OBJECTIVE: To identify and validate novel biomarkers for the early diagnosis of sepsis-associated acute kidney injury (SA-AKI) and precise continuous renal replacement therapy (CRRT) using proteomics. METHODS: A prospective multicenter cohort study was conducted. Patients with sepsis admitted to five hospitals in Taizhou City of Zhejiang Province from April 2019 to December 2021 were continuously enrolled, based on the occurrence of acute kidney injury (AKI). Sepsis patients were divided into SA-AKI group and non-SA-AKI group, and healthy individuals who underwent physical examinations during the same period were used as control (NC group). Peripheral blood samples from participants were collected for protein mass spectrometry analysis. Differentially expressed proteins were identified, and functional enrichment analysis was conducted on these proteins. The levels of target proteins were detected by enzyme linked immunosorbent assay (ELISA), and the predictive value of target protein for SA-AKI were evaluated by receiver operator characteristic curve (ROC curve). Additionally, sepsis patients and healthy individuals were selected from one hospital to externally verify the expression level of the target protein and its predictive value for SA-AKI, as well as the accuracy of CRRT treatment. RESULTS: A total of 37 patients with sepsis (including 19 with AKI and 18 without AKI) and 31 healthy individuals were enrolled for proteomic analysis. Seven proteins were identified with significantly differential expression between the SA-AKI group and non-SA-AKI group: namely cystatin C (CST3), β ₂-microglobulin (β ₂M), insulin-like growth factor-binding protein 4 (IGFBP4), complement factor I (CFI), complement factor D (CFD), CD59, and glycoprotein prostaglandin D2 synthase (PTGDS). Functional enrichment analysis revealed that these proteins were involved in immune response, complement activation, coagulation cascade, and neutrophil degranulation. ELISA results demonstrated specific expression of each target protein in the SA-AKI group. Additionally, 65 patients with sepsis (38 with AKI and 27 without AKI) and 20 healthy individuals were selected for external validation of the 7 target proteins. ELISA results showed that there were statistically significant differences in the expression levels of CST3, β ₂M, IGFBP4, CFD, and CD59 between the SA-AKI group and non-SA-AKI group. ROC curve analysis indicated that the area under the curve (AUC) values of CST3, β ₂M, IGFBP4, CFD, and CD59 for predicting SA-AKI were 0.788, 0.723, 0.723, 0.795, and 0.836, respectively, all exceeding 0.7. Further analysis of patients who underwent CRRT or not revealed that IGFBP4 had a good predictive value, with an AUC of 0.84. CONCLUSIONS Based on proteomic analysis, CST3, β ₂M, IGFBP4, CFD, and CD59 may serve as potential biomarkers for the diagnosis of SA-AKI, among which IGFBP4 might be a potential biomarker for predicting the need for CRRT in SA-AKI patients. However, further clinical validation is required.
Humans ; Sepsis/complications* ; Acute Kidney Injury/blood* ; Proteomics ; Prospective Studies ; Biomarkers/blood* ; Male ; Female ; beta 2-Microglobulin/blood* ; Middle Aged ; Cystatin C/blood* ; Aged

Objective:To investigate the diagnostic and prognostic value of systemic immune inflammatory index (SII) for severe traumatic brain injury secondary to acute lung injury (sTBI-ALI).Methods:A retrospective study was conducted on patients with severe traumatic brain injury admitted to the trauma center of the First Affiliated Hospital of Soochow University from January 2021 to November 2023. Patients received standard treatments including hemostasis and intracranial pressure management. Vital signs and blood routine data were collected upon admission. Patients were categorized into sTBI group and sTBI-ALI group based on established clinical diagnostic criteria for ALI to evaluate the diagnostic utility of SII. Subsequently, within the sTBI-ALI group, patients were stratified into survival and non-survival groups based on their 30-day outcomes to assess the prognostic value of SII.Results:A total of 260 sTBI patients were enrolled, of whom 113 developed ALI. Among the sTBI-ALI patients, 73 survived at 30 days. Compared to the sTBI group, the sTBI-ALI group exhibited significantly higher respiratory rates, heart rates, white blood cell counts, neutrophil counts, platelet counts, and SII levels (all P<0.05). Multivariate logistic regression analysis showed that SII index ( OR=1.003, 95% CI: 1.002-1.004, P<0.001) was an independent risk factor for ALI development in sTBI patients. The combined predictive model incorporating SII and heart rate yielded an AUC of 0.801 (95% CI: 0.740-0.862). The non-survival group had significantly higher neutrophil counts and SII levels, and significantly lower Glasgow Coma Scale scores than the survival group (all P<0.05). Multifactorial regression analysis indicated that SII index ( OR=1.002, P=0.004, 95% CI: 1.000-1.003) served as an independent risk factor for 30-day mortality in sTBI-ALI patients. The combined predictive model of SII and GCS achieved an AUC of 0.904 (95% CI: 0.848-0.960). Conclusions:SII demonstrates potential as a biomarker for predicting the development of ALI following sTBI. Furthermore, incorporating SII into predictive models significantly enhances the ability to forecast mortality risk in sTBI-ALI patients.

