Valproic acid regulates the miR-155/Jarid2 axis by affecting miR-155 promoter methylation in glioma.

The most frequent primary brain tumor in adults is glioma, yet no effective curative treatments are currently available. Our previous study demonstrated the enhancing effects of JARID2 on glioma sensitivity to TMZ treatment. In this study, miR-155 is predicted to target JARID2. miR-155 is overexpressed in clinical glioma specimens and cell lines. miR-155 overexpression in glioma cells enhances cell viability and represses cell apoptosis. Through targeting, miR-155 inhibits JARID2 expression. miR-155 inhibition inhibits glioma cell viability and enhances cell apoptosis, whereas JARID2 knockdown enhances cell viability and inhibits cell apoptosis; JARID2 knockdown partially reverses miR-155 inhibition effects on glioma phenotypes. miR-155 inhibition reduces but knockdown of JARID2 promotes the tumor formation ability of glioma cells in vivo. Valproic acid (VPA) upregulates JARID2 expression, inhibits glioma cell viability and enhances cell apoptosis. VPA downregulates the expression level of miR-155 by increasing the methylation level of the miR-155 promoter, suggesting that the miR-155/JARID2 axis is implicated in VPA inhibition of glioma cell viability and enhancement of glioma cell apoptosis. This study demonstrates a new mechanism of VPA treatment of gliomas by affecting the miR-155/JARID2 axis, which could be regarded as a new strategy for the prevention and treatment of glioma.

Outcomes and prognostic factors in patients with locally advanced cervical cancer treated with concurrent chemoradiotherapy.

BACKGROUND: To investigate the prognostic factors affecting long-term survival in locally advanced cervical cancer (LACC) patients treated with concurrent chemoradiotherapy (CCRT). METHODS: We retrospectively analyzed 192 naive LACC (stage IIB-IVA) patients who underwent intensity-modulated radiotherapy (IMRT) with concurrent platinum-based chemotherapy in Xiangya Hospital from January 2014 to June 2017. The clinicopathological factors of all patients were collected. To explore the relationship between factors and prognosis, survival rates were estimated by the Kaplan-Meier method. Univariate and multivariate Cox proportional hazards models were used to evaluate the effect of various factors on overall survival (OS) and progression-free survival (PFS). The nomogram and calibration curves were generated on the basis of survival analysis. RESULTS: The median follow-up time was 39.5 months. There-year rates of OS and PFS were 89.1% and 82.8%. LACC patients with non-squamous cell carcinoma [NSCC, including adenocarcinoma or adenosquamous carcinoma (AC/ASC)], advanced stage (IIIA-IVA), initially positive lymph node (pelvic or para-aortic lymph node, PLN/PALN), and a lower pretreatment hemoglobin (HGB) level (< 126 g/L) had lower survival rates. In univariate analysis, patients with NSCC, advanced stage, PLN or PALN metastasis had worse OS. Patients with NSCC, advanced stage, PLN or PALN metastasis, and a lower pretreatment HGB level had worse PFS. In multivariate analysis, NSCC and PALN metastasis were independent prognostic parameters of OS. NSCC, PALN metastasis and a lower pretreatment HGB level were independent prognostic parameters of PFS. CONCLUSIONS: NSCC and PALN metastasis were poor prognostic factors of OS and PFS, a lower pretreatment HGB level was an independent prognostic factor of PFS in LACC patients treated with CCRT.

[Retracted] MicroRNA­181 serves an oncogenic role in breast cancer via the inhibition of SPRY4.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that the Transwell cell migration assay data shown in Figs. 3B and 9B were strikingly similar to data appearing in different form in other articles by different authors. Owing to the fact that the contentious data in the above article had already been published elsewhere, or were already under consideration for publication, prior to its submission to Molecular Medicine Reports, the Editor has decided that this paper should be retracted from the Journal. After having been in contact with the authors, they agreed with the decision to retract the paper. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 18: 5603­5613, 2018; DOI: 10.3892/mmr.2018.9572].

LncRNA H19 Promotes Cell Proliferation, Migration, and Angiogenesis of Glioma by Regulating Wnt5a/ß-Catenin Pathway via Targeting miR-342.

