Objective: To examine the involvement of methylation of p16 gene on the p16 status and tumor progression in primary breast cancer and the role of p16 protein in primary breast cancer.Methods: DNA was extracted from 45 cases of primary breast carcinomas and 15 cases of benign breast lesions.Polymerase chain reaction(PCR),using the extracted DNA and methylation-sensitive restriction enzymes(HpaⅡ?SacⅡ),was performed to detect the methylation of 5′CpG island in the p16 gene.The results were compared with the P16 expression by immunostaining.Results:(1) p16 was overexpressed in breast cancer.With the increase of histological grade of breast carcinomas,p16 expression decreased.In addition,the low expression of p16 in primary breast cancer was associated with tumor size and lymph node involvement.(2) Methylation of the CpG island of the p16 gene was inversely correlated with p16 expression in a subset of the primary breast carcinomas.With the increase of histological grade of breast carcinomas,DNA methylation was enhanced.Conclusion:Methylation of p16 gene may be the cause of loss of p16 expression.

Objective:To explore the effect of radiotherapy on the FHIT protein expression and cell apoptosis of cervical squamous carcinoma and discuss the relationship between FHIT protein expression and cell apoptosis. Methods:Expression of FHIT protein was measured by immunohistochemical method and cell apoptosis was detected by TdT-mediated dUTP terminal nick end labeling (TUNEL) staining in 50 cases of squamous cell cervical carcinoma at ⅡB-ⅢB stages before, during (Dt 10 Gy and Dt 30 Gy), and after radiotherapy. Results:Of the 50 patients, the positive rates of the expression of FHIT protein was 56% at Dt 10 Gy, 68% at Dt 30 Gy, and 84% after radiotherapy, which were significantly increased compared with that before radiotherapy (36%, P<0.05). The positive rates of cell apoptosis was 52% at Dt 10 Gy, 64% at Dt 30 Gy and 78% after radiotherapy, which were significantly elevated compared with that before radiotherapy (28%, P<0.05). In the process of radiotherapy, cell apoptosis was positively related to the expression of FHIT protein (P<0.05). Conclusion:Radiotherapy reinforces the expression of FHIT protein and induces apoptosis cocurrently. FHIT protein has regulatory effects in cell apoptosis induced by radiotherapy.

Objective To detect epidermal growth factor receptor (EGFR) 19, 21 exon gene mutation and gene copy number in lung adenocarcinoma tissue, and to analyze the relationship of EGFR 19, 21 mutation with copies number. MethodsEGFR mutations and gene copy number in the tissue samples embedded by paraffin and fixed by for marlin from 58 cases of lung adenocarcinoma were detected by RT-PCR and FISH. The statistical data were analyzed by chi-square test.ResultsOf 58 cases, the overall single mutation rate of EGFR exon 19, 21 was 43.1% (25/58), and 2 cases contained both types of the mutation.The overall FISH positive rate of EGFR was 51.7 % (30/58), including 8 positive amplification and 22 highly ploidy amplification. The testing results showed that there had no statistically differences in FISH positive rates of EGFR mutation among different differentiation lung adenocarcinoma tissues(P ＞0.05), and the FISH positive rates of EGFR mutation in poorly differentiated cancer were lower than those in moderatedly differentiated and well-differentiated cancer (P ＜0.05). EGFR mutation was closely related to EGFR gene copies (P ＜0.01). ConclusionThere are high EGFR mutation frequencies and FISH positive rates in lung adenocarcinoma tissue; Combined detection of EGFR mutation and gene copy number may provide a better approach in selecting patients who may benefit from anti-EGFR target therapy.

BACKGROUND/AIMS: The current study aims to investigate the protective effects of Bifidobacterium infantis on the abnormal immune response to inflammatory bowel disease (IBD) in dextran sodium sulfate (DSS)-induced colitis. METHODS: Eight-week-old BALB/c mice were separated into five groups at random (control, DSS, DSS+B9 [B. infantis 1×10⁹ CFU], DSS+B8 [B. infantis 1×10⁸ CFU], and DSS+B7 [B. infantis 1×10⁷ CFU]). Colitis was induced by 5% DSS ad libitum for 7 days, at which time we assessed weight, the disease activity index (DAI) score, and the histological damage score. The nuclear transcription factor Foxp3 (a marker of Treg cells), cytokines interleukin-10 (IL-10) and transforming growth factor β1 (TGF-β1), and related proteins (programmed cell death ligand 1 [PD-L1] and programmed cell death 1 [PD-1]) were detected by an immunohistochemical method and Western blot. RESULTS: B. infantis increased weight, decreased DAI scores and histological damage scores, increased the protein expression of Foxp3 (p<0.05) and cytokines IL-10 and TGF-β1 in mouse colon tissue (p<0.05), and increased the expression of PD-L1 in the treatment groups relative to that in the DSS group (p<0.05). The effect of B. infantis on Foxp3 and PD-L1 was dose dependent in the treatment groups (p<0.05). PD-L1 was positively correlated with Foxp3, IL-10, and TGF-β1. CONCLUSIONS: In a mouse model of IBD, B. infantis can alleviate intestinal epithelial injury and maintain intestinal immune tolerance and thus may have potential therapeutic value for the treatment of immune damage in IBD.
Animals ; Bifidobacterium ; Blotting, Western ; Cell Death ; Colitis ; Colon ; Cytokines ; Dextrans ; Immune Tolerance ; Inflammatory Bowel Diseases ; Interleukin-10 ; Methods ; Mice ; Models, Theoretical ; Sodium ; T-Lymphocytes, Regulatory ; Transcription Factors ; Transforming Growth Factors

