Nasopharyngeal carcinoma （NPC）， characterized by insidious onset and non-specific features at the initial stage， is usually diagnosed at middle or late stage with metastasis. The invasion and metastasis of NPC involve complex biological processes， which are affected by many factors. The regulation of gene expression is involved in the invasion and metastasis of NPC， which has become a hot topic. Micro-ribonucleic acids （miRNAs） are short （about 22 nucleotides long） non-coding ribonucleic acids （RNAs） that participate in each step of invasion and metastasis of malignant tumor cells and play an important regulatory role by modulating the transcription and translation of target genes. Abnormal expression of miRNAs has been found in NPC， which regulates the invasion and metastasis of NPC cells. This paper summarized the regulatory mechanisms of different miRNAs as tumor suppressor genes in the migration and invasion of NPC cells， including the modulation of target genes， migration-and invasion-related proteins， and important signaling pathways， which involve biological processes such as epithelial-mesenchymal transition （EMT）， neovascularization and lymphatic vessels， tumor stem cells， and resistance to radiotherapy and chemotherapy. As Chinese medicine shows remarkable efficacy in the prevention and control of NPC， especially in the alleviation of adverse reactions and reduction of metastasis rate after radiotherapy and chemotherapy， we also summed up the effect and mechanism of Chinese medicine and the active components in inhibiting the migration and invasion of NPC cells by miRNAs. Thereby， this review is expected to lay a theoretical basis for further research and development of new drugs against NPC.

Objective:To study the biological behavior of nasopharyngeal carcinoma stem cells and to explore the activation of Ras signaling pathway regulated by CD44.Methods:CNE2-SC and 5-8F-SC were nasopharyngeal carcinoma stem cells and obtained by serum-free suspension culture. Cell counting kit-8 (CCK-8) assay, colony formation assay, Transwell migration assay, cell adhesion array were used to investigate the growth, proliferation, migration and adhesion of nasopharyngeal carcinoma stem cells. Western blot test was used to detect the expressions of Ras signaling pathway related proteins and siRNA-mediated interference was used to determine the activation of Ras signaling pathway regulated by CD44.Results:The growth rates of CNE2-SC and 5-8F-SC cells were significantly lower than those of nasopharyngeal carcinoma cells at 24, 48 and 72 hours after inoculation ( P<0.05). After 14 days of implantation, the colony formation rates of CNE2-SC (44.5±1.9)% and 5-8F-SC (47.4±1.8)% were higher than those of CNE2 (34.9±1.5)% and 5-8F (37.2±1.7)%, respectively( P<0.01). The migration cell number of CNE2-SC was (87.6±7.8), 3.97 times higher than that of CNE2 ( P<0.01). The migration cell number of 5-8F-SC was (67.2±5.7), 3.07 times higher than 5-8F ( P<0.01). The adhesion rates of CNE2-SC and CNE2 cells were (42.1±7.6)% and (8.9±2.0)%, respectively at 3 hours after inoculation and were (82.4±5.0)% and (12.1±2.2)% at 6 hours after inoculation, respectively. The adhesion rate of CNE2-SC cells was higher than that of CNE2 cells (all P<0.01). The adhesion rates of 5-8F-SC and 5-8F cells were (53.6±6.1)% and (7.3±1.5)% at 3 hours after inoculation, and (90.7±3.6)% and (11.0±1.2)% at 6 hours after inoculation, respectively. The adhesion rate of 5-8F-SC cells was higher than that of 5-8F cells ( P<0.01). The expression levels of CD44, Ras and N-cadherin were significantly higher, while phosphatase and tensin homolog deleted on chromosome 10 (PTEN), E-cadherin in nasopharyngeal carcinoma stem cells were lower than those of the nasopharyngeal carcinoma cells. Furthermore, the levels of phosphorylated mitogen extracellular kinase1/2 (p-MEK1/2) and phosphorylated extracellular signal-regulated protein kinase1/2 (p-ERK1/2)were significantly increased in nasopharyngeal carcinoma stem cells ( P<0.01). Correlation analysis showed that the protein expression levels of CD44 was highly positively correlated with RAS in nasopharyngeal carcinoma stem cells( r=0.985, P=0.002; r=0.962, P=0.038). Deletion of CD44 in CNE2-SC decreased the expression levels of HER-2, Ras and p-ERK1/2, p-Akt and phosphorylated protein kinase C-δ(p-PKCδ) ( P<0.01). Conclusions:Despite compare to the nasopharyngeal carcinoma cell, nasopharyngeal carcinoma stem cells grows at a relatively slow rate, the capacities of clone formation, migration, adhesion are promoted. This may be related to the CD44-regulated abnormal activation of Ras signaling pathway.

