OBJECTIVE To develop a method for the determination of tacrolimus （TAC） concentration in human whole blood and to apply it in clinical therapeutic drug monitoring. METHODS Whole blood samples were processed by protein precipitation with methanol. The determination was performed using liquid chromatography-tandem mass spectrometry （LC-MS/MS）， with ascomycin serving as the internal standard. Chromatographic separation was carried out on a Kinetex F5 100Å column with a mobile phase consisting of 0.1 mmol/L ammonium acetate containing 0.2 mmol/L formic acid and methanol. Gradient elution was performed at a flow rate of 0.4 mL/min. The injection volume was 5 μL. Detection was conducted using multiple reaction monitoring （ m / z 821.6→768.6 for TAC； m / z 809.4→756.1 for ascomycin） with an electrospray ionization source in positive ion mode. The study focused on 86 whole blood samples collected from 83 pedi atric patients who received TAC therapy at Children’s Hospital of Nanjing Medical University from September 1 to 30， 2025. The aforementioned method was employed to measure the TAC concentration in the whole blood samples. The correlation and agreement between the aforementioned method and the traditional enzyme multiplied immunoassay technique （EMIT） were evaluated through Spearman correlation analysis， Bland-Altman analysis， and Passing-Bablok regression analysis. RESULTS The linear range of TAC was 0.5-100 ng/mL； the evaluation results for accuracy， precision， extraction recovery， matrix effect， and stability tests all met the relevant requirements. Clinical application results showed that the median concentration of TAC in pediatric whole blood measured by LC-MS/MS and EMIT methods were 4.4 and 4.0 ng/mL， respectively. Moreover， the two methods exhibited a strong correlation （correlation coefficient of 0.848 1） and good agreement （average relative deviation of 6.5%）. CONCLUSIONS A reliable LC-MS/MS method for the determination of TAC concentration in human whole blood is successfully established. This method demonstrates strong correlation and good agreement with the EMIT method， making it suitable for clinical therapeutic drug monitoring.

Objective: To investigate the damaging effects of red blood cell supernatant (RBC-S) stored for different durations (7 d, 14 d, and 28 d) on vascular endothelial cells, and to explore the underlying mechanisms using bioinformatics analysis, so as to provide references for optimizing red blood cell transfusion strategies. Methods: Human umbilical vein endothelial cells (HUVECs) were co-cultured with RBC-S stored for 7, 14 and 28 days, designated as the 7 d group, 14 d group and 28 d group respectively, which were collectively defined as the experimental groups. Cell damage was evaluated by cell proliferation assay (Cell Counting Kit8, CCK8), lactate dehydrogenase (LDH) release assay, 4′, 6diamidino2phenylindole (DAPI) staining, and flow cytometry for apoptosis and reactive oxygen species (ROS) levels. The damage degree of RBC-S on vascular endothelial cells was assessed by statistical analysis of damage data among different groups. Since the damage effect reached a plateau at all time points, the 28 d storage group was selected as the representative for further mechanistic studies. Transcriptomic analysis was performed to explore the role of frizzled class receptor 1 (FZD1) and Wnt signaling pathway in red blood cell storagerelated endothelial dysfunction. Results: Compared with the control group, the storage groups treated with 7 d, 14 d, and 28 d RBC-S showed significantly decreased cell proliferation rates [control group 100%, 7 d group (69.51±2.30)%, 14 d group (74.54±2.89)%, 28 d group (73.59±2.36)%, P<0.05], significantly reduced numbers of DAPI-stained cell nuclei [control group (213±12.5) per field, 7 d group (140.33±17.04) per field, 14 d group (152.00±23.72) per field, 28 d group (144.33±19.09) per field, P<0.05] and significantly increased LDH release [control group (1), 7 d group (8.33±1.41), 14 d group (9.23±0.83), 28 d group (9.16±0.60), P<0.05]. There was no significant difference in the degree of damage caused by RBC-S among different storage groups (P>0.05). With the prolongation of storage time, free hemoglobin (FHb) gradually increased [control group (not detected), 7 d (16.57±6.38) mg/L, 14 d (76.80±22.83) mg/L, 28 d (286.97±29.02) mg/L, P<0.05]. The apoptotic rate (20.53±2.94)% and ROS relative intensity (5.13±0.91) in the 28 d storage group were significantly higher than those in the control group (P<0.05). Transcriptomic analysis showed that FZD1 played a key role in vascular endothelial dysfunction induced by red blood cell storage and was closely related to the Wnt signaling regulatory network. Conclusion: RBC-S stored for 7 d, 14 d, or 28 d can all significantly damage vascular endothelial cells, and the damaging effect reaches a plateau at 7 d of storage. Mechanistic investigation of the 28 d group indicated that the downregulation of the FZD1/Wnt signaling pathway may play a critical role in vascular endothelial dysfunction induced by red blood cell storage, providing a theoretical basis for further optimizing red blood cell storage and transfusion strategies.

