.063, χ2=32.108, all P<0.001). There were significant differences between the two groups (all P<0.05). T2DM+NAFLD group was the highest, followed by T2DM group, and the control group was the lowest. Partial correlation analysis showed that carotid IMT was positively correlated with systolic blood pressure, fasting blood glucose, HbA1c, HOMA-IR, triglyceride and LDL-C ( r=0.356, 0.572, 0.575, 0.620, 0.172, 0.291, all P<0.05), and negatively correlated with HDL-C and CTRP3 ( r=-0.335, -0.675, all P<0.001). Multivariate linear regression analysis showed that HbA1c, HDL-C and ctrp3 were the influencing factors of carotid atherosclerosis ( t=2.621, -3.764, -7.280, all P<0.05) Conclusion:Serum CTRP3 is associated with carotid atherosclerosis in T2DM patients with NAFLD,and may have a protective effect on vascular lesions in T2DM patients with NAFLD.

ObjectiveTo explore the role of interleukin-6 （IL-6）/signal transducer and activator of transcription 3 （STAT3） pathway in the balance of T helper 17 （Th17）/regulatory T （Treg） cells in ulcerative colitis （UC） with internal dampness-heat accumulation syndrome and the intervention mechanism of Shaoyaotang. MethodA total of 60 SD rats were randomized into blank group （equivalent volume of normal saline）， model group （equivalent volume of normal saline）， western medicine control group （0.42 g·kg^-1 mesalazine）， and low-dose （11.1 g·kg^-1）， medium-dose （22.2 g·kg^-1）， and high-dose （44.4 g·kg^-1） Shaoyaotang groups. UC with internal dampness-heat accumulation syndrome was induced in rats with the compound method except for the blank group. The administration lasted 14 days for each group. At 24 h after the last administration， rats were killed and the spleen and colon tissues were separated. The histopathological changes of colon were observed based on hematoxylin and eosin （HE） staining and the levels of interleukin-17 （IL-17） and transforming growth factor-β₁ （TGF-β₁） in colon tissue were detected by immunohistochemistry （IHC）. Flow cytometry was employed to determine the levels of Th17/Treg cells in the spleen， and Western blot to measure the levels of IL-6 and STAT3 proteins in colon tissue. ResultCompared with the blank group， the model group had lesions such as congestion and erosion， low percentage of spleen Treg cells （P<0.01）， high percentage of Th17 cells （P<0.01）， and high levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. Compared with the model group， the administration groups showed alleviation of colon injury， high percentage of spleen Treg cells （P<0.05， P<0.01）， low percentage of Th17 cells （P<0.01）， and low levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. ConclusionShaoyaotang regulates the balance of Th17/Treg by inhibiting the IL-6/STAT3 pathway， thereby relieving the pathological damage of UC rats with internal dampness-heat accumulation syndrome and affecting their immune function.

Objective: To investigate the prevalence of hyperuricemia and its correlation among adolescents aged 13-16 in Hanjiang district of Yangzhou city. Methods: Five hundred and nine adolescents aged 13-16 years in Hangjiang district of Yangzhou city were recruited. Their fasting serum uric acid level were measured by Benecheck premium UA monitoring system. The demographic imformation, family history, eating and living habits of the respondents were collected by questionnaires. Results: The average serum uric acid level of 509 adolescents in Hangjiang district of Yangzhou city was (374±98) μmol/L, and (420±103) μmol/L in males, higher than that of females [(329±70) μmol/L](t=11.69, P<0.05). The overall prevalence of HUA was 36.9%(188/509), and 43.2%(108/250) in males, higher than in females 30.8%(80/259)(χ²=4.16, P<0.05). Multivariate Logistic regression analysis showed that male [OR=1.70, 95%CI(1.18, 2.45)], obese [OR=8.99, 95%CI(4.58, 17.63)], high-salt oily diet [OR=1.67, 95%CI(1.05, 2.61)], soy products [OR=2.66, 95%CI(1.45, 4.88)], animal offal [OR=7.81, 95%CI(3.42, 17.83)], seafood [OR=4.80, 95%CI(2.83, 8.12)], sugary drinks [OR=6.49, 95%CI(3.08, 13.64)] and time spent on electronic products [OR=1.98, 95%CI(1.07, 3.65)] might be risk factors for hyperuricemia (P<0.05), while fruit [OR=0.52, 95%CI(0.32, 0.85)] and outdoor activities [OR=0.55, 95%CI(0.33, 0.92)] might be protective factors for HUA (P<0.05). Conclusion There is a high prevalence of hyperuricemia among adolescents aged 13-16 in Hangjiang district of Yangzhou city. The level of serum uric acid in adolescents significantly increased. Male, obesity, high salt and oily diets, soy products, animal offal, seafood, and sugary drinks may be risk factors for HUA. Fruits and outdoor activities may be protective factors for HUA. Relevant disciplines can take corresponding measures to prevent adolescent HUA.