PURPOSE: MicroRNAs (miRs) were recently recognized to be important for immune cell differentiation and immune regulation. However, whether miRs were involved in allergen-specific immunotherapy (SIT) remains largely unknown. This study sought to examine changes in miR-146a and T regulatory cells in children with persistent allergic rhinitis (AR) after 3 months of subcutaneous immunotherapy (SCIT) and sublingual immunotherapy (SLIT). METHODS: Twenty-four HDM-sensitized children with persistent AR were enrolled and treated with SCIT (n=13) or SLIT (n=11) for 3 months. Relative miR-146a and Foxp3 mRNA expression, the TRAF6 protein level, and the ratio of post-treatment to baseline IL-10+CD4+ T cells between the SCIT and SLIT groups were examined in the peripheral blood mononuclear cells (PBMCs) of AR patients using quantitative reverse transcription polymerase chain reaction (qRT-PCR), flow cytometry, and Western blot analysis, respectively. Serum levels of IL-5 and IL-10 were determined using ELISA. RESULTS: After 3 months of SIT, both the TNSS and INSS scores were significantly decreased compared to the baseline value (P<0.01). The relative expression of miR-146a and Foxp3 mRNA was significantly increased after both SCIT and SLIT (P<0.01). The ratio of post-treatment to baseline IL-10+CD4+ T cells and the serum IL-10 level were significantly increased in both the SCIT and SLIT groups (P<0.01), whereas the TRAF6 protein level and serum IL-5 level were significantly decreased (P<0.01). No significant differences in these biomarkers were observed between the SCIT and SLIT groups. CONCLUSIONS: Our findings suggest that miR-146a and its related biomarkers may be comparably modulated after both SCIT and SLIT, highlighting miR-146a as a potential therapeutic target for the improved management of AR.
Biomarkers ; Blotting, Western ; Cell Differentiation ; Child* ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Humans ; Immunotherapy* ; Interleukin-10 ; Interleukin-5 ; MicroRNAs ; Polymerase Chain Reaction ; Reverse Transcription ; Rhinitis* ; RNA, Messenger ; Sublingual Immunotherapy ; T-Lymphocytes ; TNF Receptor-Associated Factor 6

Objective To understnad the defense mechanisms of patients with borderline personality disorder (BPD) and to explore its relationship with childhood abuses.Methods One hundred and forty-two outpatients with BPD (group A),63 outpatients with other personality disorders (group B) and 20 outpatients without any Axis Ⅱ diagnoses (group C) were enrolled and completed Chinese versions of the Childhood Experience of Care and Abuse Questionnaire (CECA.Q) and the Defensive Style of Questionnaire (DSQ).Multivariate analysis of variance and Pearson correlation analysis were used for data analysis.Results Compared to group B and C,the score of defense mechanisms reported by group A were significantly increased in all eight premature defense mechanisms,such as projection(F =26.5,P ＜ 0.05),passive aggression (F =18.1,P ＜ 0.05),acting out (F =28.3,P ＜ 0.05),complaint (F =16.7,P ＜0.05),fantasy(F =16.3,P ＜ 0.05),splitting (F =28.8,P ＜ 0.05),withdrawal (F =8.3,P ＜ 0.05),somatization(F=7.6,P ＜ 0.05),and part of the middle defense mechanisms,such as omnipotence-devaluation(F =9.2,P ＜ 0.05),isolation(F =4.7,P ＜ 0.05) and identification (F =10.0,P ＜ 0.05).At the same time,the BPD patients got lower scores in the mature defense mechanisms,such as sublimation (F =4.1,P ＜ 0.05) and repression(F =3.9,P ＜ 0.05).Those who experienced physical abuse use more complaint (F =6.9,P ＜ 0.05),denial mechanisms (F =5.2,P ＜ 0.05) and less anticipation one (F =4.6,P ＜ 0.05).Those who experienced sexual abuse use more somatization (F =4.4,P ＜ 0.05) and isolation mechanism (F =3.6,P ＜ 0.05).Conclusions BPD patients would like to use more immature and middle defense mechanisms and less mature ones.Emotional,physical and sexual abuse from parents and other adult care takers have a correlation with immature defense mechanisms,which indicates the importance of care about parental rearing patterns and mental health of children.

