In this study, we aimed to investigate the molecular mechanism of Krüppel-like factor 7 (KLF7) in colorectal cancer (CRC) cell invasion and migration. The expression pattern of KLF7 in CRC tissues and the correlation between KLF7 expression and clinical symptoms of CRC were analyzed. CRC cell lines were transfected with si-KLF7, followed by qRT-PCR or western blot detection of KLF7, miR-139-5p, and tumor protein D52 (TPD52) expression, cell counting kit-8 (CCK-8) assay to detect cell viability, and transwell detection of invasion and migration. Chromatin immunoprecipitation (ChIP) analyzed the enrichment KLF7 in the miR-139-5p promoter. The dual-luciferase reporter assay verified the binding relationship between KLF7 and miR-139-5p, and between miR-139-5p and TPD52. In the subcutaneous tumorigenesis experiment, tumor growth was observed and ki67-positive expression was detected. KLF7 is abundantly expressed in CRC cells KLF7 silencing inhibits CRC cell viability, invasion, and migration. KLF7 represses miR-139-5p expression by binding to the miR-139-5p promoter. miR-139-5p targets TPD52 expression. miR-13-5p inhibition or TPD52 overexpression partially counteracted the effect of KLF7 silencing in CRC cells. KLF7 silencing suppresses tumor growth in vivo. In conclusion, KLF7 suppresses miR-139-5p expression by binding to the miR-139-5p promoter, thereby upregulating TPD52 expression and enhancing CRC cell invasion and migration.

BACKGROUND: The TGF-β gene is a gemcitabine (GEM) resistance gene; however, the mechanism by which it regulates GEM resistance in pancreatic cancer remains unclear.
METHODS: The PANC-1 cell line was treated with GEM and then stimulated with TGF-β. Subsequently, we constructed GEM-resistant pancreatic cancer cell lines, knocked down TGF-β in these cell lines, and detected changes in the proliferation and apoptosis of drug-resistant cancer cells. In addition, the protein expression levels of KLF-4, GFI-1, and ZEB-1 were determined. The xenograft tumor models of nude mice were constructed by subcutaneously injecting GEM-resistant PANC-1 cells into mouse axilla. The tumors were removed, dissected, and weighed after 6 weeks. The protein levels of KLF-4, GFI-1, and ZEB-1 in tumor tissues were quantified. In addition, the percentage of M2 macrophages in tumor tissues was determined using flow cytometry.
RESULTS: The protein levels of TGF-β in pancreatic cancer cells were significantly decreased after GEM treatment. The protein expression of KLF-4 was downregulated, whereas the expressions of GFI-1 and ZEB-1 were upregulated after TGF-β stimulation. Apoptosis increased and proliferation decreased after TGF-β knockdown in GEM-resistant pancreatic cancer cells, moreover, silencing TGF-β promoted the expression of Caspase 3 and Cleaved caspase 3. In addition, the protein expression of KLF-4 was upregulated, whereas the expressions of GFI-1 and ZEB-1 were downregulated. Further, the volume and weight of the transplanted tumor decreased after TGF-β knockdown. The protein expression of KLF-4 was upregulated, whereas the expressions of GFI-1 and ZEB-1 were downregulated in tumor tissues. In addition, the percentage of M2 macrophages decreased in tumor tissues after TGF-β knockdown.
CONCLUSIONS: The knockdown of TGF-β inhibits epithelial-to-mesenchymal transition, suppresses the proliferation and promotes the apoptosis of drug-resistant cancer cells, and decreases the macrophage polarization to the M2 phenotype, consequently ameliorating GEM resistance in pancreatic cancer.

