N7-methylguanosine-related gene decapping scavenger enzymes as a novel biomarker regulating epithelial cell function in diabetic foot ulcers

Figure 7 Assessment of normal epidermal keratinocyte function following decapping scavenger enzyme knockdown in vitro. Cells were treated with 30 mmol/L glucose for 48 hours before being exposed to siDCPS [small interfering RNA targeting decapping scavenger enzyme (DCPS)] for 24 hours. A: Dot blot assay showed downregulation of N7-methylguanosine under diabetic condition; B: Efficient DCPS knockdown by small interfering RNA (n = 4). Evaluate cell cycle distribution changes induced by DCPS knockdown; C and D: DCPS knockdown induced significant G2 phase arrest in cell cycle, n = 3; E-G: DCPS knockdown reduced cyclin-dependent kinase 6 and cyclin D1 protein expression, n = 3; H and I: DCPS knockdown compromises RNA stability of cyclin D1 and cyclin-dependent kinase 6 transcripts. Assess the impact of DCPS knockdown on cellular proliferation; J and K: 5-ethynyl-2’-deoxyuridine staining and statistical analyses in normal epidermal keratinocytes (NHEKs), n = 4 (scale bar: 100 μm); L and M: DCPS knockdown attenuated cellular proliferation in flow cytometry assays, n = 3; N and O: DCPS knockdown enhanced apoptosis in flow cytometry, n = 3. DCPS knockdown inhibits NHEK migration; P and Q: Transwell assays were conducted for evaluation of NHEK migration, n = 3 (scale bar: 100 μm); R-U: Cell scratch assays were observed in living cells at 0, 12, 24 and 48 hours, n = 3 (scale bar: 200 μm). The results were expressed as mean ± SD. NC: Negative control; HG: High glucose; m7G: N7-methylguanosine; siDCPS: Small interfering RNA targeting decapping scavenger enzyme; RMSD: Root mean square deviation; CDK6: Cyclin-dependent kinase 6; 7-AAD: 7-Aminoactinomycin D. ^aP < 0.001, ^bP < 0.01, and ^cP < 0.05.