PAX1 and SOX1 Gene Methylation as a Detection and Triage Method for Cervical Intraepithelial Neoplasia Diagnosis.

INTRODUCTION: Methylation assays have demonstrated potential as dependable and high-precision approaches for identifying or triaging individuals with cervical cancer (CA) or cervical intraepithelial neoplasia (CIN). Our investigation aimed to assess the efficacy of the diagnosis and triage of the PAX1/SOX1 methylation panel in detecting CIN or CA. METHODS: A total of 461 patients with abnormal high-risk human papillomavirus (hrHPV) or cytology test results were recruited for this study. Each patient underwent an assortment of assessments, comprising a cytology test, hrHPV test, colposcopy examination, and PAX1 and SOX1 methylation tests. RESULTS: The extent of methylation of both genes demonstrates a positive correlation with the severity of CIN lesions and CA. To determine the correlation for patients with CIN2 or worse (CIN2+), the area under curve was 0.821 (95% CI: 0.782-0.853) for PAX1 and 0.800 (95% CI: 0.766-0.838) for SOX1, while for CIN3 or worse (CIN3+), 0.881 (95% CI: 0.839-0.908) for PAX1 and 0.867 (95% CI: 0.830-0.901) for SOX1. The PAX1/SOX1 methylation marker panel performed sensitivity and specificity of 77.16% and 91.67% for CIN2+, 84.76% and 90.50% for CIN3+, respectively. Regarding triaging hrHPV+ patients, the PAX1/SOX1 methylation test only referred 11.83% of the patients who are unnecessary for colonoscopy examination, which is comparatively lower than cytology, thereby signifying a promising triage strategy for hrHPV-positive women. Furthermore, we observed that the positive PAX1/SOX1 methylation test result for untreated CIN1 or fewer patients would result in a higher likelihood of progression upon a 24-month follow-up visit. CONCLUSION: The present investigation demonstrates that the PAX1/SOX1 methylation marker panel exhibits favorable diagnostic performance in CIN detection and holds the potential to be employed for individual CIN tests or hrHPV-positive triage.

Survival Outcomes of Patients With Stage IB3 Cervical Cancer Who Undergo Abdominal Radical Hysterectomy Versus Radiochemotherapy.

Objective: This study aimed to compare the survival outcomes among stage IB3 cervical cancer patients who undergo abdominal radical hysterectomy (ARH)+pelvic lymphadenectomy ± para-aortic lymph node dissection versus radiochemotherapy (R-CT). Methods: Based on the large number of diagnoses and treatments for cervical cancer in the Chinese database, propensity score matching (PSM) was used to compare the 5-year overall survival (OS) and disease-free survival (DFS) rates of the ARH group and R-CT group. Results: There were 590 patients with stage IB3 cervical cancer according to the FIGO 2018 staging system, with 470 patients in the ARH group and 120 patients in the R-CT group. The ARH and R-CT groups showed different 5-year OS and DFS rates in the total study population, and the 5-year OS and DFS rates in the R-CT group (n = 120) were lower than those in the ARH group (n = 470) (OS: 78.1% vs. 92.1%, p < 0.001; DFS: 71.6% vs. 90.3%, p < 0.001). R-CT was associated with a worse 5-year OS rate (hazard ratio [HR] = 3.401; 95% confidence interval [CI] = 1.875-6.167; p < 0.001) and DFS rate (HR = 3.440; 95% CI = 2.075-5.703; p < 0.001) by Cox multivariate analysis. After 1:3 PSM, the 5-year OS and DFS rates in the R-CT group (n = 108) were lower than those in the RH group (n = 280) (OS: 76.4% vs. 94.0%, p < 0.001; DFS: 69.3% vs. 92.6%, p < 0.001, respectively). R-CT was associated with a worse 5-year OS rate (HR = 4.071; 95% CI = 2.042-8.117; p < 0.001) and DFS rate (HR = 4.450; 95% CI = 2.441-8.113; p < 0.001) by Cox multivariate analysis. Conclusion: Our study found that for FIGO 2018 stage IB3 cervical cancer patients, ARH resulted in better OS and DFS than R-CT.

Discussion on the Treatment Strategy for Stage â ¡A1 Cervical Cancer (FIGO 2018).

