ObjectiveTo investigate the effects of Huanglian Jiedutang （HLJDT） on cognitive function in mice with ischemic stroke （IS） and to elucidate whether its neuroprotective effects are mediated by inhibition of the nuclear factor-κB （NF-κB） signaling pathway and subsequent suppression of NF-κB-regulated neuronal apoptosis. MethodsAn IS model was established using middle cerebral artery occlusion （MCAO）. Sixty C57BL/6J mice were randomly assigned to five groups （n =12 per group）， i.e.， sham operation， model， HLJDT low-dose （3.9 g·kg^-1·d^-1）， HLJDT high-dose （7.8 g·kg^-1·d^-1）， and Ginkgo biloba extract （GBE， 31.2 mg·kg^-1·d^-1）. Post-operatively， neurological deficit scores （Longa score）， cerebral infarct volume assessed by 2，3，5-triphenyltetrazolium chloride （TTC） staining， and brain water content were evaluated. Learning and memory were assessed using new object recognition （NOR） and fear conditioning （FC） tests. Hippocampal pathology was examined via hematoxylin and eosin （HE） staining. Immunofluorescence detected expression of glial fibrillary acidic protein （GFAP， astrocyte marker）， cellular oncogene Fos （c-Fos， neuronal activation marker）， and glutamate decarboxylase 65 （GAD65）. Western blot measured nuclear factor-κB inhibitor protein α （IκBα）， phosphorylated IκBα （p-IκBα）， NF-κB p65， phosphorylated NF-κB p65 （p-NF-κB p65）， ionic calcium binding adapter molecule 1 （Iba-1）， tumor necrosis factor （TNF）-α， interleukin （IL）-1β， and apoptosis-related proteins， such as cleaved cysteinyl aspartate-specific protease 3 （Caspase-3）， B-cell lymphoma 2 （Bcl-2）， and Bcl-2-associated X protein （Bax）. Real-time quantitative PCR （Real-time PCR） was used to assess mRNA levels of Iba-1， TNF-α， IL-1β， NF-κB p65， cleaved Caspase-3， Bax， and Bcl-2. ResultsCompared with the sham group， the model group exhibited significantly increased neurological deficit scores， brain water content， and cerebral infarct volume （P<0.01）. Hippocampal CA1 neurons were disorganized， showing nuclear pyknosis and karyolysis. NOR exploration time and FC freezing time were significantly reduced （P<0.01）. GFAP and c-Fos expression were increased， while GAD65 expression was decreased （P<0.01）. Cleaved Caspase-3 and Bax were upregulated， Bcl-2 was downregulated， and the Bax/Bcl-2 ratio was elevated （P<0.01）. Expression levels of p-IκBα， p-NF-κB p65， IL-1β， TNF-α， and Iba-1 were significantly increased （P<0.01）. Compared with the model group， HLJDT high-dose， low-dose， and GBE groups showed significant improvements in all parameters （P<0.01）. Among them， the HLJDT high-dose group showed the most pronounced neuronal structural recovery and superior performance in NOR and FC tests （P<0.01）. In this group， GFAP and c-Fos decreased， GAD65 increased （P<0.01）， apoptosis-related protein expression was reversed， and NF-κB signaling and related inflammatory factor expression were suppressed （P<0.01）. ConclusionHLJDT ameliorates cognitive dysfunction in mice after IS， potentially by inhibiting the NF-κB signaling pathway， thereby reducing neuroinflammation and hippocampal neuronal apoptosis.