To introduce and analyze guideline terminology related to clinical question formulation, evidence retrieval and appraisal, and recommendation development.

OBJECTIVES: Hypertriglyceridemic acute pancreatitis (HTG-AP) has a rapid onset and is associated with a high risk of progression and recurrence. Early identification of patients at risk of severe disease can help reduce the likelihood of multiple organ failure and mortality. This study aims to investigate the changes in inflammatory composite markers and D-dimer (D-D) levels in young and middle-aged/elderly patients with HTG-AP and to evaluate their predictive value for disease progression. METHODS: A total of 230 patients with HTG-AP admitted to Chongqing University Jiangjin Hospital (Jiangjin Central Hospital) between 2017 and 2023 were retrospectively enrolled. Patients were first divided into a young group (≤45 years) and a middle-aged/elderly group (>45 years), and then stratified into mild and severe groups based on disease severity. Inflammatory composite markers, neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), monocyte-to-lymphocyte ratio (MLR), C-reactive protein-to-lymphocyte ratio (CLR), systemic inflammation response index (SIRI), systemic immune inflammation index (SII), as well as D-D levels, were compared among groups. Least absolute shrinkage and selection operator (LASSO) regression and Logistic regression were used to identify independent risk factors for disease progression in each age group. Receiver operating characteristic (ROC) curves and the DeLong test were used to assess and compare the predictive performance (area under the curve, AUC) of risk factors. Internal validation was performed using the bootstrap method (n=1 000). RESULTS: No significant differences in NLR, PLR, MLR, SIRI, SII, CLR, or D-D levels were observed between the young (n=127) and middle-aged/elderly (n=103) groups (all P>0.05). Among young patients, the severe group (n=59) had significantly higher NLR, SIRI, SII, CLR, and D-D levels compared to the mild group (n=68) (all P<0.05). Among middle-aged/elderly patients, CLR and D-D levels were significantly higher in the severe group (n=49) than in the mild group (n=54) (P<0.05). LASSO and Logistic regression analyses identified elevated D-D as an independent risk factor for disease progression in young patients (P=0.007, OR=1.458, 95% CI 1.107 to 1.920), while both D-D (P=0.001, OR=2.267, 95% CI 1.413 to 3.637) and CLR (P=0.003, OR=1.007, 95% CI 1.003 to 1.012) were independent risk factors in middle-aged/elderly patients. ROC analysis showed that D-D predicted disease progression in young and middle-aged/elderly patients with AUCs of 0.653 and 0.741, sensitivities of 67.8% and 57.1%, and specificities of 72.1% and 88.9%, respectively. CLR predicted progression in middle-aged/elderly patients with an AUC of 0.687, sensitivity of 63.3%, and specificity of 70.4%. DeLong test showed no significant difference in AUC between D-D and CLR for middle-aged/elderly patients (Z=0.993, P=0.321). Internal validation via bootstrap analysis yielded a D-D AUC of 0.732, with sensitivity and specificity of 68.1% and 91.0%, respectively. CONCLUSIONS Differences in inflammatory response and coagulation function exist across age groups and disease severities in HTG-AP patients. Elevated D-D is an independent predictor of disease progression in both young and middle-aged/elderly patients, while CLR also predicts progression in the latter group. D-D, in particular, demonstrates strong predictive value for severe disease in middle-aged/elderly patients with HTG-AP.
