KLF2 Antibody - #DF13602

Fig. 6: TRβ modulates KLF2 and CEBPA to drive the expression of AT1 cell marker genes. a Principal component analysis (PCA) of TFs in bulk RNA-Seq of mouse lung (same as Fig. 3c, n = 3). The lower and upper bounds of the boxplot correspond to the first and third quartiles (the 25th and 75th percentiles); whiskers represent minima/maxima or 1.5*IQR. b Histogram showing the contributions of TFs in PC.2 and PC.5. c, d Protein and mRNA levels (n = 3) of KLF2 and CEBPA in the mouse lung, as shown in Fig. 3a. e qPCR analysis of KLF2 and CEBPA expression in isolated AT2 cells from mice (n = 3). f, g Immunoblotting and qPCR analysis (n = 3) of KLF2 and CEBPA in A549 and MLE12 with a GC-1 gradient. h The mRNA level of Klf2 and Cebpa in primary AT2 cells after 10 nM GC-1 treatment for 6 h (n = 3). i–l Protein (i, k) and mRNA (j, l) tests of AT1 cell markers, KLF2, and CEBPA expression in A549 after KLF2 (i, j) and CEBPA (k, l) overexpression (n = 3). m Luciferase activity of KLF2, CEBPA, and AT1-marker promoters cloned in pGL3.0 after THRB transfection for 36 h with or without GC-1 10 nM for 12 h (n = 3). Values were normalized to the transfection vector. n, o Luciferase activity of indicated promoters after KLF2 (n) and CEBPA (o) transfection with 600 ng plasmid for 36 h (n = 3). p–r TRβ (p), KLF2 (q), and CEBPA (r) bind to the promoter regions (pro) of KLF2, CEBPA, and AT1 cell genes in ChIP q-PCR assays (n = 3). KRT5 promoter as negative control. Values were normalized to IgG. s Luciferase activity of AT1-maker promoters with KLF2 (300 ng) and CEBPA (300 ng) co-transfection for 36 h, as shown in (n, o) (n = 3). t IF image of KLF2 and CEBPA after co-transfection in A549. Scale bar, 10 μm. u CoIP of KLF2 and CEBPA overexpressed in A549. v Illustration of the regulatory processes of GC-1 on AT1 cell markers. The value of n indicates biologically independent samples. Similar results were repeated in two biologically independent experiments. The statistical tests used were one-way ANOVA (d, e, and g), two-tailed unpaired Student’s t test (h), and two-way ANOVA (j, l, m–s). Data are mean ± SEM.

Figure 6 FGF2 reduces oxidative stress and ferroptosis via KLF2-NFE2L2 pathway in vivo. (A) Skeletal muscle specimens were used for the KLF2 and NFE2L2 WB assay. (B) Protein quantification of KLF2 and NFE2L2 from (A), normalised to GAPDH band density and histone 3 band density, respectively. (C) Skeletal muscle specimens showing HO-1, SOD1, SLC7A11, 4-HNE, and GPX4 on a WB assay. (D) Quantification of the GAPDH band density-normalized HO-1 and SOD1 protein levels from (C). (E) Quantification of protein level of SLC7A11, 4-HNE and GPX4 from (C) with normalized to GAPDH band density. (F) DAPI staining shows the nuclei in these micrographs of skeletal muscle slices that have been immunostained for KLF2 and CD31. Scale bars: 100 μm. (G) Micrographs of skeletal muscle slices with NFE2L2 and CD31 immunostaining; DAPI labelling shows nuclei. Scale bars: 100 μm. (H) Quantification of cells that co-express KLF2 and CD31; proportions of co-expressing cells to all CD31-positive cells are shown. (I) Quantification of NFE2L2 and CD31 double-positive cells, the ratio of co-expressing cells to all CD31-positive cells is shown. (J) Micrographs of skeletal muscle slices with nuclei identified by DAPI labelling and DHE and 4-HNE staining. Scale bars: 100 μm. Based on the immunofluorescence data in (J), the ROS level was quantified in (K). Based on the immunofluorescence results in (J), 4-HNE fluorescence intensity was quantified in (L). (M) Content of Fe2+ was plotted as a histogram. (N) The concentration of MDA is shown as a histogram. (O) The concentration of GSH is shown as a histogram. Data are provided as mean ± SD (n = 3-5 per group). Significance: ns, not significant; *P < 0.05.

Figure 3 KLF2 inhibits ccRCC migration in vitro and KLF2 protein expression is decreased in high pathological grade tissue (II + III). (A) Expression of KLF2 mRNA in ccRCC and normal HK-2 cells. (B) Western blotting was used for assessment of KLF2 protein levels in ccRCC and normal HK-2 cells. (C) Western blotting assessment of KLF2 OE in 769-P and 786-O cells. (D) Migration of 769-P and 786-O cells after wounding (magnification, x40). (E) compared with NC. (F) Immunohistochemistry and (G) analysis of the KLF2 expression in ccRCC tissue (n=20). **P<0.01 vs. stage I and ***P<0.001 vs. NC. KLF, Krüppel-like factor; ccRCC, clear cell renal cell carcinoma; NC, negative control; OE, overexpression.