Development and external validation of a nomogram for the early prediction of acute kidney injury in septic patients: a multicenter retrospective clinical study.

Background and purpose: Acute kidney injury (AKI) is a common serious complication in sepsis patients with a high mortality rate. This study aimed to develop and validate a predictive model for sepsis associated acute kidney injury (SA-AKI). Methods: In our study, we retrospectively constructed a development cohort comprising 733 septic patients admitted to eight Grade-A tertiary hospitals in Shanghai from January 2021 to October 2022. Additionally, we established an external validation cohort consisting of 336 septic patients admitted to our hospital from January 2017 to December 2019. Risk predictors were selected by LASSO regression, and a corresponding nomogram was constructed. We evaluated the model's discrimination, precision and clinical benefit through receiver operating characteristic (ROC) curves, calibration plots, decision curve analysis (DCA) and clinical impact curves (CIC) in both internal and external validation. Results: AKI incidence was 53.2% in the development cohort and 48.2% in the external validation cohort. The model included five independent indicators: chronic kidney disease stages 1 to 3, blood urea nitrogen, procalcitonin, D-dimer and creatine kinase isoenzyme. The AUC of the model in the development and validation cohorts was 0.914 (95% CI, 0.894-0.934) and 0.923 (95% CI, 0.895-0.952), respectively. The calibration plot, DCA, and CIC demonstrated the model's favorable clinical applicability. Conclusion: We developed and validated a robust nomogram model, which might identify patients at risk of SA-AKI and promising for clinical applications.

Sequence Type 5 (ST5) as a Possible Predictor of Bacterial Persistence in Adult Patients with Methicillin-Resistant Staphylococcus aureus Pneumonia Treated with Vancomycin.

Vancomycin remains the mainstay of treatment for methicillin-resistant Staphylococcus aureus (MRSA) pneumonia. This study assessed risk factors for vancomycin failure in 63 patients with MRSA pneumonia through detailed clinical, microbiological, pharmacokinetic/pharmacodynamic, and genetic analyses of prospective multicenter studies conducted from February 2012 to July 2018. Therapeutic drug monitoring was performed during vancomycin treatment, and the 24-h area under the curve (AUC0-24) was calculated. All baseline strains were collected for MIC determination, heterogeneous vancomycin-intermediate S. aureus (hVISA) screening, and biofilm determination. Whole-genome sequencing was performed on the isolates to analyze their molecular typing and virulence and adhesion genes. Clinical signs and symptoms improved in 44 patients (44/63, 69.8%), with vancomycin daily dose (P = 0.045), peak concentration (P = 0.020), and sdrC (P = 0.047) being significant factors. Isolates were eradicated in 51 patients (51/63, 81.0%), with vancomycin daily dose (P = 0.009), cardiovascular disease (P = 0.043), sequence type 5 (ST5; P = 0.017), tst (P = 0.050), and sec gene (P = 0.044) associated with bacteriological failure. Although the AUC0-24/MIC was higher in the groups with bacterial eradication, the difference was not statistically significant (P = 0.108). Multivariate analysis showed that no variables were associated with clinical efficacy; ST5 was a risk factor for bacterial persistence (adjusted odds ratio, 4.449; 95% confidence interval, 1.103 to 17.943; P = 0.036). ST5 strains had higher frequencies of the hVISA phenotype, biofilm expression, and presence of some adhesion and virulence genes such as fnbB, tst, and sec than non-ST5 strains. Our study suggests that ST5 is a possible predictor of bacterial persistence in MRSA pneumonia treated with vancomycin. IMPORTANCE Few studies have simultaneously examined the influence of clinical characteristics of patients with pneumonia, the vancomycin pharmacokinetic/pharmacodynamic (PK/PD) index, and the phenotypic and genetic characteristics of methicillin-resistant Staphylococcus aureus (MRSA) strains. We assessed risk factors for vancomycin failure in patients with MRSA pneumonia by analyzing these influences in a prospective multicenter study. Sequence type 5 (ST5) was a possible predictor of bacterial persistence in adult patients with MRSA pneumonia (adjusted odds ratio, 4.449). We found that this may be related to ST5 strains having higher levels of vancomycin heterogeneous resistance, biofilms, and the presence of adhesion and virulence genes such as fnbB, tst, and sec.

