Objective:To investigate the role and related mechanisms of the LiaSR two-component system in acid tolerance and biofilm formation abilities of Streptococcus mutans (Sm) 593. Methods:The growth curves of various Sm strains in pH=5.5 brian heart infusion (BHI) medium were analyzed. And colony forming unit (CFU) was also performed to evaluate the acid tolerance of Sm. Laurdan probe, H +-K +adenosine triphosphate (ATP)ase activity analysis kit, proton permeability assay and real-time fluorescence quantitative PCR (RT-qPCR) were conducted to detect the acid tolerant mechanisms of LiaSR two-component system in Sm. Crystal violet staining, CFU, SYTOX probe and anthrone-sulfuric method were used to analyze the properties and structures of the Sm biofilms. RT-qPCR was conducted to detect the expression levels of underlying regulated genes. Results:The growth of mutants in acidic BHI were inhibited ( P<0.05). The acid tolerance of mutants significantly decreased compared to the wild-type strain ( P<0.05). In mutants, the activity of H +-ATPase (917.06±59.53 and 469.53±47.65) were elevated by 7.22-folds and 3.70-folds compared to the wild-type strain (127.00±50.71) ( P<0.001, P<0.001) and the encoded gene atpD (3.39±0.21 and 1.94±0.17) were also elevated by 3.39-folds and 1.94-folds compared to the wild-type strain (1.00±0.15) ( P<0.001, P=0.001). The Laurdan generalized polarization of mutants (0.18±0.04 and 0.18±0.05) increased significantly compared to the wild-type strain (0.08±0.05) ( P=0.006, P=0.003) and the expression levels of fabM gene were decreased in mutants (0.52±0.11 and 0.57±0.05) by 1/2 ( P=0.014, P=0.022). In liaR deletion mutant, the reduced terminal pH (4.76±0.01) can also be observed ( P<0.001). The total amount of the biofilms of three Sm didn't show significant differences ( P>0.05). But the number of viable bacteria of mutants′ biofilms were decreased [Sm 593: (12.00±2.80)×10 7 CFU/ml; Sm ΔliaS: (2.95±1.13)×10 7 CFU/ml; Sm ΔliaR: (7.25±1.60)×10 7 CFU/ml] ( P=0.001, P=0.024). The extracellular DNA were increased by 18.00-folds and 6.50-folds in mutants′ biofilms (128.73±15.65 and 46.38±5.52) compared to the wild-type strain (7.16±3.62) ( P<0.001, P=0.003). Water-soluble exopolysaccharides could be found up-regulated in liaS deletion mutant [(138.73±10.12) μg/ml] ( P=0.003) along with the expression level of gtfC gene (1.65±0.39) ( P=0.014). The expression level of gtfD were elevated by 47.43-folds and 16.90-folds in mutants ( P<0.001, P=0.010). Conclusions:The LiaSR two-component system can promote the expression of fabM gene and increase the fluidity of Sm which contributes to acid tolerance. The LiaR can also decrease the proton permeability and restrict the entrance of H +. The LiaSR two-component system can negatively regulate the production of the extracellular matrix in Sm biofilm.

Objective:To evaluate the feasibility of using positive anti-M-type phospholipase A 2 receptor(PLA 2R)antibody detection as a surrogate for renal biopsy in the diagnosis of idiopathic membranous nephropathy(IMN)in elderly Chinese. Methods:From June 2021 to March 2022, clinicopathological data of ninety-six elderly patients(≥60 years old)with positive anti-PLA 2R antibody detection(ELISA≥14 RU/ml), complete renal pathology records, and the exclusion of secondary disease and diabetes mellitus were collected from the Department of Nephrology, The First Affiliated Hospital of Zhengzhou University.Patients were divided into high eGFR group(≥60 ml·min -1·1.73 m -2)and low eGFR group(<60 ml·min -1·1.73 m -2), and the data were analyzed retrospectively. Results:Of the 96 patients, 95(99.0%)had IMN(1 in stage Ⅰ, 59 in stage Ⅱ, 34 in stage Ⅲ, and 1 in stage Ⅳ), and 1(1.0%)had atypical membranous nephropathy(AMN).For the IMN patients, 81(85.3%)had high eGFR and most IMN patients(66.7%)were in stage Ⅱ, with 10(12.3%)showing additional pathological findings, including mild mesangial proliferative IgA nephropathy(2 cases)and subacute tubulointerstitial nephropathy(2 cases).There were 14 patients(14.7%)with low eGFR, and most of them(57.1%)had stage Ⅲ IMN, with 10(71.4%)showing additional pathological findings.This percentage was higher than that in the high eGFR group( χ2=21.642, P<0.05).The most common additional pathological findings were acute tubular injury(4 cases)and ischemic kidney injury(2 cases). Conclusions:Positive anti-PLA 2R antibody detection is highly predictive of IMN in elderly Chinese patients, but it often co-exists with other pathological entities.The advantages of renal biopsy in detecting other pathological conditions and the risks associated with the procedure should be evaluated.