Glioma is the most common malignant brain tumor and long non-coding RNAs (lncRNAs) have been reported to play an important role in the growth and angiogenesis of glioma. However, the potential mechanisms of lncRNA H19 in glioma remain unclear. In the present study, the effects of lncRNA H19 on glioma cell proliferation, migration, and angiogenesis were evaluated. The expression levels of H19, miR-342, and Wnt5a in glioma tissues and cells were detected by RT-qPCR or Western blotting. Dual luciferase reporter assay confirmed the interaction between H19, miR-342, and Wnt5a. Cell proliferation, migration, and angiogenesis were analyzed by colony formation, transwell, and tube formation assays, respectively. IHC was performed to test the angiogenesis-related factor CD31. H19 and Wnt5a expression were remarkably upregulated in glioma tissues and cells, whereas miR-342 expression was downregulated. Moreover, functional analysis confirmed that knockdown of H19 or overexpression of miR-342 suppressed glioma cell proliferation, migration, and angiogenesis in vitro. Besides, H19 was found to directly target miR-342 to promote Wnt5a expression and activate ß-catenin pathway in glioma cells. Moreover, suppression of miR-342 or overexpression of Wnt5a reversed the inhibitory effect of sh-H19 on glioma growth and metastasis. Additionally, we verified that H19 promoted glioma cell proliferation, migration, and angiogenesis via miR-342/Wnt5a/ß-catenin axis in vivo. H19 regulates glioma cell growth and metastasis through miR-342 to mediate Wnt5a/ß-catenin signaling pathway, which provides new therapeutic targets for glioma treatment.

The function and mechanism of the JARID2/CCND1 axis in modulating glioma cell growth and sensitivity to temozolomide (TMZ).

A maximal surgical resection followed by radiotherapy and chemotherapy with temozolomide (TMZ) as the representative agent is the standard therapy for gliomas. However, tumor cell resistance to radiotherapy and chemotherapy leads to poor prognosis and high mortality in patients with glioma. In the present study, we demonstrated that JARID2 was downregulated and CCND1 was upregulated within glioma tissues of different grades and glioma cells. In tissue samples, JARID2 was negatively correlated with CCND1. JARID2 overexpression significantly inhibited glioma cell viability, promoted glioma cell apoptosis upon TMZ treatment, and increased p21, cleaved-PARP, and cleaved-caspase3 in TMZ-treated glioma cells. JASPAR tool predicted the possible binding sites between JARID2 and CCND1 promoter regions; through direct binding to CCND1 promoter region, JARID2 negatively regulated CCND1 expression. Under TMZ treatment, JARID2 overexpression inhibited CCND1 expression, promoted glioma cell apoptosis, and increased p21, cleaved-PARP, and cleaved-caspase3 in glioma cells treated with TMZ; meanwhile, CCND1 overexpression exerted opposite effects on glioma cells treated with TMZ and partially reversed the effects of JARID2 overexpression. In conclusion, JARID2 targets and inhibits CCND1. The JARID2/CCND1 axis modulates glioma cell growth and glioma cell sensitivity to TMZ.

Multiple extracranial metastases from glioblastoma multiforme: a case report and literature review.

Extracranial metastasis from glioblastoma multiforme (GBM) is rare, especially multi-site metastases without intracranial recurrence. However, the metastatic mechanism of GBM remains unknown and there is currently no consensus regarding the best therapeutic regimen. We report the case of a 46-year-old man with primary GBM who developed scalp metastases and subsequent multiple pulmonary metastases. He was treated with the Stupp regimen after surgery for the intracranial tumor. However, a series of soft masses in the scalp were subsequently identified, and new nodules were found in his left eyebrow arch during chemoradiotherapy. Despite salvage chemotherapy and targeted therapy, the patient eventually died of respiratory failure with multiple pulmonary metastases. This case highlights the need for rigorous follow-up, including brain magnetic resonance imaging, in patients with GBM. The occurrence of extra-central nervous system symptoms indicates the possibility of metastasis, and the relevant examinations should be conducted promptly. Positive therapies may help to relieve symptoms and prolong survival in patients with metastatic GBM.

Downregulation of iASPP Expression Suppresses Proliferation, Invasion and Increases Chemosensitivity to Paclitaxel of Head and Neck Squamous Cell Carcinoma In Vitro.