ERBB2 Mutations as Potential Predictors for Recurrence in Colorectal Serrated Polyps by Targeted Next-Generation Sequencing.

Background: Follow-up guidelines for serrated polyps (SPs) are mainly based on factors such as histology and size with limited evidence. The underlying genomic mechanism of SPs in relation to recurrence risks is utterly unknown. Methods: We applied targeted next-generation sequencing (NGS) approach on two groups of SPs [polyp-relapsed SPs (PRSPs) vs. polyp-free SPs (PFSPs)] based on the surveillance outcomes to compare differences of DNA variants in 71 colorectal cancer-associated genes. A multicenter validation cohort was established longitudinally from 2016 to 2019 to confirm the relevant results. Results: Among the 96 NGS samples, at least one mutant after filtration was detected in 90 samples (94%). Molecular profiling presented BRAF, KRAS, and APC as top 3 mutated genes. FBXW7, MSH2, and ERBB2 might be recurrence-relevant, while DMD, BRCA1, and BRCA2 might be negatively correlated with recurrence. Notably, ERBB2 mutants (R678Q and V842I) (n = 5) had higher risks of polyp recurrence than the wild types (n = 85), with a median polyp-free interval of 15 months compared to 26 months [P < 0.001; hazard ratio (HR) = 4.9; 95% confidence interval (CI) = 1.9-12.8]. Furthermore, a multicenter cohort composed by 321 SPs verified that ERBB2-mutated SPs had increased risks of polyp recurrence (P < 0.001; HR = 3.7; 95% CI = 2.3-6.0) and advanced neoplastic lesion (ANL) recurrence (P < 0.001; HR = 10.0; 95% CI = 2.7-36.9) compared with wild-type SPs, respectively. Conclusions: Our results are emphasizing that SP individuals with ERBB2 mutants are at higher risks of subsequent colorectal neoplasms. ERBB2 mutants might work as facilitated markers for prediction of high-risk SPs and might implicate a potential mechanism in the serrated pathway to colorectal carcinoma (CRC).

m6A modification regulates lung fibroblast-to-myofibroblast transition through modulating KCNH6 mRNA translation.

Idiopathic pulmonary fibrosis (IPF) is a chronic, fatal lung disease characterized by progressive and non-reversible abnormal matrix deposition in lung parenchyma. Myofibroblasts originating mainly from resident fibroblasts via fibroblast-to-myofibroblast transition (FMT) are the dominant collagen-producing cells in pulmonary fibrosis. N6-methyladenosine (m6A) modification has been implicated in various biological processes. However, the role of m6A modification in pulmonary fibrosis remains elusive. In this study, we reveal that m6A modification is upregulated in a bleomycin (BLM)-induced pulmonary fibrosis mouse model, FMT-derived myofibroblasts, and IPF patient lung samples. Lowering m6A levels through silencing methyltransferase-like 3 (METTL3) inhibits the FMT process in vitro and in vivo. Mechanistically, KCNH6 is involved in the m6A-regulated FMT process. m6A modification regulates the expression of KCNH6 by modulating its translation in a YTH-domain family 1 (YTHDF1)-dependent manner. Together, our study highlights the critical role of m6A modification in pulmonary fibrosis. Manipulation of m6A modification through targeting METTL3 may become a promising strategy for the treatment of pulmonary fibrosis.

Relationship between early serum sodium and potassium levels and AKI severity and prognosis in oliguric AKI patients.

