RESUMO
OBJECTIVE: To determine the regulating role of phosphatidylinositol 3-kinase( PI3K) / protein kinase B( Akt)signaling pathway in the autophagy activity of rat NR8383 cells exposed to silicon dioxide( SiO_2). LY294002 was used to block PI3 K pathway. METHODS: i) The normal NR8383 cells were used and divided into blank group and silica exposure group( final concentrations of SiO_2 suspension were 0 and 50 mg / L respectively). They were cultured for 3,6,12,20 and24 hours. The enzyme linked immunosorbent assay( ELISA) was used to assess the amount of tumor necrosis factor-α( TNF-α) and transforming growth factor-β1( TGF-β1) in supernatants of cultured cells,and then the optimal time of cells exposed to dust was determined. ii) NR8383 cells were divided into control group( treated with a same volume of F-12 K medium without serum),silica group( treated with SiO_2 suspension,final concentration 50 mg / L) and intervention group( treated with SiO_2 suspension and PI3 K inhibitor LY294002,final concentration 50 mg / L and 20 μmol / L,respectively).Cells were harvested following incubation. ELISA was used to detect the levels of TNF-α and TGF-β1 at the time point of20 hours after incubation. To reveal the autophagy status of cells,Western blotting was used to detect Akt and microtubuleassociated proteins 1 light chain 3( LC3) protein at time point of 20 hours; laser scanning confocal microscope( LSCM)was used to observe the immunofluorescence expression of autophagy at time points of 3,6,12 and 20 hours. The cells were also treated with the lysosomal inhibitor chloroquine diphosphate( CDP) at the same time of SiO_2 treatment. RESULTS: i) The time point of 20 hours was confirmed to be the best dust exposure time for in vitro cell model of NR8383 cells.ii) The levels of TNF-α and TGF-β1 of supernatant in the silica group were higher than those of the control group( P <0. 05). The levels of TNF-α and TGF-β1 of supernatant in the intervention group were higher than those of the control group and silica group( P < 0. 05). The Akt protein expression of the intervention group was lower than those in the control group and the silica group,respectively. The LC3 Ⅱ / Ⅰ protein level of the silica group was higher than those of the control group and intervention group( P < 0. 05),but no statistical significance was found between the control group and intervention group( P > 0. 05). LSCM results indicated that autophagy expression at time points of 3 and 6 hours were stronger than those of 12 and 20 hours in control group; autophagy expression at time point of 12 hours was stronger than those of 3 and 6 hours in the silica group,while the autophagy expression at time point of 20 hours was slightly weaker than that of 12 hours,but still stronger than those of 3 and 6 hours. Compared with the same time point in control group,autophagy expression at 3 and 6 hours were weaker in the silica group,while the expressions increased obviously at time points of 12 and 20 hours. Autophagy expression at all time points decreased in the intervention group compared with silica group,especially at the time point of 20 hours. The autophagy expression in each group increased in varying degrees after added with CDP blocking. CONCLUSION: Silica dust exposure can induce autophagy in rat NR8383 cells. PI3 K inhibitor LY294002 can reduce the autophagy expression indicating that the PI3 K / Akt signaling pathway might participate in the autophagy process of silica dust inducing autophagy in alveolar macrophages.
RESUMO
<p><b>OBJECTIVE</b>To investigate the cerebral lesions of diffusion weighted imaging (DWI) hyperintensity in patients with subacute stroke with intravoxel incoherent motion (IVIM) technique.</p><p><b>METHODS</b>The clinical data of 20 patients with ischemic stroke (3 to 7 d after onset) who underwent DWI and IVIM scanning between June 2014 and July 2015, were retrospectively analyzed. The parameters from IVIM including slow diffusion coefficient (D), fast diffusion coefficient (D(*)) and perfusion fraction (f) were processed. DWI hyperintensity was segmented by its signal intensity greater than the mean+2 standard deviations of the value in the homologous contralateral region. Then, DWI hyperintensity was classified into two regions of interest (ROIs): infarction core and peri-core with the ADC threshold of 0.55 × 10⁻³ mm²/s. The mirrored ROIs of infarction core and peri-core were also obtained. Then, we measured the values of ADC and D, D(*) and f in these ROIs. The ratios of ADC (rADC), D (rD), D(*) (rD(*)) and f (rf) were also calculated (e.g., rADC=ADCinfarction core/ADCmirrored region).</p><p><b>RESULTS</b>Compared with mirrored region, ADC, D and f in the infarction core region decreased by 45% (P<0.001), 42% (P<0.001) and 32% (P<0.001), respectively; while ADC, D and f in the peri-core region decreased by 22% (P<0.001), 32% (P<0.001) and 8% (P=0.009), respectively. The values of rADC, rD, rD(*) and rf in the infarction core region were significantly lower than those in the peri-core region (all P<0.001). Pearson analysis showed that rADC was positively correlated with rf in the peri-core region (r=0.467, P=0.038).</p><p><b>CONCLUSION</b>During subacute stage of stroke, compared to the infarction core region within DWI hyperintensity, D and f increase in the peri-core region of DWI hyperintensity, reflecting the increased water diffusion in microstructure and perfusion volume in microvasculature. This result shows that the potential reason for the heterogeneous ADC signal is associated with the disappearance of cellular edema and microvascular compensatory with increased blood volume.</p>