The objective of this study was to elucidate the protective role of quercetin in atherosclerosis by examining its effect on the phenotypic switch of vascular smooth muscle cells (VSMCs) to macrophage-like cells and the underlying regulatory pathways. Aorta tissues from apolipoprotein E-deficient (ApoE KO) mice fed a high-fat diet (HFD), treated with or without 100 mg/kg/day quercetin, were analyzed for histopathological changes and molecular mechanisms. Quercetin was found to decrease the size of atherosclerotic lesions and mitigate lipid accumulation induced by HFD. Fluorescence co-localization analysis revealed a higher presence of macrophage-like vascular smooth muscle cells (VSMCs) co-localizing with phospho-Janus kinase 2 (p-JAK2), phospho-signal transducer and activator of transcription 3 (p-STAT3), and Krüppel-like factor 4 (KLF4) in regions of foam cell aggregation within aortic plaques. However, this co-localization was reduced following treatment with quercetin. Quercetin treatment effectively inhibited the KLF4-mediated phenotypic switch in oxidized low-density lipoprotein (ox-LDL)-loaded mouse aortic vascular smooth muscle cells (MOVAS), as indicated by decreased expressions of KLF4, LGALS3, CD68, and F4/80, increased expression of alpha smooth muscle actin (α-SMA), reduced intracellular fluorescence Dil-ox-LDL uptake, and decreased lipid accumulation. In contrast, APTO-253, a KLF4 activator, was found to reverse the effects of quercetin. Furthermore, AG490, a JAK2 inhibitor, effectively counteracted the ox-LDL-induced JAK2/STAT3 pathway-dependent switch to a macrophage-like phenotype and lipid accumulation in MOVAS cells. These effects were significantly mitigated by quercetin but exacerbated by coumermycin A1, a JAK2 activator. Our research illustrates that quercetin inhibits the KLF4-mediated phenotypic switch of VSMCs to macrophage-like cells and reduces atherosclerosis by suppressing the JAK2/STAT3 pathway.

Human umbilical cord mesenchymal stem cells-derived small extracellular vesicles (MSC-sEV) provide a pragmatic solution as a cell-free therapy for patients with diabetic kidney disease (DKD). However, the underlying protective mechanisms of MSC-sEV remain largely unknown in DKD. Invivo and in vitro analyses demonstrated that MSC-sEV attenuated renal fibrosis and inflammation of DKD. The underlying mechanism of the MSC-sEV-induced therapeutic effect was explored by high-throughput sequencing, which identified the unique enrichment of a set of miRNAs in MSC-sEV compared with human skin fibroblasts-sEV (HSF-sEV). Vitro experiments demonstrated that the protective potential was primarily attributed to miR-23a-3p, one of the most abundant miRNAs in MSC-sEV. Further, overexpression or knockdown analyses revealed that miR-23a-3p, and its target Krüppel-like factor 3 (KLF3) suppressed the STAT3 signaling pathway in high glucose (HG) induced HK-2 cells were essential for the renal-protective property of MSC-sEV. Moreover, we found that miR-23a-3p was packaged into MSC-sEV by RNA Binding Motif Protein X-Linked (RBMX) and transmitted to HG-induced HK-2 cells. Finally, inhibiting miR-23a-3p could mitigate the protective effects of MSC-sEV in db/db mice. These findings suggest that a systemic administration of sEV derived from MSC, have the capacity to incorporate into kidney where they can exert renal-protective potential against HG-induced injury through delivery of miR-23a-3p.

Iron homeostasis is of critical importance to living organisms. Drosophila melanogaster has emerged as an excellent model to study iron homeostasis, while the regulatory mechanism of iron metabolism remains poorly understood. Herein, we accidently found that knockdown of juvenile hormone (JH) acid methyltransferase (Jhamt) specifically in the fat body, a key rate-limiting enzyme for JH synthesis, led to iron accumulation locally, resulting in serious loss and dysfunction of fat body. Jhamt knockdown-induced phenotypes were mitigated by iron deprivation, antioxidant and Ferrostatin-1, a well-known inhibitor of ferroptosis, suggesting ferroptosis was involved in Jhamt knockdown-induced defects in the fat body. Further study demonstrated that upregulation of Tsf1 and Malvolio (Mvl, homolog of mammalian DMT1), two iron importers, accounted for Jhamt knockdown-induced iron accumulation and dysfunction of the fat body. Mechanistically, Kr-h1, a key transcription factor of JH, acts downstream of Jhamt inhibiting Tsf1 and Mvl transcriptionally. In summary, the findings indicated that fat body-derived Jhamt is required for the development of Drosophila by maintaining iron homeostasis in the fat body, providing unique insight into the regulatory mechanisms of iron metabolism in Drosophila.

