Objective To explore the phenotypic conversion of regulatory T cells (Tregs) in the lungs of mice with bronchopulmonary dysplasia (BPD)-affected mice. Methods A total of 20 newborn C57BL/6 mice were divided into air group and hyperoxia group, with 10 mice in each group. The BPD model was established by exposing the newborn mice to hyperoxia. Lung tissues from five mice in each group were collected on postnatal days 7 and 14, respectively. Histopathological changes of the lung tissues was detected by HE staining. The expression level of surfactant protein C (SP-C) in the lung tissues was examined by Western blot analysis. Flow cytometry was performed to assess the proportion of FOXP3⁺ Tregs and RORγt⁺FOXP3⁺ Tregs in CD4⁺ lymphocytes. The concentrations of interleukin-17A (IL-17A) and IL-6 in lung homogenate were measured by using ELISA. Spearman correlation analysis was used to analyze the correlation between FOXP3⁺Treg and the expression of SP-C and the correlation between RORγt⁺FOXP3⁺ Tregs and the content of IL-17A and IL-6. Results The hyperoxia group exhibited significantly decreased levels of SP-C and radical alveolar counts in comparison to the control group. The proportion of FOXP3⁺Tregs was reduced and that of RORγt⁺FOXP3⁺Tregs was increased. IL-17A and IL-6 concentrations were significantly increased. SP-C was positively correlated with the expression level of RORγt⁺FOXP3⁺ Tregs. RORγt⁺FOXP3⁺ Tregs and IL-17A and IL-6 concentrations were also positively correlated. Conclusion The number of FOXP3⁺ Tregs in lung tissue of BPD mice is decreased and converted to RORγt⁺ FOXP3⁺ Tregs, which may be involved in hyperoxy-induced lung injury.
Animals ; Mice ; Mice, Inbred C57BL ; Bronchopulmonary Dysplasia ; T-Lymphocytes, Regulatory ; Interleukin-17 ; Nuclear Receptor Subfamily 1, Group F, Member 3 ; Hyperoxia ; Interleukin-6 ; Forkhead Transcription Factors ; Lung

Background::The incidence of well-differentiated gastric neuroendocrine tumors (G-NET) is increasing annually, and while they have a good prognosis and low mortality rate, their high recurrence rate makes treatment options controversial. This study aims to determine the relationship between individualized treatment plans and the recurrence of G-NET.Methods::We performed a multicenter, retrospective study of 94 patients with highly differentiated G-NET and treated at Peking Union Medical College Hospital, Yantai Yuhuangding Hospital, and Beijing Zhong-Neng-Jian Hospital from November 2015 to September 2023. Risk factors for recurrence of G-NETs were investigated using chi-squared test and multifactorial logistic regression analysis.Results::After a median follow-up of 49 months, the overall recurrence rate among the 94 G-NET patients was 14% (13/94). The recurrence rates of endoscopic mucosal resection (EMR), endoscopic submucosal dissection (ESD), somatostatin analog (SSA) therapy, and surgery were 43% (6/14), 10% (5/49), 5% (1/22), and 11% (1/9), respectively. Post-treatment recurrence rates were significantly different ( P = 0.014) among four treatments (EMR, ESD, SSA, and surgery), and further subgroup comparisons revealed lower recurrence rates in the ESD and SSA groups than in the EMR group. From the second month onward, SSA therapy considerably reduced the gastrin levels from 1081.0 (571.5, 2472.8) pg/mL to 461.5 (255.3, 795.0) pg/mL ( Z = -3.521, P <0.001). Both chi-squared test and multifactorial logistic regression analysis suggested that among the clinicopathological parameters studied, only the pre-treatment gastrin level ( P = 0.018 and 0.005) and the type of treatment ( P = 0.014 and 0.017) were significantly associated with G-NET recurrence. Conclusions::Individualized treatment strategies may reduce the risk of relapse after G-NET treatment. Long-term SSA therapy may be a secure and efficacious treatment option for type 1 G-NET with more than six lesions, and it substantially decreases the incidence of post-treatment recurrence.