The identification of clinical questions for clinical practice guidelines of Chinese patent medicine （CPM） is important for subsequent evidence retrieval， evaluation of evidence quality， formation of recommendations. This paper described a methodological proposal for the identification of clinical questions for CPM guidelines to highlight the characteristics of Chinese patent medicine and reflect its effect in specific stage of the disease. Considering four aspects， namely， the drug of Chinese patent medicine （D）， the specific disease stage （S）， comparison （C）， and specific outcome （O）， DSCO framework has been proposed to formulate the clinical questions. Multi-source information through scientific research， policy or standard documents， and clinical data are suggested for collecting clinical questions， and clear selection criteria should be set to finalize the clinical questions to be addressed by the guideline. In addition， the above process needs to be transparently and publicly reported in order to ensure the clarity and completeness of the guidelines.

Objective:To investigate the therapeutic efficacy and underlying mechanisms of the traditional Chinese medicine formula "Hua Xian Fang" (HXF) in the treatment of radiation-induced pulmonary fibrosis (RIPF).Methods:In vivo experiment, 36 male specific pathogen free (SPF)-grade C57BL/6 mice aged 6-8 weeks were randomly divided into the control, irradiation (17 Gy thoracic irradiation), and irradiation+HXF groups (17 Gy thoracic irradiation+HXF). After 16 weeks, lung coefficient, HE staining and Masson staining were used to evaluate the degree of pulmonary inflammation and fibrosis. Immunohistochemistry was performed to measure the expression levels of α-smooth muscle actin (α-SMA) in lung tissues. Quantitative real-time PCR (qPCR) was performed to detect the mRNA expression levels of interferon-γ (IFN-γ). Enzyme linked immunosorbent assay (ELISA) was conducted to determine the levels of IFN-γ in serum and bronchoalveolar lavage fluid (BALF). During in vitro experiment, NIH/3T3 fibroblasts were stimulated with IFN-γ after 6 Gy irradiation, followed by 48 hours of culture. qPCR, immunofluorescence staining, and Western blot were used to assess the expression of α-SMA and collagen Ⅰ at the transcription and protein levels. One way ANOVA was used for multiple group comparisons, and Tukey test was used for inter group multiple comparisons. Results:Compared to the control group, mice in the irradiation group showed significant increases in lung coefficient, Szapiel score, Ashcroft score, and α-SMA expression in lung tissues (all P<0.001). Compared to the irradiation group, the irradiation+HXF group exhibited significant decreases in the above indicators (all P<0.001). qPCR demonstrated that the mRNA expression levels of IFN-γ were significantly higher in the irradiation+HXF group than that in the irradiation group ( P=0.001). ELISA results showed that the levels of IFN-γ in serum and BALF were significantly elevated in the irradiation+HXF group compared to those in the irradiation group ( P=0.032, 0.037). In vitro experiment revealed that after irradiation, the expression levels of α-SMA and collagen Ⅰ mRNA and protein in NIH/3T3 cells were significantly increased, while decreased after IFN-γ stimulation. Conclusion:HXF effectively alleviates RIPF, probably by the upregulation of IFN-γ in blood and tissues and inhibition of fibroblast activation.

