Lentivirus-mediated SMO RNA interference inhibits SMO expression and cell proliferation,and affects the cell cycle in LNCaP and PC3 cancer cell lines

Smoothened （SMO） is an important member of the Hedgehog signaling pathway. We constructed a specific recombinant lentiviral vector for RNA interference,targeting the SMO gene （NM_005631） to observe its effect on SMO expression,cell proliferation and the cell cycle in the human androgen-sensitive prostate cancer cell line,LNCaP,and in the androgen-independent prostate cancer cell line,PC3. Four siRNA sequences were designed and inserted into a lentiviral vector pGCSIL-GFP to construct four recombinant vectors. The vector with the highest interfering efficiency was co-transfected with packaging vectors （pHelper1.0 and pHelper2.0） in 293T cells to assemble lentivirus particles by liposome for infecting LNCaP and PC3 cell lines,respectively. The expression level of SMO mRNA,tumor cell proliferation and cell cycle were measured by quantitative realtime polymerase chain reaction （qRT-PCR）,3-（4,5）-dimethylthiahiazo （-z-yl）-3,5-di-phenytetrazoliumromide （MTT） assay and flow eytometry,respectively. Sequence results showed that recombinant lentiviral vectors were constructed successfully.pGCSIL-GFP-723 had the highest interfering efficiency,named Lv-SIL-SMO723 after co-transfection,with which LNCaP and PC3 cell lines were infected. Compared with the control groups,results showed significantly decreased （P〈0.05） SMO mRNA expressions of LNCaP and PC3,lower mean percentage of S-phase cells and higher mean percentage of G_2/M phase cells,as well as obviously slow proliferation （P〈0.01） of LNCaP in the infected group. Yet,the proliferation of PC3 was not altered （P〉0.05）. In conclusion,the recombinant lentivirus particles were able to suppress SMO expression,regulate the cell cycle in the LNCaP and PC3 cell lines and markedly inhibit proliferation of LNCaP cells but not PC3 cells.

Respiratory face mask:a novel and cost-effective device for use during the application of myocardial ischemia in rats

To shorten operation time and improve survival rate of rats with myocardial ischemia or myocardial infarction,we use a novel device comprised of a face mask and a head/neck retainer in this study.We report the basic design of the novel respiratory face mask(RFM) and evaluate its performance in a rat model of myocardial ischemia.The device is cost-effective and easier to handle than other devices,such as tracheal intubation.Compared with conventional tracheal intubation,we found that RFM shortens operation time significantly while keeping blood indices normal;the mean operation time for rats in the mask group was(32±3) min,and that for the intubation group was(45±7) min(P<0.05).Moreover,the size and shape of the RFM can be changed according to the body weight of rats.In conclusion,RFM is an appropriate device for the establishment of myocardial infarction or ischemia-reperfusion in rats.