PURPOSE: Acute kidney injury (AKI) is a common organ dysfunction in ICU and up to now there is no good way to predict the AKI progression and patient prognosis. Blood electrolyte tests are common in ICU, but there are few studies on early blood electrolytes and the AKI progression and patient prognosis. Therefore, we concentrated on the serum sodium and potassium levels before AKI diagnosis and evaluated the relationship between serum sodium and potassium levels and the severity and prognosis of AKI. METHODS: This study included data of all patients from the MIMIC-III. We used the urine output criteria in the KDIGO as diagnostic criteria for oliguric AKI. Patients admitted to the ICU several times only included their initial ICU admission results. Patients younger than 18 years old, diagnosed with AKI stage 3, ICU stays less than 24 h or without corresponding laboratory results or data were excluded. The included patients were divided into four groups based on the interquartile range of serum sodium and potassium. We evaluated the serum sodium and potassium levels before AKI diagnosis and AKI severity and prognosis through retrospective analysis. RESULTS: Patients with serum potassium > 4.6 mmol/L were more likely to progress to AKI stage 3 or death than patients with serum potassium ≤ 4.6 mmol/L (overall p < 0.0001). Patients with sodium < 137 mmol/L or > 141 mmol/L had a higher risk of progressing to AKI stage 3 (overall p = 0.00023) and risk of death (overall p < 0.0001) than other patients. In the Cox regression model, after adjusting for age, sex, and BMI, serum sodium or potassium were associated with AKI progression and prognosis (p < 0.01). After continuing to adjust for comorbidities, serum potassium was still associated with AKI progression and prognosis (p < 0.01), but serum sodium was only associated with prognosis (p = 0.027). After adjusting for other indicators, there was no statistically significant correlation between serum sodium or potassium and AKI progression and prognosis. After adjusting for serum sodium or potassium, the corresponding results were not significantly different from those before adjustment. CONCLUSION: This study found that abnormal serum sodium or potassium levels before AKI diagnosis were more likely to lead to AKI progression and poor prognosis, of which lower serum sodium and higher serum potassium were more likely to progress to AKI stage 3 or death.

Exosomes as mediators and biomarkers in fibrosis.

Fibrosis is characterized by aberrant myofibroblast accumulation and excessive extracellular matrix deposition, which leads to organ failure and significantly contributes to mortality worldwide. Exosomes, which are extracellular nanovesicles with a diameter of 30-100 nm that are secreted into the extracellular space by various types of cells, facilitate intercellular communication by delivering different cargos such as proteins, mRNAs and microRNAs. Growing evidence indicates that exosomes play an important role in various fibrotic diseases. A deeper understanding of the effects of exosomes in fibrosis may help in exploring new diagnostic and therapeutic targets. In this review, we summarize recent findings on exosomes in fibrotic diseases, with a special focus on exosomal cargo dysregulation and their potential diagnostic and therapeutic value in fibrosis.

Preoperative D-dimer level is an independent prognostic factor for non-small cell lung cancer after surgical resection: a systematic review and meta-analysis.

BACKGROUND: Whether high preoperative D-dimer level has any impact on long-term survival of patients with surgically treated non-small cell lung cancer (NSCLC) remains unclear. Therefore, we conducted the first meta-analysis focusing specifically on prognostic value of high preoperative D-dimer level in NSCLC patients after surgical resection comprehensively. METHODS: We conducted a systematic search for relevant studies in PubMed, Embase, and Web of Science on January 28, 2019. Data for analysis consisted of hazard ratio (HR) with 95% confidence interval (CI) of overall survival (OS) and disease-free survival (DFS) from multivariate analysis and were analyzed by using the STATA 12.0 package. RESULTS: Finally, we included a total of 6 cohort studies consisting of 1,817 patients with surgically treated NSCLC for analysis. Our meta-analysis found that NSCLC patients with high preoperative D-dimer level had a significantly worse OS (random effects: HR =2.04; 95% CI: 1.30-3.20; P=0.002; I2=67.4%) and DFS (fixed effects: HR =1.98; 95% CI: 1.41-2.78; P<0.001; I2=0.0%) than these with normal preoperative D-dimer level after surgery. However, potential heterogeneity and publication bias was observed during analysis. CONCLUSIONS: High pretreatment level of D-dimer remains to be an independent predictor of poor prognosis in NSCLC patients after surgery. Further well-conducted studies with appropriate adjustments are needed to confirm and update our conclusions.

circHIPK3 regulates lung fibroblast-to-myofibroblast transition by functioning as a competing endogenous RNA.