Objective:To develop and validate a nomogram model for predicting microvascular invasion (MVI) in hepatocellular carcinoma (HCC) based on preoperative enhanced computed tomography imaging features and clinical data.Methods:The clinical data of 210 patients with HCC undergoing surgery in the Second Affiliated Hospital of Anhui Medical University from May 2018 to May 2022 were retrospectively analyzed, including 172 males and 38 females, aged (59±10) years old. Patients were randomly divided into the training group ( n=147) and validation group ( n=63) by systematic sampling at a ratio of 7∶3. Preoperative enhanced computed tomography imaging features and clinical data of the patients were collected. Logistic regression was conducted to analyze the risk factors for HCC with MVI, and a nomogram model containing the risk factors was established and validated. The diagnostic efficacy of predicting MVI status in patients with HCC was assessed by receiver operating characteristic (ROC) curve, calibration curves, decision curve analysis (DCA), and clinical impact curve (CIC) of the subjects in the training and validation groups. Results:The results of multifactorial analysis showed that alpha fetoprotein ≥400 μg/ml, intra-tumor necrosis, tumor length diameter ≥3 cm, unclear tumor border, and subfoci around the tumor were independent risk factors predicting MVI in HCC. A nomogram model was established based on the above factors, in which the area under the curve (AUC) of ROC were 0.866 (95% CI: 0.807-0.924) and 0.834 (95% CI: 0.729-0.939) in the training and validation groups, respectively. The DCA results showed that the predictive model thresholds when the net return is >0 ranging from 7% to 93% and 12% to 87% in the training and validation groups, respectively. The CIC results showed that the group of patients with predictive MVI by the nomogram model are highly matched with the group of patients with confirmed MVI. Conclusion:The nomogram model based on the imaging features and clinical data could predict the MVI in HCC patients prior to surgery.

Objective: To investigate the expression and clinical significance of PD-1 molecule in tumor cells (T-ALL cells) derived from the patient with T-cell acute lymphoblastic leukemia (T-ALL). Methods: T-ALL cells from one patient and PBMCs from four healthy volunteers provided by the Department of Hematology in Jiangsu Provincial Hospital of Traditional Chinese Medicine in December 2015, and human 293T/PD-1 cells provided by Persongen Bio Therapeutics (Suzhou) Co., Ltd. were used for this study. The mouse T-ALL xenograft model was constructed by injecting T-ALL cells into tail vein of B-NDG mice, and flow cytometry was used to verify whether the cells obtained from the spleen of transplanted mice were mainly consisted of T-ALL cells. Flow cytometry was used to study the protein expression of PD-1 in T-ALL cells, and RT-PCR was applied to further verify the mRNA expression of PD-1 in T-ALL cells. The PD-1 gene in T-ALL cells was sequenced by SNP genotyping to detect whether the DNA sequence of PD-1 gene changed. PD-1 inhibitor was used in vitro to study their effects on proliferation, apoptosis, and the mRNA expression levels of related factors in T-ALL cells. Results: The mouse T-ALL xenograft model was successfully constructed and verified by flow cytometry as T-ALL. PD-1 was highly expressed at both mRNA and protein levels in T-ALL cells (all P<0.01). A C-to-T mutation was detected in the fifth exon of the PD-1 gene. PD-1 inhibitor had no significant effect on proliferation and apoptosis of T-ALL cells in vitro; PD-1 inhibitor up-regulated the mRNA expression of tumor-suppressor protein IGFBP3 and decreased the mRNA expression of oncoprotein SULT1A3 (all P<0.01). Conclusion: PD-1 is highly expressed in T-ALL cells, and PD-1 could be used as a target for clinical diagnosis and treatment for T-ALL.

