BACKGROUND: How to acquire high-purity epidermal stem cells (ESCs) and maintain ESC phenotype and in vitro proliferation characteristics are urgently solved problems in the in vitro culture of ESCs.OBJECTIVE: To establish methods to in vitro isolate and culture human ESCs.DESIGN,TIME AND SETTING: Cell observation was performed in the Department of Bum,First Affiliated Hospital of Nanchang University.MATERIALS: Foreskin was obtained front 2-12-year-old patients who were subjected to circumcision.These patients did not suffer from urinary system infection.METHODS: Keratinocytes and fibroblasts were isolated from foreskin using Dispase Ⅱ and trypsin.ESCs were isolated by rapidly adhering to type Ⅳ collagen.The supematant fluid of fibroblasts of passages 2-3,which were in the exponential growth phase,was collected.After screening,the supematant fluid was mixed with serum-free medium of keratinocytes at the proportion of 1:1.Simultaneously,fetal bovine serum,epidermal growth factor,and bovine pituitary extract were added to prepare medium of ESCs,which was used for in vitro proliferation of human ESCs.Homologous keratinocytes were used as controls.MAIN OUTCOME MEASURES: After 2 passages,cloning efficiency and cloning sustaining time were calculated by plate clone formation assay.Expression levels of β 1 integrin and keratin 19 were detected by immunohistochemistry.RESULTS: The cloning efficiency of human ESCs was significantly increased,and the cloning sustaining time was significantly prolonged as compared with homologous keratinocytes cultured in the same batch (P ＜ 0.01).lmmunohistochemistry results demonstrated that both β 1 integrin and keratin 19 were positive in the human ESCs.CONCLUSION: Keratinocytes could be successfully isolated by means of rapid adherence to type Ⅳ collagen and cultured with conditioned medium in vitro.

Objective To investigate the clinical significance of pulse high-volume hemofiltration (PHVHF) on expressions of Toll-like receptor (TLR) 2 and TLR4 mRNA in peripheral blood mononuclear cells (PBMCs) in patients with severe sepsis.Methods Forty patients with severe sepsis were divided into conventional treatment group (n =20) and PHVHF group (n =20) according to random number table.Another fifteen healthy volunteers served as controls.TLR2 and TLR4 mRNA expressions in PBMCs were detected using RT-PCR and plasma concentrations of TNF-α and IL-6 were measured using ELISA method before therapy and at 24 h,48 h and 72 h after therapy.Vital signs,BIL,Cr,BUN,Lac,PaO2/FiO2,acute physiology,chronic health evaluation Ⅱ (APPACHE Ⅱ),sequential organ failure assessment (SOFA) and prognosis were compared among the groups.Besides,complications associated with PHVHF therapy were monitored.Results Expressions of TLR2 and TLR4 mRNA in PBMCs and concentrations of TNF-α and IL-6 were significantly higher in patients with severe sepsis than in the controls (P ＜ 0.01).At 72 hours after therapy,PHVHF group showed significantly lower concentrations of TNF-α and IL-6 than those before therapy (P ＜ 0.01) as well as significantly lower expressions of TLR2 and TLR4 mRNA in PBMCs than those in conventional treatment group (P ＜ 0.O1).However,no significant decline in the levels of TNF-α and IL-6 and the expressions of TLR2 and TLR4 mRNA in PBMCs were shown in conventional treatment group after therapy.At 72 hours after therapy,PHVHF group showed significant increases of MAP and PaO2/FiO2 and significant decreases of Cr,BUN,Lac,APACHE Ⅱ and SOFA as compared to those before therapy (P ＜ 0.05).Moreover,the differences of MAP,PaO2/FiO2,Cr,BUN,Lac,APACHE Ⅱ and SOFA were statistically significant between and conventional treatment group at 72 hours after therapy (P ＜ 0.05).Conclusion PHVHF achieves a reduced systemic inflammatory response,improved major organ functions,shortened length of stay in ICU and down-regulated TLR2 and TLR4 expressions in PBMCs that may be a novel mechanism of PHVHF in treatment of severe sepsis.