Myofibroblasts predominantly emerging through fibroblast-to-myofibroblast transition (FMT) are considered to be the key collagen-producing cells in pulmonary fibrosis. Circular RNAs (circRNAs) are important players involved in many biological processes. circHIPK3 has been identified as the one of the most abundant circRNAs in human lung. In this study, we characterized the role of circHIPK3 in pulmonary fibrosis. We revealed that circHIPK3 is upregulated in bleomycin-induced pulmonary fibrosis mice model, FMT-derived myofibroblasts. circHIPK3 silencing can ameliorate FMT and suppress fibroblast proliferation in vivo and vitro. Fundamentally, circHIPK3 regulates FMT by functioning as an endogenous miR-338-3p sponge and inhibit miR-338-3p activity, thereby leading to increased SOX4 and COL1A1 expression. Moreover, dysregulated circHIPK3 expression was detected in the clinical samples of patients with idiopathic pulmonary fibrosis. Intervention of circHIPK3 may represent a promising therapy for pulmonary fibrosis.

Somatic mutation profiling of liver and biliary cancer by targeted next generation sequencing.

Liver and biliary cancers are highly lethal cancer types lacking effective treatments. The somatic mutations, particularly those with low mutant allele frequencies, in Chinese patients with liver and biliary cancer have not been profiled, and the frequency of patients benefiting from targeted therapy has not been studied. The present study evaluated the tumor tissues of 45 Chinese patients with hepatocellular carcinoma (HCC) and 12 Chinese patients with biliary tract cancer (BTC) by targeted next generation sequencing, with an average coverage of 639×, to identify alterations in 372 cancer-related genes. A total of 263 variants were identified in 139 genes, with 85.6% of these variants not previously reported in the Catalogue Of Somatic Mutations In Cancer database, and the mutation profile was different from the current datasets, including The Cancer Genome Atlas dataset and the National Cancer Center Japan (NCC_JP) dataset. Patients with hepatitis B virus (HBV) infection harbored more mutations than those without HBV infection, and the mutations in HBV carriers occurred preferentially in genes involved in vascular endothelial growth factor signaling pathways. Mutations in fibroblast growth factor and RAS signaling pathways were enriched in patients with cirrhosis, and alterations in interleukin and transforming growth factor signaling pathways were more frequently identified in individuals with abnormal bilirubin expression. Of all the patients, 7% exhibited variants in the target of sorafenib, and 42% harbored variants in the targets of drugs that have been approved to treat other types of cancer. These findings indicate diverse HCC/BTC variants patterns in different populations, and that the mutation load and patterns are correlated with clinical features. Further clinical studies are now warranted to evaluate the efficacies of other targeted drugs besides sorafenib in the treatment of patients with liver and biliary cancer.

Ventilator management for acute respiratory distress syndrome associated with avian influenza A (H7N9) virus infection: A case series.

BACKGROUND: Data on the mechanical ventilation (MV) characteristics and radiologic features for the cases with H7N9-induced ARDS were still lacking. METHODS: We describe the MV characteristics and radiologic features of adult patients with ARDS due to microbiologically confirmed H7N9 admitted to our ICU over a 3-month period. RESULTS: Eight patients (mean age 57.38±16.75; 5 male) were diagnosed with H7N9 in the first quarter of 2014. All developed respiratory failure complicated by acute respiratory distress syndrome (ARDS), which required MV in ICU. The baseline APACHE II and SOFA score was 11.77±6.32 and 7.71±3.12. The overall CT scores of the patients was 247.68±34.28 and the range of CT scores was 196.3-294.7. The average MV days was 14.63±6.14, and 4 patients required additional rescue therapies for refractory hypoxemia. Despite these measures, 3 patients died. CONCLUSION: In H7N9-infected patients with ARDS, low tidal volume strategy was the conventional mode. RM as one of rescue therapies to refractory hypoxemia in these patients with serious architectural distortion and high CT scores, which could cause further lung damage, may induce bad outcomes and requires serious consideration. Prone ventilation may improve mortality, and should be performed at the early stage of the disease, not as a rescue therapy.

Effect of Platelet-derived P-selectin on Neutrophil Recruitment in a Mouse Model of Sepsis-induced Acute Kidney Injury.

