OBJECTIVETo determine the regulatory effects of the circadian clock gene Per1 on cell cycle-related genes and its influence on the proliferation, apoptosis, cycle, and tumorigenicity in vivo of human oral squamous cell carcinoma SCC15 cells.

METHODSThree groups of the short hairpin RNA (shRNA) of lentivirus recombinant plasmids were designed against the RNA of Per1 and then transfected to the SCC15 cells. The optimum interference group was screened through Western blot and quantitative real-time PCR (qRT-PCR) and assigned as the experimental group. The transfected lentivirus plasmid without an interference effect on any gene was set as the control group (Control-shRNA). Untreated SCC15 cells were set as the blank group. The mRNA expressions of cell cycle-related genes, namely, Per1, p53, Cyclin D1, Cyclin E, Cyclin A2, Cyclin B1, CDK1, CDK2, CDK4, CDK6, p16, p21, Wee1, cdc25, E2F, and Rbl1 in each group were detected through qRT-PCR. The cell proliferation, apoptosis, and cell cycle distribution in each group were evaluated through flow cytometry. The cells of the experimental group and the blank group were subcutaneously inoculated in nude mice to observe tumorigenesis.

RESULTSThree groups of Per1-shRNA lentivirus plasmids were constructed successfully. Among the groups, the Per1-shRNA- I group exhibited the highest interference effect, as indicated by qRT-PCR and Western blot analysis. As such, this group was set as the experimental group. The mRNA expression levels of CyclinD1, CyclinE, CyclinB1, CDK1, and Wee1 gene in the Per1-shRNA-I group were significantly higher than those in the Control-shRNA group and the SCC15 group (P < 0.05). By contrast, the mRNA expression levels of p53, Cyclin A2, p16, p21, and cdc25 in the Per1-shRNA-I group were significantly lower than those in the Control-shRNA group and the SCC15 group (P < 0.05). The mRNA expression levels of each gene between the Control-sLRNA group and the SCC15 group did not significantly differ (P > 0.05). The mRNA expression levels of CDK2, CDK4, CDK6, E2F, and Rb1 did not significantly differed in the three groups (P > 0.05). The proliferation index of the Perl-shRNA-I group was significantly higher than those of the Control-shRNA group and the SCC15 group (P < 0.05). The apoptosis index of the Per1-shRNA-I group was significantly lower than those of the Control-shRNA group and the SCC15 group (P < 0.05). The number of S-phase cells in the Per1-shRNA-I group was significantly lower than those of S-phase cells in the Control-shRNA group and the SCC15 group (P < 0.05). The number of G2/M-phase cells in the Per1-shRNA-I group was significantly higher than those of G2/M-phase cells in the Control-shRNA group and the SCC15 group (P < 0.05). Conversely, the proliferation index, apoptotic index, and cell cycle distribution of the cells in the Control-shRNA group did not significantly differ from those of the SCC15 group (P > 0.05). The tumorigenic ability in vivo was significantly enhanced in the Per1-shRNA-I group (P < 0.05).

CONCLUSIONPer1 is an important tumor suppressor gene. Per1 can regulate a large number of downstream cell cycle-related genes. The alteration of its expression can affect cell cycle progression, proliferation, apoptosis imbalance, and tumorigenic ability in vivo. Further studies on Per1 may elucidate cancer development and provide novel effective molecular targets for cancer treatment.

Animals ; Apoptosis ; Carcinoma, Squamous Cell ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Circadian Clocks ; genetics ; Cyclin D1 ; Humans ; Mice ; Mice, Nude ; Mouth Neoplasms ; Period Circadian Proteins ; genetics ; Plasmids ; RNA, Small Interfering ; Real-Time Polymerase Chain Reaction ; Transfection

Objective To investigate the differences between animal temperature controller (ATC) and artificial climate chamber (ACC) used for the establishment of classical heat stroke (CHS) rat model.Methods Twenty-four male specific pathogen-free Wistar rats were randomly (random number) and equally assigned to three groups,namely room temperature control (C-C) group,heat stroke under conscious state (HS-C) group,and heat stroke under anesthesia (HS-A) group.Rats of HS-C or HS-A group were placed into ACC or ATC,then exposed to 35 ℃ heat stress.The systolic blood pressure (SBP) and core body temperature (Tc) were monitored.The time required for onset of HS was recorded.The white blood cell count (WBC) in peripheral blood and serum levels of C-reactive protein (CRP),tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) were measured.The histopathological changes of major organs were also confirmed by hematoxylin-eosin (HE) staining.Results The onset time in HS-A group was significantly shorter thanthatin HS-C group [(40.0 ± 4.3) minvs.(110.1 ± 5.3) min,P＜0.01].The SBP and Tc at this moment were lower in HS-A group [(159.1 ± 5.91) mmHg vs.(174.54 ± 5.77) mmHg,P＜0.01;(43.5 ± 0.4)℃ vs.(44.4 ± 0.2)℃,P＜0.01].TheWBC,CRP,TNF-α and IL-1 β levels of these two HS groups were dramatically elevated compared with C-C group (P ＜0.01).The inflammatory cytokines levels in HS-A group were significantly lower than those in HS-C group (P ＜ 0.01),but there was no difference in WBC between them (P ＞ 0.05).However,there was no obvious difference in histopathological change in major organ observed between HS-A and HS-C groups.Conclusions In comparison of these two methods,ATC is similar to ACC in respect of the establishment of CHS rat model.ATC is quicker in onset of HS,and more simplified and economical than ACC and could substitute ACC.