Objective To establish the reference interval of urine Alzheimer-associated neuronal thread protein(AD7c-NTP)for healthy adults in Mianyang area using the indirect method.Methods The detection results of urine AD7c-NTP from 5 093 healthy in-dividuals were collected from the information management database of Medical Laboratory Department of Sichuan Science City Hospital from March 2017 to March 2022.Skewness-kurtosis and Kolmogorov-Smirnov tests were used to determine whether the data followed a normal distribution.After removing outliers using the Box Plots method,the enrolled subjects were grouped by gender and age.The Mann-Whitney U or Kruska-Wallis H tests were used to analyze the between-group differences of urine AD7c-NTP in healthy individu-als with different genders and ages.The adjacent age groups without statistically significant difference(P＞0.05)were combined,and the indirect method(non-parametric test method)was used to calculate the reference intervals for different gender and age groups.Results Skewness-kurtosis and Kolmogorov-Smirnov tests showed that the data followed a non-normal distribution.After removing 293 outliers using the Box Plots method,a total of 4 800 subjects,including 3 199 males and 1 601 females,were enrolled.The enrolled subjects were grouped by gender and age,and the non-parametric test method were used to establish the reference intervals of urine AD7c-NTP in healthy populations with different genders.The Mann-Whitney U test confirmed that urine AD7c-NTP levels existed gen-der differences(Z=14.09,P＜0.01),and the reference intervals for males and females were≤1.10 ng/mL and≤1.40 ng/mL,re-spectively.There were also statistical differences in urine AD7c-NTP levels among different age groups of the same gender.After combi-ning adjacent age groups without statistically significant difference(P＞0.05),the reference intervals of urine AD7c-NTP in healthy populations with different genders and ages were established by the non-parametric test method,which were≤1.00 ng/mL for male 20-39 years old group,≤1.10 ng/mL for male 40-79 years old group,≤1.60 ng/mL for male≥80 years old group,≤1.30 ng/mL for female 20-69 years old group,and≤1.60 ng/mL for female≥70 years old group,respectively.The established reference intervals of urine AD7c-NTP were further verified by healthy individuals,and the results met the standards.Conclusion The reference intervals of urine AD7c-NTP in healthy populations with different genders and ages in Mianyang area are established successfully using the indi-rect method,which may help to predict the risk of Alzheimer's disease in clinical practice and provide support for the diagnosis and treatment of the disease.

Objective To establish the reference interval of urine Alzheimer-associated neuronal thread protein(AD7c-NTP)for healthy adults in Mianyang area using the indirect method.Methods The detection results of urine AD7c-NTP from 5 093 healthy in-dividuals were collected from the information management database of Medical Laboratory Department of Sichuan Science City Hospital from March 2017 to March 2022.Skewness-kurtosis and Kolmogorov-Smirnov tests were used to determine whether the data followed a normal distribution.After removing outliers using the Box Plots method,the enrolled subjects were grouped by gender and age.The Mann-Whitney U or Kruska-Wallis H tests were used to analyze the between-group differences of urine AD7c-NTP in healthy individu-als with different genders and ages.The adjacent age groups without statistically significant difference(P＞0.05)were combined,and the indirect method(non-parametric test method)was used to calculate the reference intervals for different gender and age groups.Results Skewness-kurtosis and Kolmogorov-Smirnov tests showed that the data followed a non-normal distribution.After removing 293 outliers using the Box Plots method,a total of 4 800 subjects,including 3 199 males and 1 601 females,were enrolled.The enrolled subjects were grouped by gender and age,and the non-parametric test method were used to establish the reference intervals of urine AD7c-NTP in healthy populations with different genders.The Mann-Whitney U test confirmed that urine AD7c-NTP levels existed gen-der differences(Z=14.09,P＜0.01),and the reference intervals for males and females were≤1.10 ng/mL and≤1.40 ng/mL,re-spectively.There were also statistical differences in urine AD7c-NTP levels among different age groups of the same gender.After combi-ning adjacent age groups without statistically significant difference(P＞0.05),the reference intervals of urine AD7c-NTP in healthy populations with different genders and ages were established by the non-parametric test method,which were≤1.00 ng/mL for male 20-39 years old group,≤1.10 ng/mL for male 40-79 years old group,≤1.60 ng/mL for male≥80 years old group,≤1.30 ng/mL for female 20-69 years old group,and≤1.60 ng/mL for female≥70 years old group,respectively.The established reference intervals of urine AD7c-NTP were further verified by healthy individuals,and the results met the standards.Conclusion The reference intervals of urine AD7c-NTP in healthy populations with different genders and ages in Mianyang area are established successfully using the indi-rect method,which may help to predict the risk of Alzheimer's disease in clinical practice and provide support for the diagnosis and treatment of the disease.