The PCR-amplified severe fever with thrombocytopenia syndrome virus(SFTSV)Gn-DⅢ-Ⅲ gene was inserted into the pET-30a(+)prokaryotic expression vector to generate the re-combinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ.The plasmid was transformed into E.coli BL21(DE3)for Gn-DⅢ-m protein expression and the expression conditions were optimized.The Gn-DⅢ-Ⅲ protein purified with Ni-NTA column affinity chromatography was applied as the captured antigen to establish an indirect ELISA method for the detection of SFTSV antibody.The results demonstrated that the recombinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ was successfully constructed as identified by PCR and sequencing.The recombinant protein SFTSV Gn-D m-Ⅲ was soluble ex-pression in E.coli under the optimal induction conditions of 0.4 mmol/L IPTG at 25 ℃ for 4 h,and the protein purity was 91.77％after purification by Ni-NTA column.The optimal reaction con-ditions for the indirect ELISA of SFTSV antibody were as follows:coating antigen concentration(5 μg/mL),primary antibody(incubation at 37 ℃ for 1.5 h),and secondary antibody(diluted 1:10 000 and incubated at 37 ℃ for 1 h).The established method had no cross-reactivity with Rift Valley fever virus(RVFV),Ebola virus(EBOV),and tick-borne encephalitis virus(TBEV)posi-tive sera.The method had a high sensitivity,with P/N＞2.1 for SFTSV-positive sera diluted to 81920.Coefficients of variation for intra-and inter-batch reactions were less than 10％.Detection of four SFTSV-infected human clinical serum samples showed the serum samples from patients in re-mission were tested as positive(P/N＞2.1),while serum samples from patients with multiple or-gan failure were detected as negative(P/N＜2.1).The results indicated that the SFTSV Gn-D Ⅲ-Ⅲ protein was successfully expressed and purified,and it was used as the coating protein to estab-lish an indirect ELISA assay for SFTSV antibody,which possesses good specificity,sensitivity and reproducibility.This method might be applied to detect human SFTSV clinical serum samples.

Objective To evaluate the risk of bias and reporting quality in randomized controlled trials(RCTs)of the Chinese medicine injection for acute cerebral infarction in the last five years.Methods RCTs literature on Chinese medicine injection in the treatment of acute cerebral infarction was systematically searched in CNKI,Wanfang Data,VIP,China Biology Medicine Database(CBM),PubMed,Embase and Cochrane Library from April 20,2018 to April 20,2023.The risk of bias and reporting quality of included RCTs were evaluated using the Cochrane Risk of Bias Tool(ROB 1.0)and CONSORT-CHM Formulas 2017,respectively.Results A total of 4 301 articles were retrieved,and 408 RCTs were included according to inclusion and exclusion criteria.The ROB evaluation results showed that the the majority of studies were rated as having an unclear risk of bias due to the lack of reporting on allocation concealment,blind method,trial registration information,and funding sources.The evaluation results of CONSORT-CHM Formulas 2017 showed that the number of reported papers of 17 items was greater than or equal to 50%,and the number of reported papers of 25 items was less than 10%,and most of the RCTs did not show the characteristics of TCM syndrome differentiation and treatment.Conclusion The quality of Chinese medicine injection in the treatment of acute cerebral infarction RCTs is generally low.It is recommended that researchers refer to the methodology design of RCTs and international reporting standards,improve the trial design,standardize the trial report,and highlight the characteristics of TCM syndrome differentiation and treatment.

Objective:To explore the risk factors related to delayed pleural effusion in multiple trauma patients.Methods:A retrospective cohort study was conducted to analyze the clinical data of 145 multiple trauma patients admitted to the First Affiliated Hospital of Soochow University from January 2022 to October 2023, including 99 males and 46 females, aged 18-81 years [56.0(46.5, 64.5)years]. Based on whether delayed pleural effusion developed after injury, the patients were divided into delayed pleural effusion group ( n=66) and non-delayed pleural effusion group ( n=79). The clinical data of the patients in both groups were collected, including gender, age, underlying disease (diabetes mellitus and hypertension), cause of injury (traffic injury, blow injury, fall from height, and others), comorbid injuries (traumatic brain injury, maxillofacial fracture, clavicular fracture, scapular fracture, sternal fracture, spinal fracture, multiple rib fracture, pneumothorax, lung contusion, and pelvic fracture), severity of injury [injury severity score (ISS) and abbreviated injury scale (AIS) score for the chest], location and number of rib fractures, vital signs at admission (body temperature, heart rate, respiration, systolic blood pressure, diastolic blood pressure), and clinical test indices [white blood cells (WBC), hemoglobin (Hb), platelets (PLT), total protein (TP), albumin (ALB), C-reactive protein (CRP), procalcitonin (PCT), fibrinogen (FIB), fibrin degradation product (FDP), D-dimer (D-D), aspartate transaminase (AST), alanine transferase (ALT), and creatinine (Cr)]. Univariate analysis was conducted to assess the correlation between the forementioned factors and the development of delayed pleural effusion after multiple traumas. Multivariate Logistic regression analysis was used to determine the independent risk factors for the development of delayed pleural effusion after multiple traumas. Results:The results of univariate analysis showed that multiple rib fracture, pneumothorax, pulmonary contusion, chest AIS score, posterior rib fracture, number of rib fractures, TP, ALB, CRP, PCT and FDP were correlated with delayed pleural effusion in multiple trauma patients ( P<0.05 or 0.01); whereas gender, age, underlying disease, cause of injury, sternal fracture, spinal fracture, clavicular fracture, scapular fracture, pelvic fracture, maxillofacial fracture, traumatic brain injury, anterior rib fracture, ISS, vital signs at admission, WBC, Hb, PLT, FIB, D-D, AST, ALT, and Cr were not correlated with delayed pleural effusion in multiple trauma patients ( P>0.05). The results of multivariate Logistic regression analysis revealed that lung contusion ( OR=3.96, 95% CI 1.59, 9.85, P<0.01), ALB ( OR=0.79, 95% CI 0.66, 0.94, P<0.01), and CRP ( OR=1.02, 95% CI 1.01, 1.03, P<0.01) were significantly correlated with delayed pleural effusion in multiple trauma patients. Conclusion:Lung contusion, ALB, and CRP are the independent risk factors for delayed pleural effusion in multiple trauma patients.