In this study,the current status for breast diseases in a region with high-incidence of cervical cancer were epidemiologically investigated.From March to August,2009,17618 women,from Wufeng area of Hubei province,China,were recruited to screen breast diseases by using breast infrared diagnostic apparatus.Other diagnostic methods,such as B-mode ultrasound,X-ray mammography,needle biopsy and pathological examination were,if necessary,used to further confirm the diagnosis.The screening showed that 5990 of 17618 cases (34.00％) had breast diseases,5843 (33.16％) had mammary gland hyperplasia,48 (0.27％) had breast fibroadenoma,ll (0.06％) had breast carcinoma,and 88 (0.50％) had other breast diseases.The peak morbidity of breast cancer was found in the women aged 50-0 ages.The morbidity of breast cancer was significantly increased in women elder than or equal to 50 years old (n=8,0.157％) in comparison with that in the subjects younger than 50 years old (n=3,0.024％) (u=2.327,P＜0.05).It was shown that the occurrence of breast diseases was concentrated in women aged 20-40 years,while the total morbidity reached its peak at the age of 30 years and then decreased sharply after age of 40.Compared with the patients elder than or equal to 40 years old (n=3289,27.46％),the morbidity rate of breast diseases was significantly increased in women less than 40 years old (2648 cases,47.18％; P＜0.001).However,there was no significant difference in the morbidity of breast diseases between the age group of 20-29 years and that of 30-39 years (P=0.453),and both of them were high.There was no significant association between the morbidity of breast diseases and cervical cancer.Since the morbidity of breast diseases was higher among young women,more attention should be paid to the screening of breast diseases among young women for early diagnosis.

By using a yeast two-hybrid system, a yeast two-hybrid bait vector was constructed and identified for screening of the HPV18 E6-interacting proteins, and its effects on the growth of yeast cells and the activation of reporter genes were investigated. Total mRNA extracted from Hela cells was reversely transcribed into cDNA. Fragment of HPV18 E6 cDNA was amplified using RT-PCR and directly ligated to the pGBKT7 vector. The recombinant plasmid was confirmed by restriction endonuclease analysis and DNA sequencing. The recombinant pGBKT7-HPV18 E6 plasmid and empty pGBKT7 vector were transformed into the yeast cell AH109, respectively. After they were cultured respectively in YPDA liquid medium and nutrition-deficient culture medium, their toxicity and transcriptional activation were tested by both the phenotype assay and the color assay. The bait plasmid HPV18 E6 was successfully obtained. After being cultured in YPDA liquid medium for 16h, the A (600 nm) values of two yeast fluids were 0.98+/-0.03 and 0.99+/-0.02, respectively. The recombinant pGBKT7-HPV18 E6 plasmid and empty pGBKT7 vector could grow to white colonies on SD/-Trp/X-alpha-gal plates, while no colony could survive on SD/-His/-Trp/X-alpha-gal, SD/-Ade/-Trp/X-alpha-gal plates, indicating that the bait plasmid pGBKT7-HPV18 E6 was constructed successfully and expressed correctly, and could not activate the transcription of reporter gene alone. The yeast two-hybrid GAL4 system 3 can be utilized to find HPV18 E6 interacting proteins.

By using a yeast two-hybrid system,a yeast two-hybrid bait vector was constructed and identified for screening of the HPV18 E6-interacting proteins,and its effects on the growth of yeast cells and the activation of reporter genes were investigated.Total mRNA extracted from Hela cells was reversely transcribed into cDNA.Fragment of HPV18 E6 cDNA was amplified using RT-PCR and directly ligated to the pGBKT7 vector.The recombinant plasmid was confirmed by restriction endonuclease analysis and DNA sequencing.The recombinant pGBKT7-HPV18 E6 plasmid and empty pGBKT7 vector were transformed into the yeast cell AH109,respectively.After they were cultured respectively in YPDA liquid medium and nutrition-deficient culture medium,their toxicity and transcriptional activation were tested by both the phenotype assay and the color assay.The bait plasmid HPV18 E6 was successfully obtained.After being cultured in YPDA liquid medium for 16h,the A600 nm values of two yeast fluids were 0.98±0.03 and 0.99±0.02,respectively.The recombinant pGBKT7-HPV18 E6 plasmid and empty pGBKT7 vector could grow to white colonies on SD/-Trp/X-α-gal plates,while no colony could survive on SD/-His/-Trp/X-α-gal,SD/-Ade/-Trp/X-α-gal plates,indicating that the bait plasmid pGBKT7-HPV18 E6 was constructed successfully and expressed correctly,and could not activate the transcription of reporter gene alone.The yeast two-hybrid GAL4 system 3 can be utilized to find HPV18 E6 interacting proteins.