Aristolochic acid (AA), extracted from Aristolochiaceae plants, plays an essential role in traditional herbal medicines and is used for different diseases. However, AA has been found to be nephrotoxic and is known to cause aristolochic acid nephropathy (AAN).AA-induced acute kidney injury (AKI) is a syndrome in AAN with a high morbidity that manifests mitochondrial damage as a key part of its pathological progression. Melatonin primarily serves as a mitochondria-targeted antioxidant. However, its mitochondrial protective role in AA-induced AKI is barely reported. In this study, mice were administrated 2.5 mg/kg AA to induce AKI. Melatonin reduced the increase in Upro and Scr and attenuated the necrosis and atrophy of renal proximal tubules in mice exposed to AA. Melatonin suppressed ROS generation, MDA levels and iNOS expression and increased SOD activities in vivo and in vitro. Intriguingly, the in vivo study revealed that melatonin decreased mitochondrial fragmentation in renal proximal tubular cells and increased ATP levels in kidney tissues in response to AA. In vitro, melatonin restored the mitochondrial membrane potential (MMP) in NRK-52E and HK-2 cells and led to an elevation in ATP levels. Confocal immunofluorescence data showed that puncta containing Mito-tracker and GFP-LC3A/B were reduced, thereby impeding the mitophagy of tubular epithelial cells. Furthermore, melatonin decreased LC3A/B-II expression and increased p62 expression. The apoptosis of tubular epithelial cells induced by AA was decreased. Therefore, our findings revealed that melatonin could prevent AA-induced AKI by attenuating mitochondrial damage, which may provide a potential therapeutic method for renal AA toxicity.

ObjectiveTo investigate the Alzheimer-associated neurofilament protein （AD7c-NTP） in urine of middle-aged and elderly people and its correlation between common metabolites. MethodsA total of 1 150 middle-aged and elderly people who did their physical exmanination in the health examination center of the Sichuan Science City Hospital and the Third Hopital of Mianyang were recruited from March 2017 to March 2020. The level of urine AD7c-NTP were measured by enzyme-linked immunosorbent assay （ELISA）， and common metabolites in blood were measured by biochemical analyzer. Based on urine AD7c-NTP level ≤1.5 ng/mL， the objects was divided into normal group （n=956） and elevated group （n=194）. Thier demographic data and blood biochemical indicators were collected. ResultsThe urine AD7c-NTP level in middle-aged and elderly people was 0.60（0.30~1.20） ng/mL. The urine AD7c-NTP level was higher in women than that in men ［1.04（0.40~1.30） ng/mL vs. 0.84（0.30~1.00） ng/mL， Z=4.202， P˂0.01］. And the urine AD7c-NTP level was lower in the normal group than that in the elevated group ［0.50（0.30~0.90） ng/mL vs. 2.10（1.70~2.10） ng/mL， Z=22.035， P˂0.01］. The results of the univariate comparison showed that， the differences between the two groups in age （Z=6.545）， fasting glucose （Z=3.506）， blood uric acid （Z=2.574）， urea nitrogen （Z=2.891）， creatinine （Z=2.243）， total bilirubin （Z=3.936）， glutathione （Z=0.969）， total cholesterol （t=3.956） and low density lipoprotein （Z=-5.678） were were statistically significant （P˂0.05 or 0.01）. Spearman correlation analysis showed that， the urine AD7c-NTP level was positively correlated with age and the levels of urea nitrogen， glucose， total cholesterol and low density lipoprotein （r=0.177， 0.178， 0.171， 0.109， 0.149， P˂0.01）， and negatively correlated with the level of total bilirubin （r=-0.172， P˂0.01）. Conclusionthe urine AD7c-NTP level in middle-aged and elderly females was signifitcantly higher than in middle-aged and elderly males.The urine AD7c-NTP level of middle-aged and elderly people was positively correlated with age， urea nitrogen， glucose， total cholesterol and low density lipoprotein， and negatively correlated with total bilirubin.

Objective To study the effect of endovenous laser ablation combined with subfascial endoscopy therapy for venous ulcer of the lower limbs.Methods Clinical data of 112 patients undergoing endovenous laser ablation combined with subfascial endoscopy therapy for venous ulcer of the lower limbs from Oct 2011 to Feb 2016 was retrospectively evaluated.Results Patients had average 2-6 perforating veins in their affected lower limbs.Procedures were successful in all the patients,all the superficial varicose veins disappeared after 1 month and 1 year of ultrasound follow-up,perforating veins closed,ulcer healed,and chromatosis alleviated.Conclusion Endovenous laser ablation combined with subfascial endoscopy therapy for venous ulcer of the lower limbs is effective,safe and quick recovery.