Humans ; Fibrin Fibrinogen Degradation Products/metabolism* ; Disease Progression ; Middle Aged ; Pancreatitis/etiology* ; Male ; Female ; Retrospective Studies ; Adult ; Biomarkers/blood* ; Hypertriglyceridemia/blood* ; Acute Disease ; Predictive Value of Tests ; Aged ; Inflammation ; C-Reactive Protein/analysis* ; Neutrophils ; Age Factors

Objective To investigate the effects of a continuous-dose administration versus different dosage regimens of polyethylene glycol electrolyte solution(PEG)taken in two doses with a 12-hour interval on bowel cleansing efficacy,with the goal of optimizing bowel preparation protocols and improving patient tolerability.Methods 232 patients who underwent painless colonoscopy and used PEG as a bowel cleanser from June 2024 to September 2024 were selected as study subjects.Participants were divided into three groups:the control group(3.00 L PEG continuous dose),experimental group A(0.75 L+2.25 L PEG),and experimental group B(1.50 L+1.50 L PEG).All patients underwent painless colonoscopy within 4～6 h after completing PEG intake.The interval between the two doses of PEG in group A and group B was 12 h.The bowel cleansing efficacy was assessed by using the Boston bowel preparation scale(BBPS),and the rates of colon polyp detection,adverse reactions,sleep duration,and tolerability were recorded.Results There were no significant statistical differences in BBPS scores and colon polyp detection rates among the three groups(P>0.05).Experimental group B experienced the least adverse reactions,followed by experimental group A,while the control group reported the most significant adverse reactions(P<0.05).The timing of PEG administration did not have a significant impact on sleep duration among the three groups(P>0.05).Patients in experimental group B showed good tolerability to PEG and were willing to accept this bowel preparation regimen,followed by group A,while the control group exhibited the poorest tolerability,with significant statistical differences among the three groups(P<0.05).Conclusion The continuous administration and divided administration of PEG have no significant impact on the effectiveness of intestinal cleansing and the detection rate of colonic polyps.However,the divided PEG regimen with a 12 h interval results in fewer adverse reactions and better tolerance,especially the optimal regimen of taking 1.50 L PEG in two doses with a 12 h interval.

Objective To sort,summarize,and introduce key terms related to guideline reporting,evaluation,dissemination,implementation,and updating.Methods We systematically searched guideline de-velopment manuals and methodological literature from database inception to October 25,2024.Terms related to guideline reporting,updating,evaluation,and implementation were extracted,standardized,and finalized through a structured consensus process.Results A total of 13 guideline manuals and 32 methodological articles were included,yielding 14 core terms with standardized definitions.Conclusions This article introduces key terms such as reporting standards,external review,and research gaps across guideline development phases to promote concept application and deepen readers'understanding of guideline development.

