Objective:To investigate the methylation status of N-myc downstream regulated gene-1(NDRG-1) gene in breast cancer and the effects of methylation enzyme inhibitor 5-Aza-2'-deoxycytidine (5-Aza-CdR) on growth and expression of NDRG-1 mRNA in human breast cancer cell line T47D.Methods:Sensitive methylation-specific (MSP)-PCR was used to detect the methylation status in the promoter regions of NDRG-1 gene in 47 samples of breast cancer and tumor adjacent tissues and 15 cases of benign breast disease. The change in expression of the tumor suppressor gene NDRG-1 mRNA in cultured T47D cells was detected by RT-PCR before and after 5-Aza-CdR treatment. Cell proliferation was observed by MTT assay.Results:Hypermethylation frequencies of NDRG-1 gene promoter were 46.8% in breast cancer tissues and 21.3% in tumor adjacent tissues. No hypermethylation of NDRG-1 gene was observed in the tissues of breast benign disease. The growth of T47D cells was suppressed obviously after 5-Aza-CdR treatment compared with the control group. RT-PCR showed that compared with the control group, NDRG-1 mRNA expression was increased at different concentrations in 5-Aza-CdR treatment group. Conclusion:The promoter methylation status of NDRG-1 gene was significantly related with the occurrence of breast carcinoma. 5-Aza-CdR could effectively reverse the methylation of NDRG-1 gene and recover its expression, thereby inhibiting the growth of tumor cells.

Background and purpose:CpG methylation in promoter region is an essential mechanism for the dysfunction of the tumor suppression gene.Folate metabolism provides active methyl for organism methylation.Methionine synthase(MS) plays a vital role in the process of folate metabolism.This study aimed to explore the genetic polymorphism of MS, CHD5 methylation and their association with breast cancer morbidity.Methods:Fortyseven patients with primary breast cancer, 52 healthy subjects and 15 breast hyperplasia patients were enrolled in this experiment.The mRNA expression of CHD5 was determined by reverse transcription PCR(RT-PCR).Methylationspecific PCR(MSP) was used to detect the methylation status of CHD5.Polymorphisms in the MS gene were analyzed through the polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP).Results:The mRNA expression of CHD5 in breast cancer tissues(0.27?0.19) and adjacent non-cancerous tissues(0.33?0.17) were both significantly lower than that in the breast hyperplasia tissues(0.67?0.14)(P

Objective:To explore the effects of problem-solving based telephone and texting interventions for medication adherence of discharged patients with schizophrenia.Method:A total of 178 discharged patients with schizophrenia were randomly assigned to telephone intervention (TI)group (n =63),text-messaging intervention (TMI)group (n =61),and control group (n =54).All patients were routinely treated,and patients in TI and TMI groups were given problem-solving based interventions.Medication adherence and psychotic symptoms were as-sessed by pill counting and the Positive and Negative Symptom Scale (PANSS)at baseline and the first,third, sixth,ninth and twelfth month.Results:Overall,patients with TI had higher medication adherence than those with TMI (P <0.001),and patients with TMI had higher medication adherence than that in control group (P <0.001). PANSS scores in TI and TMI groups were not significantly different (P >0.05),but significantly lower than control group (P <0.01).There was higher rate of re-admission in control group in comparison with TI and TMI groups (9.3% vs.[0% and 1.6%],P <0.05).Conclusion:It suggests that problem-solving based telephone and texting interventions are effective for improving medication adherence in patients with schizophrenia after their discharge from hospital,so as to reducing psychotic symptoms and lowering re-hospitalization risk.In comparison with TMI, TI has a better effect in improving medication adherence.

Aim Toinvestigatetheinducementeffect of isatin on apoptosis of breast cancer cell line MCF-7 , andexploreitsdetailedmechanism.Methods MCF-7 cell lines were exposed to isatin at different concentra-tions(0,50,100,200 μmol·L-1 )for 48 h.Apop-totic features were demonstrated by nuclei staining with Hoechst 33258.Bcl-2,Bax and p53 mRNA were ana-lyzed by RT-PCR.Caspase-9 activation and mitochon-drial depolarization were assayed by flow cytometry. Bcl-2,Bax,p53 and cytochrome c proteins were ana-lyzedbyWesternblot.Results Isatininducesapopto-sis of MCF-7 cells.Furthermore,Bcl-2 expression was decreased and the ratio of Bcl-2 to Bax was significant-ly decreased by isatin.While,p53 expression relative-ly decreased.The mitochondrial transmembrane poten-tial was markedly reduced and the release of cyto-chrome c into the cytosol was increased after treatment with isatin.Simultaneously,caspase-9 was activated. Conclusions Isatinsignificantlyinducedtheapopto-sis of MCF-7 cells in vitro.These results strongly sug-gest that the p53 dependent mitochondrial pathway is involved in apoptosis.