Objective:To analyze the clinical features of severe refractory mycoplasma pneumoniae pneumonia (SRMPP) in children, and explore its risk factors complicated with extrapulmonary organ dysfunction.Methods:The clinical data of children with SRMPP who were admitted to the Department of Critical Care Medicine of Shanghai Children's Hospital from July 2017 to June 2019 were retrospectively summarized. The patients were divided into two groups according to the occurrence of extrapulmonary organ dysfunction: the extrapulmonary organ dysfunction group and the respiratory dysfunction group. The differences of clinical features and laboratory indexes between the two groups were compared, and the risk factors of extrapulmonary organ dysfunction were screened out by logistic regression analysis.Results:A total of 107 cases with SRMPP were admitted to the Pediatric Intensive Care Unit during the past two years, and there were 44 cases (41.1%) complicated with pleural effusion, 17 cases (15.9%) with plastic bronchitis, 104 cases (97.2%) with positive results for macrolide resistance genes (2063, 2064), with an in-hospital mortality rate of 2.8% (3/107). Among 107 children with SRMPP, there were 51 cases (47.7%) with extrapulmonary organ dysfunction, 43 cases (40.2%) with cardiovascular dysfunction, 13 cases (12.1%) with coagulation dysfunction, 11 cases (10.3%) with gastrointestinal dysfunction, 4 cases (3.7%) with renal dysfunction, 4 cases (3.7%) with brain dysfunction, 3 cases (2.8%) with liver dysfunction, and 16 cases (15.0%) with multiple organ dysfunction. Compared with the respiratory dysfunction group, the incidence of capillary leak syndrome was higher (52.9% vs. 17.9%, P < 0.001), the capillary leak index was increased [11.71 (4.63, 27.07) vs. 5.78 (2.07, 15.71), P =0.019], serum albumin was decreased [(32.2 ± 5.6)g/L vs. (34.7 ± 6.7)g/L, P=0. 041], and prothrombin time was prolonged significantly [12.7 (11.7, 13.8)s vs. 12.0 (11.4, 13.0)s, P=0. 009]. Logistic regression analysis showed that capillary leak syndrome ( OR=0. 278, 95% CI 0.102-0.759, P=0. 013) and prolonged prothrombin time ( OR=1. 443, 95% CI 1.018-2.046, P=0. 039) were independent risk factors for SRMPP complicated with extrapulmonary organ dysfunction. Conclusions:Approximately 50% of children with SRMPP have dysfunction of extrapulmonary organs, such as circulation, coagulation and gastrointestinal disorders. Capillary leak syndrome and prolonged prothrombin time are independent risk factors for SRMPP complicated with extrapulmonary organ dysfunction.