To address the increased energy demand, tumor cells undergo metabolic reprogramming, including oxidative phosphorylation (OXPHOS) and aerobic glycolysis. This study investigates the role of Kruppel-like factor 4 (KLF4), a transcription factor, as a tumor suppressor in hepatocellular carcinoma (HCC) by regulating ATP synthesis. Immunohistochemistry was performed to assess KLF4 expression in HCC tissues. Functional assays, such as CCK-8, EdU, and colony formation, as well as in vivo assays, including subcutaneous tumor formation and liver orthotopic xenograft mouse models, were conducted to determine the impact of KLF4 on HCC proliferation. Luciferase reporter assay and chromatin immunoprecipitation assay were utilized to evaluate the interaction between KLF4, miR-206, and RICTOR. The findings reveal low KLF4 expression in HCC, which is associated with poor prognosis. Both in vitro and in vivo functional assays demonstrate that KLF4 inhibits HCC cell proliferation. Mechanistically, it was demonstrated that KLF4 reduces ATP synthesis in HCC by suppressing the expression of RICTOR, a core component of mTORC2. This suppression promotes glutaminolysis to replenish the TCA cycle and increase ATP levels, facilitated by the promotion of miR-206 transcription. In conclusion, this study enhances the understanding of KLF4\'s role in HCC ATP synthesis and suggests that targeting the KLF4/miR-206/RICTOR axis could be a promising therapeutic approach for anti-HCC therapeutics.

Rationale: CD8+ T cells undergo a series of metabolic reprogramming processes during their activation and proliferation, including increased glycolysis, decreased aerobic oxidation of sugars, increased amino acid metabolism and increased protein synthesis. However, it is still unclear what factors regulate these metabolic reprogramming processes in CD8+ T cells in the tumor immune microenvironment. Methods: T cell chromobox protein 4 (CBX4) knock-out mice models were used to determine the role of CBX4 in CD8+ T cells on the tumor immune microenvironment and tumor progression. Flow cytometry, Cut-Tag qPCR, Chip-seq, immunoprecipitation, metabolite detection, lentivirus infection and adoptive T cells transfer were performed to explore the underlying mechanisms of CBX4 knock-out in promoting CD8+ T cell activation and inhibiting tumor growth. Results: We found that CBX4 expression was induced in tumor-infiltrating CD8+ T cells and inhibited CD8+ T cell function by regulating glucose metabolism in tumor tissue. Mechanistically, CBX4 increases the expression of the metabolism-associated molecule aldolase B (Aldob) through sumoylation of trans-acting transcription factor 1 (SP1) and Krüppel-like factor 3 (KLF3). In addition, Aldob inhibits glycolysis and ATP synthesis in T cells by reducing the phosphorylation of the serine/threonine protein kinase (Akt) and ultimately suppresses CD8+ T cell function. Significantly, knocking out CBX4 may improve the efficacy of anti-PD-1 therapy by enhancing the function of CD8+ T cells in the tumor microenvironment. Conclusion: CBX4 is involved in CD8+ T cell metabolic reprogramming and functional persistence in tumor tissues, and serves as an inhibitor in CD8+ T cells\' glycolysis and effector function.