Objective: To investigate the birth defect condition in Haidian district of Beijing city, 61 272 live-born infants who were delivered in Haidian Maternal and Child Health Hospital from 2003 to the March of 2009 are analyzed. Methods: Data was collected from the hospital' s medical records and from the birth defect surveillance. Results: Among the newborns studied, 1 076 were found having birth defect (17.56‰). The most common birth defects are congenital heart defect, followed by dysmorphosis of external ear, polydactyly, hypospadia, cleft lip and palate. In addition, three birth defects that are not included in the birth defect surveillance list were enorchia, renal agenesis and giant hemangioma. The birth defect rates of preterm and small for gestational age infants are significantly higher than it of the term infants. The birth defect patterns for these two types of abnormal infants are distinct. Conclusion: We have determined the pattern of birth defects in Beijing, which may help in policy-making regarding the prevention and intervention of birth defects.

Objective To prepare the human papillomavirus (HPV) 16 peptide vaccine and explore the effect in vitro and in vivo. Methods (1) Prediction of the major histocompatibility complex (MHC) class I restricted T cell epitopes by bioinformatics target at transporter associated with antigen processing (TAP) and named by E7Pa, E7Pb, E7Pc separately. (2)In vivo, the C57BL/6 mice were divided into five groups with same amounts randomly after loading with TC-1 cells (HPV 16 positive tumor cells from C57BL/6 mouse), named as E7Pa + CpG,E7Pb + CpG,E7Pc + CpG (as experiment groups, and added 50 μg/ml E7Pa, E7Pb, E7Pc, respectively), CpG(as positive control group and added Con A with 12 mg/L final concentration) and blank control group (without any treatment). The T cell proliferation was detected by methyl thiazolyl tetrazolium (MTT) assay at different time points;the lactate dehydrogenase (LDH) delivery method was used to test the cytolytie T lymphocyte (CTL) activity of mouse splenic lymphocyte in different ratio of effector cells and target cells (E:T);the related cytokines in tumor tissue and mouse peripheral blood were evaluated by real-time PCR and enzyme-linked immunosorbent assay (ELISA), respectively. The tumor volumes were measured to contrast the therapeutic effect in different groups. Results (1) Three peptide named E7Pa, E7Pb, E7Pc were successfully preparated which had high affinity and specificity. (2) After vaccination of 24, 48, 72,96 hours, MTT results shown that the proliferation rate in E7Pa + CpG group were(131±32)%, (302±15)%, (552±28)%, (731±24)% individually, which were much higher than those in blank control [(72± 15) %, (120 ± 57) %, (176 ±41)%, (288±29)% ;P<0.01], and the other groups i. e. E7Pb + CpG,E7Pc +CpG and CpG groups all proliferated much higher than those in blank control group with statistic signification (P<0. 05), but there was no significant difference between groups(P>0.05);the LDH delivery assay showed that when the ratio of E:T was 100:1, the activity of CTL in the E7Pa + CpG group was most powerful than the other groups with statistic signification (P<0. 01). Meanwhile, the ratio of E:T was concentration-dependent. Compared E7Pb + CpG, E7Pc + CpG or CpG groups with blank control group, there were significantly difference(P<0. 05) ,while there was no significant difference between groups(P >0. 05). The mRNA levels of interferon γ (IFN-γ), interleukin-2 (IL-2) in tumor tissue and peripheral blood in E7Pa + CpG group were significantly higher than those in blank control group (P<0. 01), which was the similar results when compared E7Pb + CpG, E7Pc + CpG or CpG groups with control group (P < 0. 05), and without significant difference between groups(P > 0. 05). The tumor volumes were suppressed obviously in all the experiment groups, especially at the 60th days, the volumes in ETPa + CpG group were much smaller than that in blank control group with statistic signification (P < 0. 01),which was the similar results that E7Pb + CpG, E7Pc + CpG or CpG groups had difference than blank control group with statistic signification (P < 0. 05), and without significant difference between groups(P >0. 05). Conclusion The HPV16 E7 peptide target at TAP combination with CpG as a vaccine could treat effectively the HPV16 E7 positive tumor in experiment.