Objective:To Explore the changes of annexin A2 and vascular endothelial cadherin (VE-Cad) in patients with cerebral infarction (CI) undergoing emergency thrombolysis, and analyze their relationship with progression within 10 d.Methods:Using a prospective research method, 78 patients with CI were selected from October 2019 to June 2022 in Xi'an International Medical Center Hospital, and all patients were treated with emergency thrombolysis. The serum levels of annexin A2 and VE-Cad before and after thrombolysis were measured by enzyme-linked immunosorbent assay, and the National Institute stroke scale (NIHSS) was used to assess patients' neurologic impairment. The baseline data, imaging findings at admission and routine laboratory examination indexes were recorded. The progression within 10 d after thrombolysis was recorded. Pearson correlation analysis was used to analyze the correlation between annexin A2, VE-Cad and NIHSS score. Multivariate Logistic regression was used to analyze the independent risk factors of progression within 10 d after thrombolysis in patients with CI. The value of annexin A2 and VE-Cad in predicting the progression within 10 d after thrombolysis in patients with CI was evaluated by the receiver operating characteristics (ROC) curve. A restricted cubic spline model was drawn to evaluate the dose-response relationship between annexin A2, VE-Cad and the progression within 10 d after thrombolysis in patients with CI.Results:Compared with before thrombolysis, the annexin A2 after thrombolysis was significantly higher: (24.50 ± 3.27) μg/L vs. (20.86 ± 3.84) μg/L, the VE-Cad and NIHSS score were significantly lower: (4.72 ± 1.05) mg/L vs. (6.81 ± 1.31) mg/L and (8.64 ± 2.35) scores vs. (13.01 ± 2.86) scores, and there were statistical differences ( P<0.01). Before and after thrombolysis, Pearson correlation analysis result showed there was a negative correlation between annexin A2 and NIHSS score ( r =-0.796 and - 0.568, P<0.01), and a positive correlation between VE-Cad and NIHSS score ( r = 0.820 and 0.502, P<0.01). Among 78 patients with CI treated with emergency thrombolysis, 7 cases (8.97%) experienced progression within 10 d. There were statistical differences in hypertension, diabetes, hyperlipidemia, onset to thrombolysis time, infarct site, systolic blood pressure, triacylglycerol, high-density lipoprotein cholesterol, and the NIHSS score, annexin A2, VE-Cad before and after thrombolysis between patients with progression within 10 d after thrombolysis and patients without progression within 10 d after thrombolysis ( P<0.05 or <0.01); there were no statistical differences in gender composition, age, body mass index, coronary heart disease, atrial fibrillation, smoking, alcohol consumption, family history of stroke, carotid plaques, blood glucose, diastolic blood pressure, white blood cell count, platelet count, total cholesterol low-density lipoprotein cholesterol between the two groups ( P>0.05). After adjusting for hypertension, diabetes and hyperlipidemia, multivariate Logistic regression analysis result showed that the infarction site, onset to thrombolysis time, VE-Cad after thrombolysis and annexin A2 after thrombolysis were still independent factors of progression within 10 d after thrombolysis in patients with CI ( OR = 2.570, 2.496, 3.147 and 0.352; 95% CI 1.285 to 5.139, 1.303 to 4.781, 1.629 to 6.080 and 0.158 to 0.782; P<0.05 or <0.01). ROC curve analysis results showed that the area under the curve of annexin A2 combined with VE-Cad after thrombolysis to predict the progression within 10 d after thrombolysis in patients with CI was significantly larger than that of annexin A2 and VE-Cad after thrombolysis alone (0.898 vs. 0.822 and 0.799, χ2 = 2.17 and 1.98, P = 0.039 and 0.048). The optimal cutoff values of annexin A2 and VE-Cad after thrombolysis were <23.27 μg/L and >4.92 mg/L, with a sensitivity of 88.24%, and a specificity of 77.05%. The restricted cubic spline analysis result showed that the continuous changes in annexin A2 after thrombolysis were roughly negatively correlated with the progression within 10 d after thrombolysis in patients with CI ( OR = 0.720, 95% CI 0.561 to 0.930, P = 0.010), the continuous changes in VE-Cad after thrombolysis were roughly positively correlated with the progression within 10 d after thrombolysis in patients with CI ( OR = 1.450, 95% CI 1.126 to 1.188, P = 0.004). When annexin A2<23.80 ng/L and VE-Cad>5.25 mg/L after thrombolysis, the risk of progression within 10 d after thrombolysis in patients with CI significantly increased. Conclusions:The expression of annexin A2 increases and VE-Cad decreases after emergency thrombolysis in patients with CI, and the expression levels of both are closely related to the degree of neurologic impairment, and the risk of progression within 10 d after thrombolysis could be determined clinically by detecting their changes.

