OBJECTIVETo investigate the effect of SHIP gene mutation on the cell cycle and its related gene expression in K562 cells.

METHODSThe recombined green fluorescent protein (GFP) containing FIV-SHIP gene was transfected into K562 cells. The transfection efficiency and cell cycle of K562/SHIP were assessed by flow cytometry (FCM). The proliferation of K562 cells was detected by MTT assay, the mRNA levels of SHIP by real-time fluorescent relative-quantification reverse transcriptional PCR (FQ-PCR), and the protein levels of SHIP, Cyclin D1, p21(WAF1/CIPI) and p27(KIP1) by Western blot.

RESULTSWild type SHIP inhibited K562 cell proliferation and caused a G(0)/G(1) arrest \[(34.2 +/- 7.8)% vs (0.7 +/- 8.3)% (P < 0.01)\]; while the point mutation of SHIP gene did not show such effect. Western blot results showed that the Akt phosphorylation and cyclin D1 expression was significantly decreased (P < 0.01), and the expression of p27(KIP1) and p21(WAF1/CIPI) increased. Site-directed mutation of SHIP gene SH2 domain (TTC-->CTC, Phe-->Leu) did not influence the Akt phosphorylation and cyclins (P > 0.05).

CONCLUSION(1) wtSHIP gene can down-regulate Akt phosphorylation and result in inhibition of cyclin D1 expression, up-regulating p27(KIP1) and p21(WAF1/CIPI) expression, finally leading to the reduction of K562 cell proliferation, and inducing G(0)/G(1) phase arrest. (2) SHIP gene suppresses the proliferation of K562, being dependent on its intact structure and function.

Cell Cycle Proteins ; metabolism ; Humans ; Inositol Polyphosphate 5-Phosphatases ; K562 Cells ; Mutation ; Phosphoric Monoester Hydrolases ; genetics ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Transfection

The src homology 2 (SH2)-domain containing inositol-5-phosphatase (SHIP) is another recently identified lipid phosphatase after phosphatase and tensin homology deleted on chromosome ten gene (PTEN). It plays an important role in negatively regulating the proliferation of hematopoietic cells. The relationship between SHIP and the inhibition of tumor proliferation is rarely reported. The purpose of this study is to evaluate the apoptosis induced by SHIP gene in K562 cell line and to explore the involved signaling pathway. The K562 cells were transfected with human SHIP gene by using the lentiviral vector containing SHIP, and the transfection was verified by fluorescent quantitative PCR (FQ-PCR) and Western blot. Then the effects of SHIP protein expression on cell growth and apoptosis were measured. The levels of p-Akt, bcl-2 family, caspase and the activity of NFkappaB were assayed by Western blot and ELISA, respectively. The results are as follows: (1) Human leukemia cell line K562 was SHIP-negative; (2) Transfection with SHIP gene led to the re-expression of SHIP mRNA and protein in K562, as shown by FQ-PCR and Western blot; (3) The expression of SHIP protein inhibited cell growth and significantly increased apoptosis in K562 cells; (4) Compared to that in control group, the expression level of p-Akt-308 and p-Akt-473 in SHIP-expressing cell group decreased significantly (P<0.01); SHIP activated caspase-9, caspase-3, up-regulated protein levels of bad, p27, down-regulated expression of bcl-xL, while it had no effect on the expression of bcl-2 and bax. Furthermore, the inhibition of NF-kappaB was achieved along with the inactivation of Akt. These data suggest that SHIP gene has potential abilities to inhibit K562 leukemic cell proliferation and induce its apoptosis via inactivating PI3K/Akt pathway. The loss of SHIP might be the explanation of aberrant high-level p-Akt in human leukemia. It may be at least one of the mechanisms by which the loss of SHIP expression contributes to leukemia progression.
Apoptosis ; Apoptosis Regulatory Proteins ; metabolism ; Cell Proliferation ; Down-Regulation ; Genetic Vectors ; Humans ; Inositol Polyphosphate 5-Phosphatases ; K562 Cells ; NF-kappa B ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Phosphoric Monoester Hydrolases ; genetics ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; Transfection

OBJECTIVETo explore the effect of mutation in PxxP domain of SHIP on migration and invasion of leukemia cells and its mechanism.

METHODSThe lentiviral vector mediated wild type SHIP (wtSHIP) and mutant SHIP (muSHIP) plasmids were transfected into K562 cells through gene transfection techniques. Expression of SHIP at mRNA and protein level was detected by real-time PCR and Western blot, respectively. Transwell assay was used to analyze the difference between the migration and invasion ability of the K562/wtSHIP and the K562/muSHIP cells after transfection. Primary migration associated factor FAK, MMP and NF-κB were assayed by Western blot.