Objective:To investigate the prognostic value of cardiac ultrasound left ventricular ejection fraction (LVEF) on admission in patients with septic cardiomyopathy.Methods:A retrospective cohort study was conducted. The patients with septic cardiomyopathy hospitalized in the intensive care unit of Zhoupu Hospital Affiliated to Shanghai Health College from January 2019 to March 2023 were enrolled. The general information including gender and age, LVEF on admission, severity of illness scores within 24 hours after admission [acute physiology and chronic health evaluationⅡ (APACHEⅡ) score and sequential organ failure assessment (SOFA) score], procalcitonin (PCT), cardiac biomarkers [N-terminal pro-brain natriuretic peptide (NT-proBNP), cardiac troponin T (cTnT), and MB isoenzyme of creatine kinase (CK-MB)], mitochondria related indicators [aspartate aminotransferase (AST), AST/alanine aminotransferase (ALT) ratio], blood lactate (Lac), the usage of vasoactive drugs and mechanical ventilation, and the prognosis during hospitalization were collected. The differences in above clinical data between the two groups were compared. The variables with statistically significant differences in univariate analysis were incorporated into multivariate Logistic regression analysis to analyze the independent risk factors for death during hospitalization in patients with septic cardiomyopathy. Receiver operator characteristic curve (ROC curve) was drawn to evaluate the prognostic value of LVEF by echocardiography on admission in patients with septic cardiomyopathy during hospitalization.Results:A total of 62 patients were enrolled, including 36 males and 26 females. Thirty-nine cases died and 23 cases survived during hospitalization, and the mortality was 62.90%. Compared with the survival group, the LVEF of patients on admission was lower in the death group [0.51 (0.40, 0.57) vs. 0.56 (0.51, 0.63), P < 0.01], APACHEⅡ score, SOFA score, Lac, NT-proBNP, CK-MB within 24 hours after admission were higher [APACHEⅡ score: 22.18±8.38 vs. 17.39±8.22, SOFA score: 9.90±3.87 vs. 7.09±3.27, Lac (mmol/L): 5.10 (2.63, 11.50) vs. 2.00 (1.40, 5.00), NT-proBNP (μg/L): 5.24 (2.84, 11.29) vs. 2.53 (0.35, 6.63), CK-MB (U/L): 1.88 (0.21, 5.33) vs. 0.17 (0.02, 1.62), all P < 0.05], and the proportion of vasoactive drug application was higher (82.05% vs. 47.83%, P < 0.01). Multivariate Logistic regression analysis showed that LVEF on admission was an independent risk factor for predicting the prognosis of patients with septic cardiomyopathy during hospitalization [odds ratio ( OR) = 0.920, 95% confidence interval (95% CI) was 0.855-0.990, P = 0.025]. ROC curve analysis showed that the area under the ROC curve (AUC) of LVEF on admission for predicting the death of patients with septic cardiomyopathy was 0.715 (95% CI was 0.585-0.845, P = 0.005). When LVEF ≤ 0.52, the sensitivity was 73.9%, and the specificity was 61.5%. Conclusions:The lower cardiac ultrasound LVEF on admission, the worse the prognosis of patients with septic cardiomyopathy. The cardiac ultrasound LVEF on admission can be used as a clinical index to evaluate the severity of the condition and predict the prognosis of patients with septic cardiomyopathy.

Objective:To evaluate the prognostic value of Karnofsky performance scores (KPS) in elderly patients with sepsis, so as provide a basis for clinical evaluation of the condition, prognosis and corresponding treatment measures.Methods:A retrospective cohort study was conducted to collect the general information, clinical data, and follow-up data of limb motor function status and self-care ability of elderly patients with sepsis who were hospitalized in the Intensive Care Unit of our hospital from January 2018 to June 2021. Patients were divided into the survival group and death group according to whether they survived the hospitalization. Statistical analysis was performed using t-test, chi-square test, and Mann-Whitney test. The KPS score before admission, disease severity scores (APACHEⅡ and SOFA), serum procalcitonin (PCT), N-terminal pro-brain natriuretic peptide (NT-proBNP), and arterial blood lactate level on admission were compared between the two groups. Then, the factors with significance in univariate analysis were analyzed by Logistic regression method, the independent risk factors for predicting in-hospital mortality were determined, and the receiver operating characteristic (ROC) curve was drawn to evaluate the prognostic value of KPS score in elderly patients with sepsis.Results:A total of 135 patients were collected. There were 85 males and 50 females, 60 died and 75 survived during hospitalization, with a mortality rate of 44.4%. The preadmission KPS score of elderly patients with sepsis in the death group was significantly lower than that in the survival group [30 (30, 40) vs. 70 (50, 90), P<0.001]. Multivariate logistic regression analysis showed that KPS score ( OR=0.938, 95% CI: 0.914-0.963, P<0.001), SOFA score ( OR=1.255, 95% CI: 1.066-1.451, P=0.002) and arterial blood lactate ( OR=1.219, 95% CI: 1.059-1.404, P=0.006) were independent risk factors for predicting the prognosis during hospitalization. ROC curve analysis of mortality showed that compared with SOFA score and blood lactate, the area under the curve of KPS score was the largest, with AUC of 0.830 (95% CI: 0.756-0.890, P<0.001). In addition, the combination of KPS, SOFA and blood lactate had a greater predictive value for the prognosis of elderly patients with sepsis than that of the single index, with an AUC of 0.883 (95% CI: 0.826-0.940, P<0.001). Conclusions:The lower the KPS score, the worse the prognosis of elderly patients with sepsis. The KPS score can be used as a clinical indicator to predict the prognosis of elderly patients with sepsis.