Objective: This study aimed to explore the best treatment strategy for International Federation of Gynecology and Obstetrics (FIGO) 2018 stage IIA1 cervical cancer patients by comparing the survival outcomes of two treatment methods: abdominal radical hysterectomy (ARH) with standard postoperative therapy and radio-chemotherapy (R-CT). Methods: Patients with FIGO2018 stage IIA1 cervical cancer who underwent ARH or received R-CT were screened from the clinical diagnosis and treatment for cervical cancer in China (Four C) database. The recurrence cases between the two groups were analyzed. The 5-year overall survival (OS) and disease-free survival (DFS) of patients diagnosed with stage IIA1 cervical cancer in 47 hospitals in mainland China between 2004 and 2018 were compared by using propensity score matching (PSM). Results: A total of 724 patients met the inclusion criteria. In the total study population, The R-CT group had higher recurrence (22.8% for the R-CT group and 11.2% for the ARH group, P<0.001) rates compared to the ARH group.The 5-year OS and DFS of the ARH group (n=658) were significantly higher than those of the R-CT group (n=66) (OS: 85.9% vs. 71.2%, P=0.009; DFS: 79.2%vs. 70.5%, P=0.027). R-CT was associated with worse 5-year OS (HR=3.19, 95% CI: 1.592-6.956, P=0.001) and DFS (HR=2.089, 95% CI: 1.194-3.656, P=0.01). After 1:2 PSM, the 5-year OS and DFS of the ARH group (n=126) were significantly higher than those of the R-CT group (n=64) (OS:88.9% vs. 70.1%, P=0.04; DFS:82.8% vs. 69.8%, P=0.019). R-CT was still associated with worse 5-year OS (HR=2.391, 95% CI: 1.051-5.633, P=0.046) and DFS (HR=2.6, 95% CI: 1.25-5.409, P=0.011). Conclusion: Our study demonstrated that for stage FIGO2018 stage IIA1 cervical cancer patients, ARH offers better oncological outcomes than R-CT.

Comparison of survival outcomes of abdominal radical hysterectomy and radiochemotherapy IIA2 (FIGO2018) cervical cancer: a retrospective study from a large database of 63,926 cases of cervical cancer in China.

AIM: This study aimed to compare the 5-year overall survival (OS) and 5-year DFS disease-free survival (DFS) of abdominal radical hysterectomy (ARH) and radiochemotherapy (R-CT) for stage IIA2 (FIGO 2018) cervical cancer patients. METHODS: Based on this multicenter, retrospective cohort study based on data from the clinical diagnosis and treatment of cervical cancer in China (Four C) database, 609 cases with 2018 FIGO stage IIA2 cervical cancer from 2004 to 2018 were reviewed. The 5-year OS and 5-year DFS of patients with either of the two treatment methods were compared by means of a multivariate Cox regression model and the log-rank method in the total study population and after propensity score matching (PSM). RESULTS: We selected 609 of 63,926 patients and found that R-CT was associated with a significantly worse 5-year OS (71.8% vs. 95.3%, P < 0.001; hazard ratio (HR) = 6.596, 95% CI 3.524-12.346) and 5-year DFS (69.4% vs. 91.4%, P < 0.001; HR = 4.132, 95% CI 2.570-6.642, P < 0.001) than ARH in the total study population. After matching (n = 230/230), among FIGO 2018 IIA2 patients, the 5-year OS and DFS were lower in the R-CT group than in the ARH group (OS: 73.9% vs. 94.7%, P < 0.001; HR = 5.633, 95% CI 2.826-11.231, P < 0.001; DFS: 69.2% vs. 91.1%, P < 0.001; HR = 3.978, 95% CI 2.336-6.773, P < 0.001, respectively). CONCLUSIONS: In patients with stage FIGO 2018 IIA2 cervical cancer, ARH offers better 5-year OS and DFS outcomes than R-CT; however, due to the inherent biases of retrospective studies, this needs to be confirmed by randomized trials.

CCR7 Has Potential to Be a Prognosis Marker for Cervical Squamous Cell Carcinoma and an Index for Tumor Microenvironment Change.

The tumor microenvironment (TME) has an essential role in the development of cervical squamous cell carcinoma (CSCC); however, the dynamic role of the stromal and immune cells is still unclear in TME. We downloaded data from The Cancer Genome Atlas (TCGA) database and applied ESTIMATE and CIBERSORT algorithms to measure the quantity of stromal and immune cells and the composition of tumor-infiltrating immune cell (TIC) in 253 CSCC cases. The protein-protein interaction (PPI) network and Cox regression analysis presented the differentially expressed genes (DEGs). Then, C-C chemokine receptor type 7 (CCR7) was screened out as a prognostic marker by the univariate Cox and intersection analysis of PPI. Further analysis showed a positive correlation between the expression of CCR7 and the survival of CSCC patients. The result of the Gene Set Enrichment Analysis (GSEA) of genes in the high CCR7 expression group displayed a predominant enrichment in immune-related pathways. An enrichment in metabolic activities was observed in the low CCR7 expression group. CIBERSORT analysis showed a positive correlation between Plasma cells, CD8+ T cells, and regulatory T cells and the CCR7 expression, suggesting that CCR7 might play a crucial role in maintaining the immunological dominance status for TME. Therefore, the expression level of CCR7 might help predict the survival of CSCC cases and be an index that the status of TME transitioned from immunological dominance to metabolic activation, which presented a new insight into the treatment of CSCC.