RESULTSAfter transfection, the SHIP expression in transfected K562 cells were significantly increased. Compared with the migration ability of K562/wtSHIP\[(15.8 ± 1.4)%\], that of K562/muSHIP cells \[(54.3 ± 2.4)% \] increased greatly and almost at the same level of that of K562/pFIV\[(50.3 ± 3.8)%\] (P < 0.01). The invasion assay also showed that K562/wtSHIP\[(32 ± 6)/HP\] has a lower invasion ability than that of the K562/muSHIP group \[(83 ± 16)/HP\] and K562/pFIV group \[(78 ± 13)/HP\] (P < 0.01). Western blot analysis showed that the expression of p-FAK and NF-κB was up-regulated in K562/muSHIP group compared to that of the K562/wtSHIP group.

CONCLUSIONSThe results confirmed that mutation in PxxP domain of SHIP gene played an important role in negative regulating function of SHIP gene. The mutation affects the cell migration and invasion ability through increase in MMP-9 expression, FAK phosphorylation and NF-κB activation. It suggested that the mutation of PxxP domain in SHIP gene might be pathogenic, and be one of the reasons for SHIP abnormality in leukemia.

Cell Movement ; Genetic Vectors ; Humans ; Inositol Polyphosphate 5-Phosphatases ; K562 Cells ; Leukemia ; pathology ; Mutation ; Phosphoric Monoester Hydrolases ; genetics ; Plasmids

OBJECTIVETo analyze the direct and indirect outcomes of influencing factors on smoking behaviors among adolescents.

METHODSSelf-administered questionnaires were used to collect smoking-related information from 2021 respondents. Measurement models were built by confirmatory factor analysis and parameters were estimated by ERLS method. The final structural equation model was determined by comprehensive evaluation and necessary modification.

RESULTSThree latent variables were extracted from 10 manifest variables of environment, while only one latent variable was identified from 9 manifest variables of attitudes. The goodness of fit for the structural equation model was satisfactory that all indices had met corresponding requirements. The final model could explain 38.8% of the variance of smoking behaviors. Four factors (smoking environment, smoking restriction from parents and teachers, determination of cigarette refusal and attitudes toward smoking) were directly affecting the smoking behaviors, while another three factors (grade, health knowledge and school environment) had indirect impacts. According to the percentages of their contribution, the risk factors were ranked as follows: smoking environment (45.76%), attitudes toward smoking (19.88%) and grade at school (0.44%). Similarly, the top protective factor were: determination of cigarette refusal (16.61%), followed by smoking restriction from parents and teachers (10.51%), health knowledge (3.89%) and school environment (2.92%).

CONCLUSIONSHeath knowledge had minor effect on smoking in adolescents but could indirectly affect their smoking behaviors through changing their belief. Grade at school had a doubled influence on smoking, but mainly served as a risk factor. Tobacco control measures for adolescents should not only be limited to health education but environment factors as well.

Adolescent ; Adolescent Behavior ; China ; Humans ; Male ; Smoking ; psychology ; Students

OBJECTIVES: This cohort study investigated the correlation between Parkinson’s disease (PD) risk and chronic obstructive pulmonary disease (COPD) risk under particulate matter with an aerodynamic diameter ≤2.5 μm (PM2.5) exposure. METHODS: Data from the National Health Research Institutes of Taiwan were used in this study. The Environmental Protection Administration of Taiwan established an air quality monitoring network for monitoring Taiwan’s general air quality. COPD was indicated by at least 3 outpatient records and 1 hospitalization for COPD. After the implementation of age, sex, and endpoint matching at a 1:4 ratio, 137 patients and 548 patients were included in the case group and control group, respectively. Based on the 2005 World Health Organization (WHO) standards, monthly air particle concentration data were classified into the following 4 groups in analyses of exposure–response relationships: normal level, and 1.0, 1.5, and 2.0 times the WHO level ([concentration ≥2]×25 μg/m3×number of exposure months). RESULTS: A multivariate logistic regression revealed that the 1.0 and 1.5 WHO level groups did not significantly differ from the normal level group, but the 2.0 WHO level did (odds ratio, 4.091; 95% confidence interval, 1.180 to 14.188; p=0.038). CONCLUSIONS Elevated PM2.5 concentrations were significantly correlated with an increased risk of PD among patients with COPD. Furthermore, exposure to high PM2.5 levels can further increase the risk of PD.

OBJECTIVETo investigate the methylation of CpG island in the SHP-1 gene promoter and its significance in lymphoma. To evaluate the effects of As2O3 on demethylation of SHP-1 in human lymphoma cell line T2 and on proliferation of T2 cells.