OBJECTIVE: To investigate the diagnostic value of Hepcidin as a sepsis biomarker in critically ill adults. METHODS: An observational study was conducted. The patients with suspected or proven infection admitted to intensive care unit (ICU) of Zhoupu Hospital Affiliated to Shanghai University of Medicine and Health Sciences from March 2016 to November 2017 were enrolled. According to the third international consensus definitions for sepsis and septic shock (Sepsis-3), the patients were divided into non-sepsis group and sepsis group, and the septic patients were subdivided into general sepsis subgroup and septic shock subgroup according to the severity of disease. The differences in serum Hepcidin, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), procalcitonin (PCT), C-reactive protein (CRP), white blood cell (WBC), neutrophil granulocytes (NEUT) and lactic acid (Lac) within 1 hour after ICU admission between non-sepsis and sepsis groups and among the sepsis subgroups were compared. The acute physiology and chronic health evaluation II (APACHE II) within 24 hours after ICU admission and sequential organ failure score (SOFA) were recorded, and the mortality rate was followed up for 28 days. Receiver operation characteristic curve (ROC) was used to evaluate and compare the diagnostic value of Hepcidin and PCT, CRP, WBC for sepsis. Logistic regression model was used to estimate the association between Hepcidin and sepsis. Spearman correlation analysis was used to analyze the correlation between Hepcidin and other parameters of sepsis patients. RESULTS: A total of 183 patients were enrolled, 93 in the non-sepsis group and 90 in the sepsis group (48 with general sepsis and 42 with septic shock). (1) The levels of Hepcidin, IL-6, TNF-α, PCT, Lac in serum, and APACHE II and SOFA scores in the sepsis group were significantly higher than those in the non-sepsis group. ROC analysis showed that the area under the ROC curve (AUC) of Hepcidin and PCT for sepsis diagnosis were 0.865 [95% confidence interval (95%CI) = 0.807-0.911] and 0.848 (95%CI = 0.788-0.897), respectively, without statistical significance (Z = 0.443, P = 0.657). Furthermore, the AUC of Hepcidin for sepsis diagnosis was significantly higher than that of the conventional biomarkers CRP and WBC [AUC was 0.530 (95%CI = 0.455-0.604) and 0.527 (95%CI = 0.452-0.601), respectively] with statistical significance (both P < 0.01). When Hepcidin > 54.00 μg/L, its sensitivity for sepsis diagnosis was 95.56%, specificity was 66.67%, positive and negative predictive value was 73.51% and 93.94%, respectively. Parallel test was conducted for combination of Hepcidin and PCT, which showed that the AUC was 0.885, and the sensitivity and negative predictive value was significantly improved to 98.96% and 98.36%, respectively. Logistic regression analysis demonstrated that after adjusted for PCT, Hepcidin > 54.00 μg/L was also associated with sepsis independently, with odds ratio (OR) of 1.011 (95%CI = 1.008-1.015, P < 0.001), indicating that Hepcidin and PCT were not completely overlapped in the diagnosis of sepsis. (2) With the increase in infection severity, serum Hepcidin, PCT, IL-6, TNF-α, Lac, APACHE II, SOFA score and 28-day mortality all showed an increasing trend in patients. There was a significantly positive correlation between Hepcidin and IL-6, TNF-α, PCT, APACHE II, and SOFA in the sepsis patients (r value was 0.526, 0.449, 0.591, 0.359, and 0.374, respectively, all P < 0.01), but no correlation was found between Hepcidin and Lac (r = 1.104, P > 0.05). CONCLUSIONS: Serum Hepcidin is a useful biomarker for the diagnosis of sepsis, and it is correlated to the severity of the sepsis. The combination of Hepcidin and PCT can improve the accuracy of diagnosis of sepsis. CLINICAL TRIAL REGISTRATION China Clinical Trial Registration Center, ChiCTR-DDD-16008522.