Long non-coding RNA HAND2-AS1 delays cervical cancer progression via its regulation on the microRNA-21-5p/TIMP3/VEGFA axis.

Cervical cancer is a common cause of cancer-related mortality in women. Mounting evidence suggests that long non-coding RNAs (lncRNAs) function vitally in many cancers. In this study, we discovered that the regulation of the heart and neural crest derivatives expressed 2-antisense RNA 1 (HAND2-AS1) in cervical cancer. RT-qPCR was conducted to detect the expression of HAND2-AS1 and microRNA-21-5p (miR-21-5p). The relationship of HAND2-AS1 and miR-21-5p was identified by dual-luciferase reporter gene assay. The roles of HAND2-AS1, miR-21-5p and tissue inhibitor of metalloproteinases-3 (TIMP3) in cervical cancer were accessed via gain- and loss-of-function approaches. The expression of related proteins in the vascular endothelial growth factor A (VEGFA) signaling pathway was detected through Western blot analysis. Finally, xenografts of cervical cancer in nude mice were established to assess the effect of HAND2-AS1 on tumorigenesis in vivo. HAND2-AS1 and TIMP3 were downregulated in cervical cancer, which were identified to be associated with a poor prognosis of patients with cervical cancer. Moreover, HAND2-AS1 was upregulated the expression of TIMP3 through competitively binding to miR-21-5p. Overexpressed HAND2-AS1 or downregulated miR-21-5p inhibited cell proliferation, migration, and invasion while promoting cell apoptosis, in association with increased expression of proteins in VEGFA signaling pathway. These changes were reversed by silencing of TIMP3. Overexpressed HAND2-AS1 reduced the tumor formation ability in nude mice. In summary, HAND2-AS1 may exert inhibitory effects on cervical cancer cell growth and cervical cancer development through its regulation on the miR-21-5p/TIMP3/VEGFA axis. This highlights that HAND2-AS1 may serve as a potential target for cervical cancer diagnosis and treatment.

Risk factors and long-term impact of urologic complications during radical hysterectomy for cervical cancer in China, 2004-2016.

OBJECTIVE: The primary objective was to describe the incidence and risk factors of urologic complications during radical hysterectomy for cervical cancer. The secondary objective was to investigate the impact of urologic complications on long-term survival. METHODS: Patients who underwent radical hysterectomy for cervical cancer from 2004 to 2016 were identified in the MSCCCC (Major Surgical Complications of Cervical Cancer in China) database. Data on demographic characteristics, clinical characteristics, hospital characteristics and urologic complications were collected. Multivariable logistic regression was used to assess the risk factors of urologic complications and Cox proportional hazards models were performed to identify prognostic factors. RESULTS: A total of 21,026 patients undergoing radical hysterectomy for cervical cancer were identified. The incidence of any urologic complications was 1.54%: 83 (0.39%) ureteral injuries, 17 (0.08%) bladder injuries, 1 (0.005%) ureteral injury combined with bladder injury, and 223 (1.05%) genitourinary fistulas. In a multivariable analysis, surgery at a women and children's hospital (OR = 2.26, 95% CI 1.47-3.48), surgery at a facility in a first-tier city (OR = 2.08, 95% CI 1.24-3.48), and laparoscopic surgery (OR = 4.68, 95% CI 3.44-6.36) were associated with a higher risk of urologic complications. Cox proportional hazards models revealed that the occurrence of urologic complications was a significant predictor of 2-year overall survival (OR = 1.78, 95% CI = 1.09-2.92), but was not a predictor of 5-year overall survival (OR = 1.27, 95% CI = 0.83-1.94). CONCLUSION: The incidence of urologic complications during radical hysterectomy is low. The risk of urologic complications may be higher for patients who are treated at a women and children's hospital, are treated in first-tier city hospitals, and receive laparoscopic surgery. Urologic complications have an impact on short-term survival, but not on long-term survival.

GINS2 regulates cell proliferation and apoptosis in human epithelial ovarian cancer.