BACKGROUND: Acute kidney injury (AKI) is a severe disease in critically ill patients. Neutrophil infiltration into kidney was associated with the development of AKI, and P-selectin may be involved in the process of neutrophil recruitment in kidney. This study aimed to explore the potential effect of platelet-derived P-selectin on neutrophil recruitment in a mouse model of sepsis-induced AKI. METHODS: A total of 30 C57BL/6 male mice were divided into five groups (n = 6 in each): sham group, sepsis group, anti-Ly6G group, anti-P-selectin group, and platelet depletion group. Sepsis was induced by cecal ligation and puncture. Serum creatinine concentration and platelet activity were measured by biochemical detector and flow cytometry, respectively. Histological and pathological features were analyzed using hematoxylin-eosin (H&E) and immunohistochemistry (IHC) staining, respectively. Myeloperoxidase (MPO) activity was detected with MPO assay. Unpaired t-test was used for data analysis. RESULTS: Serum creatinine increased significantly in septic group compared to sham group (2.68 ± 0.27 mg/dl vs. 0.82 ± 0.19 mg/dl, t = 12.06, P = 0.0000) but attenuated in antibodies-treated animals compared to septic group (anti-Ly6G: 1.62 ± 0.30 mg/dl vs. 2.68 ± 0.27 mg/dl, t = 5.76, P = 0.0004; anti-P-selectin: 1.76 ± 0.31 mg/dl vs. 2.68 ± 0.27 mg/dl, t = 4.92, P = 0.0012; and platelet depletion: 1.93 ± 0.29 mg/dl vs. 2.68 ± 0.27 mg/dl, t = 4.14, P = 0.0032). Platelet amount significantly decreased compared to sham group (658.20 ± 60.64 × 109/L vs. 822.00 ± 48.60 × 109/L, t = 4.71, P = 0.0015) in septic mice, especially in platelet depletion group (240.80 ± 44.98 × 109/L vs. 822.00 ± 48.60 × 109/L, t = 19.63, P = 0.0000). P-selectin activity was significantly increased in septic group compared to sham group (16.54 ± 1.60% vs. 1.90 ± 0.29%, t = 15.64, P = 0.0000) but decreased significantly in platelet depletion group compared to septic group (3.62 ± 0.68% vs. 16.54 ± 1.60%, t = 12.89, P = 0.0002). IHC analysis shown that neutrophil infiltration increased in septic mice compared to sham group (36.67 ± 3.79% vs. 9.17 ± 1.61%, t = 11.58, P = 0.0003) and function-blocked groups (anti-Ly6G: 36.67 ± 3.79% vs. 15.33 ± 1.53%, t = 9.05, P = 0.0008; anti-P-selectin: 36.67 ± 3.79% vs. 21.33 ± 1.53%, t = 6.51, P = 0.0029; and platelet depletion: 36.67 ± 3.79% vs. 23.33 ± 3.06%, t = 4.75, P = 0.0090). MPO increased significantly in septic group compared to control (49.73 ± 1.83 ng/mg prot vs. 13.04 ± 2.16 ng/mg prot, t = 19.03, P = 0.0000) but decreased in function-blocked groups compared to septic group (anti-Ly6G: 26.52 ± 3.86 ng/mg prot vs. 49.73 ± 1.83 ng/mg prot, t = 9.59, P = 0.0000; anti-P-selectin: 33.06 ± 6.75 ng/mg prot vs. 49.73 ± 1.83 ng/mg prot, t = 4.85, P = 0.0013; and platelet depletion: 33.37 ± 2.25 ng/mg prot vs. 49.73 ± 1.83 ng/mg prot, t = 5.33, P = 0.0007). CONCLUSION: Platelets-derived P-selectin may be involved in the development of septic AKI through inducing neutrophil infiltration into kidney.

Comparison of surgical fixation and nonsurgical management of flail chest and pulmonary contusion.