Objective:To evaluate the predictive value of lung ultrasound on mortality in children with severe acute respiratory distress syndrome (ARDS) undergoing extracorporeal membrane oxygenation (ECMO) support.Methods:A prospective observational study was used to enroll patients with severe ARDS who met the Berlin criteria in the Pediatric Intensive Care Unit of Children’s Hospital of Shanghai Jiao Tong University from January 2016 to December 2019. Patients with ECMO support <3 d, lack of appropriate acoustic windows, with severe pneumothorax, and secondary to congenital heart disease or chronic lung disease were excluded. ECMO parameters, respiratory mechanics parameters and outcome were collected and analyzed. Lung ultrasound score (LUS) was measured at the initiation of ECMO as LUS-0 h, then at 24 h, 48 h, 72 h, and 7 d after ECMO support as the value of LUS-24 h, LUS-48 h, LUS-72 h, LUS-7 d, as well as after weaning ECMO as LUS-w. The patients were divided into survivors and non-survivors according to hospital survival status. Receiver operating characteristic (ROC) curve and Kaplan-Meier survival analysis curve were performed to explore the predictive value of lung ultrasound on mortality in patients with severe ARDS undergoing ECMO.Results:A total of 26 patients were enrolled in this study, of which 18 patients survived and 8 died. There were no significant differences in PRISM Ⅲ, dynamic pulmonary compliance (Cdyn), oxygenation index, PaO 2/FiO 2, and PaCO 2 on PICU admission between the two groups (all P>0.05). The values of LUS-72 h and LUS-w in non-survivors were significantly higher than those in survivors [26 (24, 29) vs16 (13, 19), P<0.01] and [30 (26, 35) vs11 (10, 13), P<0.01]. The values of Cdyn-72 h, Cdyn-7 d and Cdyn-w in survivors were significantly higher than those in non-survivors [0.48 (0.42, 0.54)mL/cmH 2O·kg vs 0.36 (0.29, 0.40) mL/cmH 2O·kg, P<0.01; 0.60 (0.52, 0.67) mL/cmH 2O·kg vs 0.27 (0.13, 0.30) mL/cmH 2O·kg, P<0.01, and 0.66 (0.62, 0.70) mL/cmH 2O·kg vs 0.30 (0.13, 0.35) mL/cmH 2O·kg, P<0.01]. ROC curve analysis showed that an area under ROC curve (AUC) of LUS-72 h for predicting PICU mortality was 0.955 (95% CI: 0.864-1.000; P<0.01). The cutoff value of LUS-72 h was 24 with a sensitivity of 87.5% and a specificity of 100.0%. Kaplan-Meier survival analysis showed that PICU mortality of patients with LUS-72 h≥24 was significantly higher than that in patients with LUS-72 h < 24 ( P<0.01) . Conclusions:Lung ultrasound is an effective tool for monitoring progress of children with severe ARDS received ECMO support. LUS-72 h >24 is an index to predict the worsen outcome in children with severe ARDS under ECMO support.

Objective:To investigate the effects of therapeutic plasma exchange(TPE)as adjuvant therapy in children with myasthenia gravis(MG)in pediatric intensive care unit(PICU).Methods:A retrospective study was conducted in 7 children with MG admitted to PICU at Shanghai Children′s Hospital from January 2016 to December 2019.TPE was performed on unsatisfactory effect of acetylcholinesterase inhibitors, glucocorticoids or IVIG.The TPE dose was 50-70 mL/kg for 2 to 3 times for each case.The clinical symptoms, anti-acetylcholine antibody(AChR-Ab)level and prognosis were measured before and after TPE.Results:Seven children with myasthenia gravis admitted to PICU from January 2016 to December 2019, including 4 cases of systemic myasthenia gravis(1 case of myasthenia crisis with respiratory failure)and 3 cases of ocular myasthenia gravis.The AChR-Ab level decreased from 1.66(0.99, 3.33)nmol/L before TPE to 0.66(0.40, 10.97)nmol/L after TPE( Z=-2.545, P=0.011). The symptoms of muscle weakness and blepharoptosis were partially or completely relieved in 7 cases.There were no significantly changes in the levels of circulating immune complex, complement C3, CD4 + , CD8 + and NK cells before and after TPE(all P>0.05). During the process of TPE, 2 cases had mild rash, and 1 case had hypotensive shock, which were recovered after timely treatment.After TPE, the fibrin levelsdecreased from 1.90(1.40, 2.40)g/L to 1.10(1.00, 1.30)g/L( Z=-3.092, P=0.002). Conclusion:TPE reduce the AChR-Ab levels and improve the short-term symptoms in children with myasthenia gravis who have failed conventional treatment.TPE may be an optional therapy for pediatric severe MG.