Objective:To study the biological behavior of nasopharyngeal carcinoma stem cells and to explore the activation of Ras signaling pathway regulated by CD44.Methods:CNE2-SC and 5-8F-SC were nasopharyngeal carcinoma stem cells and obtained by serum-free suspension culture. Cell counting kit-8 (CCK-8) assay, colony formation assay, Transwell migration assay, cell adhesion array were used to investigate the growth, proliferation, migration and adhesion of nasopharyngeal carcinoma stem cells. Western blot test was used to detect the expressions of Ras signaling pathway related proteins and siRNA-mediated interference was used to determine the activation of Ras signaling pathway regulated by CD44.Results:The growth rates of CNE2-SC and 5-8F-SC cells were significantly lower than those of nasopharyngeal carcinoma cells at 24, 48 and 72 hours after inoculation ( P<0.05). After 14 days of implantation, the colony formation rates of CNE2-SC (44.5±1.9)% and 5-8F-SC (47.4±1.8)% were higher than those of CNE2 (34.9±1.5)% and 5-8F (37.2±1.7)%, respectively( P<0.01). The migration cell number of CNE2-SC was (87.6±7.8), 3.97 times higher than that of CNE2 ( P<0.01). The migration cell number of 5-8F-SC was (67.2±5.7), 3.07 times higher than 5-8F ( P<0.01). The adhesion rates of CNE2-SC and CNE2 cells were (42.1±7.6)% and (8.9±2.0)%, respectively at 3 hours after inoculation and were (82.4±5.0)% and (12.1±2.2)% at 6 hours after inoculation, respectively. The adhesion rate of CNE2-SC cells was higher than that of CNE2 cells (all P<0.01). The adhesion rates of 5-8F-SC and 5-8F cells were (53.6±6.1)% and (7.3±1.5)% at 3 hours after inoculation, and (90.7±3.6)% and (11.0±1.2)% at 6 hours after inoculation, respectively. The adhesion rate of 5-8F-SC cells was higher than that of 5-8F cells ( P<0.01). The expression levels of CD44, Ras and N-cadherin were significantly higher, while phosphatase and tensin homolog deleted on chromosome 10 (PTEN), E-cadherin in nasopharyngeal carcinoma stem cells were lower than those of the nasopharyngeal carcinoma cells. Furthermore, the levels of phosphorylated mitogen extracellular kinase1/2 (p-MEK1/2) and phosphorylated extracellular signal-regulated protein kinase1/2 (p-ERK1/2)were significantly increased in nasopharyngeal carcinoma stem cells ( P<0.01). Correlation analysis showed that the protein expression levels of CD44 was highly positively correlated with RAS in nasopharyngeal carcinoma stem cells( r=0.985, P=0.002; r=0.962, P=0.038). Deletion of CD44 in CNE2-SC decreased the expression levels of HER-2, Ras and p-ERK1/2, p-Akt and phosphorylated protein kinase C-δ(p-PKCδ) ( P<0.01). Conclusions:Despite compare to the nasopharyngeal carcinoma cell, nasopharyngeal carcinoma stem cells grows at a relatively slow rate, the capacities of clone formation, migration, adhesion are promoted. This may be related to the CD44-regulated abnormal activation of Ras signaling pathway.