Background:Identifying missed diagnoses of polyps is significant in reducing the incidence of colorectal cancer.Until now,the role of repeat colonoscopy during colorectal polypectomy in reducing themissed diagnoses of polyps is sparsely studied.Aims:To analyze the value of repeat colonoscopy during colorectal polypectomy in reducing themissed diagnoses of polyps.Methods:One hundred and forty-six patients in the first affiliated hospital of Ningbo University who underwent polypectomy were consecutively recruited for repeat colonoscopy from January 2020 to January 2021.The number,size,and location of missed polyps were recorded and analyzed during colorectal polypectomy.Univariate logistic regression analysis was used to analyze the independent risk factors of missed polyps.Results:There were 86 males and 60 females among the 146 enrolled patients,which aged(55.54±10.51)years.The overall missed polyps'rate was 27.17%.We found that all missed polyps were sessile polyps and<10 mm in size,the difference was significant in the left colon.Univariate logistic regression analysis showed that the endoscopist' experience was an independent risk factor for missed polyps.Conclusions:Endoscopist'experience was an independent risk factor for missed polyps.Repeat colonoscopy could reduce the missed polyps.

Objective:To study the origin traceability of anemarrhenae rhizoma from Bozhou and Hebei based on multi-element fingerprints technology , and establish a discrimination model .Methods:The contents of 48 elements were determined by using induc-tively coupled plasma mass spectrometry ( ICP-MS) for 44 samples of anemarrhenae rhizome from Bozhou and Hebei province .Princi-pal component analysis ( PCA) and orthogonal partial least squares discrimination analysis ( OPLS-DA) were applied in the data analy-sis to screen out the significant elements .And then Fisher linear discrimination analysis was used to determine the origin of anemarrhe-nae rhizoma and the discrimination models were developed .Results:Two discrimination models were developed by the discrimination a-nalysis of the whole model method with nine significant elements identified by PCA and OPLS -DA, and 100％correct classification and 95.5％cross validation were achieved by the models .Conclusion: It is a promising approach to classify the geographical origin of anemarrhenae rhizome based on multi-element fingerprints analysis combined with multivariate statistical analysis .The discrimination models are good enough to be applied in the origin traceability of anemarrhenae rhizome.

AIM:To investigate the role of ClC-3 chloride channel in the promotion of radio sensitization of na-sopharyngeal carcinoma CNE-2Z cells by dihydroartemisinin(DHA).METHODS:MTT was used to detect the inhibito-ry effect of DHA on the viability of CNE-2Z cells and normal nasopharyngeal epithelial NP69-SV40T cells,the radio sensi-tization effect of DHA on CNE-2Z cells was detected by cloning assay,the expression of ClC-3 protein was detected by Western blot,the expression of ClC-3 protein was down-regulated by siRNA technology,and the chlorine current of cells was recorded by whole cell patch-clamp technology.RESULTS:(1)Compared with NP69-SV40T cells,DHA selective-ly inhibited the proliferation of CNE-2Z cells,with IC10 values of(13.020±4.831)μmol/L and(5.244±1.050)μmol/L,respectively(P＜0.01).(2)The results of clonal formation experiments showed that DHA had a radio sensitizing effect on CNE-2Z cells,with a radio sensitization ratio of 1.9.(3)DHA could activate the chlorine channel of CNE-2Z cells and produce an outward chlorine current,but had no effect on the chlorine channel of NP69-SV40T cells.(4)DHA promoted the expression of ClC-3 chloric channel protein in CNE-2Z cells(P＜0.01).(5)Chlorine channel blocker NPPB could in-hibit the radio sensitizing effect of DHA on CNE-2Z cells by 1.84 times,and also inhibited the chlorine current activated by DHA.(6)the down-regulation of CNE-2Z ClC-3 protein could inhibit the radio sensitization effect of DHA on CNE-2Z cells by 4.19 times,and the activation of chlorine current by DHA on CNE-2Z cells was no longer produced.CONCLU-SION:DHA has a radio sensitizing effect on nasopharyngeal carcinoma CNE-2Z cells,which is likely to be related to the activation of ClC-3 chloride channel.

