Fig. 5 | Scientific Reports

Fig. 5

From: Role of B1 antisense RNA on the proliferation and killing tumor ability of aged mouse spleen lymphocytes

Fig. 5The alternative text for this image may have been generated using AI.

Effects of B1 asRNA on ZFP92 protein expression. (A) Representative images of ZFP92 IF microscopy (magnification 25 × 10). The fluorescence intensity of ZFP92 in B1asRNA culture group was higher than that in the PHA culture group and siZFP92#3 significantly reduced the expression level of ZFP92 protein induced by B1asRNA. (B) The fluorescence intensity of each group was calculated by Image J software. The fluorescence intensity of ZFP92 in B1 asRNA group was significantly higher than that in PHA group (mean ± SD of three independent experiments). **P < 0.01. (C) Staining of euchromatin and heterochromatin regions in the nucleus of lymphocytes observed by laser confocal microscopy (magnification 100 × 10). The DAPI-labeled nuclear regions and FITC-labeled ZFP92 protein regions did not overlap. ZFP92 protein fluorescence in large lymphoblastoid cells was stronger than that in small lymphocytes, but its DAPI staining was lighter than in small lymphocytes. DAPI: Results of DAPI staining of nuclei; ZFP92: ZFP92 staining results; Merge: Results of overlapping DAPI and ZFP92 stains; White: White light. The arrow indicates the large lymphocyte. (D) CCK8 assay showed that the cell proliferation activity significantly decreased when the lymphocytes were treated with ZFP92 siRNA (mean ± SD of three independent experiments). *P < 0.05. siZFP92#3 had the strongest effect in reducing the proliferation activity of lymphocytes. Therefore, siZFP92#3 was used in the experiment shown in (A), and it was also continued to be used in the experiments of (E). (E) LDH release assay shows that siZFP92#3 significantly decreased the ability of killing S180 cells of lymphocytes (means of three independent experiments, statistical differences were calculated using one‑way ANOVA with Bonferroni). *P < 0.05. (F) Effects of mouse B1 asRNA on DNA expression of transcription factors in spleen lymphocytes. ChIP-qPCR was used to detect the effect of B1 asRNA treatment on the binding ability of ZFP92 protein to transcription factors (Nanog, Oct4, Sox2, Klf4, c-Myc) in spleen lymphocytes from aged mice. Nanog, Oct4, Sox2, Klf4 and Myc DNA enrichment efficiency of ZFP92 protein in B1 asRNA treated group was higher than that in conventional culture group (PHA group) (mean ±SD of three independent experiments). *P < 0.05, **P < 0.01.

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