Objective To observe the relationship of efficacy of high-intensity focused ultrasound (HIFU) in treatment of hysteromyoma and quantitative parameters and enhance manifestations of CEUS.Methods Forty patients with single hysteromyoma underwent CEUS before HIFU,immediately after and 6 months after treatment,respectively.Time to peak (TTP),peak intensity (PI) of time-intensity curve were analyzed,and the enhance manifestations were observed.The ablation ratio of hysteromyoma immediately after HIFU and the ratio of reduction volume 6 months after treatment were also analyzed.Results Among 40 hysteromyomas,21 evenly enhanced (even enhance group) and 19 unevenly enhanced (uneven enhance group).The ablation rate greater than or equal to 80 ％ were observed in 22 hysteromyomas (ablation rate ≥80％ group),while ablation rate less than 80％ were observed in 18 lesions (ablation rate ＜80％ group).There were significant differences of TTP ([27.48 ± 7.89] s vs [21.78 ± 4.76] s;t =2.69,P =0.01),PI ([44.29 ± 3.74] dB vs [47.39±4.68]dB;t=-2.34,P=0.02) and the ratio of reduction volume 6 months after HIFU ([59.47±16.21]％ vs [37.30±11.89]％;t=4.83,P＜0.01) between ablation rate ≥80％ group and ablation rate ＜80％ group.The ablation rate ([65.03±18.39]％ vs [41.64±18.78]％;t=-4.49,P＜0.01) and the ratio of reduction volume 6 months after HIFU ([84.74±7.76]％ vs [58.18±12.94]％;t=-3.21,P＜0.01) were significantly different between even enhance group and uneven enhance group.TTP and the ratio of reduction volume 6 months after HIFU positively correlated with the ablation rate (r =0.448,0.660,both P＜ 0.01),while PI and enhance manifestations negatively correlated with ablation rate (r =-0.460,rs =-0.614,both P＜ 0.01).Conclusion TTP,PI of time-intensity curve and enhance patterns of hysteromyoma correlated with the ablation rate of HIFU,which having certain clinical value in evaluation of therapeutic efficacy.

Objective: To investigate the optimal duration of pegylated-alpha interferon (Peg-INFα) combined with ribavirin (RBV) in treating chronic hepatitis C infection in human immunodeficiency virus (HIV)-infected patients. Methods: A multicenter prospective study was conducted. The study subjects were divided into two groups; HIV/HCV co-infections (Group A, n = 158) and control with HCV-monoinfections (Group B, n = 60). All recruited patients received standard Peg-INFα plus RBV therapy. Group A was divided into 3 subgroups according to CD4⁺ cell counts: A1 subgroup, 79 cases, CD4⁺ counts > 350 cells /μl, who received anti-HCV before combination antiretroviral therapy(cART); A2 subgroup, 45 cases, CD4⁺ counts between 200 and 350 cells/μl, who did not start anti-HCV until they could tolerate cART well; A3 subgroup, 34 cases, CD4⁺ counts < 200 cells /μl, cART was administered first, and anti-HCV therapy was started when CD4⁺ counts > 200 cells/μl. The anti-HCV efficacy of two groups and 3 subgroups were compared. Statistical analysis for normal distribution and homogeneity of variance data was calculated by t-test and the counting data was analyzed by χ ² test. The Mann-Whitney U test was used for non-normal data. A one-way analysis of variance (ANOVA) was used for the comparison of multiple groups, followed by SNK method. Multiple independent samples were used for non-parametric tests. Results: There was no significant difference in age and baseline HCV RNA levels between groups and subgroups (P > 0.05). By an intent-to-treat (ITT) analysis, in Group A, the ratio of complete early virological response (cEVR) rate was 75.3% (119/158), the ratio of end of treatment virological response (eTVR) rate was 68.4% (108/158), and the ratio of sustained virological response (SVR) rate was 48.7% (77/158); in Group B, the ratio of cEVR rate was 93.3% (56/60), the ratio of eTVR rate was 90.0% (54/60), and the ratio of SVR rate was 71.7% (43/60); The therapeutic index of Group A were lower than those of Group B (P≤0.05). By per-protocol (PP) analysis, the ratio of cEVR rate in Group A [75.2% (88/112)] was still lower than that in Group B [93.3% (56/60)], but no significant differences were found in the ratio of eTVR rate and SVR rate between 2 groups (P > 0.05). Comparing the efficacy of subgroups (A1, A2 and A3) by ITT analysis, the ratios of cEVR rate were respectively 78.5% (62/79), 75.6% (34/45) and 67.6% (23/34); the ratios of eTVR rate were respectively 68.4%(54/79), 80.0%(36/45)and 52.9%(18/34); and the ratios of SVR rate were respectively 41.8%(33/79), 64.4%(29/45)and 44.1%(15/34). The ratio of eTVR in subgroup A2 was obviously higher than that in subgroup A3 and the ratio of SVR in subgroup A2 was statistically higher than that of subgroup A1(P≤0.05). However, by PP analysis, no significant differences of the therapeutic indexes were found among the respective subgroups (P > 0.05). Conclusion HIV-HCV co-infected patients would have better anti-HCV efficacy with Peg-INFα-2a plus RBV than HCV- monoinfected patients. The best time for initiating anti-HCV therapy in HIV-HCV co-infected patients is when CD4⁺ counts 200 cells/ μl.