Objective:To investigate the mRNA expression of the WIF-1 gene and the methylation of its promoter in breast can-cer, and to determine the correlation between the epigenetic aberrant WIF-1 DNA methylation and the clinicopathological significance of WIF-1 in breast cancer . Methods:RT-PCR and sensitive methylation-specific-PCR (MSP) were used to detect WIF-1 mRNA ex-pression and the methylation of the WIF-1 promoter in 30 breast cancer samples as well as in tumor-adjacent tissue samples and 9 be-nign breast tissues. Results:The WIF-1 mRNA expression in 30 breast cancer samples significantly decreased compared with those of the other two groups. In addition, WIF-1 methylation was more frequent in breast-tumor tissues compared with those in tumor-free tis-sues. Meanwhile, WIF-1 mRNA expression in breast cancer tissues involved the abnormal methylation of its promoter. Clinicopatholog-ical correlation analysis showed that the abnormal methylation of the WIF-1 gene promoter was not associated with age, TNM stage, histotype, or lymph node metastasis. Conclusion:WIF-1 mRNA expression loss due to abnormal methylation may be a crucial factor in breast cancer development and can thus be used in the prognosis and progression of the disease.

Objective To investigate the effects of simulated microgravity by rotary cell culture system (RCCS) on expression profiles of miRNA in mouse fibroblasts L929 cell line.Methods L929 cells were cultured in vitro and divided into simulated microgravity (SMG) group and normal gravity (NG) group according to the simple random method.Samples of two groups were collected on 7th day of culture and the total RNAs were extracted,labeled,and hybridized in sequence.Feature Extraction Software was used to collect the array images and get raw data,which were analyzed by Genespring Software.Differentially expressed miRNAs were identified and then validated by qRT-PCR.Target genes of differentially expressed miRNAs were predicted by the databases of Targetscan and microRNAorg,and the intersections of databases were identified as potential regulatory target genes.Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) analysis were applied to determine the roles of these target genes.Relevant biological functions and/or signaling pathways of the regulated genes especially related with wound healing process were categorized based on their enrichments.Then integration predictions of the miRNA and mRNA expression profiles had been proposed to refine the functional miRNA-mRNA relationships.The miRNA-mRNA functional network and miRNA-mRNA control network were constructed.Results Four miRNA genes were up-regulated significantly including mmumiR-669j,-122-5p,-30a-3p,-6516-3p,among which mmu-miR-669j was up-regulated at 52.84 folds with the greatest significance (P ＜ 0.05).Seventeen miRNA genes were down-regulated significantly including mmu-miR-21a-3p,-miR-28a-5p,-218-5p,-210-3p,-miR-19a-3p,-miR-31-3p,and-miR-19b-3p,among which mmu-miR-28a-5p was down-regulated at 15.47 folds with the greatest significance (P ＜ 0.05).The qRT-PCR showed a high concordance with the microarray results (P ＜ 0.05).Target gene prediction and functional enrichment analysis suggested that a variety of biological processes and signaling pathways involved in wound repair were significantly enriched (P ＜ 0.05).Function network and regulation network of miRNA-mRNA covered all the differentially expressed miRNAs,which suggested that miR-21 a-3p and predicted target gene Smad3 might play an important role in wound healing under microgravity.Conclusions Simulated microgravity by RCCS can significantly affect the expression of stress-related miRNAs in mouse fibroblasts L929.The miRNA target gene prediction and functional enrichment analysis based on gene chip technology may provide theoretical basis for illustrating the mechanism and management of weightlessness stress injury.