METHODST2 cells were treated with AsO3. Methylation specific PCR was used to detected the status of SHP-1 methylation in newly diagnosed lymphoma tissues and the T2 cells. The mRNA and protein expression of SHP-1 were determined by FQ-PCR and Western blot. The expression of phospha-c-kit was examined by Westren blot. MTT and flow cytometry were used to determine the growth and apoptosis in T2 cells.

RESULTST2 cells contained completely methylated SHP-1. Furthermore, there was constitutive c-kit phosphorylation. The expression of SHP-1 was recoverd when the cells exposed to AsO3, and concomitant with increasing SHP-1, a parallel down-regulation of phosphorylated c-kit occurred, so that by day 3 phosphorylated c-kit was barely detectable. As2O3 inhibited the cell growth, and the effects were dose- and time-dependent. As2O3 also increased apoptosis rate of T2 cells in a dose- and time-dependent manner, too, and on the 1, 2, 3 d treatment with AsO3 (2.5 micromol/L), the apoptosis rates were 6.12%, 26.53%, 50.90%, respectively. The frequency of methylation in SHP-1 gene promoter in lymphoma tissues was 87.5% (28/32). In the control group, however, 12 specimens of benign lymph node proliferation showed no methylation in CpG island of SHP-1 gene promoter.

CONCLUSIONHypermethylation of SHP-1 gene promoter in lymphoma indicates the inactivation of SHP-1 gene and its possible role in the tumorigenesis of lymphoma. As2O3 can effectively cause demethylation and inhibit the growth of tumor by reactivating the SHP-1 gene transcription. SHP-1 methylation leading to epigenetic activation of c-kit may have a tentative role in the pathogenesis of lymphoma. Therefore, As2O3 is potentially useful in the treatment of lymphoma as a demethylating agent.

Antineoplastic Agents ; administration & dosage ; pharmacology ; Apoptosis ; drug effects ; Arsenicals ; administration & dosage ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; CpG Islands ; DNA Methylation ; drug effects ; Dose-Response Relationship, Drug ; Gene Expression Regulation, Neoplastic ; Humans ; Lymphoma ; metabolism ; pathology ; Lymphoma, Non-Hodgkin ; genetics ; metabolism ; pathology ; Oxides ; administration & dosage ; pharmacology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; genetics ; metabolism ; Proto-Oncogene Proteins c-kit ; metabolism ; RNA, Messenger ; metabolism ; Transcriptional Activation ; drug effects ; Up-Regulation

OBJECTIVETo explore the relationship between psychological stress, fatigue and different smoking behaviors among undergraduate students.

METHODS8138 undergraduate students in grade 1 to 3 were sampled with cluster sampling method from a comprehensive university in Guangzhou, and were studied on their psychological stress, peer and self smoking behaviors, as well as socio-demographic factors with a self-administered questionnaire. Physical and mental fatigue was measured by Chalder fatigue scale. A multinomial logistic regression analysis was performed to explore the association between psychological stress, fatigue and different smoking behaviors with 'non-smokers' defined as control group while 'triers' and 'current smokers' as case group, respectively.

RESULTSThere were students trying to smoke or becoming current smokers on university campus. After adjusting the effect of relevant socio-demographic factors and peer smoking behavior, physical fatigue (OR = 1.044, 95% CI: 1.013-1.075) and 'worry about current academic and future work' (OR = 1.020, 95% CI: 1.001-1.040) were the risk factors of trying to smoke among male students, while 'study stress (OR = 1.063,95% CI: 1.012-1.117) was the risk factor for female students when start smoking. 'Personnel relationship and environment stress' (OR = 1.152, 95% CI: 1.012-1.312) were the risk factors for female students' current smoking behavior.

CONCLUSIONPsychological stress and fatigue were moderately associated with smoking behavior among undergraduate students and the impact was different on different gender and smoking behaviors that suggesting that some specific psychological measures should be taken so as to control different smoking behaviors among undergraduate students.

Fatigue ; psychology ; Female ; Humans ; Logistic Models ; Male ; Sex Factors ; Smoking ; psychology ; Stress, Psychological ; psychology ; Students ; psychology ; Universities ; Young Adult

OBJECTIVETo explore the influencing factors of smoking behavior transition at different stage of smoking among undergraduate students.

METHODS8138 undergraduate students from grade 1 to 3 were selected using cluster sampling method from an university in Guangzhou with their smoking behaviors and relevant factors studied through a self-administered questionnaire. Beginners, un-regular smokers and regular smokers were defined as "case group", while non-smokers, beginners and un-regular smokers were defined as "control group" accordingly. Three multivariable logistic regressions were separately conducted to explore the influencing factors of behavioral transition at different stage of smoking.