Objective To evaluate the value of superb micro-vascular imaging( SMI) in evaluating the efficacy of uterine fibroids treated with high intensity focused ultrasound( HIFU) . Methods Forty patients with single fibroid were selected before and after HIFU treatment ,color Doppler flow imaging(CDFI) ,SMI and contrast-enhanced ultrasound ( CEUS ) pattern were used to detect the lesions . CEUS was used as standard . The correlation of different blood flow levels in the fibroids with SMI and the efficacy of HIFU were evaluated . Results Before HIFU treatment ,the blood flow signals of different degrees were found in the uterine fibroids . SMI showed that 4 fibroids( 10 .0％ ) were in the first degree ,21 fibroids( 52 .5％ ) were in the second degree and 15 fibroids ( 37 .5％ ) were in the third degree . CEUS showed that 8 fibroids ( 20 .0％ ) were hypo-enhanced ,19 fibroids( 47 .5％ ) were iso-enhanced and 13 fobroids(32 .5％ ) were hyper-enhanced . The correlation analysis showed that there was close relationship between the results of SMI and CEUS( Kappa = 0 .754 , P = 0 .00) . After HIFU treatment ,SMI and CEUS had no statistical difference in evaluating the efficacy of HIFU( P > 0 .05) . The ratio of non-perfused volume and the ratio of the volume reduction at 6 months after HIFU in the third degree of SMI were lower than those in the first and second degrees( P < 0 .05) . Conclusions There are some relationships between SMI and CEUS in evaluating the blood flow signal of uterine fibroids . SMI can prompt therapeutic efficacy of uterine fibroids ablated by HIFU and provide some clinical reference values .

OBJECTIVETo investigate cytokine level in bronchoalveolar lavage fluid (BALF) in children with refractory Mycoplasma pneumoniae pneumonia (RMPP) and the effects of methylprednisolone on RMPP.

METHODSixty cases with RMPP and 20 cases with bronchial foreign body with no respiratory tract infection as control group hospitalized in Department of Pulmonary Diseases, the Children's Hospital Affiliated to Medical School of Zhejiang University from February 2012 to February 2013 were enrolled. The RMPP patients were divided into two groups randomly (30 cases in each). Steroid group were given methylprednisolone 2 mg/(kg·d) intravenously for 3 days, and the cases in non steroid group were not given steroid therapy. Patients whose fever relieved after steroid treatment were classified as defervesced group while the others were classified as non defervesced group. Each patient was examined with fiberoptic bronchoscopy and bronchoalveolar lavage 3 days after admission and cytokine level in BALF of each patient was detected.

RESULT(1) In steroid group, the proportion of patients whose fever disappeared within 3 days after steroid therapy was 9/30 cases (30%), and in non steroid group no one responded within 3 days after medication, showing statistically significant difference (χ² = 14.073, P=0.002), at the same time, the duration of cough in steroid group was significantly shorter than that in non steroid group (5.1 d vs. 7.0 d, t=-2.276, P=0.027). The total fever time of steroid group was 4.7 days, which as compaired with non steroid group (6.7 days) was shorter, but the difference was not significant (t=-1.351, P=0.134). (2) IL-1 β, IL-4, IL-6, IL-8, IL-10, IFN-γ in BALF of steroid group and non steroid group were both significantly higher than that of control group. But the same comparison between steroid group and non steroid group showed no significant difference. (3) In steroid group, IL-2 and IL-8 in BALF of patient whose fever disappeared after steroid therapy were both significantly lower than that of patients who still had fever (t=2.771, 2.054, P=0.010, 0.049) , but no significant difference was found between the two groups in BALF IL-1 β, IL-4, IL-6, IL-10, IFN-γ levels (P>0.05).