Go-Ichi-Ni-San 2 (GINS2), also known as partner of Sld five 2, is involved in the initiation of DNA replication and cell cycle progression. GINS2 is abundantly expressed in a number of malignant solid tumors, including breast cancer, melanoma and hepatic carcinoma. However, the functions of GINS2 in epithelial ovarian cancer (EOC) remain unclear. The aim of the present study was to investigate these functions. GINS2 expression was detected in EOC and normal ovarian tissues using immunohistochemistry. To investigate the functions of GINS2 in EOC, GINS2 expression was stably knocked down in SKOV-3 cells using lentiviral short hairpin RNA (shRNA). The expression of GINS2 mRNA and protein in SKOV-3 cells was examined using reverse-transcription quantitative polymerase chain reaction (RT-qPCR) and western blot analyses, respectively. Cell proliferation was determined using high-content screening and MTT assays. Cell cycle progression and apoptosis were detected using flow cytometry. Compared with normal ovarian tissues, EOC tissues expressed increased levels of GINS2 expression (16.7 vs. 58.3%). Increased expression of GINS2 mRNA was also observed in SKOV-3 and OVCAR3 cells. In the investigation of GINS2 functions in EOC, GINS2 expression at the mRNA and protein levels was significantly inhibited by specific GINS2 shRNA. GINS2 knockdown significantly inhibited the proliferation and viability of SKOV-3 cells and induced cell cycle arrest in S phase. Furthermore, GINS2 knockdown in SKOV-3 cells significantly increased cell apoptosis. GINS2 is markedly expressed in EOC tissues and cell lines. Stable GINS2 knockdown in SKOV-3 cells significantly inhibited cell proliferation and induced cell cycle arrest and cell apoptosis. Therefore, GINS2 may be involved in EOC progression.

Long noncoding RNA BDNF-AS is downregulated in cervical cancer and has anti-cancer functions by negatively associating with BDNF.

PURPOSE: We investigated expression and mechanism long noncoding RNA BDNF-AS in human cervical cancer (CC). METHODS: BDNF-AS expressions were examined by qPCR in CC cell lines and human CC tumors. CC cell lines, SiHa and DoTc2-4510 were transduced with lentivirus to ectopically overexpress BDNF-AS. Possible anti-cancer effects of BDNF overexpression were examined on CC in vitro proliferation and migration, and in vivo transplantation. Human BDNF gene expression was also examined in CC cell lines and tumors. In CC cells with overexpressed BDNF-AS, BDNF was upregulated to examine its direct effect in NDNF-AS-modulated CC proliferation and migration. RESULTS: BDNF was downregulated in both CC cells and human CC tumors. In CC cells, BDNF-AS overexpression is anti-cancer by inhibiting proliferation and migration in vitro, and transplantation in vivo. BDNF was inversely expressed as BDNF-AS in CC. Upregulation of BDNF in BDNF-AS-overexpressed CC cells reversed the anti-cancer effects of BDNF-AS. CONCLUSION: BDNF-AS is downregulated in CC. Overexpressing BDNF-AS may inhibit CC, possibly through inverse regulation on BDNF.

MicroRNA-137 is negatively associated with clinical outcome and regulates tumor development through EZH2 in cervical cancer.

We intend to evaluate the expression, clinical relevance, and functional role of microRNA-137 (miR-137) in human cervical cancer (CC). MiR-137 expressions were assessed by qPCR in CC cell lines and human CC tumors. The correlation between endogenous miR-137 expression and CC patients' postoperative overall survival was examined statistically. CC cell lines, Ca-Ski, and SiHa cells were transduced with lentivirus to ectopically upregulate endogenous miR-137 expressions. Possible inhibitory effects of miR-137 upregulation on CC in vitro proliferation and migration, as well as in vivo transplantation were evaluated. Targeting of enhancer of zeste homolog 2 (EZH2) gene by miR-137 in CC was assessed by dual-luciferase activity assay and qPCR. In CC cells with upregulated miR-137, EZH2 was overexpressed to assess its direct function in miR-137 mediated CC proliferation and migration. MiR-137 was downregulated in both CC cells and human CC tumors. Downregulation of endogenous miR-137 was significantly correlated with CC patients' short overall survival. In CC cells, miR-137 upregulation is tumor-suppressive by inhibiting proliferation and migration in vitro, and transplantation in vivo. EZH2 was a direct downstream target gene of miR-137 in CC. Forced overexpression of EZH2 in miR-137-upregulated CC cells reversed the tumor-suppression induced by miR-137. MiR-137 is lowly expressed in CC and possibly acting as a negative biomarker for CC patients' clinical outcome. MiR-137 upregulation may suppress CC, very likely by inversely regulating EZH2.

[Inhibitors of JAK2/STAT Signaling Pathway and Hematologic Malignancies-Review].

Jauns kinase (JAK)/transducer and activator of transcription（STAT） pathway is a classical approach to study the rapid changes of the gene expression in specific target cells by a variety of extracellular signals. The JAK and STAT transfer cytokine receptor signaling plays a unique role in multiple cellular and molecular biological changes.The abnormal signal of JAK/STAT pathway will lead to the hematopoietic abnormalities.Studies had shown that the abnormal activation of JAK2/STAT signaling pathway are in many kinds of malignant hematological diseases, such as in acute lymphoblastic/myeloid leukemia, chronic myeloid leukemia, lymphoma, myelodysplastic syndromes, myeloprofilerative neoplasm, especially in the patients of myeloproliferative neoplasm(MPN) with JAK gene mutation(JAK2V617F), this mutation has an important value for MPN diagnosis. At present, the effect of the specific inhibitors of JAK2 has showed good perspective, which had been applied to clinic treatment and achieved remarkable curative effect. In this review, the JAK2/STAT signaling transduction, the JAK2 signal and hematologic malignancies, the kagulation of signaling pathway and the inhibitors of JAK2/STAT signaling pathway are summarized.