BACKGROUND:The cultivation of mammary gland stem cel s is of great significance for the development of mammary gland and breast cancer.
OBJECTIVE:To seek an easy method to isolate and culture mammary gland stem cel in vitro, and verify the safety of cel s.
METHODS:Mammary epithelial cel s were isolated from normal tissues surrounding breast cancer, and CD49f-and EPCAM-positive cel s were sorted using flow cytometry fol owed by surface marker analysis and cel colony formation ability analysis. Afterwards, real-time fluorescent quantitative PCR was used to detect C-erbB-2 and Maspin mRNA expression in mammary gland stem cel s, breast cancer tissues and normal tissues surrounding breast cancer.
RESULTS AND CONCLUSION:Human mammary gland stem cel s were successful y cultured and highly expressed CD49f and EPCAM, with the presence of mixed colony, pleural epithelial cel colony, and myoepithelial cel colony. c-erbB-2 was lowly expressed while Maspin highly expressed in mammary gland stem cel s. Our experimental findings indicate that the mammary gland stem cel s derived from normal tissue surrounding breast cancer have biological safety.

Objective To judge the effect of plasma exchange (PE) to the patients with severe hepatopath of nonage according to evaluating the change of the liver function of reserve with 13C-methacetin breath test. Methods There are two groups: the case group and the control group. Each group has 30 patients. The patients in the case group were treated by PE. All the patients received 13C-methacetin breath test at before or one week after treatment. MVmax40, CUM40 and CUM120 were present. At the same time, clinical symptoms, glutamate-pyruvate transaminase (ALT), total bilirubin (TBiL) and prethrombin active (PTA) were observed. Results MVmax40, CUM40, CUM120 and PTA were higher, ALT and TBiL were lower in the case group after treatment (t=4.679, 4.752, 5.048, 5.413, 6.208, 7.413, P=0.000,P<0.01). After a week, MVmax40, CUM40, CUM120 and PTA were higher, ALT and TBiL were lower in the case group than that in the control group (t=2.260, 2.247, 2.476, 4.017, 3.250, 3.658, P<0.05). The total effective rates in the case group and the control group were 83.3 % (25/30) and 43.3 % (13/30),which are significant different(χ2 10.335,P<0.01). Conclusion PE can im-prove the liver reservation functionin the severe hepatopath of nonage.

AIM: To study the influence of glycine(GLY) on lipopolysaccharide-binding protein(LBP) mRNA expression induced by LPS. METHODS: The level of LBP mRNA expression in liver tissues of rats was examined by RT-PCR,and the effects of glycine on LBP mRNA expression in liver tissues of rats induced by LPS were investigated. RESULTS: The level of LBP mRNA expression in hepatic tissue of rats in the LPS group was significantly higher than that in the control group( P

Objective To investigate the effects of estrogen on mismatch repiar gene expression in colonic mucosa in vivo. Methods A total of 42 healthy individuals underwent colonoscopy were enrolled in the study. Half an hour before colonoscopy examination, blood sample was taken for determining the serum estradiol (E2) level. N ormal colonic mucosal tissues determined by naked eye under colonoscopy examination were taken in the right hemi colon to detect HMLH1 and hMSH2 gene expression by semi-quantitative RT-PCR and immunohistochemistry staining. Then the correlation of serum E2 levels with hMLH1 and hMSH2 expression in colonic mucosa was analyzed. Results A bimodal curve was presented for the correlation between serum E2 level in healthy individuals and hMLH1 expression in colonic mucosa. A strong positive correlation of E2 level with hMLH1 expression in normal colonic mucosa was observed when serum E2 level was more than 45 pg/ml (For mRNA, P=0. 003, r=0. 701; for immunohistochemistry positivity index, P=0. 000, r=0. 874).However there was no correlation between E2 level and hMSH2 expression. Conclusion High serum E2 level might increase the hMLH1 gene expression in colonic mucosa in vivo.