Objective To explore the status of infection, biotype and resistant background of epi-demic strains of Haemophilus influenza ( Hi ) in neonates, and the clinical features of neonatal pneumonia caused by Hi. Methods The multicenter prospective epidemiological cross-sectional design was used; four hospitals in west Sichuan China were chosen as research field,sputum bacterial culture was done and biologi-cal typing,PCR identification and drug sensitivity test of Hi epidemic strains were carried out among 0 to 28 days hospitalized neonates with infectious pneumonia in four hospitals located in west Sichuan China. The ca-ses with discharge diagnosis of neonatal infectious pneumonia with Hi positive separation were assumed as case group,and the same number of cases with Hi negative separation were assumed as control group accord-ing to 1∶1 extraction at the same time. Results Totally 757 cases with admitting diagnosis of neonatal infec-tious pneumonia in four hospitals were investigated in west Sichuan from November 2014 to October 2015, and the rate of sputum culture was 95. 51%(723/757). The total pathogenic bacteria positive rate of sputum culture was 15. 63%(113/723),and Hi positive rate was 1. 94%(14/723),Hi accounting for 12. 39%(14/113) of the pathogenic bacteria in respiratory system. All the Hi strains(100%) were non-typeable Hae-mophilus influenzae( NTHi) indentified by PCR. The main biotypes of 14 Hi strains were typeⅠwith 57. 1%(8/14),type Ⅲ with 14. 3%(2/14) and type Ⅳ with 28. 6%(4/14). The total of 35. 7%(5/14) bacterial strains of β-lactamase distributed in four hospitals,7. 1%(1/14) bacterial strains of β-lactamase-nonproduc-ing-ampicillin-resistant,and 35. 7%(5/14) bacterial strains of β-lactamase-positive-ampicillin-resistant were found in four hospitals. The rate of resistance and mediation to cefuroxime were 14. 2%(2/14) respectively, the resistance rate to cefaclor was 35. 7%( 5/14 ) , and 21. 4%( 3/14 ) to ofloxacin. None of the 14 strains was resistant to amoxicillin clavulanic acid and cefotaxime. The 1∶1 matching analysis had been done for 10 cases with discharge diagnosis of neonatal pneumonia caused by Hi. There were no statistical differences in general conditions,main symptoms, lung signs, X-ray appearance, classification of leukocyte and C-reactive protein levels between case group and control group(P>0. 05). Conclusion All the Hi isolated from spu-tum were NTHi among 0 to 28 days inpatients with neonatal pneumonia and the main biotype were typeⅠ, type Ⅲand typeⅣin west Sichuan China. There were no significant differences in the clinical manifestations of neonatal pneumonia with NTHi infection and other infectious pneumonia.

Objective To evaluation the precision performance of XE-2100 Automatic Blood Cell Analyzer using EXCEL Mi-crosoft with reference to CLSI EP15-A3 .Methods According to CLSI EP15-A3 ,high level and low level (BIO-RAD company) of quality control materials were measured using XE-2100 Automatic Blood Cell Analyzer ,and analyzed by one-factor analysis of vari-ance of EXCEL 2003 and equation editor .The within-run coefficient of variation and within-laboratory coefficient of variation were calculated and compared with national industry standards and manufacturer statement .Results For high and low level of quality control materials ,the within-run coefficient of variation and within-laboratory coefficient of variation of white blood cell ,red blood cell ,hemoglobin and platelet were less than national industry standards and manufacturer statement .Conclusion The precision ver-ification scheme of CLSI EP15-A3 with EXCEL Microsoft has a relatively good maneuverability and statistical performance ,which could be used in verification of the precision performance of quantitative testing items in clinical laboratories .

Objective To perform the methodological evaluation on Beckman Immage 800 automatic special protein analyzer for determining serum freeκ,λlight chains (sFLC) .Methods The Beckman Immage800 automatic special protein analyzer was used to quantitatively detect sFLC values for investigating its precision and accuracy ,analyzing its detecting range ,interference and residual contamination rate ,meanwhile verifying its reference intervals .Results The low and high values of within‐run precision coefficients of variation(CV) forκsFLC were 7 .84% and 2 .95% respectively ;which of between‐run precision CV were 7 .38% and 5 .57% re‐spectively .The within‐run precision CV for λFLC were 4 .59% and 3 .94% respectively ,which of between‐run precision CV were 3 .97% and 2 .01% respectively ;bias for detecting theκFLC andλFLC fixed quality serum and the target values was less than 5% , when the analytical detection ranges of κFLC andλFLC were 10 .8-128 .0 mg/L and 10 .1-121 .0 mg/L ,a value was 0 .95-1 .05 , r2 ≥0 .98;the carry‐over rates of κFLC andλFLC were 0 .411% and 0 .216% respectively .The interference test results showed that when free hemoglobin≤342 .0 μmol/L ,conjugated bilirubin ≤342 .0 μmol/L ,hemoglobin≤5 g/L and chyle ≤2 400 turbidness , which had no influence on sFLC results ;among 40 healthy subjects undergoing the physical examination ,1 case of κFLC detection results exceeded the reference interval recommended by the manufacturer ,while theλFLC detection values in these 40 cases were all within the reference interval recommended by the manufacturer .Conclusion The main analytical performance of the Beckman Im‐mage800 automatic special protein analyzer for detecting sFLC meets the requirements of quality objectives and can provides the sci‐entific and precise evidence of diagnosis and treatment for clinic .