Objective:To investigate the efficacy of extracorporeal membrane oxygenation (ECMO) in rescuing refractory severe hypoxic respiratory failure in children.Methods:Patients with refractory hypoxic respiratory failure who treated with ECMO from July 2016 to June 2019 in the Department of Intensive Medicine (PICU) of Shanghai Children's Hospital Affiliated to Shanghai Jiaotong University were enrolled in this study. Their oxygen index, initial ECMO timing, organ function, in-hospital survival rate, and ECMO-related complications were collected and analyzed.Results:Totally 30 patients with severe refractory hypoxemia who received ECMO therapy were included. The mean age of patients was 19 ( IQR 9.75, 52) months. Twenty-one patients (70%) treated with veno-arterial (VA-ECMO), 7 patients (23.3%) received veno-venous ECMO (VV-ECMO), and mixed model [one patient (3.3%) VV to VA-ECMO, and one patient (3.3%) VA to VV-ECMO]. The mean duration of ECMO was 161.5 ( IQR 91, 284) h. The total in-hospital survival rate was 66.7%, of which the VA-ECMO survival rate 59.1% (13/21) and the VV-ECMO survival rate 85.7% (6/7) without significant difference (χ 2=1.365, P=0.243). The mean oxygenation index (OI) at initial ECMO intervention was 43 ( IQR 35.3, 60.8) in the non-survival group, which was significantly higher than that in the survival group [26.5 ( IQR 20, 45.3), z=-2.267, P =0.023]. The course of confirmed respiratory failure at ECMO intervention was 129 ( IQR 90.25, 197) h in the non-survival group, which was significantly longer than that in the survival group [54.5 ( IQR 16.25, 121.75) h, z=-2.2464, P =0.014]. Conclusions:Compared with VA-ECMO, the survival rate has a better tendency in VV-ECMO treated patients with refractory severe hypoxic respiratory failure. And patients with OI >43 or severe hypoxic respiratory failure diagnosed for more than 5 days with ECMO support may have worsen prognosis.

Hypoglycemia crisis and hyperglycemia crisis are the most common endocrine and metabolic emergencies in PICU.Hypoglycemic crisis is caused by low blood glucose level(<2.2 mmol/L) and/or rapid reduction in blood glucose, which leads to autonomic neurogenic symptoms and central nervous system injury, even causes permanent brain injury and death.Early identification and timely glucose infusion are the key to improving prognosis.Hyperglycemic crisis is caused by abnormally increased blood glucose levels(>11 mmol/L), including diabetic ketoacidosis and hyperglycemic hyperosmolar state.The treatment principle of diabetic ketoacidosis and hyperglycemic hyperosmolar state include rapid fluid resuscitation to restore blood volume, control blood glucose, correct electrolyte and acid-base balance disorders, discover and removal of incentives and prevent complications.Cerebral edema and osmotic demyelination syndrome are the most fatal complications of hyperglycemic crisis.Early identification and active treatment can reduce the mortality.

Objective To investigate the effect of pyrin domain 3 (NLRP3) inflammasome in the process of contrast induced human kidney cell apoptosis.Methods Human kidney 2 (HK-2) cells were cultured in DMEM-F12 medium with 5％ FBS.Cells were divided into control group,Contrast group (O group),NLRP3-siRNA+Iohexol group (si-NLRP3+O group),ASC-siRNA+Iohexol group (si-ASC+O group),and mannitol group (M group).Different concentrations of hypotonic contrast agent were added to HK-2 cell culture plates for 24,48 and 72 h.Flow cytometry was used to detect apoptosis.NLRP3 and ASC mRNA expressions were detected by RT-PCR.The expressions of NLRP3,ASC,caspase-8/cleaved caspase-8,Bcl-2/Bax,caspase-1/cleaved caspase-1,and caspase-3/cleaved caspase-3 protein were detected by Western blot.The levels of interleukin (IL) 1β and IL-18 in supernatant were detected by ELISA.Results Compared with the control group,the rate of apoptotic cells,as well as the expressions of NLRP3,ASC and cleaved caspase-1 proteins were increased in HK-2 cells of contrast group.The expressions of NLRP3 and ASC mRNA in the contrast group also increased,so did IL-1β and IL-18 levels (all P＜0.05),suggesting that NLRP3 inflammasome in HK-2 cells was activated by contrast.Compared with the control group,the expressions of cleaved caspase-8,Bax and cleaved caspase-3 protein were increased,and the expression of anti-apoptotic protein Bcl-2 was decreased (all P ＜ 0.05).Compared with the contrast group,the rate of apoptotic cells in the si-NLRP3 + contrast group and si-ASC + contrast group was significantly decreased;the expression of cleaved caspase-1 was decreased;the expressions of Bax and cleaved caspase-3 were decreased,and Bcl-2 level was increased.The expressions of IL-1β and IL-18 in the supernatant of cells were decreased (all P ＜ 0.05).Conclusion Contrast agent can activate the NLRP3 pathway in HK-2 cells and induce apoptosis,which could be reduced by blocking the NLRP3 pathway.