OBJECTIVE: The objective of this study is to compare the clinical efficacy of surgical fixation and nonsurgical management of flail chest and pulmonary contusion (FC-PC) and to compare the diverse timings of surgery to discuss case management in FC-PC. METHODS: The data of 39 patients diagnosed with FC-PC were obtained from the intensive care unit of Shanghai First People's Hospital and analyzed retrospectively from July 2010 to Dec 2013. The patients required ventilator support and were divided into a surgical group and a nonsurgical group, according to the treatment method. The clinical data, such as mortality, the duration of mechanical ventilation (DMV), intensive care unit length of stay, hospital length of stay (HLOS), days of antibiotic use, transfusion volume, medical expense as well as the incidence of tracheotomy, pleural effusion, and incidence of ventilator associated pneumonia, were collected for all subjects. The surgical group was further divided into 2 groups according to the surgery timing. Surgery within 7 days of admission was defined as early surgery, and all other times were defined as late surgery. The clinical data and incidence of incision infection were collected and compared. RESULTS: The patients in the surgical group had a slightly shorter HLOS. No differences were noted in mortality and the other clinical data between the groups. The early surgical group had a shorter DMV and less incidence of tracheotomy. The other parameters had no differences. CONCLUSIONS: Surgery for FC-PC could reduce the HLOS, and early surgery could decrease the DMV and the need for tracheotomy.

Overexpressed miRNA-155 dysregulates intestinal epithelial apical junctional complex in severe acute pancreatitis.

AIM: To investigate whether miRNA-155 (miR-155) dysregulates apical junctional complex (AJC) protein expression in experimental severe acute pancreatitis (SAP). METHODS: Twenty-four male BALB/c mice were randomly assigned to two groups: the SAP group (n = 12) receiving sequential intraperitoneal injection of 50 µg/kg caerulein and 10 mg/kg lipopolysaccharide over 6 h, and the control group (n = 12) receiving intraperitoneal injection of normal saline. Animals were sacrificed 3 h following the last injection for collection of blood samples and pancreas and distal ileal segment specimens. Routine pancreas and intestine histology was used to assess SAP pathology and intestinal epithelial barrier damage. Levels of serum amylase, diamine oxidase (DAO), and tumor necrosis factor (TNF)-α were determined using commercial kits. Total RNA samples were isolated from intestinal epithelial specimens and reversely transcribed into cDNA. miR-155 and RhoA mRNA expression profiles were determined using quantitative real-time polymerase chain reaction. Target genes for miR-155 were predicted using the miRTarBase database, RNA22 and PicTar computational methods. Western blotting was performed to quantitate the protein expression levels of the target gene RhoA, as well as zonula occludens (ZO)-1 and E-cadherin, two AJC component proteins. RESULTS: Intraperitoneal injection of caerulein and lipopolysaccharide successfully induced experimental acute pancreatic damage (SAP vs control, 10.0 ± 2.0 vs 3.2 ± 1.2, P < 0.01) and intestinal epithelial barrier damage (3.2 ± 0.7 vs 1.4 ± 0.7, P < 0.01). Levels of serum amylase (21.6 ± 5.1 U/mL vs 14.3 ± 4.2 U/mL, P < 0.01), DAO (21.4 ± 4.1 mg/mL vs 2.6 ± 0.8 mg/mL, P < 0.01), and TNF-α (61.0 ± 15.1 ng/mL vs 42.9 ± 13.9 ng/mL, P < 0.01) increased significantly in SAP mice compared to those in control mice. miR-155 was significantly overexpressed in SAP intestinal epithelia (1.94 ± 0.50 fold vs 1.03 ± 0.23 fold, P < 0.01), and RhoA gene containing three miR-155-specific binding sites in the three prime untranslated regions was one of the target genes for miR-155. RhoA (22.7 ± 5.8 folds vs 59.6 ± 11.6 folds, P < 0.01), ZO-1 (46 ± 18 folds vs 68 ± 19 folds, P < 0.01), and E-cadherin proteins (48 ± 15 folds vs 77 ± 18 folds, P < 0.01) were underexpressed in SAP intestinal epithelia although RhoA mRNA expression was not significantly changed in SAP (0.97 ± 0.18 folds vs 1.01 ± 0.17 folds, P > 0.05). CONCLUSION: TNF-α-regulated miR-155 overexpression inhibits AJC component protein syntheses of ZO-1, and E-cadherin by downregulating post-transcriptional RhoA expression, and disrupts intestinal epithelial barrier in experimental SAP.

Expression and significance of HIF-1α in pulmonary fibrosis induced by paraquat.