In order to investigate the role of the PTEN expression in carcinogenesis and development of endometrial carcinoma and clarify whether and how PTEN and PI3K/Akt pathway relate to endometrial carcinoma, the expression of PTEN and phospho-Akt was detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) methods and Western-blot from 24 cases of endometrial carcinoma, 10 cases of endometrial atypical hyperplasia, 10 cases of endometrial hyperplasia, and 10 cases of normal endometrium. SP immunohistochemical methods were used to measure levels of PTEN protein expression in following 5 study groups: 31 cases of endometrium in proliferative phase, 30 cases of endometrium in secretory phase, 71 cases of endometrial hyperplasia, 25 cases of atypical hyperplasia and 73 cases of endometrial carcinoma. Immunostaining score of PTEN was 3.39+/-0.15 in proliferative phase, 1.90+/-0.21 in secretory phase, 3.34+/-0.29 in endometrial hyperplasia, 0.62+/-0.11 in atypical hyperplasia, and 0.74+/-0.19 in endometrial carcinoma, respectively. PTEN mRNA relative value in normal endometrium, endometrial hyperplasia, endometrial atypical hyperplasia, and endometrial carcinoma was 2.45+/-0.51, 2.32+/-0.32, 0.46+/-0.11, and 0.35+/-0.13 respectively. The expression levels of PTEN mRNA and protein in patients with endometrial carcinoma and atypical hyperplasia were significantly lower than in those of proliferative phase and with endometrial hyperplasia. The level of PTEN expression in patients with endometrial carcinoma was significantly related to tissue type (P<0.005), differentiation (P<0.05) and clinical stage (P<0.05), but not to depth of myometrium invasion (P>0.05). Western blot analysis revealed that Phospho-Akt level in PTEN negative cases was significantly higher, and there was a negative correlation between PTEN and phospho-Akt (r=-0.8973, P<0.0001). It was suggested that loss of PTEN expression was an early event in endometrial tumorigenesis. The phosphorylation of Akt induced by the loss of PTEN took part in the tumorigenesis and development of endometrial carcinoma.

In order to investigate the role of the PTEN expression in carcinogenesis and develop-ment of endometrial carcinoma and clarify whether and how PTEN and PI3K/Akt pathway relate to endometrial carcinoma,the expression of PTEN and phospho-Akt was detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) methods and Western-blot from 24 cases of endomctrial carcinoma,10 cases of endometrial atypical hyperplasia,10 cases of endometrial hy-perplasia,and 10 cases of normal endometrium.SP immunohistochemical methods were used to measure levels of PTEN protein expression in following 5 study groups:31 cases of endometrium in proliferative phase,30 cases of endometrium in secretory phase,71 cases of endometrial hyperplasia,25 cases of atypical hyperplasia and 73 cases of endometrial carcinoma.Immunostaining score of PTEN was 3.39±0.15 in proliferative phase,1.90±0.21 in secretory phase,3.34～0.29 in endometrial hyperplasia,0.624±0.11 in atypical hyperplasia,and 0.74±0.19 in endometrial carcinoma,respectively.PTEN mRNA relative value in normal endometrium,endometrial hyperplasia,endometrial atypical hyperplasia,and endometrial carcinoma was 2.45±0.51,2.32±0.32,0.46±0.11,and 0.35±0.13 respec-tively.The expression levels of PTEN mRNA and protein in patients with endometrial carcinoma and atypical hyperplasia were significantly lower than in those of proliferative phase and with endo-metrial hyperplasia.The level of PTEN expression in patients with endometrial carcinoma was sig-nificantly related to tissue type (P<0.005),differentiation (P<0.05) and clinical stage (P<0.05),but not to depth of myometrium invasion (P>0.05).Western blot analysis revealed that Phospho-Akt level in PTEN negative cases was significantly higher,and there was a negative correlation between PTEN and phospho-Akt (r=- 0.8973,P<0.0001).It was suggested that loss of PTEN expression was an early event in endometrial tumorigenesis.The phosphorylation of Akt induced by the loss of PTEN took part in the tumorigenesis and development of endometrial carcinoma.

To screen for novel binding proteins interacting with high-risk HPV 18 E6 oncogene, the strain AH109 was transformed with pGBKT7-HPV18 E6 plasmid, and subsequent transference was utilized to screen for interacting proteins with HPV 18 E6 in human Hela cDNA library. HPVl8 E6 mRNA was expressed in yeast and there was no self-activation and toxicity in AH109. Seven proteins that interacted with HPV18 E6, including transmembrane protein 87B, phosphonoformate im- muno-associated protein 5, vimentin, KM-HN-1 protein, dedicator of cytokinesis 7, vaccinia related kinase 2 and a hypothetical protein, were identified. It was suggested that yeast two-hybrid system is an efficient for screening interacting proteins. The high-risk HPV 18 E6 oncogene may interact with the proteins, which may be associated with signal transduction and transeriptional control, epithelial cell invasion and migration, as well as humoral and cellular immune etc. This investigation provides functional clues for further exploration of potential oncogenesis targets for cancer biotherapy.