Objective To investigate the multi-detector computed tomography (MDCT) evaluating the clinical staging of adenocarcinoma of the esophagogastric junction (AEG) after neoadjuvant chemotherapy.Methods The retrospective cross-sectional study conducted.The clinicopathological data of 46 AEG patients who were admitted to the Peking University Cancer Hospital between January 2016 and April 2018 were collected.All patients underwent MDCT before and after neoadjuvant chemotherapy and at preoperative 2 weeks,the distance between tumor center and boundary of esophagogastric junction (EGJ) was judged through coronal measured values and axial formula method.Patients underwent radical resection of gastric cancer + D2 lymph node dissection after neoadjuvant chemotherapy,pathologists reviewed the distance between center of AEG and boundary of EGJ,T staging (ycT) and N staging (ycN) of clinical staging,T staging (ypT) and N staging (ypN) of pathological staging after neoadjuvant chemotherapy were determined according to TNM staging of American Joint Committee on Cancer (AJCC) (8th edition),and tumor regression grading (TRG) was determined according to the criterion established by National Comprehensive Cancer Network.Observation indicators:(1) CT examination after neoadjuvant chemotherapy;(2) clinical staging after neoadjuvant chemotherapy;(3) postoperative pathological examination;(4) postoperative pathological staging;(5) accuracy of clinical staging after neoadjuvant chemotherapy;(6)relationship between imaging changes of CT examination and pathological reactions.Count data were described as absolute number or percentage,and comparisons among groups were analyzed by the chi-square test.Comparisons of ordinal data were analyzed by the non-parametric test.Results (1) CT examination after neoadjuvant chemotherapy:5 of 46 AEG patients,coronal images of CT showed whole tumor and boundary of EGJ,axial images of CT showed EGJ wall thickening,heterogeneous enhancement in all layers of lesions,and unsmooth serosal surface;the distance between tumor center and boundary of EGJ is less than 2 cm by direct measurement,5 patients were confirmed as esophageal cancer staging.For 41 patients,the same coronal image of CT cannot showed whole tumor and boundary of EGJ,axial images of CT showed EGJ wall thickening,heterogeneous enhancement in all layers of lesions,and irregular-shaped serosal surface;27 patients whose calculated values were negative based on formula method used esophageal cancer staging,and 14 patients whose calculated values were positive used gastric staging.(2) Clinical staging after neoadjuvant chemotherapy:among 46 AEG patients,ycT staging:staging ycT1,ycT2,ycT3,ycT4a and ycT4b were respectively detected in 1,6,31,6 and 2 patients;ycN staging:staging ycN0,ycN1,ycN2 and ycN3a were respectively detected in 5,14,23 and 4 patients.(3) Postoperative pathological examination:of 46 patients,38,3,3 and 2 were respectively confirmed as adenocarcinoma,adenocarcinoma with signet-ring cell carcinoma,adenocarcinoma with neuroendocrine carcinoma and adenocarcinoma with squamous carcinoma.Of 46 patients,the distance between tumor center and boundary of EGJ can be observed in 14 patients by gastric cancer staging and 32 patients by esophageal cancer staging.(4) Postoperative pathological staging:ypT staging:1,3,5,29,7 and 1 patients were respectively detected in staging ypT0,ypT1,ypT2,ypT3,ypT4a and ypT4b;ypN staging:17,4,15,9 and 1 patients were respectively detected in staging ypN0,ypN1,ypN2,ypN3a and ypN3b.One,3,16 and 26 patients were confirmed as staging TRG 0,TRG 1,TRG 2 and TRG 3,including 20 patients tumor regression and 26 patients without tumor regression.(5) Accuracy of clinical staging after neoadjuvant chemotherapy:the accuracies of ycT staging and ycN staging were 78.3％ (36/46) and 54.3％ (25/46).(6) Relationship between imaging changes of CT examination and pathological reactions:of 46 patients,33 and 13 had respectively reduced and stable gastric wall thickness of primary lesion.Among 20 patients with tumor regression,17 and 3 had respectively reduced and stable gastric wall thickness of primary lesion;of 26 patients without tumor regression,reduced and stable gastric wall thickness of primary lesion were respectively in 16 and 10 patients,with no statistically significant difference (x2 =3.069,P＞0.05).Of 46 patients,31,14 and 1 had respectively reduced,stable and increased sum of minor diameters of suspicious celiac lymph nodes.The reduced,stable and increased sum of minor diameters of suspicious celiac lymph nodes were detected in 16,4,0 of 20 patients with tumor regression and 15,10,1 of 26 patients without tumor regression,respectively,with no statistically significant difference (Z =-1.629,P＞ 0.05).The changes of gastric wall thickness of primary lesion and sum of minor diameters of celiac lymph nodes before operation were not consistent to that after operation in 3 patients.CT examination showed gastric wall thickness of primary lesion reduced after chemotherapy,and sum of minor diameters of celiac lymph nodes didn't change;pathological staging and clinical staging were respectively in staging ypN0 and ycN1.Conclusion According to the TNM staging of AJCC (Sth edition),the distance between tumor center and boundary of EGJ is judged through coronal measured values and axial formula method and therefore determining to select staging system of esophageal cancer or gastric cancer,meanwhile,rectifying over T3 staging of Siewert Ⅱ gastric cancer and increasing overall accuracy of clinical staging.