It is commonly accepted that epithelial-mesenchymal transition contributes to fibrotic remodeling, but the molecular pathways involved in paraquat (PQ)-induced epithelial-mesenchymal transition remain uncharacterized. The objective of this study was to evaluate the potential involvement of HIF-1α in TGF-ß1/ß-Catenin and Snail pathway after PQ poisoning. In our study, 86 Spragne-Dawley rats were randomly divided into control group and PQ group, which received intragastric infusion of 20% PQ solution 50 mg/kg. Rats in the PQ group were subsequently divided into eight subgroups (10 for each subgroup) and samples were collected at different predetermined time points (2, 6, 12, 24, 48, 72, 96 h and 7 d). Fibrosis markers, including ß-catenin, snail and α-SMA, were measured by western blot. The activity of HIF-1α was determined by western blot and immunofluorescence. We found that in PQ-induced pulmonary fibrosis, the level of PaO2 was significantly reduced in the 6-h subgroup, when compared to the control group (P < 0.01). Interestingly, between 6 and 72 h, there was no significant difference in PaO2. On the other hand, the level of PaCO2 started to increase from 72-h subgroup (P < 0.01). Fibrosis markers including ß-catenin, snail and α-SMA, measured by western blot, were significantly increased at 2 h, while the level of p-GSK-3ß was increased at 6 h. And the level of GSK-3ß showed significant reduction beginning at 24 h. The activity of HIF-1α measured by western blot assays was significantly increased starting from 2 h with sustained expression. The result of Pearson coefficient analysis showed that HIF-1α was positively correlated with Snail (r = 0.935, P < 0.01) and ß-catenin (r = 0.761, P < 0.05). Meanwhile, immunofluorescent analysis of HIF-1α revealed partial staining appearing from 2 h. Our data illustrated a positive correlation between Snail, ß-catenin signaling and HIF-1α, suggesting a potential synergistic role of HIF-1α in PQ-induced pulmonary fibrosis, which may be independent of GSK-3ß. It might also represent a potential therapeutic window for treatment of paraquat poisoning.

[The relationship of endoplasmic reticulum stress with paraquat induced lung fibrosis in rats].

OBJECTIVE: To investigate the relationship between pulmonary fibrosis and endoplasmic reticulum stress (ERS) in rats with paraquat poisoning. METHODS: One hundred male Sprague-Dawley (SD) rats were randomly divided into control group (n=10) and paraquat poisoning group (n=90). The animals were sacrificed by exsanguination at 2, 6, 12, 24, 48, 72, 96, 168, 336 hours after administration with 20% parquat solution. The paraffin sections of lung tissue were stained with hematoxylin-eosin (HE) and Masson trichrome to observe the pathological changes. Glucose-regulated protein 78 (GRP78) was determined by immunohistochemistry, and malondialdehyde (MDA) of lung tissue was measured. The total protein of tissue was abstracted, and the α-smooth muscle actin (α-SMA) and GRP78 was detected by Western blotting. RESULTS: HE and Masson staining demonstrated inflammatory infiltration and collagen deposition in the lung after paraquat administration with a tendency of exaggeration with time, and finally resulted in fibrosis. The expressions of MDA, α-SMA and GRP78 in the lung tissue were significantly increased 2 hours after paraquat administration compared with those of control group (MDA: 1.38 ± 0.18 nmol/mg vs. 0.85 ± 0.05 nmol/mg, α-SMA: 0.23 ± 0.01 vs. 0.14 ± 0.03, GRP78: 0.72 ± 0.02 vs. 0.37 ± 0.06, P<0.05 or P<0.01), and the expressions of MDA and α-SMA were gradually increased with time. GRP78 protein expression was decreased at 72 hours after paraquat administration. CONCLUSIONS: The results reveal that the expressions of α-SMA and GRP78 in paraquat poisoning group are up-regulated, suggesting ERS plays an important role in paraquat induced-pulmonary fibrosis.

[Hypoxia-inducible factor-1α expression in renal tissue of rats with paraquat poisoning].