Objective To discover and certify the minimum required months of IQC ( Internal Quality Control ) data which were used to quantify the imprecision To identify the impact of test items , different concentrantions and years on the defective rate .Methods IQC data involving 20 analysis items ( including Albumin, Creatine kinase, Total bilirubin, Alanine aminotransferase, Aspartate aminotransferase, Lactate dehydrogenase,γ-Glutamyl transferase, Alkaline phosphatase, Calcium, Chlorine, Creatinine, Glucose, Potassium, Sodium, Phosphorus, Triglyceride, Total cholesterol, Total protein, Uric acid, Urea) and six measurement systems (including Roche, Architect, Hitachi, Dade/Behring, Beckman, Olympus) were collected from hospitals in Shanghaibetween 2009 and 2011.A total of 3 534 groups, referred to one year laboratory′s IQC data of one concentration range , were analysed to find the minimum required months in each group when the cumulative months were compared with the population by using T test.The correlation coefficient of hospital′grades, measurement levels and test items were evaluated by u test, and the percentage of groups of P>0.05 were collected.The cumulative IQC data′coefficient variation ( CV) of six months and eleven months were compared with the total CV, respectively .Imprecision higher than the professional specification was regarded as unqualified .Difference of unqualified rate among test items, concentrations and years were expolored .Results Rates of hospital′grades, measurement levels , items are 94.2%,100%, 100%respectively , 88%of groups′imprecision became stable in ten months .25%groups reach the criteria that the relative bias is ≤5% when calculated the cumulative IQC data′CV of six months, while 88%groups do when calculated the cumulative IQC data′CV of ten months.The unqualified rate is 20%and the most unqualified item is Ca , the unqualified groups of low level are larger .With increase of year , the unqualified rate showed a downtrend .Conclusions Ten months′IQC data is more reliable to quantify imprecision .Unqualified rates are different according to test items , measure levels and years , the establishment of professional specification should consider them.

Objective To investigate the action and mechanism of NF-κB pathway in up-regulating B cell-activating factor receptor (BAFF-R) expression in multiple myeloma cells induced by IFN-γ.Methods Activated NF-κB were detected with Western blot, while the expression of BAFF-R were measured with RT-PCR and ELISA, and investigated the effect of BAY11-7082 on transcription of BAFF-R mRNA and translation of protein in multiple myeloma cells stimulated by IFN-γ. Results IFN-γ can induce the degradation of IκB-α in time-dependent and dosage-dependent manner, and up-regulated BAFF-R expression in multiple myeloma cells. BAY11-7082, an NF-κB inhibitor, inhibited not only the transcription of BAFF-R mRNA but also the protein of regulated by IFN-γin dosage-dependent manner. Conclusion NFκB may play an important role in high expression of BAFF-R in multiple myeloma cells induced by IFN-γ.

Objective To construct and screen siRNA targeting a proliferation-inducing ligand (APRIL) gene in a mouse colorectal cancer celline, CT-26. To investigate the effects to the cell growth and migrant capacity of CT-26 after knockdown APRIL gene, lay the foundation for molecular targeted therapy to colorectal cancer. Methods Four pairs of APRIL siRNA were designed and chemically synthesized. And disorder sequences were synthesized as a negative control. These sequences were transfected with LipofectAMINE 2000 into CT-26 cells, which high-expressed APRIL gene. The transfection efficency rate of 6-FAM labelled control siRNA was detected by fluorescence microscope. The inhibition effectiveness of APRIL mRNA and protein was analyzed by FQ-RT-PCR and Western blot, respectively. Cell proliferation activity was analyzed by cell counting kit-8, cell migration capacity was detected by the repair of cell damage, and MMP-2 together with TIMP-1, two important regulatory genes in cell metastasis, were measured by RT-PCR.Results The different kinds of APRIL siRNA effectively suppressed the level of APRIL mRNA and the protein expression in CT-26 (P ＜ 0.05 ). Cell proliferation and metastasis ability were repressed after APRIL siRNA transfection( P ＜ 0.05 ), compared with random siRNA control and nontransfected control. The mRNA levels of MMP-2 and TIMP-1 genes wre significantly altered among APRIL siRNA groups and two control groups ( P ＜ 0.05). Conclusion We have constructed and screened a kind of siRNA (APsi737) targeting APRIL gene in a mouse colorectal cancer cell line, CT-26. APRIL siRNA can effectively inhibit the cell growth and migration capacity, maybe be regulated by MMP-2 and TIMP-1.