Objective To assess left ventricular (LV) global systolic function in patients with essential hypertension with normal geometric LV by 3-dimensional ultrasound speckle tracking imaging(3D-STI). Methods Fifty patients with essential hypertension were enrolled in this study, 29 normal subjects matched with age and sex were selected as control groups. LV global longitudinal peak systolic strain (GLS), radial peak systolic strain (GRS), circumferential peak systolic strain (GCS), LV global 3-dimentional radial peak systolic strain (3DGRS) were measured in all subjects by 3D-STI from the apical full-volume image and compared between groups. LV ejection fraction (LVEF) was acquired from 3D-STI.Results Compared with controls, LV GLS, GRS, GCS and 3DGRS were significantly reduced in patients with hypertension ( P ＜ 0.05 for all). A Pearson correlate revealed that LV GCS, GLS and GRS corresponded with LVEF( r1 =0.930, P1 ＜0.001; r2 = 0.705, P2 ＜0.001; r3 =0.474, P3 =0.001,respectively) in patients with hypertension, and LV GCS, GLS correlated with LVEF( r1 = 0. 838, P1 ＜0. 001; r2 = 0. 697, P2 ＜ 0. 001, respectively) in normal subjects. Conclusions LV global 3D strain decreases in patients with hypertension in the early period,3D-STI could evaluate the early change of heart function in patients with hypertension. LV circumferential movement plays a major role in the LV 3D movement and impacts on LVEF.

Objective To investigate the value of left ventricular global two-dimensional strain and strain rate index measured by two-dimensional speckle tracking imaging (2D-STI) in assessing myocardial injury in various degree of rats following acute myocardial infarction. Methods Fifty-five Wistar rats were randomly divided into myocardial infarction(MI) group ( n =45) and sham-operation(SO) group ( n = 10).To establish rats acute myocardial infarction model with different infarct extent, MI group were randomly divided into MI15 group,MI30 group and ML60 group( n = 15,respectively) which underwent occlusion of left anterior descending coronary artery for 15 minutes, 30 minutes and 60 minutes respectively. Echocardiography was performed at baseline and 24 hours after reperfusion. High frame rate twodimensional images were recorded from the left ventricular short-axis views at the papillary muscle level.Left ventricular global circumferential strain(GSc) and strain rate(GSRc) were measured using EchoPAC work station. Left ventricular internal diameter at diastole (LVIDd) and systole ( LVIDs), fractional shortening(FS) and ejection fraction(EF) were measured by anatomical M-model echocardiography. Area of necrosis(AN) of each segment was measured after triphenyl tetrazolium chloride(TTC) staining. Results ① Compared with baseline and SO group, LVIDd and LVIDs of MI15, MI30 and MI60 group significantly increased respectively,whereas FS and EF significantly decreased( P <0. 05). Compared with MI15 group and MI30 group, LVIDd and LVIDs of MI60 group significantly increased, whereas FS and EF significantly decreased(P <0. 05). ② Compared with baseline and SO group,GSc and GSRc of MI15 group, MI30 group and MI60 group significantly decreased. GSc and GSRc of MI group decreased with ischemia duration ( P <0.05). ③ GSc and GSRc significantly correlated with AN respectively ( P <0. 01) while the correlation coefficient was 0. 90 and 0. 88 respectively, and GSc and GSRc were significantly predictors of AN( P <0.01) while the Beta was 0.558 and 0.491 respectively.④AN increased with ischemia duration( P <0.05). Conclusions Left ventricular global circumferential strain and strain rate index measured by 2D-STI,which decreased significantly as the area of necrosis increased, can accurately assess myocardial injury after myocardial infarction in various degree.

Objective To analyze MRI features of different cervical flexion positions in Hirayama disease (HD) and discuss the effects on these features by different cervical flexion angles.Methods The cervical MR images of neutral and different flexion positions (20°, 25°, 30°, 35° ,40°) of 20 patients, who were clinically diagnosed as HD,were studied.At flexion positions, the appearance of anterior shifting of the posterior wall of the cervical dural canal and widening of epidural space was recorded.The maximum sagittal diameters (d) of widened cervical epidural space and the cervical canal sagittal diameters (D) on the same level were measured to calculate d/D value for quantitative evaluation of the two signs.Comparisons of appearance of the signs among different flexion positions were made using F/sher's exact test.Repeated measures analysis of variance (rmANOVA) was used to compare mean d/D values among groups with different positions, and paired comparisons were also performed.Results The appearance of anterior shifting of the posterior wall of the cervical dural canal were different between 20° group (70%, 14/20) and other 4 larger angles groups (100%) (χ2 =5.76, P=0.020).The d/D values were 0.51±0.06,0.54±0.08,0.57±0.09,0.61±0.09,0.59±0.07 respectively at abovementioned 5 flexion positions, which were different among groups( F = 3.450 ,P = 0.013 ).The value was greater at 35° than that at 20° and 25°( P <0.05 ), and it was also greater at 40° than that at 20° ( P < 0.05 ).Conclusion Cervical flexion angle has an effect on anterior shifting of the posterior wall of the cervical dural canal and widening of epidural space.

With the development of the medicine industry and the expansion of global commu-nication and cooperation , more and more Chinese higher vocational college students who major in pharmaceutical management and administration are aware of the demands and are trying to master English for Specific Purpose (ESP) for their better future development. The necessity, course orienta-tion, curriculum design and teaching method for opening ESP course in pharmaceutical management and administration major of higher vocational college were studied in this paper.