Objective To explore the computed tomography (CT) features of gastric cancer invasion to the pancreas and significance in the assessment of resectability of primary lesions.Methods The retrospective cohort study was conducted.The clinical data of 31 gastric cancer patients who were admitted to the Peking University Cancer Hospital between February 2011 and August 2016 were collected.Of 31 patients receiving CT examinations,11 who were diagnosed with suspected pancreas invasion by preoperative CT examinations but operation confirmed no invasion were allocated into the pancreas negative (PN) group,11 who were confirmed as pancreas invasion and under vent radical gastrectomy of gastric cancer combined with pancreas resection were allocated into the pancreas invasion (PI) group,and 9 who were confirmed as pancreas invasion and had unresectable primary lesions were allocated into the pancreas invasion non-resected (PI-NR) group.Observation indicators:(1) morphologic type of contact surface between gastric cancer and pancreas;(2) comparison of CT findings among the 3 groups:primary lesion location,tunor thickness,Borrmann type,serosa pattern of gastric cancer,judging obvious region invaded by gastric cancer,contact or invasion site with pancreas,contact length between gastric cancer and pancreas,pattern,clarity and CT values of contact surface or peripancreas invaded and normal peripancreas;(3) treatment or follow-up situations.All the patients underwent radical resection and palliative resection for gastric cancer or non-operation according to results of exploration.Telephone interview was performed to detect the survival of patients up to February 2017.Measurement data with skewed distribution were described as M (Qn),and comparisons among groups were done by the Kruskal-Wallis test.Comparison of count data were done by the Fisher exact probability.Results (1) Morphologic type of contact surface between gastric cancer and pancreas:there were 4 types according to results of CT examination.Type Ⅰ.pancreas contacted with gastric cancer and there was no change in the morphology and radian of contact surface.Type Ⅱ:pancreas contacted with gastric cancer and radian of contact surface became flattened or shallow depression.Type Ⅲ:contact surface showed a inserted sign or obvious depression.Type Ⅳ:pancreas didn't contact with gastric cancer and there was increased density in fat space between pancreas and gastric cancer,with a smudge sign or strip-and sheet-like opacity.Of 31 patients,type Ⅰ,Ⅱ,Ⅲ and Ⅳ were detected in 5,10,4 and 12 patients,respectively.(2) Comparison of CT findings among the 3 groups:nodular protrusion,spiculation and strip shape,clounding patch opacity of serosa panern of gastric cancer were detected in 1,6,4 patients in the PN group and 5,4,2 patients in the PⅠ group and 0,2,7 patients in the PI-NR group,respectively,with a statistically significant difference (X2=10.054,P＜0.05).Two,8 and 8 patients in the PN,PI and PI-NR groups had obvious tumor invasion located at a adjacent region between stomach and pancreas,with a statistically significant difference (X2 =11.259,P＜0.05).Contact or invasion site with pancreas located at head,body and tail of pancreas was detected in 6,5,0 patients in the PN group and 1,7,3 patients in the PI group and 5,4,0 patients in the PI-NR group,respectively,with a statistically significant difference (X2=8.390,P＜0.05).Type Ⅰ,Ⅱ,Ⅲ and Ⅳ of contact surface between gastric cancer and pancreas were detected in 5,6,0,0 patients in the PN group and 0,4,4,3 patients in the PI group and 0,0,0,9 patients in the PI-NR group,respectively,with a statistically significant difference (X2=29.291,P＜0.05).Number of patients with clear and ambiguous contact surface was 10,1 patients in the PN group and 0,11 patients in the PI group and 0,9 patients in the PI-NR group,respectively,with a statistically significant difference (X2 =26.227,P＜ 0.05).CT values of contact surface or peripancreas invaded were-46 HU (-57 HU,-20 HU) in the PN group and-34 HU (-41 HU,-25 HU) in the PI group and-10 HU (-15 HU,-10 HU) in the PI-NR group,respectively,with a statistically significant difference (Z=15.306,P＜0.05).CT values of normal peripancreas were-87 HU (-96 HU,-76 HU) in the PN group and-88HU (-70 HU,-1 HU) in the PI group and-83 HU (-98 HU,-74 HU) in the PI-NR group,respectively,with statistically significant differences in CT values between contact surface or peripancreas invaded and normal peripancreas among the 3 groups (Z=12.581,13.780,7.793,P＜0.05).(3) Treatment or followup situations:of 31 patients,22 underwent radical gastrectomy and 9 underwent simplex exploration or short surgery.All the 31 patients were followed up for 6.0-71.0 months,with a median time of 13.5 months.Postoperative 1-and 2-year survival rates were 82.6％ and 77.1％.Conclusions There are significant differences in pancreatic invasion and resectability between CT features of contact surface of gastric cancer and pancreas and tumor classification.CT features include that pancreas contacts with gastric cancer in the PN group,radian of contact surface becomes flattened and with a inserted sign in the PI group,and there are increased density in fat space between pancreas and gastric cancer and a smudge sign or strip-and sheet-like opacity in the PI-NR group.

To construct a bait expression vector containing the duck circovirus Cap gene for use in the yeast two-hybrid system, the whole cap codon-optimized gene was inserted into pGBKT7 vector and confirmed by PCR, restriction enzyme digestion, and sequence analysis. After transformation into a Y2HGold yeast strain, the expression of Cap protein was analyzed by Western blotting. Toxicity and self-activation of the bait protein were detected using different dropout minimal base. PCR reaction, restriction enzyme digestion, and sequencing analyses indicated that the duck circovirus Cap gene was correctly inserted into pG- BKT7. Western blotting showed that the whole Cap protein was expressed. The recombinant bait protein had no toxicity and self-activation. Therefore, the bait vector with the Cap gene was constructed successfully, providing a foundation for future screening for interacting proteins in the yeast two-hybrid system.
Animals ; Capsid Proteins ; genetics ; metabolism ; Circovirus ; classification ; genetics ; isolation & purification ; Cloning, Molecular ; Ducks ; Genetic Vectors ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; metabolism ; Saccharomyces cerevisiae ; genetics ; metabolism ; Two-Hybrid System Techniques

Objective The aim of this study is to investigate the expression of Survivin in laryngeal car-cinoma and to elucidate the relationship between cell proliferation and the expression of Survivin .Methods The expression and DNA quantification of Survivin in 63 cases of laryngeal carcinoma and 15 cases of normal laryngeal tissues were detected by immunohistochemical staining ( SP) and flow cytometry .Results Forty-five sections of laryngeal carcinoma were positive for survivin as determined by immunohistochemical staining ,but Survivin was undetected in normal laryngeal tissues .The intensity of Survivin expression was significantly increased with tumor differentiation and lymph node metastasis (P<0.05),and was negatively correlated with age ,gender and clinical stage of patients(P>0.05).High DNA index(DI)and proliferation index(PI)were detected in laryngeal carcino-ma(P<0.05),and PI was positively correlated with expression of Survivin (P<0.05).Conclusion Survivin overexpression triggers laryngeal carcinoma cell hyperproliferation ,especially in development of laryngeal carcino-ma.These data identify Survivin as an important target in laryngeal carcinoma and provide a translational pathway for developing new therapies against this target .