CONCLUSION(1) Three days of 2 mg/(kg·d) methylprednisolone therapy had the antipyretic effect in children with RMPP, and could shorten the length of cough. (2) Incresed BALF IL-1 β, IL-4, IL-6, IL-8, IL-10, IFN-γ levels were observed in RMPP and high level of BALF IL-2 and IL-8 might have some relevance with persistent fever of RMPP in children.

Bronchoalveolar Lavage Fluid ; chemistry ; Bronchoscopy ; Child ; Cytokines ; chemistry ; Fever ; Humans ; Methylprednisolone ; therapeutic use ; Mycoplasma pneumoniae ; Pneumonia, Mycoplasma ; drug therapy

Objective To investigate the role of immunohistochemistry in the differential diagnosis between small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC).Methods The protein expressions of CD56, Syn, TTF1, CK5/6, CK14, P63, CK7, and NapsinA in percutaneous lung biopsy and bronchoscopic biopsy specimens which were suspected as SCLC were examined by immunohistological streptavidin-peroxidase ( SP) method to analyze the pathological characteristics , immunological pheno-typical features , and differential diagnosis between SCLC and NSCLC .Results Among 72 cases of lung cancer patients ,there were 27 cases of SCLC,17 cases of low differentiated squamous cell carcinoma ( SCC) and 28 cases of low differentiated adenocarcinoma ( ADC) .Conclusions It is the different therapy between SCLC and NSCLC , immunohistochemistry analysis of biopsy can provide ac-curate diagnosis of SCLC and NSCLC , which will result in less misdiagnosis and provide an important valuable in the selection of clini -cal treatment protocols for lung cancer patients .

Objective To establish a model of rat endotoxic pulpitis induced by Lipopolysaccharide (LPS);to study the dynamic expression and location of Oct-4B in rat pulpitis model at different time;To evaluate the role of Oct-4B on pulp injury and reparation in rat endotoxic pulpitis. Method The slides were made and the dynamic expression and location of Oct-4B was examined by immunohistochemistry. Results Oct-4B was moderately positive in the pulp fibroblasts and odontoblasts in 2 weeks. After 2 weeks, it was still positive in odontoblasts and fibroblasts. Up to 3 weeks, it was weakly positive in odontoblasts and fibroblasts. Conclusion Oct-4B is dynamically expressed in the rats with pulpitis, and may play an important role in the incidence,development and recovery of pulpitis.

Objective To explore the effect of Yuduqing on the apoptosis of CML K562 cells cultured in vitro and its molecular mechanism.Methods In serologic pharmacological test,the K562 cells were divided into 8 different groups.Serum with imatinib plus Yuduqing (high-dose,middle-dose,low-dose) were added into the cells respectively in the 8 groups of K562 cells.Morphological assessment of apoptosis was performed with optical microscope,the rates of apoptosis and the cell cycles analysis was performed with flow cytometry at 12 h,24 h,48 h and 72 h time points respectively after the intervention.Results The primary cell of normal control group had a low rate of apoptosis,while the blank control group K562 cell apoptosis rate was lower,the difference is significant (P＜0.05).The differences between the rates of apoptosis in high-dose,middle-dose and low-dose Yuduqing groups and those in normal control group and blank control group were significant in 12 hours or 24 hours (P＜0.05).Drug groups showed significant differences of pair-comparison in groups and a certain time dose dependence.But the rate of apoptosis(31.48± 6.58) in k562 cells in high-dose Yuduqing group did not increase further at 72 hours after the intervention and it was not statistically different from that of 48 hours,nor statistically different from that of middle-dose group at 72 hours(27.54±5.89) after the intervention (P＞0.05).The rate ofapoptosis in k562 cells in imatinib group (23.80±6.94) was relatively high at 12 hours after the intervention and it was significantly different from that in blank control group (P＜0.05).The rates of apoptosis in imatinib and Yuduqing (high-dose,middle-dose,and low-dose) groups were significantly higher than those in imatinib group or Yuduqing high-dose,middle-dose,and low-dose)groups (P＜0.05).Conclusion Serum with Yuduqing could induce apoptosis of K562 cells cultured in vitro and its action was dose-time dependent; Serum with Yuduqing (high-dose and middle-dose) was similar to serum with imatinib in inducing apoptosis of K562 cells cultured in vitro; Yuduqing could enhance the efficacy of imatinib.