Objective Our previous study has revealed that iASPP is elevated in human head and neck squamous cell carcinoma (HNSCC) and iASPP overexpression signifcantly correlates with tumor malignant progression and poor survival of HNSCC. This study investigated the function of iASPP playing in proliferation and invasion of HNSCC in vitro. Methods HNSCC cell line Tu686 transfected with Lentiviral vector-mediated iASPP-specific shRNA and control shRNA were named the shRNA-iASPP group and shRNA-NC group, respectively. The non-infected Tu686 cells were named the CON group. CCK-8 assay, flow cytometry, transwell invasion assay were performed to detect the effects of iASPP inhibition in vitro. Results Our results demonstrated that the proliferation of shRNA-iASPP cells at the time of 72 h (F=32.459, P=0.000), 96 h (F=51.407, P=0.000), 120 h (F=35.125, P=0.000) post-transfection, was significantly lower than that of shRNA-NC cells and CON cells. The apoptosis ratio of shRNA-iASPP cells was 9.42% ± 0.39% (F=299.490, P=0.000), which was significantly higher than that of CON cells (2.80% ± 0.42%) and shRNA-NC cells (3.18% ± 0.28%). The percentage of shRNA-iASPP cells in G0/G1 phase was 74.65% ± 1.09% (F=388.901, P=0.000), which was strikingly increased, compared with that of CON cells (55.19% ± 1.02%) and shRNA-NC cells (54.62% ± 0.88%). The number of invading cells was 56 ± 4 in the shRNA-iASPP group (F=84.965, P=0.000), which decreased significantly, compared with the CON group (111 ± 3) and the shRNA-NC group (105 ± 8). The survival rate of shRNA-iASPP cells administrated with paclitaxel was highly decreased, compared with CON cells and shRNA-NC cells (F=634.841, P=0.000). Conclusion These results suggest iASPP may play an important role in progression and aggressive behavior of HNSCC and may be an efficient chemotherapeutic target for the treatment of HNSCC.

Long non-coding RNA DLX6-AS1 silencing inhibits malignant phenotypes of gastric cancer cells.

It has been revealed that long non-coding RNAs (lncRNAs) serve a key role in various malignancies, including gastric cancer (GC). In the present study, the expression and function of lncRNA distal-less homeobox 6 antisense 1 (DLX6-AS1) in GC was investigated. The data revealed that the expression of DLX6-AS1 was significantly upregulated in GC tissues compared with adjacent paired noncancerous tissues. Furthermore, the expression of DLX6-AS1 was higher in advanced GC tissue samples (III/IV) compared with the expression in early-stage samples (I/II). Furthermore, the current study demonstrated that a high expression of DLX6-AS1 was significantly associated with advanced clinical stage, lymph node metastasis and distant metastasis. Compared with patients with a low DLX6-AS1 expression, DLX6-AS1 expression in patients with GC was associated with decreased survival. In vitro experimental data indicated that DLX6-AS1 was upregulated in GC cell lines and that the inhibition of DLX6-AS1 markedly reduced GC cell proliferation, colony formation, cell cycle progression, migration and invasion. Further investigation revealed that knockdown of DLX6-AS1 inhibited EMT in GC cells. In summary, the present study demonstrated that lncRNA DLX6-AS1 was upregulated and serves an oncogenic role in GC, indicating that DLX6-AS1 may be a novel therapeutic target for GC treatment.

MicroRNA­181 serves an oncogenic role in breast cancer via the inhibition of SPRY4.

Numerous microRNAs (miRs) have been implicated in breast cancer; however, the molecular mechanism is not fully understood. The present study examined the function and regulatory mechanism of miR­181 in breast cancer. Reverse transcription­quantitative polymerase chain reaction and western blot analysis were used to examine the RNA and protein expression. MTT assay, wound healing assay and transwell assay were conducted to study cell proliferation, migration and invasion. Luciferase reporter gene assay was used to confirm targeting relationship. The results suggested that the miR­181 expression levels were significantly higher in breast cancer cell lines and clinical tissue samples. The increased expression of miR­181 was markedly associated with higher clinical stage and lymph node metastasis. The patients with high miR­181 expression demonstrated worse prognosis compared with those with a low expression of miR­181. Small interfering RNA­induced miR­181 downregulation significantly inhibited breast cancer cell proliferation, migration and invasion in vitro, and tumor growth in vivo. Protein sprouty homolog 4 (SPRY4), downregulated in breast cancer tissues and cell lines, was observed to be a novel target gene of miR­181. Downregulation of SPRY4 was significantly associated with breast cancer progression in addition to poor prognosis. Knockdown of SPRY4 rescued the inhibitory effects of miR­181 downregulation on the malignant phenotypes of breast cancer cells. Thus, the present study demonstrated that miR­181 serves a promoting role in breast cancer at least in part through the inhibition of SPRY4 expression. The present results expand the understanding of the miR­181/SPRY4 axis' function during for the malignant progression of breast cancer.