Adult ; Biomarkers ; C-Reactive Protein ; Calcitonin ; Calcitonin Gene-Related Peptide ; China ; Critical Illness ; Hepcidins ; Humans ; Prognosis ; Protein Precursors ; ROC Curve ; Sepsis

Objective:To investigate the effect of botulinum toxin type A on children with odorihidrosis.Methods:From March 2017 to February 2021, 121 children with odorihidrosis, including 48 males and 73 females, aged 13 to 17 (15.9±1.2) years, were admitted to the Burn and Plastic Surgery Department of the 980 Hospital of PLA. There were 24 cases in mild group, 50 cases in moderate group and 47 cases in severe group. Botulinum toxin A was injected into 20-50 points on each side, and 1 U was injected into each point. The total amount of botulinum toxin A was 50-100 U on both sides.Results:Three groups of children were evaluated for efficacy, 24 cases of mild group was significantly effective in 23 cases, accounting for 95.8%. In the moderate group, 46 (92.0%) of 50 cases showed obvious effect. 49 cases (98.0%) were effective; In the severe group, 40 cases (85.1%) showed obvious effect and 45 cases (95.7%) were effective. Three groups of children with different efficacy had no statistical significance ( P>0.05). The significant efficiency in mild and moderate groups was higher than that in severe group, and the difference was statistically significant ( P<0.05). Conclusions:Botulinum toxin type A is effective in the treatment of children with mild and moderate bromhidrosis, and is worthy of clinical application.

Objective: To investigate the diagnostic value of Hepcidin as a sepsis biomarker in critically ill adults. Methods: An observational study was conducted. The patients with suspected or proven infection admitted to intensive care unit (ICU) of Zhoupu Hospital Affiliated to Shanghai University of Medicine & Health Sciences from March 2016 to November 2017 were enrolled. According to the third international consensus definitions for sepsis and septic shock (Sepsis-3), the patients were divided into non-sepsis group and sepsis group, and the septic patients were subdivided into general sepsis subgroup and septic shock subgroup according to the severity of disease. The differences in serum Hepcidin, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), procalcitonin (PCT), C-reactive protein (CRP), white blood cell (WBC), neutrophil granulocytes (NEUT) and lactic acid (Lac) within 1 hour after ICU admission between non-sepsis and sepsis groups and among the sepsis subgroups were compared. The acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ) within 24 hours after ICU admission and sequential organ failure score (SOFA) were recorded, and the mortality rate was followed up for 28 days. Receiver operation characteristic curve (ROC) was used to evaluate and compare the diagnostic value of Hepcidin and PCT, CRP, WBC for sepsis. Logistic regression model was used to estimate the association between Hepcidin and sepsis. Spearman correlation analysis was used to analyze the correlation between Hepcidin and other parameters of sepsis patients. Results: A total of 183 patients were enrolled, 93 in the non-sepsis group and 90 in the sepsis group (48 with general sepsis and 42 with septic shock).① The levels of Hepcidin, IL-6, TNF-α, PCT, Lac in serum, and APACHEⅡand SOFA scores in the sepsis group were significantly higher than those in the non-sepsis group. ROC analysis showed that the area under the ROC curve (AUC) of Hepcidin and PCT for sepsis diagnosis were 0.865 [95% confidence interval (95%CI) = 0.807-0.911] and 0.848 (95%CI = 0.788-0.897), respectively, without statistical significance (Z = 0.443, P = 0.657). Furthermore, the AUC of Hepcidin for sepsis diagnosis was significantly higher than that of the conventional biomarkers CRP and WBC [AUC was 0.530 (95%CI = 0.455-0.604) and 0.527 (95%CI = 0.452-0.601), respectively] with statistical significance (both P < 0.01). When Hepcidin > 54.00 μg/L, its sensitivity for sepsis diagnosis was 95.56%, specificity was 66.67%, positive and negative predictive value was 73.51% and 93.94%, respectively. Parallel test was conducted for combination of Hepcidin and PCT, which showed that the AUC was 0.885, and the sensitivity and negative predictive value was significantly improved to 98.96% and 98.36%, respectively. Logistic regression analysis demonstrated that after adjusted for PCT, Hepcidin > 54.00 μg/L was also associated with sepsis independently, with odds ratio (OR) of 1.011 (95%CI = 1.008-1.015, P < 0.001), indicating that Hepcidin and PCT were not completely overlapped in the diagnosis of sepsis. ② With the increase in infection severity, serum Hepcidin, PCT, IL-6, TNF-α, Lac, APACHEⅡ, SOFA score and 28-day mortality all showed an increasing trend in patients. There was a significantly positive correlation between Hepcidin and IL-6, TNF-α, PCT, APACHEⅡ, and SOFA in the sepsis patients (r value was 0.526, 0.449, 0.591, 0.359, and 0.374, respectively, all P < 0.01), but no correlation was found between Hepcidin and Lac (r = 1.104, P > 0.05). Conclusions: Serum Hepcidin is a useful biomarker for the diagnosis of sepsis, and it is correlated to the severity of the sepsis. The combination of Hepcidin and PCT can improve the accuracy of diagnosis of sepsis. Clinical trial registration China Clinical Trial Registration Center, ChiCTR-DDD-16008522.