Objective To explore the relationship between waist-hip ratio （WHR） and the risk of acute ischemic stroke （AIS） in elderly population and its clinical significance.Methods Demographic information，disease diagnosis and blood biochemical data of 8375 residents who were followed up in Huamu Community in 2019 were retrospectively analyzed. Patients were divided into AIS group（506 cases）and non AIS group（7869 cases）according to whether residents had newly developed AIS during the follow up period. The data between the two groups were compared to analyze the correlation between WHR and AIS. Multivariate binary Logistic regression was used to analyze the influence of WHR on the risk of AIS.Results In this study，the percentages of residents in the AIS group and non-AIS group were 6.04% （506/8375） and 93.96%（7869/8375） respectively. Mean WHR of residents in the AIS group and non-AIS group was（0.91±0.07）and （0.90±0.07）respectively，and the difference between the two groups was statistically significant（t=2.715，P=0.007）.There were statistical differences in age，BMI，waist circumference，WHR，meat diet，hypertension，diabetes，coronary heart disease，serum creatinine，low density lipoprotein and total cholesterol between 2 groups（P<0.05）.There were no significant differences in gender，systolic blood pressure，diastolic blood pressure，hip circumference，smoking history，drinking history，atrial fibrillation，platelets，hemoglobin，serum glutamic pyruvic transaminase，blood urea nitrogen，fasting blood glucose，high density lipoprotein and triglyceride between the two groups （P>0.05）.The correlation between WHR and AIS risk［OR（95%CI）P］was 5.07（1.31，19.57） 0.018 before，and 4.34 （1.08，17.39）0.038 after adjusting for confounding factors.Conclusions Elevated WHR increases the risk of AIS in elderly population.These findings have certain significance for advocating weight control and prevention of cerebrovascular diseases in elderly population.

Objective:To investigate the changes of various cytokines and coagulation function in B cell acute lymphoblastic leukemia(ALL) patients with different CRS scores during CD19-CAR-T cell immunotherapy.Methods:87 patients with B-ALL hospitalized in the First Affiliated Hospital of Soochow University and 30 normal controls were enrolled into this study from July 2018 to October 2020. The age of the patients was 32(20, 56) years old and 36(41.4%) were female. All these coagulation indicators, prothrombin time (PT), activated partial thromboplastin time (APTT), D-dimer, fibrinogen (Fg) were analyzed by automatic blood coagulation in B-ALL patients before and after treated with CAR-T cell. The ratio of CD19-CAR-T cells and the expression of IL-6, IL-10, IFN-γ, TFN-α, IL-2, IL-4, and IL-17A were analyzed using flow cytometry. The patients′ clinical parameters were detected, and the CRS classification of severity was made according to the standard of consensus.Results:Patients with CRS>3 had prolonged PT and APTT, increased D-dimer, and decreased fibrinogen ( P<0.05). The levels of cytokines of IFN-γ, IL-6, and IL-10 were significantly higher in patients with CRS>3 than that in controls ( P<0.05).The D-dimer level is positively correlated with IL-10. Conclusion:Patients with severe CRS grading have significant coagulation dysfunction in CD19-CAR-T cell immunotherapy. Cytokines IFN-γ, IL-6, and IL-10 may affect coagulation function and CRS grading during CD19-CAR-T cell immunotherapy.

OBJECTIVETo observe the effects of 2-deoxyglucose inhibiting synovial pannus of adjuvant arthritis rats and to explore its potential mechanism of inhibiting angiogenesis by investigating proliferation, migration and matrigel tube formation assay .

METHODSThe effect of 2-DG on synovial pannus was evaluated by histopathology of HE staining; HUVEC proliferation was determined by CCK-8 method; migration of FLS were determined by transwell; matrigel tube formation assay was made for assessing tube number of HUVEC; p-AMPK and Bcl-2 were detected by Western blot assay; AMPK signaling pathway in HUVEC was inhibited by compound C, which is an inhibitor of AMPK activation.

RESULTS2-DG (200 mg/kg) obviously decreased appearance of synovial pannus ( < 0.01); , 2-DG (0.5 mmol/L and/or 5 mmol/L) obviously inhibited proliferation, migration and tube number of HUVEC ( < 0.01 or < 0.001), and its effects on HUVEC were reversed by using AMPK antagonist (Compound C); Western blot showed that 2-DG (5 mmol/L) increased expression of p-AMPK and decreased expression of Bcl-2 ( < 0.05).

CONCLUSIONSActivating AMPK pathway and decreasing expression of Bcl-2 may the potential mechanism by which 2-DG contributes to anti-angiogenesis and effects of inhibiting proliferation, migration and tube number of HUVEC.