Objective:To elucidate the effect of bepridil, an Na +/Ca 2+ exchanger (NCX) inhibitor, on the proliferation, migration and apoptosis of melanoma cells, and to explore their potential underlying mechanisms. Methods:Six paraffin-embedded tissue specimens were collected from 3 patients histopathologically diagnosed with melanocytic nevi and 3 patients histopathologically diagnosed with melanoma in the Hospital for Skin Diseases, Chinese Academy of Medical Sciences from January to December 2023. NCX1 expression in tissues was determined by immunohistochemical staining, and Western blot analysis was performed to verify the expression of NCX1 in primary melanocytes and melanoma cell lines A375, A875, SKMEL-28, M14, MV3, and SK-MEL-5. Cell counting kit 8 (CCK8) assay was conducted to evaluate the effect of bepridil at different concentrations on the viability of melanoma cells, and the proliferation curve was drawn to calculate the half inhibitory concentration (IC50) of bepridil. Some melanoma cells were then treated with bepridil at IC50 (bepridil groups), and cells treated with dimethyl sulfoxide-containing media served as control groups. Intracellular Ca 2+ levels were assessed in A375 and SK-MEL-28 cells using the Fluo-4 calcium assay kit, the migration and apoptosis of A375, SK-MEL-28, and A2058 cells were estimated by Transwell assay and flow cytometry respectively. The effects of bepridil treatment on gene expression and pathways in A375 cells were evaluated by transcriptome sequencing, Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The content of reactive oxygen species (ROS) in melanoma cells after the bepridil treatment was detected by flow cytometry, and the expression of endoplasmic reticulum stress- and mitochondrial apoptotic pathway-related molecules was determined by Western blot analysis. Comparisons between two groups were performed by t test. Results:Immunohistochemical assay showed that the expression of NCX1 was significantly higher in the melanoma tissues (0.320 ± 0.020) than in the melanocytic nevus tissues (0.235 ± 0.008, t = 4.04, P = 0.016) ; Western blot analysis showed that the NCX1 protein bands were darker in color in the melanoma cell lines than in the primary melanocytes. CCK8 assay showed a gradual decrease in melanoma cell viability with increasing concentrations of bepridil. In the A375 and SK-MEL-28 cells, the fluorescence intensity of calcium was higher in the bepridil groups after the treatment with bepridil at IC50 (25 μmol/L) (64.82 ± 2.98, 75.84 ± 2.07, respectively) than in the corresponding control groups (37.10 ± 2.33, 66.54 ± 1.47, respectively, both P < 0.05) ; in the A375, SK-MEL-28, and A2058 cells, the migration ability was lower in the bepridil groups (103.00 ± 9.07, 67.33 ± 7.22, 61.33 ± 1.76, respectively) than in the corresponding control groups (400.00 ± 25.17, 276.70 ± 14.63, 116.00 ± 10.69, respectively, all P < 0.05), while their apoptosis rates were higher in the bepridil groups (5.72% ± 0.06%, 13.58% ± 0.86%, 25.76% ± 1.95%, respectively) than in the corresponding control groups (3.99% ± 0.50%, 6.47% ± 0.88%, 8.01% ± 0.36%, respectively, all P < 0.05). Transcriptome sequencing revealed 119 up-regulated genes and 164 down-regulated genes in bepridil-treated A375 cells compared with control cells, and the differentially expressed genes were mainly associated with metabolic pathways, endoplasmic reticulum stress, and tumor-related pathways. The ROS levels were higher in bepridil-treated A375, SK-MEL-28, and A2058 cells (1 907 ± 33, 7 607 ± 535, 3 380 ± 300, respectively) than in the corresponding control groups (1 646 ± 16, 4 386 ± 163, 2 110 ± 66, respectively, all P < 0.05). Western blot analysis showed that the expression of C/EBP homologous protein and activating transcription factor 4 was higher in bepridil-treated A375 and SK-MEL-28 cells than in the corresponding control groups, but was lower in the A375 and SK-MEL-28 cells treated with bepridil and BAPTA (a calcium chelator) than in the corresponding cells treated with bepridil alone. Conclusion:The NCX inhibitor bepridil could increase intracellular Ca 2+ levels, suppress the proliferation and migration, and promote the apoptosis of melanoma cells, which may be closely related to biological processes such as endoplasmic reticulum stress.

