The Radiosensitizing Effect of Resveratrol on Hopypharyngeal Carcinoma Cell Line FADU and its Effect on the Cell Cycle / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the radiosensitizing effect of resveratrol on hypopharyngeal carcinoma cell line FADU in vitro.</p><p><b>METHODS</b>Hypopharyngeal carcinoma cell line FADU was cultured in in vitro DMEM. Its inhibition on cell proliferation was detected using cytotoxicity test (MTT assay). The cell survival curve was drawn using clone formation to obtain sensitive enhancement ratio (SER). Changes of the cell cycle and cell apoptosis were analyzed using flow cytometry (FCM).</p><p><b>RESULTS</b>Results of MTT showed the inhibition of resveratrol on FADU cells increased along with its concentrations (P < 0.05). Results of clone formation indicated the surviving fraction at 2 Gy (SF2) was 0.717 ± 0.062 in the irradiation group, and 0.426 ± 0.035 in the resveratrol plus irradiation group (with SER ranged 1.684 ± 0.178) with statistical difference (P = 0.007). Results of FCM showed that after radiation of 4 Gy radiation, cells at G2/M phase arrest increased, but cells at G1 decreased. After radiation of resveratrol for 24 h, cells at G1 decreased, but cells at G2/M phase and S phase arrest increased. When 4 Gy radiation combined resveratrol was used, cells at G2/M phase arrest significantly increased, but cells at G1 significantly decreased. The apoptosis rate was 1.94% ± 1.65% in the control group, 4.56% ± 0.92% in the irradiation group, 2.03% ± 1.46% in the resveratrol group, and 23.11% ± 7.22% in the resveratrol plus irradiation group. There was statistical difference between the resveratrol plus irradiation group and the rest 3 groups (P < 0.05).</p><p><b>CONCLUSION</b>Resveratrol could enhance the radiosensitivity of hypopharyngeal carcinoma FADU cells in vitro possibly by inducing cell apoptosis and causing changes in the cell cycle distribution.</p>

Inhibitory effect of lentiviral vectors expressing small interfering RNA targeting survivin gene on Hep-2 cell growth in vitro and in nude mice / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the effect of the lentiviral vectors expressing small interfering RNA (siRNA) for survivin gene knockdown in inhibiting Hep-2 cell growth in vitro and its tumorigenicity in nude mice.</p><p><b>METHODS</b>The tumorigenicity of Hep-2 cells transfected with the siRNA mediated by the lentiviral vectors was tested in nude mice. The expression of survivin gene of the transfected cells at the mRNA and protein levels were detected by RT-PCR and Western blotting, respectively, and the cell cycle changes were analyzed by flow cytometry.</p><p><b>RESULTS</b>Transfection of the siRNA targeting survivin significantly decreased the expression of survivin mRNA and protein in Hep-2 cells in vitro by 60%-85% and 70%, respectively, resulting also in increased cell apoptosis as shown by flow cytometry (P<0.01). The transfection significantly lowered the tumorigenicity of the cells in nude mice.</p><p><b>CONCLUSION</b>The lentiviral vectors expressing survivin siRNA can significantly inhibit survivin gene expression in Hep-2 cells and induce the cell apoptosis in vitro, and suppress the tumorigenicity of the cells in nude mice.</p>

Effect of recombinant caspase-3 gene transfection on CNE2 cell apoptosis / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the apoptosis-inducing effect of caspase-3 on human nasopharyngeal carcinoma cells (CNE2).</p><p><b>METHODS</b>Recombinant caspase-3 was subcloned into the eukaryotic expression vector PEGFP-C1 containing the reporter gene using DNA recombinant technique. CNE2 cells were transfected with the recombinant caspase-3 gene via lipofectamine 2000 and the expression of caspase-3 mRNA was detected by reverse transcription-polymerase chain reaction. The cell morphological changes were observed under fluorescence microscope and electron microscope and the cell survival rate after the transfection was assessed by MTT assay.</p><p><b>RESULTS</b>Transfection with the recombinant caspase-3 gene induced significant apoptosis in CNE2 cells, which exhibited obvious morphological changes typical of apoptotic cells.</p><p><b>CONCLUSION</b>The recombinant caspase-3 gene can inhibit the growth and effectively induce apoptosis of CNE2 cells.</p>

Adenovirus-mediated antisense HSP70 cDNA transfection inhibits the growth of laryngeal carcinoma Hep-2 cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct a recombinant adenovirus vector carrying antisense heat shock protein 70 (HSP70) cDNA and observe its effect on inhibiting the growth of laryngeal carcinoma Hep-2 cells.</p><p><b>METHODS</b>The HSP70 gene fragment encoding the 5' region was cloned reversely into the shuttle plasmid PAdTrack-CMV, and the resultant plasmid was recombined with the backbone plasmid PadEasy-1 in the E.coli Bj5183 cells to generate the recombinant adenovirus vector. The adenovirus were then packaged and amplified in 293 cells, and the viral titer was determined using GFP.</p><p><b>RESULTS</b>The recombinant adenovirus vector carrying antisense HSP70 cDNA was constructed successfully with a viral titer of 8 x 10(9). HSP70 expression of Hep-2 cells was obviously blocked by antisense HSP70 RNA, and Western blotting and immuohistochemistry demonstrated that cells transfected with antisense HSP70 did not express or express HSP70 at low levels. Flow cytometry presented apoptotic peak in the antisense HSP70-transfected cells, but not in the control cells.</p><p><b>CONCLUSION</b>The recombinant adenovirus vector containing antisense HSP70 cDNA can effectively deliver antisense HSP70 gene into Hep-2 cells, suggesting the great potential of this gene therapy strategy in management of human laryngeal carcinoma.</p>

Protective effect of adeno-associated virus-mediated neurotrophin-3 on the cochlea of guinea pigs with gentamicin-induced hearing loss / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the protective effect of local gene therapy with adeno-associated virus (AAV)-mediated neurotrophin-3 (NT-3) on the function and morphology of the cochlea of guinea pigs with gentamicin-induced hearing loss.</p><p><b>METHODS</b>Hearing loss was induced with gentamicin (80 mg.kg(-1).day(-1) injected intramuscularly) in 18 pigmented guinea pigs 4 days prior to gene transfer. The guinea pigs were then divided into groups A, B, and C for AAV-mediated NT-3 gene transfer (n=7), AAV infection (n=7) or no particular intervention (n=4), respectively. Mini-Osmotic pump were implanted in either side of the ears in groups A and B, and the guinea pigs were injected with gentamicin (80 mg.kg(-1).day(-1)) intramuscularly since the operation day for 10 consecutive days. In group C, only gentamicin was administrated. Before and 14 days after gentamicin administration, auditory brainstem response audiometry (ABR) and distort-product otoacoustic emissions (DPOAE) were recorded, and the animals sacrificed to observe the morphological changes of the cochlear microscopically.</p><p><b>RESULTS</b>Compared with groups B and C, the animals in group A showed better auditory ability (ABR and DPOAE) and significantly higher surviving rate of the outer hair cells (P<0.05).</p><p><b>CONCLUSION</b>AAV-mediated NT-3 gene transfer may protect and repair the cochlear hair cells and auditory function damaged by aminoglycoside ototoxicity in guinea pigs, and aseptic procedure is of vital importance in cochlear local gene therapy.</p>