[Effect of IFN-α2b on COX-2 and Angiogenesis in JAK2V617F Mutation Myeloproliferative Neoplasms].

OBJECTIVES: To investigate the influence of interferon-alpha-2b (IFN-α2b) with JAK2 kinase, COX-2 and microvessel density in patients of MPN and the relation of JAK2V617F and COX-2 in human erythroleukemia cell line (HEL) cells. METHODS: Forty-two cases of MPN patients with JAK2V617F mutation of initial treatment were collected from the Frist hospital of Baoding, including the IFN-α2b treatment group with 17 cases and untreated group with 25 cases. 10 cases of idiopathic immune thrombocytopenic purpura (ITP) patients synchronization were enrolled as controls. JAK2V617F/JAK2 mutation burden of MPN patients was detected by real time PCR (qRT-PCR);the expression levels of p-JAK2, COX-2 and microvascular density (MVD) marked with CD105 inpathological tissues of bone marrow in patients of MPN and ITP were detected by immunohistochemistry. The HEL cells were treated with different concentrations of IFN-α2b. The cell proliferation inhibition rate was calculated by CCK-8 test;the apoptosis rate was detected by flow cytometry; cell migration ability was tested by transwell chambers. JAK2 and COX-2 mRNA were detected by semi-quantitative PCR; p-JAK2 and COX-2 protein in HEL cells were detected by Western blotting. RESULTS: The expression levels of p-JAK2, COX-2 protein and MVD in untreated group were significantly higher than those of control groups. p-JAK2, COX-2 and MVD levels were significantly reduced in patients treated with IFN-α2b. Cell growth inhibition rates and apoptosis rates raise up by dose of IFN-α2b in HEL cells at 48 h.The mRNA expression levels of JAK2 and COX-2 as well as protein expression levels of p-JAK2 and COX-2 had a decreasing tendency with the increase of IFN-α2b concentration at 48 h.The migration capacity level of HEL cells which treated with 0.5×10 4 U/L IFN-α2b after 24 h was lower than that of control group. CONCLUSIONS: Angiogenesis of MPN and COX-2 were inhibited by IFN-α2b which regulates JAK2 signal pathway.

[The Effect of Ruxolitinib on the Expression of VEGF and HIF-1α in Leukemia HEL Cells].

OBJECTIVES: To investigate the effect of Ruxolitinib on the expression of vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1 α (HIF-1α) in HEL cells. METHODS: he HEL cells were treated with Ruxolitinib in different concentrations (1 nmol/L, 5 nmol/L, 10 nmol/L, 50 nmol/L, 100 nmol/L, 500 nmol/L). The growth inhibition of Ruxolitinib on HEL cells was detected by CCK-8 assay;the mRNA expression level ofJAK2 were measured by RT-PCR and the protein level of p-JAK2, VEGF, HIF-1α were observed by Western blot after treated with Ruxolitinib for 24,48,72 h. Chick chorioallantoic membrane (CAM) test was used to testify the effect of Ruxolitinib on angiogenesis. RESULTS: Ruxolitinib with different concentrations could inhibit HEL cells proliferation. RT-PCR showed that the mRNA level ofJAK2 decreased in a concentration-dependent manner and Western blot demonstrated that the expression levels of p-JAK2, VEGF and HIF-1α were lower in Ruxolitinib treatment groups than those in control group (P<0.05) after HEL cells were treated with different concentrations of Ruxolitinib for 24,48,72 h. Ruxolitinib significantly suppressed blood vessels'formation in CAM. CONCLUSIONS: Ruxolitinib can inhibit VEGF, HIF-1α expression and angiogenesis of HEL leukemia cells by inhibiting JAK2 pathway.

[Comparison of safety and efficacy of laparoscopic versus abdominal radical hysterectomy in the treatment of patients with stage I a2-II b cervical cancer].