Objective:To investigate the correlation between systemic immune inflammatory index (SII) and other metabolic indicators and diabetic epiretinal membranes (dERM).Methods:A retrospective case-control study. From March 2022 to July 2023, 81 patients (81 eyes) with dERM in Department of Ophthalmology, Affiliated Jinhua Hospital of Zhejiang University of Medicine School diagnosed by fundus screening were included in the study. A total of 81 patients (81 eyes) with diabetes who were matched in age, gender, and duration of diabetes and had no dERM or diabetic macular edema in both eyes during fundus screening were selected as the control group. All patients underwent optical coherence tomography (OCT) examination and laboratory tests for peripheral blood neutrophil, lymphocyte, platelet counts, serum albumin, blood lipids, uric acid, and glycosylated hemoglobin (HbA1c). SII was calculated. Random urine samples were collected for urinary albumin/creatinine ratio (ACR) testing. The OCT device's own analysis software obtained the macular volume coefficient, including central foveal thickness (CMT), macular volume, and average macular thickness. The macular volume coefficient, SII, serum albumin, blood lipids, uric acid, HbA1c, and ACR between the two groups were compared using paired t tests or Mann-Whitney U tests. Conditional logistic regression analysis was performed to evaluate the risk factors for dERM; Spearman correlation test was used to analyze the correlation between CMT, SII, ACR, disorganization of retinal inner layers (DRIL), intraretinal cyst (IRC), and hyper-reflective foci (HRF) in patients with dERM. Results:There were significant differences in CMT, macular volume, average macular thickness, SII, serum albumin, and ACR between the dERM group and the control group ( Z=-7.234, -6.306, -6.400, -3.063, -2.631, -3.868; P<0.05). Conditional logistics regression analysis showed that high SII [odds ratio (OR)= 3.919, 95% confidence interval ( CI) 1.591-9.654, P=0.003] and ACR ( OR=4.432, 95% CI 1.885-10.420, P=0.001) were risk factors for dERM. Spearman correlation analysis showed that HRF, IRC, DRIL were positively correlated with CMT ( Rs=0.234, 0.330, 0.248; P=0.036, 0.003, 0.026); HRF was positively correlated with SII and ACR ( Rs=0.233, 0.278; P=0.036, 0.012). Conclusion:Elevated SII and ACR are independent risk factors for the occurrence of dERM.

Objective:To evaluate the application value of extended latissimusdorsi musculocutaneous flap combined with breast precision hand in stage Ⅰ breast reconstruction.Methods:Twelve cases of extended latissimusdorsi musculocutaneous flap combined with stage Ⅰ breast reconstruction for breast cancer precision surgery were collected from January 2022 to February 2023 in our department, the postoperative complications of surgical techniques were analyzed, and the cosmetic effects of breast reconstruction were evaluated according to Harris standards.Results:All 12 cases underwent breast reconstruction with extended latissimusdorsi musculocutaneous flap, and the survival rate of the flap was 100%. There was local cutaneous margin necrosis in 1 case of back incision, partial ischemic necrosis of the nipple in 1 case, and effusion in the donor area of the back in 5 cases. Harris evaluation of cosmetic effect of breast reconstruction was excellent in 3 cases, good in 6 cases, and fair in 3 cases. Follow-up time was 4-16 months (median was 12 months), and no local recurrence or distant metastasis was observed during follow-up.Conclusions:The extended latissimusdorsi musculocutaneous flap combined with precision mastectomy stage Ⅰ reconstruction is a safe and feasible treatment method for breast cancer.