Abnormal expression and potential clinical value of oncogenic Krüppel-like factor-5 in lung squamous cell carcinoma

Figure 1 Bioinformatics analysis of lung Krüppel-like factor-5 mRNA and its function. A: KLF5 mRNA in lung adenocarcinoma (LUAD) (n = 585) or lung squamous-cell carcinoma (LUSC) (n = 550) tissues from the Cancer Genome Atlas (TCGA). database; B: KLF5 mRNA in LUSC or normal tissues from the Gene Expression Profiling Interactive Analysis (GEPIA) (Left) or TCGA (Right) databases; C: KLF5 mRNA in LUAD or normal tissues from the GEPIA (Left) or TCGA (Right) databases; D: Biological process of KLF5; E: Molecular function of KLF5; F: KEGG analysis of KLF5. KLF5: Krüppel-like factor 5 LUAD: Lung adenocarcinoma; LUSC: Lung squamous-cell carcinoma.

Figure 2 Krüppel-like factor-5 expression in the lung tissues or sera of patients. A-C: Krüppel-like factor-5 (KLF5) in lung squamous-cell carcinoma (LUSC): B: KLF5 in Adj-Ca tissues of LUSC; C: KLF5 ng/mg wet lung tissues (n = 60), ^bP < 0.01; D-F: KLF5 in lung adenocarcinoma (LUAD): E: KLF5 in Adj-Ca tissues of LUAD; F: KLF5 ng/mg wet lung tissues (n = 60); G: The positive rates of serum KLF5 (> 800 ng/mL) in LUSC, chronic pneumonia and normal control, ^fP < 0.01; H: Receiver operating characteristic curve of circulating KLF5 Levels for LUSC diagnosis. Adj-Ca: Adjacent cancerous tissues; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; KLF5: Krüppel-like factor 5; LUAD: Lung adenocarcinoma; LUSC: Lung squamous-cell carcinoma; MIF: Multiplex immunofluorescence; LC: Lung cancer; CP: Chronic pneumonia; NC: Normal control; ROC: Receiver operating characteristic.

Figure 3 Intervening KLF5 affected the biological behaviors of lung cancer cells. A: Lung Beas-2B cells and lung cancer SPC-A-1, NCI-1650, A549 and NCI-1975 cells; B: Krüppel-like factor 5 (KLF5) at the protein level in different cells, as determined by Western blotting; C: Ratios of KLF5 to glyceraldehyde-3-phosphate dehydrogenase; D: KLF5 at the mRNA level in different cells, as determined by quantitative real-time PCR (qRT-PCR); E: Proliferating curve of A549 cells divided into blank, N-KLF5 or KLF5 intervention (KLF5i) groups and transfected with KLF5i plasmids; F: Alterations in KLF5 mRNA by qRT-PCR; G: Migrating analysis of A549 cells among different groups; H: Cell cycle alterations. GAPDH: Glyceraldehyde-3- phosphate dehydrogenase. ^aP < 0.05 and ^cP < 0.001, compared with the blank or N-KLF5 group. KLF5: Krüppel-like factor 5.

Figure 4 Interfering with KLF5 transcription affected xenograft tumor growth. A: Nude mouse xenograft tumor growth after subcutaneous injection of A549 cells transfected with or without Krüppel-like factor 5 intervention (KLF5i) plasmids; B: Xenograft tumor weight (g); C: Xenograft tumor volume; D: Krüppel-like factor 5 (KLF5) straining by IHC in the N-KLF5 group (n = 6); E: KLF5 straining by IHC in the KLF5i group (n = 6); F: KLF5 straining by MIF in the N-KLF5 group; G: KLF5 straining by MIF in the KLF5i group; H: KLF5 in xenograft tumors by Western blotting (top) and their ratios from KLF5i to GAPDH (bottom). ^cP < 0.001, compared with the blank or N-KLF5 group. GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; IHC: Immunohisto- chemistry; KLF5: Krüppel-like factor 5; MIF: Multiplex immunofluorescence.

Figure 5 Krüppel-like factor 5-related pathways and possible mechanisms in lung cancer. A-F: Biocarta analysis: A: WNT_pathway; B: MAPK_pathway; C: FAS_pathway; D: TGFB_pathway; E: AMI_pathway; F: Intrinsic_ pathway; G-L: GO analysis: G: Cytokines; H: Execution_phase_ of_apoptosis; I: Protein_acetylation; J: Regulation of RNA splicing; K: Bicellular tight junction assembly; L: MRNA splicing by splicing exosomes; M: A possible new mechanism is that abnormal transcription of Krüppel-like factor 5 promotes lung squamous-cell carcinoma progression by activating related signaling pathways (such as Wnt and so on). KLF5: Krüppel-like factor 5; Wnt3a: A member of the Wnt signaling pathway.