OBJECTIVE: To compare the safety and efficacy after laparoscopic radical hysterectomy (LRH) and abdominal radical hysterectomy (ARH) in the treatment of patients with stage I a2-II b cervical cancer. METHODS: In a retrospective study, data were analyzed from patients with International Federation of Gynecology and Obstetrics (FIGO) stage Ia2-II b cervical cancer underwent LRH or ARH at Union Hospital, Tongji Medical College, Huazhong University of Science and Technology; First Affiliated Hospital, School of Medicine, Shihezi University; and the Guizhou Provincial People's Hospital between 2000 and 2015. Perioperative outcomes and survival analysis were compared. RESULTS: (1) The FIGO stages, histotypes, metastasis of lymph nodes, lymph vascular space invasion and neoadjuvant chemotherapy significantly differed between the LRH group and the ARH group (all P<0.05). In order to eliminate the effects by the unbalanced data, stratified analysis was conducted based on FIGO stage. There were 861 patients in stage I a2-I b1 group, including 663 patients in LRH group and 198 patients in ARH group. And there were 668 patients in stage I b2-IIb group, including 389 patients in LRH group and 279 patients in ARH group. (2) In the patients with stage I a2- I b1 and I b2- II b tumor, there were no significant difference in age, histotype, differentiation degree, parametrial invasion, lymphvasular invasion space and neoadjvant chemotherapy between the LRH group and the ARH group (all P>0.05). For patients with stage I a2- I b1, the operation time in the LRH group was longer than that in the ARH group (P=0.027), and it showed less blood loss and lower blood transfusion rate in the LRH group than those in the ARH group (all P=0.000). The findings were similar in the patients with stage I b2-II b (all P=0.000). (3) There were no significant difference in intraoperative complications and postoperative complications between the LRH and the ARH group in the patients with stage I a2-I b1 and I b2-IIb, respectively (all P>0.05). (4) The median follow- up time was 24 months (range: 1 to 177 months), the recurrence rate was 3.6% (38/1 052) in LRH group and 3.1% (15/477) in ARH group,there was not significant difference (P>0.05). The estimated 3- year overall survival (OS) and the free-progression survival time (PFS) were respectively 92.4% and 91.5% in LRH group, and 91.8% and 91.5% in ARH group. There was no significant difference in the overall survival (P=0.738) or progress free survival (P=0.990) by log-rank test. Moreover, there were no significant difference in OS or PFS between the LRH group and the ARH group in patients with stage I a2- I b1 and I b2- II b, respectively (all P>0.05). CONCLUSION: LRH is safe and effective, and it could be used a routine way for the treatment of patients with stage I a2-IIb cervical cancer.

[Anti-angiogenic effect of interferon on JAK2V617F positive myeloproliferative neoplasms and its anti-angiogenic mechanisms].

OBJECTIVE: To research the suppressive effect of interferon α2b (IFN-α2b) on angiogenesis of JAK2 tyrosine kinase gene point mutation (JAK2V617F) positive myeloproliferative neoplasm (MPN) and its anti-angiogenic mechanisms. METHODS: (1) A total of 42 cases of JAK2V617F positive MPN patients in the No.1 Hospital of Baoding were selected between January 2012 and October 2014, including the newly diagnosed group before treatment (n=25) and the IFN-α2b treatment group (n=17). Ten cases of idiopathic immune thrombocytopenic purpura (ITP) patients were enrolled as controls. JAK2V617F/JAK2 ratio was detected by real-time fluorescent quantitative PCR to measure mutation; the expression levels of phosphorylated JAK2 (p-JAK2), vascular endothelial growth factor (VEGF), hypoxia-inducible factor-1α (HIF-1α) and CD105 marked microvascular density (MVD) in bone marrow were detected by immunohistochemistry. The correlations were analyzed among JAK2V617F mutation burden and VEGF, HIF-1α, and MVD. (2) Human erythroleukemia cell line HEL cells were treated with different concentrations of IFN-α2b. The apoptosis was detected by Hoechst staining method. The cell viability was detected by CCK-8 test. Cell migration ability was tested by transwell chambers. The protein expression levels of p-JAK2, VEGF, and HIF-1α were detected by Western blot. RESULTS: (1)The ratio of JAK2V617F/JAK2 in the IFN-α2b treatment group (22.69% ± 12.64%) was significantly lower than that in the newly diagnosed group (46.17% ± 19.32%) (P<0.01). The expression levels of p-JAK2, VEGF, and HIF-1α in the newly diagnosed group (82.41% ± 11.65%, 64.72% ± 25.01%, 45.12% ± 20.28%) were significantly higher than those in the IFN-α2b treatment group (60.93% ± 20.57%, 36.58% ± 15.95%, 32.15% ± 14.27%) and the control group (43.05% ± 12.59%, 25.69%± 1 3.75%, 25.07% ± 15.49%) (all P<0.01). MVD in the newly diagnosed group (26.58% ± 5.93%) was significantly higher than that in the treatment group (15.86%± 4.27%) and the control group (10.76% ± 4.01%) (P<0.01). Spearman correlation analysis showed positive correlation of JAK2V617F mutation with VEGF and MVD (r=0.589, P<0.05; r=0.577, P<0.05). (2)The apoptosis of HEL cells were increased after treated with 30 000 U/L of IFN-α2b in HEL cells after 24 h. The proliferation of HEL cell were inhibited by different concentrations of IFN-α2b in time- and dose-dependent manner. The number of membrane-permeating HEL cells after treated with 5 000 U/L of IFN-α2b in HEL cells after 24 h was significantly lower than that in the untreated cells (52.9 ± 7.5 vs 77.3 ± 6.1) (P<0.05). The expression levels of p-JAK2, VEGF, and HIF-1α were decreased in a dose-dependent manner. CONCLUSION: IFN-α2b may exert an anti-angiogenic effect by inhibiting the VEGF, HIF-1α, and MVD expression in MPN via JAK2 signal pathway.