TGF-ß1 secreted by M2 phenotype macrophages enhances the stemness and migration of glioma cells via the SMAD2/3 signalling pathway.

The positive correlation between the number of M2 phenotype TAMs (M2­TAMs) and tumour development suggests a supportive role of M2­TAMs in glioma progression. In the present study, the molecular link between glioma cells and M2­TAMs was investigated and it was demonstrated that transforming growth factor­ß1 (TGF­ß1) secreted by M2­TAMs is key in facilitating the stemness and migration of glioma cells. Cluster of differentiation (CD)133 and CD44, markers for the M2 phenotype, were assessed by western blotting. A sphere formation assay and trans­well assay were applied to test the stemness and migration abilities of glioma cells following co­cultured with M2­TAMs. Stemness markers CD133 and CD44, epithelial­mesenchymal transition­associated markers and mothers against decapentaplegic homolog (SMAD)2/3 and sex determining region Y­box 4/2 (SOX4/2) levels were also evaluated by western blotting. A xenograft tumor mouse model was used to demonstrate the tumor forming ability of glioma cells. The results showed that the U251 glioma cells co­cultured with M2­TAMs exhibited high level of sphere formation, stemness and migration ability. Recombinant TGF­ß1 protein treatment was able to achieve the same effects on U251 cells, whereas a TGF­ß pathway inhibitor reversed the stemness and migration abilities of the glioma cells induced by M2­TAMs. It was also demonstrated that TGF­ß1 secreted by M2­TAMs upregulated the phosphorylation of SMAD2/3 and the expression of SOX4/2 in glioma cells. In a mouse xenograft model, solid tumours formed by U251 cells co­cultured with M2­TAMs or pre­treated with TGF­ß1 were larger in size and had a higher growth rate. Taken together, results of the present study demonstrated that M2­TAMs promoted the stemness and migration abilities of glioma cells by secreting TGF­ß1, which activated the SMAD2/3 pathway and induced the expression of SOX4 and SOX2. These results highlight the mechanism by which M2­TAMs and glioma interact and demonstrate potential therapeutic strategies for glioma treatment.

A Study of 358 Cases of Locally Advanced Nasopharyngeal Carcinoma Receiving Intensity-Modulated Radiation Therapy: Improving the Seventh Edition of the American Joint Committee on Cancer T-Staging System.

To evaluate the rationality and limitations of the seventh edition of the American Joint Committee on Cancer (the 7th AJCC edition) T-staging system for locally advanced nasopharyngeal carcinoma (NPC). The prognosis of 358 patients with stage T3/T4 NPC treated with intensity-modulated radiotherapy (IMRT) was analyzed with the Kaplan-Meier method or the log-rank test. The 7th AJCC staging system of NPC has some limitations in that the T category is neither the significant factor in OS/LRFS nor the independent prognostic factor in OS/LRFS/DMFS/DFS (P > 0.05). After adjustment by anatomic structures, univariate analysis has shown that the adjusted-T category has statistical significance between T3 and T4 for OS (86.4% and 71.3%, P = 0.002), LRFS (97% and 90.9%, P = 0.048), DMFS (90.9% and 77.2%, P = 0.001), and DFS (86.2% and 67.5%, P = 0.000), and multivariate analysis has shown that the adjusted-T category is an independent prognostic factor for OS/DMFS/DFS (with the exception of LRFS). Then, GTV-P was taken into consideration. Multivariate analysis showed that these nT categories serve as suitable independent prognostic factors for OS/DMFS/DFS (P < 0.001) and LRFS (HR = 3.131; 95% CI, 1.090-8.990; P = 0.043). The 7th AJCC staging system has limitations and should be improved by including the modifications suggested, such as anatomic structures and tumor volume adjustment.

Aberrant expression of osteopontin and E-cadherin indicates radiation resistance and poor prognosis for patients with cervical carcinoma.