Oxidative stress due to aberrant metabolism is considered as a crucial contributor to diabetes and its complications. Hyperglycemia and hyperlipemia boost excessive reactive oxygen species generation by elevated mitochondrial respiration, increased nicotinamide adenine dinucleotide phosphate oxidase activity, and enhanced pro-oxidative processes, including protein kinase C pathways, hexosamine, polyol, and advanced glycation endproducts, which exacerbate oxidative stress. Oxidative stress plays a significant role in the onset of diabetes and its associated complications by impairing insulin production, increasing insulin resistance, maintaining hyperglycemic memory, and inducing systemic inflammation. A more profound comprehension of the molecular processes that link oxidative stress to diabetes is crucial to new preventive and therapeutic strategies. Therefore, this review discusses the mechanisms underlying how oxidative stress contributes to diabetes mellitus and its complications. We also summarize the current approaches for prevention and treatment by targeting the oxidative stress pathways in diabetes.
Oxidative Stress/physiology* ; Humans ; Diabetes Mellitus/physiopathology* ; Diabetes Complications/metabolism* ; Reactive Oxygen Species/metabolism* ; Glycation End Products, Advanced/metabolism* ; Animals

Objective:To Explore the changes of annexin A2 and vascular endothelial cadherin (VE-Cad) in patients with cerebral infarction (CI) undergoing emergency thrombolysis, and analyze their relationship with progression within 10 d.Methods:Using a prospective research method, 78 patients with CI were selected from October 2019 to June 2022 in Xi'an International Medical Center Hospital, and all patients were treated with emergency thrombolysis. The serum levels of annexin A2 and VE-Cad before and after thrombolysis were measured by enzyme-linked immunosorbent assay, and the National Institute stroke scale (NIHSS) was used to assess patients' neurologic impairment. The baseline data, imaging findings at admission and routine laboratory examination indexes were recorded. The progression within 10 d after thrombolysis was recorded. Pearson correlation analysis was used to analyze the correlation between annexin A2, VE-Cad and NIHSS score. Multivariate Logistic regression was used to analyze the independent risk factors of progression within 10 d after thrombolysis in patients with CI. The value of annexin A2 and VE-Cad in predicting the progression within 10 d after thrombolysis in patients with CI was evaluated by the receiver operating characteristics (ROC) curve. A restricted cubic spline model was drawn to evaluate the dose-response relationship between annexin A2, VE-Cad and the progression within 10 d after thrombolysis in patients with CI.Results:Compared with before thrombolysis, the annexin A2 after thrombolysis was significantly higher: (24.50 ± 3.27) μg/L vs. (20.86 ± 3.84) μg/L, the VE-Cad and NIHSS score were significantly lower: (4.72 ± 1.05) mg/L vs. (6.81 ± 1.31) mg/L and (8.64 ± 2.35) scores vs. (13.01 ± 2.86) scores, and there were statistical differences ( P<0.01). Before and after thrombolysis, Pearson correlation analysis result showed there was a negative correlation between annexin A2 and NIHSS score ( r =-0.796 and - 0.568, P<0.01), and a positive correlation between VE-Cad and NIHSS score ( r = 0.820 and 0.502, P<0.01). Among 78 patients with CI treated with emergency thrombolysis, 7 cases (8.97%) experienced progression within 10 d. There were statistical differences in hypertension, diabetes, hyperlipidemia, onset to thrombolysis time, infarct site, systolic blood pressure, triacylglycerol, high-density lipoprotein cholesterol, and the NIHSS score, annexin A2, VE-Cad before and after thrombolysis between patients with progression within 10 d after thrombolysis and patients without progression within 10 d after thrombolysis ( P<0.05 or <0.01); there were no statistical differences in gender composition, age, body mass index, coronary heart disease, atrial fibrillation, smoking, alcohol consumption, family history of stroke, carotid