Long noncoding RNA AFAP1­AS1 enhances cell proliferation and invasion in osteosarcoma through regulating miR­4695­5p/TCF4­ß­catenin signaling.

Long noncoding RNA AFAP1­AS1 has been shown to promote tumor progression in several human cancer types, such as thyroid cancer, tongue squamous cell carcinoma and lung cancer. However, the role of AFAP1­AS1 in osteosarcoma (OS) has not been investigated. In the present study, the expression of AFAP1­AS1 was significantly upregulated in OS tissues and cell lines. Moreover, AFAP1­AS1 expression was negatively correlated with OS patient prognosis. Besides, AFAP1­AS1 knockdown significantly inhibited the proliferation and invasion of OS cells in vitro. Furthermore, in vivo xenograft experiments indicated that AFAP1­AS1 depletion delayed tumor growth. Regarding the underlying mechanism, AFAP1­AS1 served as a sponge to repress the level of microRNA (miR)­4695­5p, which targeted transcription factor (TCF)4, a pivot effector of Wnt/ß­catenin signaling pathway. It was demonstrated that overexpression of AFAP1­AS1 inhibited the expression of miR­4695­5p, while miR­4695­5p overexpression decreased TCF4 expression and reduced activation of Wnt/ß­catenin pathway. Through rescue assays, it was demonstrated that restoration of TCF4 expression reversed the effects of AFAP1­AS1 knockdown or miR­4695­5p overexpression on OS cells. Taken together, these findings demonstrated that the AFAP1­AS1/miR­4695­5p/TCF4­ß­catenin axis played an important role in OS progression.

Targeted inhibition of mammalian target of rapamycin (mTOR) signaling pathway inhibits proliferation and induces apoptosis of laryngeal carcinoma cells in vitro.

AIM AND OBJECTIVE: Laryngeal carcinoma is one of the most aggressive cancers of the head and neck region. The survival rate of patients with laryngeal carcinoma is low due to its late metastases and resistance to chemotherapy and radiotherapy. It was reported that mTOR was involved in the growth and apoptosis of various cancer cells. The aim of this study was to detect the effects of mTOR inhibition by mTOR shRNA on the proliferation, apoptosis and invasive ability of Hep-2 human laryngeal carcinoma cells in vitro. METHODS AND STUDY DESIGN: mTOR shRNA was designed and transfected into Hep-2 human laryngeal carcinoma cells. Untreated cells and cells treated with control vector (non-targeted shRNA) were used as control. The proliferation and apoptosis of Hep-2 cells were detected by MTT and flow cytometry. A transwell assay was used to measure the invasive ability of Hep-2. The inhibition effects on the mTOR signaling pathway by mTOR shRNA were studied using RT-PCR and Western blot. RESULTS: Our results showed that the mRNA and protein expression of mTOR and Akt were high in laryngeal carcinoma cells and could be inhibited by mTOR shRNA. At the same time, low expression of PTEN mRNA and protein was observed in Hep-2 cells. The expression increased when the cells were transfected with mTOR shRNA. This showed that mTOR shRNA could inhibit the proliferation and invasive ability of Hep-2 cells. It also could induce the apoptosis of Hep-2 cells in vitro. CONCLUSIONS: The mTOR signaling pathway plays an important role in the development of laryngeal carcinoma. The mTOR shRNA we designed in this experiment effectively inhibited the mTOR signaling pathway. It inhibited the proliferation and invasive ability of the studied laryngeal carcinoma cells and induced their apoptosis in vitro. mTOR might therefore be a useful target in the therapy of laryngeal carcinoma.

[Experimental study on transplantation of embryonic stem cells in treating spinal cord injury].

OBJECTIVE: To observe the effect of transplantation of embryonic stem cell (ES) on neurological functional recovery of injured spinal cord in adult mouse. METHODS: The ES cells were cultured and induced in vitro. Fifty C57/BL6J mice were made animal model of semi-cut mice of T9,10. The ES cell-derived neural precursors cells were transplanted into the vertebral canal around injured spinal cord semi-cut mice. Twenty-eight C57/BL6J mice were randomly divided into three groups: sham operation group(group A, n=9), operation/cell group (group B, n = 10), and operation/DMEM group (group C, n = 9). RT-PCR analysis, X-gal staining and immunofluorescence were used to observe the cells survival and differentiation in the spinal crod. BBB test was performed to study functional improvement. RESULTS: ES cells induced and cultured in vitro displayed clonal growth with circle or ovoid shape and had one or more nucleoli. RT-PCR result showed that the induced ES cells expressed mRNA of Nestin and microtubule-associated protein, but did not express glial fibrillory acidic protein (GFAP). There was statistically significant difference in BBB scoring between group A and groups B, C after operation (P <0.01). There was statistically significant difference in BBB scoring at 1, 2 and 4 weeks of operation (P < 0.01), but no statistically significant difference at 6 and 8 weeks of operation between groups B and C (P>0.05). The X-gal staining results were positive in group B and negative in groups A and C. The immunoflurescence result showed neurofilament green fluor and no expression of GFAP in injured spinal cord region. CONCLUSION: After transplantation, ES cell-derived cells can survive, transfer into the injury position, and differentiate into neurons, but spinal cord function has no obvious improvement.