Role and action mechanism of aquaporin 3 in alleviating photoaging of skin fibroblasts by regulating hnRNPQ/p53
10.35541/cjd.20200253
- VernacularTitle:水通道蛋白3通过调控hnRNPQ/p53延缓皮肤成纤维细胞光老化的作用及机制研究
- Author:
Huaxiong ZHANG
;
Sha YAN
;
Lin HE
;
Lin LI
;
Zhaohui CHEN
;
Ji LI
- From:
Chinese Journal of Dermatology
2021;54(4):325-334
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role and action mechanism of aquaporin 3 (AQP3) in skin photoaging.Methods:Normal human skin fibroblasts (NHDF) were divided into several groups: NHDF group receiving normal culture without transfection, AQP3 cDNA group transfected with AQP3 cDNA, AQP3 siRNA group transfected with AQP3 siRNA, heterogeneous nuclear ribonucleoprotein Q (hnRNPQ) cDNA group transfected with hnRNPQ cDNA, hnRNPQ siRNA group transfected with hnRNPQ siRNA, AQP3-hnRNPQ cDNA group transfected with AQP3 and hnRNPQ cDNAs, AQP3-hnRNPQ siRNA group transfected with AQP3 and hRNPQ siRNAs, cDNA empty vector group transfected with a cDNA empty vector, and siRNA empty vector group transfected with a siRNA empty vector. Transfected or untransfected NHDFs were irradiated with ultraviolet A (UVA) at a dose of 10 J·cm -2·d -1 for 3 consecutive days to establish a model of cellular senescence, and NHDF receiving no UVA irradiation served as a control. A cell counting method was used to evaluate the cellular proliferative activity, a senescence-related β-galactosidase staining kit to evaluate the senescence level of NHDFs in each experimental group, and luciferase reporter gene technology to assess the transcriptional regulation activity of p53. Western blot analysis was performed to determine the expression of AQP3, hnRNPQ and senescence-related proteins p53 and p21 in NHDFs. Two-independent-sample t test was used for comparisons between two groups, and one-way analysis of variance for comparisons among multiple groups. Results:After 3-day consecutive irradiation with UVA, the expression of p53 and p21 in NHDFs and the percentage of β-galactosidase-positive cells significantly increased compared with the unirradiated control group (all P < 0.05) , but the expression of AQP3 and cellular proliferative activity on days 5, 6 and 7 significantly decreased in the UVA group compared with the unirradiated control group (all P < 0.05) . After 3-day consecutive irradiation with UVA, aggravated senescence-related phenotypes of UVA-induced NHDFs were observed in the AQP3 siRNA group compared with the siRNA empty vector group, and there were significant differences in the expression of p53, p21 and hnRNPQ, percentage of β-galactosidase-positive cells, p53 transcriptional regulation activity and cellular proliferative activity between the 2 groups (all P < 0.05) . Further silencing of the hnRNPQ gene could reverse the above effects. Compared with the siRNA empty vector group, the senescence-related phenotypes of UVA-induced NHDFs were attenuated in the hnRNPQ siRNA group, and significant differences were observed between the 2 groups in terms of the expression of p53, p21 and hnRNPQ, percentage of β-galactosidase-positive cells, p53 transcriptional regulation activity and cellular proliferative activity (all P < 0.05) . After 3-day consecutive irradiation with UVA, the senescence-related phenotypes of UVA-induced NHDFs were significantly attenuated in the AQP3 cDNA group compared with the cDNA empty vector group (all P < 0.05) , manifesting as significantly decreased expression of p53 (0.25 ± 0.06 vs. 0.56 ± 0.08) , p21 (0.23 ± 0.06 vs. 0.70 ± 0.07) and hnRNPQ (0.82 ± 0.09 vs. 0.92 ± 0.03) , percentage of β-galactosidase-positive cells (31.23% ± 6.54% vs. 81.53% ± 7.62%) and p53 transcriptional regulation activity (2.52 ± 0.36 vs. 7.16 ± 0.25) , but increased cellular proliferative activity ([2.93 ± 0.33]× 10 6/ml vs.[2.15 ± 0.23]× 10 6/ml) , and further overexpression of hnRNPQ could reverse the above effects. After 3-day consecutive irradiation with UVA, the expression of p53, p21, percentage of β-galactosidase-positive cells, p53 transcriptional regulation activity and cellular proliferative activity in the hnRNPQ cDNA group were 1.41 ± 0.09, 1.42 ± 0.06, 91.06% ± 4.24%, 12.35 ± 0.88 and (1.23 ± 0.41) × 10 6/ml respectively, and the senescence-related phenotypes of UVA-induced NHDFs were significantly aggravated in the hnRNPQ cDNA group compared with the cDNA empty vector group (all P < 0.05) . Conclusion:AQP3 may alleviate the UVA-induced senescence of NHDFs by regulating hnRNPQ and downregulating p53 expression.