RESULTSOf the 8138 students investigated, the prevalence rates of beginners, un-regular smokers and regular smokers were 19.6%, 2.0% and 1.1%, respectively; while all of the above said three rates increased with grades of the students. Some students might still try to smoke or become regular smoker in the university. In male undergraduate students, determinants for the transition from non-smokers to s mokers were: source of students, father' s educational level, smoking-related knowledge and belief, and smoking habit of the roommates. Risks that influencing the transition from beginners to un-regular smokers were: being non-medical students,smoking-related belief, roommates and classmates who were smokers and transition from un-regular smokers to regular smokers would include father' s educational level and average income of family. In female undergraduate students,influencing factors from non-smokers to beginners were mother's educational level, smoking-related belief, and roommates smokers, while father's educational level, roommates and classmates' smoking habits were significant associated with beginners' change to un-regular smokers.

CONCLUSIONThe influencing factors related to the transition of different stage on smoking would be different in undergraduate students,and some specific measures should be taken based on the different stage of smoking.

Age Distribution ; China ; epidemiology ; Female ; Health Knowledge, Attitudes, Practice ; Health Surveys ; Humans ; Income ; statistics & numerical data ; Male ; Smoking ; epidemiology ; Universities ; statistics & numerical data

OBJECTIVETo evaluate the long-term efficacy of SCMC-ALL-2005 protocol in treatment of low-risk childhood acute lymphoblastic leukemia (ALL).

METHODSFrom May 1, 2005 to April 30, 2009, 387 patients enrolled into SCMC-ALL-2005 protocol. Based on the characteristics of cell morphology, immunology, cytogenetics and molecular biology and treatment response, 158 patients were fit into the low-risk treatment group. All the cases were registered in pediatric oncology network database (POND). The clinical characteristics and outcome were analyzed.

RESULTSUntil December 31, 2012, the 5-year event free survival (EFS) and overall survival (OS) is (77.76±3.37)% and (89.55±2.83)%, respectively. Median follow-up time is 5.33 y (3.75-7.70 y). Five patients (3.16%) died of complication, all of them were severe infections. Twenty-seven patients (17.09%) relapsed, including 13 bone marrow relapse (8.23%), 5 testis relapse (5.32% of boys, 2 of unilateral and 3 bilateral), 6 central nerve system relapse (CNS, 3.80%), 1 relapse in both bone marrow and CNS, 1 relapse in both bone marrow and testis, and 1 right ovary and fallopian tube relapse. Relapse is related to positive minimal residual disease. Two cases (1.27%) occurred second tumors, 4 patients (2.53%) gave up treatment in complete remission without special reasons.

CONCLUSIONThe EFS and life quality of SCMC-ALL-2005 protocol in the treatment of childhood low-risk ALL is satisfactory. The treatment-related mortality rate is lower, and the long-term EFS is higher than that of XH-99 protocol.

Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Child ; Child, Preschool ; Disease-Free Survival ; Female ; Follow-Up Studies ; Humans ; Infant ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; drug therapy ; mortality ; Treatment Outcome

Huocao(a traditional Chinese herbal medicine) moxibustion is a characteristic technology in Yi medicine suitable for cold-dampness diseases. Huocao, as the moxibustion material, is confusedly used in clinical practice and little is known about its quality control. In this study, UPLC method was used to establish the chemical fingerprint of non-volatile components in Huocao, and the contents of eight phenolic acids such as chlorogenic acid were determined. Multivariate statistical analysis was performed to obtain the indicator components of Huocao for quality evaluation, and thus a comprehensive evaluation system for the quality of Huocao was built. The UPLC fingerprints of 49 batches of Huocao were established, and there were 20 common peaks, of which eight phenolic acids including neochlorogenic acid and chlorogenic acid were identified. Except for three batches of Huocao, the similarity of the other 46 batches was higher than 0.89, suggesting that the established fingerprint method could be used for quality control of the medicinal herb. The correlation coefficient between entropy weight score of the eight phenolic acids and comprehensive fingerprint score in Huocao was 0.875(P<0.01), which indicated that the eight phenolic acids could be used as indicator components for the quality evaluation of Huocao. Furthermore, in multivariate statistical analysis on the common peaks of fingerprint and the contents of the eight phenolic acids, chlorogenic acid, isochlorogenic acid A and isochlorogenic acid C were screened to be the indicator components. The results revealed that the proposed method achieved a simple and accurate quality control of Huocao based on UPLC fingerprint and multi-component content determination, which provided useful data for establishing the quality standard of Huocao.
Chlorogenic Acid ; Entropy ; Hydroxybenzoates ; Quality Control