Radiotherapy is the first-line treatment for all stages of cervical cancer, whether it is used for radical or palliative therapy. However, radioresistance of cervical cancer remains a major therapeutic problem. Consequently, we explored if E-cadherin (a marker of epithelial-mesenchymal transition) and osteopontin could predict radioresistance in patients with locally advanced cervical squamous cell carcinoma (LACSCC). Patients were retrospectively reviewed and 111 patients divided into two groups (radiation-resistant and radiation-sensitive groups) according to progression-free survival (PFS). In pretreated paraffin-embedded tissues, we evaluated E-cadherin and osteopontin expression using immunohistochemical staining. The percentage of patients with high osteopontin but low E-cadherin expression in the radiation-resistant group was significantly higher than those in the radiation-sensitive group (p<0.001). These patients also had a lower 5-year PFS rate (p<0.001). Our research suggests that high osteopontin but low E-cadherin expression can be considered as a negative, independent prognostic factor in patients with LACSCC ([Hazard ratios (95% CI) 6.766 (2.940, 15.572)], p<0.001).

[Dosimetric comparison of left-side whole breast irradiation with IMRT and hybrid IMRT].

OBJECTIVE: To evaluate the potential dosimetric benefits and optimal indications of intensity modulated radiation therapy (IMRT) and hybrid intensity modulated radiation therapy (Hybrid IMRT) for the left side breast cancer patients after breast-conservation therapy. METHODS: Eight patients with left breast carcinoma who received breast-conservation surgery were selected for this study. Two plans were designed in 3-dimensional treatment planning system. The dose distributions of target volume and normal tissues, conformal index (CI) and heterogeneous index (HI) were analyzed by dose-volume histogram (DVH). RESULTS: The PTV coverage was the same in the two radiotherapy plans. A better dose uniformity throughout the whole breast in Hybrid IMRT plan was achieved. The CI, the percentage of PTV receiving more than 105% prescribed dose (V105%), the percentage of PTV receiving more than 110% prescribed dose (V110%), and the Dmax, Dmin and Dmean of PTV were similar in the two plans. We compared the Hybrid IMRT with IMRT: V13 of the ipsilateral lung decreased from 27.66% to 20.7%, V5 of the contralateral lung decreased from 8.01% to 2.25%, V10 and V20 of the heart decreased from 35.23% and 16.77% to 19.22% and 10.6% respectively, V5 and V10 of the contralateral breast decreased from 35% and 10.39% to 20.38% and 5.7% respectively, all with significant difference. V30 and V40 of the ipsilateral lung and V40 of the heart increased by 1.28%, 1.48%, and 2.48%, with significant difference. CONCLUSION: Hybrid IMRT is a better choice for patients whose treatment position is inaccurate or cannot be repeated well.

[Inhibitive effect of E1A gene on the cell growth of human cervical carcinoma cell in vitro].

OBJECTIVE: To investigate the inhibitive effect of E1A gene carried by PEI-Fe(3)O(4) nanometer particle (NP) on the cell growth of human cervical carcinoma cell in vitro and its mechanism, and to provide the experimental evidence for the feasibility of gene therapy for human cervical carcinoma. METHODS: E1A gene conjugated to PEI-Fe3O4 NP was transfected into human cervical carcinoma cell line Hela. The cell growth curve of Hela was drawn, the doubling time and the number of colony formations on the soft agar were calculated based on the cell count. RT-PCR and Western blot were used to detect the expression of the E1A and HER-2/neu in Hela cells. RESULTS: The cell doubling time of Hela cells transfected with E1A gene (Hela-E1A) was 1.53 times and 1.58 times longer than that of the Hela transfected with blank vector (Hela-vector) and blank Hela control (Hela), respectively. The E1A transfected Hela cells grew slower than those of the control group. The cell colony formation efficiency in the Hela-E1A (6.62%) group was significantly lower than that of Hela (30.48%) and Hela-vector (28.3%) groups (P<0.05). As compared to Hela and Hela-vector, the inhibition rate of Hela-E1A was 78.28% and 76.62% respectively. RT-PCR and Western blot demonstrated that the overexpression of E1A through gene transfection significantly inhibited mRNA and protein expression of HER-2/neu in Hela cells. CONCLUSION: E1A gene can suppress the cell growth of human cervical carcinoma cell Hela in vitro. Down-regulated expression of HER-2/neu gene by E1A overexpression in Hela might contribute to the Hela growth inhibitive effect of E1A.