RESUMO
Objective To investigate the effects of aquaporin 3 (AQP3) and phospholipase D2 (PLD2) on the proliferation and apoptosis of a human cutaneous squamous cell carcinoma cell line A431.Methods Three small interfering RNAs (siRNAs) were constructed targeting the AQP3 and PLD2 genes separately,and transfected into A431 cells using liposomes.Then,fluorescence quantitative PCR was performed to find the most efficient siRNAs.Western blot was conducted to detect the protein expression levels of AQP3 and PLD2 in A431 cells after transfection with the selected AQP3-siRNA and PLD2-siRNA.Some A431 cells were divided into five groups:normal control group without any treatment,transfection reagent group treated with the oligofectamine reagent only,negative control group transfected with the negative control siRNA,AQP3-siRNA group transfected with the selected AQP3-siRNA,PLD2-siRNA group transfected with the selected PLD2-siRNA.After additional culture,cell counting kit-8 assay was performed to evaluate the proliferation of A431 cells,flow cytometry to detect the apoptosis of A431 cells after annexin V-fluorescein isocyanate/propidium iodide double-staining.Statistical analysis was carried out by the paired t test.Results The transfection with AQP3-siRNA and PLD2-siRNA induced a significant decrease in the mRNA and protein expressions of AQP3 and PLD2 respectively in A431 cells when compared with the untransfected cells.Compared with the negative control group,the proliferation of A431 cells was significantly decelerated at 24,48 and 72 hours after transfection in the AQP3-siRNA group (t =24.10,11.00,9.54,respectively,all P ＜ 0.01) and PLD2-siRNA group (t =30.47,7.02,8.73,respectively,all P ＜ 0.01).A significant increase was observed in the apoptosis of A431 cells at 48 and 72 hours after transfection with AQP3-siRNA (t =11.36,20.91,respectively,both P ＜ 0.01),and at 72 hours after transfection with PLD2-siRNA (t =4.86,P ＜ 0.05) compared with the negative control group.Conclusion The down-regulation of AQP3 and PLD2 expressions by siRNA can inhibit the proliferation,but induce the apoptosis,of A431 cells.
RESUMO
Objective To investigate the role of phosphorylated extracellular signal-regulated kinase (p-ERK), mitogen-activated protein kinase (MAPK)/ERK kinase (MEK), and nuclear factor-κB (NF-κB) in the pathogenesis of psoriasis vulgaris. Methods Immunohistochemistry and Western blot were used to detect the expression of p-MEK, p-ERK and p-NF-KB in tissue samples from 30 patients with psoriasis vulgaris and 15 normal human controls. The average optical density of immunostaining and relative grey scale of immuno-bloting were calculated. Results The average optical density of immunostaining for p-MEK, p-ERK and p-NF-KB was 0.36 ± 0.03, 0.36 ± 0.04 and 0.26 ± 0.04, respectively in lesion samples of psoriasis, significantly higher than that in normal control tissue (0.22 ± 0.02, 0.18 ± 0.03 and 0.16 ± 0.03, all P < 0.01). A significant increase was also observed in the relative grey scale of p-MEK, p-ERK and p-NF-κB in psoriatic lesions compared with the normal controls (1.41 ± 0.14 vs 0.54 ± 0.10, 2.35 ± 0.34 vs 1.86 ± 0.12, 1.07 ± 0.15 vs 0.87 ± 0.08, all P < 0.01). Conclusions The expressions of p-MEK, p-ERK and p-NF-κB are enhanced in lesions of psoriasis vulgaris, and the abnormal activation of upstream and downstream molecules in the MAPK signaling pathways might be involved in the pathogenesis of psoriasis.