Docetaxel (Doc) plays a crucial role in clinical antineoplastic practice. However, it is continuously documented that tumors frequently develop chemoresistance and relapse, which may be related to polyploid giant cancer cells (PGCCs). The aim of this study was investigate the formation mechanism and biological behavior of PGCCs induced by Doc. Ovarian cancer cells were treated with Doc, and then the effect of Doc on cellular viability was evaluated by MTT assay and microscopic imaging analysis. The biological properties of PGCCs were further evaluated by Hoechst 33342 staining, cell cycle and DNA content assay, DNA damage response (DDR) signaling detection, β-galactosidase staining, mitochondrial membrane potential detection, and reverse transcription-quantitative polymerase chain reaction. The results indicated that Doc reduced cellular viability; however, many cells were still alive, and were giant and polyploid. Doc increased the proportion of cells stayed in the G2/M phase and reduced the number of cells. In addition, the expression of γ-H2A.X was constantly increased after Doc treatment. PGCCs showed senescence-associated β-galactosidase activity and an increase in the monomeric form of JC-1. The mRNA level of octamer-binding transcription factor 4 (OCT4) and krüppel-like factor 4 (KLF4) was significantly increased in PGCCs. Taken together, our results suggest that Doc induces G2/M cell cycle arrest, inhibits the proliferation and activates persistent DDR signaling to promote the formation of PGCCs. Importantly, PGCCs exhibit a senescence phenotype and express stem cell markers.

Diabetic kidney disease (DKD) is a microvascular complication of diabetes, and glomerular endothelial cell (GEC) dysfunction is a key driver of DKD pathogenesis. Krüppel-like factor 2 (KLF2), a shear stress-induced transcription factor, is among the highly regulated genes in early DKD. In the kidney, KLF2 expression is mostly restricted to endothelial cells, but its expression is also found in immune cell subsets. KLF2 expression is upregulated in response to increased shear stress by the activation of mechanosensory receptors but suppressed by inflammatory cytokines, both of which characterize the early diabetic kidney milieu. KLF2 expression is reduced in progressive DKD and hypertensive nephropathy in humans and mice, likely due to high glucose and inflammatory cytokines such as TNF-α. However, KLF2 expression is increased in glomerular hyperfiltration-induced shear stress without metabolic dysregulation, such as in settings of unilateral nephrectomy. Lower KLF2 expression is associated with CKD progression in patients with unilateral nephrectomy, consistent with its endoprotective role. KLF2 confers endoprotection by inhibition of inflammation, thrombotic activation, and angiogenesis, and thus KLF2 is considered a protective factor for cardiovascular disease (CVD). Based on similar mechanisms, KLF2 also exhibits renoprotection, and its reduced expression in endothelial cells worsens glomerular injury and albuminuria in settings of diabetes or unilateral nephrectomy. Thus KLF2 confers endoprotective effects in both CVD and DKD, and its activators could potentially be developed as a novel class of drugs for cardiorenal protection in diabetic patients.

Chronic pancreatitis (CP) is marked by progressive fibrosis and the activation of pancreatic stellate cells (PSCs), accompanied by the destruction of pancreatic parenchyma, leading to the loss of acinar cells (ACs). Few research studies have explored the mechanism by which damaged ACs (DACs) contribute to PSCs activation and pancreatic fibrosis. Currently, there are no effective drugs for curing CP or limiting the progression of pancreatic fibrosis. In this research, co-culture with intact acinar cells (IACs) suppressed PSC activation, while co-culture with DACs did the opposite. Krüppel-like factor 4 (KLF4) was significantly upregulated in DACs and was established as the key molecule that switches ACs from PSCs-suppressor to PSCs-activator. We revealed the exosomes of IACs contributed to the anti-activated function of IACs-CS on PSCs. MiRNome profiling showed that let-7 family is significantly enriched in IAC-derived exosomes (>30% miRNome), which partially mediates IACs\' suppressive impacts on PSCs. Furthermore, it has been observed that the enrichment of let-7 in exosomes was influenced by the expression level of KLF4. Mechanistic studies demonstrated that KLF4 in ACs upregulated Lin28A, thereby decreasing let-7 levels in AC-derived exosomes, and thus promoting PSCs activation. We utilized an adeno-associated virus specifically targeting KLF4 in ACs (shKLF4-pAAV) to suppress PSCs activation in CP, resulting in reduced pancreatic fibrosis. IAC-derived exosomes hold potential as potent weapons against PSCs activation via let-7s, while activated KLF4/Lin28A signaling in DACs diminished such functions. ShKLF4-pAAV holds promise as a novel therapeutic approach for CP.