plaques, blood glucose, diastolic blood pressure, white blood cell count, platelet count, total cholesterol low-density lipoprotein cholesterol between the two groups ( P>0.05). After adjusting for hypertension, diabetes and hyperlipidemia, multivariate Logistic regression analysis result showed that the infarction site, onset to thrombolysis time, VE-Cad after thrombolysis and annexin A2 after thrombolysis were still independent factors of progression within 10 d after thrombolysis in patients with CI ( OR = 2.570, 2.496, 3.147 and 0.352; 95% CI 1.285 to 5.139, 1.303 to 4.781, 1.629 to 6.080 and 0.158 to 0.782; P<0.05 or <0.01). ROC curve analysis results showed that the area under the curve of annexin A2 combined with VE-Cad after thrombolysis to predict the progression within 10 d after thrombolysis in patients with CI was significantly larger than that of annexin A2 and VE-Cad after thrombolysis alone (0.898 vs. 0.822 and 0.799, χ2 = 2.17 and 1.98, P = 0.039 and 0.048). The optimal cutoff values of annexin A2 and VE-Cad after thrombolysis were <23.27 μg/L and >4.92 mg/L, with a sensitivity of 88.24%, and a specificity of 77.05%. The restricted cubic spline analysis result showed that the continuous changes in annexin A2 after thrombolysis were roughly negatively correlated with the progression within 10 d after thrombolysis in patients with CI ( OR = 0.720, 95% CI 0.561 to 0.930, P = 0.010), the continuous changes in VE-Cad after thrombolysis were roughly positively correlated with the progression within 10 d after thrombolysis in patients with CI ( OR = 1.450, 95% CI 1.126 to 1.188, P = 0.004). When annexin A2<23.80 ng/L and VE-Cad>5.25 mg/L after thrombolysis, the risk of progression within 10 d after thrombolysis in patients with CI significantly increased. Conclusions:The expression of annexin A2 increases and VE-Cad decreases after emergency thrombolysis in patients with CI, and the expression levels of both are closely related to the degree of neurologic impairment, and the risk of progression within 10 d after thrombolysis could be determined clinically by detecting their changes.

Objective:To explore the effect of Bian-stone warming and ironing therapy on gastrointestinal blood flow in patients with severe cerebrovascular disease undergoing therapeutic hypothermia.Methods:From January 2023 to October 2024, 92 patients with severe cerebrovascular disease in the Neurosurgical Intensive Care Unit of Xuanwu Hospital of Capital Medical University were selected for the study using convenience sampling method. The patients were divided into a control group and an experimental group of 46 cases each according to the randomized numerical table method. Control group was given routine nursing, and experimental group carried out Bian-stone warming and ironing therapy on the basis of control group. Peak systolic velocity (PSV), end-diastolic velocity (EDV), and time-averaged mean velocity (TAMV) of the superior mesenteric artery of the two groups of patients were monitored before and after five days of intervention, respectively, and the situation of gastric retention and bowel sounds were recorded and compared between the groups.Results:PSV was higher in experimental group than in control group after intervention, and the difference was statistically significant ( P<0.05). The difference in EDV between the two groups was not statistically significant ( P>0.05). TAMV of experimental group was higher than that of control group after intervention, and the difference in TAVM before and after intervention of experimental group was smaller than that of control group, and the differences were all statistically significant ( P<0.05). The number of cases of gastric retention and weakened or disappeared bowel sounds in experimental group was less than that in control group after intervention, and the differences were all statistically significant ( P<0.05) . Conclusions:Bian-stone warming and ironing therapy improves gastrointestinal blood flow rate and reduces feeding intolerance in patients with severe cerebrovascular disease undergoing therapeutic hypothermia.