Objective: To investigate the incidence and prognosis of hypophosphatemia in critically ill children treated with continuous blood purification (CBP). Methods: The medical records of the critically ill patients, who were treated with CBP, admitted to pediatric intensive care unit (PICU) of Shanghai Children's Hospital from May 2014 to April 2017 were retrospectively analyzed. The serum phosphorus levels were tested before CBP, at 48-72 h during CBP, at the end of CBP and on the next day after CBP finished. Phosphorus supplement was given to the children with severe hypophosphatemia. Results: A total of 85 patients met the inclusion criteria. The serum phosphorus levels at the 4 indicated time points were (1.4±0.5), (0.7±0.3), (0.8±0.3), (0.9±0.4) mmol/L, respectively (F=45.21, P<0.05). Among the children, 66 cases (78%) had hypophosphatemia during CBP. The incidences of moderate and severe hypophosphatemia were 32 (48%) and 9 (14%), respectively. There were 41 patients with CBP replacement rates of (35-49) ml/(kg·h), while 44 patients with CBP replacement rates of 50-70 ml/(kg·h). There were significant differences at 48-72 h during CBP, the end of CBP and on the next day after CBP ((0.8±0.4) vs. (0.5±0.2), (1.0±0.3) vs. (0.6±0.2), and (1.1±0.4) vs. (0.8±0.2) mmol/L; t=7.672, 4.060, 14.440, P<0.05). Atotal of 9 cases were treated with sodium glycerophosphate. Among the 85 children, 24 (28%) patients died while 61 (72%) survived. There were no significant differences between the two groups in serum phosphorus levels at the indicated time points ((1.4±0.5) vs. (1.4±0.5), (0.7±0.3) vs. (0.7±0.3), (0.7±0.3) vs. (0.8±0.3), and (1.0±0.3) vs. (0.9±0.3) mmol/L, respectively, P>0.05). Conclusions Hypophosphatemia is prone to occur during CBP, which probably related to the replacement rate. There was no significant relationship between hypophosphatemia and mortality in critically ill children after giving phosphorus supplementation.

Objective To investigate the association and predictive value of aspartate transaminase to platelet ratio index(APRI) in sepsis-associated liver injury(SALI). Methods We retrospectively ana-lyzed the medical records of patients with sepsis admitted to PICU in Shanghai Children′s Hospital of Shanghai Jiaotong University from April 2015 to March 2017. According to whether liver injury occurred in the sepsis patients during hospitalization,all the patients were divided into SALI group (n=34) and sepsis group(n=222). The clinical characteristics,serological indexes within 24 hours in the PICU,and the ratio of aspartate transaminase to alanine transaminase( AAR) and APRI were collected and analyzed. The receiver operating characteristic( ROC) curve was used to evaluate the power of APRI for the prediction of SALI. Results (1)A total of 256 patients were enrolled in this study. There were 34 cases with SALI,and there were 222 patients with sepsis only,the incidence of SALI was 13. 3％. (2) The values of APRI and AAR were both higher in the SALI group compared with the sepsis group[APRI:7. 12(1. 71,26. 96) vs. 0. 38 (0. 21,0. 83),P<0. 001;AAR:1. 43(0. 94,2. 69) vs. 2. 17(1. 35,2. 96),P<0. 05]. (3)The multivariate Logistic regression analysis showed that total bilirubin, APRI, AAR and platelet were the independent risk factors of SALI(P<0. 05). (4)In addition,the area under the ROC curve(AUC)for the APRI was 0. 891 (95％CI 0. 815-0. 966,P<0. 001),cut-off value was 1. 73,which was superior to total bilirubin(AUC =0. 744,95％CI 0. 634-0. 853,P<0. 001) and platelet(AUC=0. 726,95％CI 0. 611-0. 841,P<0. 001). The clinical sensitivity and specificity of the APRI for identification of SALI from sepsis was 80. 0％ and 92. 2％, respectively. Conclusion APRI is an independently risk factor for the occurrence of SALI and is a precursory marker for SALI.