Objective:To analyze food intolerance status in children in Qingdao by detecting the serum levels of food-specific IgG (sIgG).Methods:In this cross-sectional study, a total of 4 249 children aged 0 to 14 years (all were permanent residents of Qingdao City) admitted to Women and Children′s Hospital Affiliated to Qingdao University from May 2017 to December 2020 for suspected food intolerance were selected as the study objects with the whole sampling method. According to the age, the objects were divided into 4 groups: 0-<1 year group (440 cases), 1-<3 years group (1 761 cases), 3-<6 years group (1 193 cases), and ≥6 years group (855 cases). Positive condition of serum sIgG antibodies of 14 kinds of food in the children were detected by enzyme-linked immunosorbent assay. Chi-square test was used to compare the positive rate of the antibodies among different foods, gender and age groups.Results:The total positive rate of food sIgG antibody in 4 249 children was 95.32% (4 050/4 249), the highest positive rate was found in eggs (81.50%) and the lowest positive rate was found in pork (1.15%). The positive rates of sIgG antibody in milk (54.98% vs 49.69%, χ2=11.627), crab (5.59% vs 3.71%, χ2=8.049) and shrimp (4.62% vs 2.75%, χ2=9.784) in boys were significantly higher than those in girls, and the positive rates of sIgG antibody in tomato (49.19% vs 45.54%, χ2=5.510), cod (8.53% vs 5.96%, χ2=10.512) and beef (2.58% vs 1.70%, χ2=3.959) in girls were significantly higher than those in boys (all P<0.05). The total positive rate of sIgG antibody in 14 foods was the lowest in 0-<1 year group (89.09%), and it was the highest in 3-<6 years group (96.98%) ( χ2=63.950, P<0.001). The highest positive rate in 0-<1 year group was found in tomato (56.36%), and it was eggs (85.29%, 88.94%, 85.50%) in all the other 3 groups. The positive rates of corn and beef decreased with age ( χ2=44.098, 20.106, P<0.001), while those of cod and mushroom increased with age ( χ2=32.315, 40.338, P<0.001). The positive rate of wheat (57.13%, χ2=42.273), tomato (57.01%, χ2=209.862), soybean (24.99%, χ2=92.580), crab (6.81%, χ2=33.201), shrimp (6.25%, χ2=47.863) were all the highest in 1-<3 years group among the 4 groups (all P<0.001), and the positive rate of chicken was the highest in 3-<6 years group (7.88%, χ2=29.875; P<0.001). Conclusions:Children in Qingdao have a high level of food intolerance, and the highest positive rate is for eggs. Milk, crab and shrimp should be focused on for boys, while tomatoes, cod and beef shoud be paid more attention to for girls. Children of different ages have different kinds of food intolerance, and their diets should be adjusted reasonably according to the characteristics.

OBJECTIVE: To investigate the regulatory effect of metformin on regulatory T cells (Treg) in acidic environment. METHODS: CD4 CD25 Treg cells were obtained by magnetic bead sorting. Treg and conventional T cells (Tcon) cells were cultured for 24-72 h in pH 7.4 or pH 6.7 medium, and the cell proliferation, apoptosis and Foxp3 expression were detected by flow cytometry. Real-time PCR was used to detect the expression levels of the genes related with glucose metabolism. Thirty-two C57BL/6 male mouse models bearing subcutaneous prostate cancer xenograft derived from RM-1 cells were randomized into 4 equal groups for treatment with PBS, metformin, tumor vaccine, or both metformin and the vaccine. The treatment started on the 4th day following tumor cell injection, and metformin (100 mg/kg) or PBS was administered by intraperitoneal injection on a daily basis; the vaccine was intramuscularly injected every 4 days. The tumor size was continuously monitored, and the mice were euthanized on day 25 after tumor implantation to obtain tumor and blood samples. Flow cytometry was used to detect the changes in CD4, CD8, CD4Foxp3 cell subsets in the tumor tissue and peripheral blood. RESULTS: Treg cells showed significantly enhanced proliferation ( < 0.05) while the proliferation of Tcon cells was suppressed in acidic medium ( < 0.001). Treg cells cultured in acidic medium showed significantly increased expressions of OXPHOS-related genes pgc1a ( < 0.001) and cox5b ( < 0.01), which did not vary significantly in Tcon cells in acidic medium. Treg cells exhibited significantly decreased apoptosis in acidic medium ( < 0.01) with increased Foxp3 cells ( < 0.001) and intracellular alkaline levels ( < 0.01). Metformin obviously reversed the acid tolerance of Treg cells without producing significant effect on Tcon cells. In the animal experiment, both metformin ( < 0.05) and vaccine ( < 0.01) alone reduced the tumor volume, but their combined treatment more potently reduced the tumor volume ( < 0.001). Metformin alone did not obviously affect CD4 cells or CD8 cells but significantly decreased the percentage of CD4Foxp3 ( < 0.05); the vaccine alone significantly increased CD4 cells and CD8 cells ( < 0.001) and also the percentage of CD4Foxp3 cells ( < 0.05). The combined treatment, while reducing the percentage of CD4Foxp3cells to a level lower than that in the vaccine group ( < 0.01), produced the strongest effect to increase CD4 cells and CD8 cells ( < 0.01). CONCLUSIONS Metformin can inhibit the proliferation and function of regulatory T cells in an acidic environment and enhance the effect of tumor vaccine by reducing the proportion of Treg cells to achieve the anti-tumor effect.
Animals ; Cell Proliferation ; Forkhead Transcription Factors ; Male ; Metformin ; Mice ; Mice, Inbred C57BL ; T-Lymphocytes, Regulatory