OBJECTIVE: To observe the pathological changes of renal tissue in the rats with paraquat (PQ) poisoning as well as the serum creatinine (SCr) levels and expression levels of hypoxia-inducible factor-1α (HIF-1α) and transforming growth factor-ß (TGF-bgr;) in renal tissue at different time points after PQ poisoning, and to investigate the association of HIF-1α with renal injury after PQ poisoning. METHODS: Forty-eight healthy male Sprague-Dawley rats were randomly divided into control group (n = 6) and PQ group (n = 42). The control group was given a single dose of 1 ml saline by gavage; the PQ group was given a single dose of 1 ml PQ (50 mg/kg), which was prepared by diluting 20% raw liquid of PQ with saline, by gavage. The PQ group was further divided into 2, 6, 12, 24, 48, 72 and 120 h PQ subgroups (n = 6 for each subgroup) to be examined at 2, 6, 12, 24, 48, 72, or 120 h after gavage. Their arterial blood was collected for blood gas analysis as well as blood lactic acid (BLA) and SCr measurement; renal sections were subjected to HE staining; the protein expression of HIF-1α and TGF-ß in renal tissue was measured by Western blot. RESULTS: The BLA level and SCr level began to rise at 6h after poisoning. Compared with the control group, the 6, 12, 24, 48, 72 and 120 h PQ subgroups had significantly increased BLA and SCr levels (P < 0.05); the 72 and 120 h PQ subgroup showed hypoxemia (P < 0.05). The protein expression of HIF-1α in PQ group increased significantly at 6h and reached the peak level at 72 h, with a significant difference from that in the control group at 6, 12, 24, 48, 72, and 120 h (P < 0.05). The protein expression of TGF-ß in PQ group began to rise at 24 h, reached the peak level at 72 h, and declined at 120 h, with a significant difference from that in the control group at 24, 48, and 72 h (P < 0.05). The protein expression of HIF-1α was positively correlated with SCr level (r = 0.9308, P = 0.0008), uncorrelated with arterial partial pressure of oxygen (r = -0.6996, P = 0.0534), and positively correlated with BLA level (r = 0.9483, P = 0.0003). The pathological changes of renal tissue mainly included the degeneration and necrosis of renal tubular epithelial cells, which worsened as the time went on and appeared less severe at 120 h. CONCLUSION: The HIF-1α expression in renal tissue increases significantly in the early stage of PQ poisoning, which is associated with increased BLA and SCr levels and causes upregulated expression of TGF-ß that promotes renal fibrosis.

Effect of ulinastatin on paraquat-induced-oxidative stress in human type II alveolar epithelial cells.

BACKGROUND: Ulinastatin (UTI) is a urinary trypsin inhibitor extracted and purified from urine of males. This study aimed to explore the effects of UTI on paraquat-induced-oxidative stress in human type II alveolar epithelial cells. METHODS: The human type II alveolar epithelial cells, A549 cells, were cultured in vitro. The A549 cells were treated with different concentrations of paraquat (200, 400, 600, 800, 1 000, 1 200 µmol/L) and ulinastatin(0, 2 000, 4 000, 6 000, 8 000 U/mL) for 24 hours, the cell viability was measured by cell counting kit-8 and the median lethal concentration was selected. In order to establish an in vitro model of paraquat intoxication and to determine the safe dose of ulinastatin, we calculated LD50 using cell counting kit-8 to determine the survival rate of the cells. A549 cells were divided into normal control group, paraquat group and paraquat+ulinastatin group. The levels of malondialdehyde (MDA) and myeloperoxidase (MPO) were detected by biochemistry colorimetry, while the level of reactive oxygen spies (ROS) was detected by DCFH-DA assay. RESULTS: The survival rate of A549 cells treated with different concentrations of paraquat decreased in a concentration-dependent manner. Whereas there was no decrease in the survival rate of cells treated with 0-4 000 U/mL ulinastatin. The levels of MDA, MPO, and ROS were significantly higher in the paraquat group than in the normal control group after 24-hour-exposure. And the survival rate of the paraquat+ulinastatin group was higher than that of the paraquat group, but lower than that of the normal control group. The levels of MDA, MPO, and ROS were lower than those of the paraquat group. CONCLUSION: Ulinastatin can alleviate the paraquat-induced A549 cell damage by reducing oxidative stress.