[Effect of tyrosine kinase inhibitor imatinib mesylate on K562 cell invasion by PTEN pathway].

AIM: To investigate the effect of tyrosine kinase inhibitor imatinib mesylate on the PTEN signaling pathway and the cell invasion in K562 cells. METHODS: K562 cells were treated with different concentrations of imatinib mesylate. After different time periods, the mRNA levels of BCR/ABL, PTEN and FAK were detected by real-time fluorescent quantitative PCR (FQ-PCR) to analyze their relationships. The protein level of FAK was detected by immunocytochemistry. The cell invasive ability was examined by Transwell (Boyden chamber) assay. RESULTS: In the initial 36 h, the expression level of PTEN mRNA was up-regulated and the FAK mRNA was down-regulated with the reduction of BCR/ABL fusion gene expression and the cell invasive ability of K562 cells was inhibited by 2 µg/mL imatinib mesylate. 48 h later, the PTEN mRNA expression level decreased and the FAK mRNA expression level was elevated with the restore of BCR/ABL fusion gene. BCR/ABL mRNA level presented a positive correlation with PTEN mRNA expression level, and a negative correlation with FAK mRNA. CONCLUSION: Tyrosine kinase inhibitor imatinib mesylate can regulate PTEN/FAK pathway and inhibit the leukemia K562 cell invasive ability via restraining BCR/ABL fusion gene.

Effect of phosphatase and tensin homology deleted on chromosome 10 (PTEN) gene transfection on reversal of multidrug resistance in K562/ADM cells.

The phosphatase and tensin homology deleted on chromosome 10 (PTEN) gene is a novel tumor suppressor gene of the phosphatase family. Studies have shown that the PTEN gene is probably involved in human malignant disease pathogenesis, multidrug resistance, angiogenesis and extramedullary infiltration. This study was designed to investigate the effect of wild-type PTEN gene transfection on drug resistance reversal in K562/ADM leukemia cells in vitro and the possible mechanism. A recombinant adenovirus containing green fluorescent protein gene and wild-type PTEN gene (Ad-PTEN-GFP) or a recombined adenovirus containing green fluorescent protein gene only (Ad-GFP) was transfected into K562/ADM cells. These cells were then treated with different concentrations of adriamycin, cytarabine or arsenic trioxide, respectively. The half-maximal inhibitory concentration (IC(50)) of each drug was detected by MTT assay and the drug resistance reversal factor (RF) was calculated. The proliferation inhibition rate of these K562/ADM cells treated with or without the above-mentioned drugs was determined by MTT assay and the apoptosis rate was evaluated by flow cytometry. PTEN, nuclear factor-κB (NF-κB), I-κB, p53, multidrug resistance genes MDR1 and MRP, and apoptosis related genes Bcl-2, Bcl-xL and Bax mRNA levels were detected by real-time fluorescence relative-quantification reverse transcription polymerase chain reaction (FQ-PCR). PTEN, Akt, p-Akt and NF-κB (p65) protein levels were detected by Western blot. Results showed that PTEN gene transfection could increase the sensitivity of K562/ADM cells to chemotherapeutic drugs. The drug resistance reversal index of adriamycin, cytarabine and arsenic trioxide was 3.8-fold, 2.65-fold and 2.64-fold, respectively, after PTEN gene transfection. NF-κB, MDR1, Bcl-2 and Bcl-xL mRNA levels as well as p-Akt and NF-κB (p65) protein levels were down-regulated, while p53 and Bax mRNA levels were up-regulated in K562/ADM cells after transfection with Ad-PTEN-GFP. Therefore, wild-type PTEN gene transfection might increase drug sensitivity or reverse drug resistance via inhibiting the PI3K/Akt pathway and regulating downstream molecules of the cell signaling transduction pathway in K562/ADM cells, such as down-regulating NF-κB, MDR1, Bcl-2 expression but up-regulating the expression of p53 and Bax.

[PTEN: a new target in inhibiting of tumor invasion and metastasis].

Invasion and metastasis are both the main biological characteristics in malignant tumor which are influence tumor therapeutic effect and prognosis. The tumor cells interact with vascular endothelial cells and cell matrix, penetrate vascular endothelial and degrade the extracellular matrix, and metastasis to the local and distant by the interactions of a variety of signaling molecules. PTEN protein has protein phosphatase and lipid phosphatase dual activity which is produced by PTEN gene. As a tumor suppressor gene, regulates the cell signal pathways to sustain the normal physiological functions, negatively regulates of tumor cell growth and cell cycle, induces apoptosis, and inhibits invasion, infiltrating and metastasis of tumor cells. This article is reviewed about how PTEN participates in inhibiting tumor cell invasion and metastasis.

[Regulation of wild type PTEN gene on Survivin, Xiap and Smac in chronic leukemia cells].

OBJECTIVE: To explore the effects of tumor-suppressing gene wild type PTEN on the cell proliferation, apoptosis and the possible regulations of apoptosis-related molecules Survivin, Xiap and Smac gene in human chronic myeloid leukemia (CML) and cell line K562 cells. METHODS: (1) The recombinant adenovirus containing green fluorescent protein (GFP) and PTEN (Ad-PTEN-GFP) or empty vector (Ad-GFP) was transfected into K562 cells. The growth of K562 cells was observed by MTT assay while cell cycle and apoptotic rate were assessed by flow cytometry (FCM). PTEN, Survivin, Xiap and Smac mRNA levels were detected by real-time fluorescent relative-quantification reverse transcriptional PCR (FQ-PCR) while PTEN protein levels analyzed by Western blot. (2) The expression levels of PTEN, Survivin, Xiap and Smac mRNA were detected in 10 chronic myelogenous leukemia (CML) patients in chronic phase (CML-CP), 10 CML patients in blast crises (CML-BC) and 10 normal control marrow mononuclear cells (MMNC). RESULTS: The growth of K562 cells was suppressed markedly. And the maximal growth inhibition rate was 38.6% after the transfection of PTEN. Survivin, Xiap, Smac mRNA expression levels were down-regulated by around 6.14, 7.44 and 2.95 folds respectively (0.0700 ± 0.0059, 0.0089 ± 0.0006, 0.0600 ± 0.0039 vs 0.4370 ± 0.0790, 0.0661 ± 0.0072, 0.1580 ± 0.0078 vs 0.4530 ± 0.0810, 0.0700 ± 0.0079, 0.1770 ± 0.0085, all P < 0.01). The mRNA expression level of PTEN in CML-BC patients was lower than that in CML-CP patients and normal control. But Survivin, Xiap, Smac mRNA expression levels were higher in CML-BC patients than those in CML-CP and normal control. CONCLUSION: The over-expression of PTEN gene may inhibit the proliferation of K562 cells and promote cell apoptosis via the regulation of Survivin, Xiap and Smac genes.

[Effects of wild type PTEN gene on proliferation, apoptosis and the influence on apoptosis key factor Bcl-2 and Caspase family on K562 cells].

OBJECTIVE: To investigate the effect of tumor-suppressing gene,wild type PTEN gene, mediated by adenovirus vector on the cell proliferation, apoptosis and the influence on apoptosis key factor Bcl-2 and Caspase family on human chronic myeloid leukemia (CML) cell line K562 in vitro. METHODS: The recombinated Ad-PTEN gene containing green fluorescent protein gene (Ad-PTEN-GFP) or the empty vector (Ad-GFP) was transfected into K562 cells. The growth of K562 cells was evaluated by MTT assay; the transfection efficiency of Ad-PTEN-GFP, apoptosis rate and proliferation index (PI) were assessed by flow cytometry (FCM). Morphological characteristics of transfected cells under light and transmission electron microscope were applied to demonstrated the apoptosis; DNA ladder and fluorescent staining were also tested; the PTEN, Bcl-2 mRNA levels were detected by real-time fluorescent relative-quantification reverse transcriptional PCR (FQ-PCR); PTEN and Bcl-2 protein levels were detected by Western Blotting; and Caspase-3/7 and -9 protein activity were detected by corresponding kits. RESULTS: The 200 multiplicity of infection (MOI) of Ad-PTEN-GFP was applied to transfect K562 cells. The maximum growth inhibiting ratio was 37.1%. The early and advanced apoptosis rates were higher than Ad-GFP group and untransfected group (P<0.05). After 3 days transfaction of PTEN gene the Bcl-2 mRNA and protein were 0.27 fold and 0.58 fold respectively; and the Caspase-3/7 and -9 protein activity increased in time-depentend manner after transfected with PTEN gene at the first 3 days compared with Ad-GFP group and untransfected group. CONCLUSION: Over expression of PTEN gene can inhibit K562 cells proliferation and promote cell apoptosis probability via inhibiting Bcl-2 expression and up-regulating the Caspase-3/7 and -9 ability.