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Introduction: Trophoblast homeostasis and differentiation require a proper endoplasmic reticulum (ER) function. The Krüppel-like factor-6 (KLF6) transcription factor modulates trophoblast migration, differentiation, and reactive oxygen species (ROS) production. Since ROS may impact on ER homeostasis, we assessed whether downregulation of KLF6 altered the unfolded protein response (UPR) and cellular process associated with ER homeostasis., Materials and Methods: Protein and RNA expression were analyzed by Western blot and qRT-PCR, respectively, in extravillous trophoblast HTR-8/SVneo cells silenced for KLF6. Apoptosis was detected by flow cell cytometry using Annexin V Apoptosis Detection Kit. Protein trafficking was assessed by confocal microscopy of a reporter fluorescent protein whose release from the ER was synchronized., Results: KLF6 downregulation reduced the expression of BiP, the master regulator of the UPR, at protein, mRNA, and pre-mRNA levels. Ire1α protein, XBP1 splicing, and DNAJB9 mRNA levels were also reduced in KLF6-silenced cells. Instead, PDI, Ero1α, and the p-eIF2α/eIF2α ratio as well as autophagy and proteasome dependent protein degradation remained unchanged while intracellular trafficking was increased. Under thapsigargin-induced stress, KLF6 silencing impaired BiP protein and mRNA expression increase, as well as the activation of the Ire1α pathway, but it raised the p-eIF2α/eIF2α ratio and CHOP protein levels. Nevertheless, apoptosis was not increased., Discussion: Results provide the first evidence of KLF6 as a modulator of the UPR components. The increase in protein trafficking and protection from apoptosis, observed in KLF6-silenced cells, are consistent with its role in extravillous trophoblast migration and differentiation., Competing Interests: Declaration of competing interest Lucille T. Kourdova, Andrea L. Miranda, Milagros Ovejero, Agustín Anastasía, Susana Genti-Raimondi, Ana C. Racca, and Graciela M. Panzetta-Dutari declare no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Diabetic kidney disease (DKD) is the main cause of chronic kidney disease worldwide. While injury to the podocytes, visceral epithelial cells that comprise the glomerular filtration barrier, drives albuminuria, proximal tubule (PT) dysfunction is the critical mediator of DKD progression. Here, we report that the podocyte-specific induction of human KLF6, a zinc-finger binding transcription factor, attenuates podocyte loss, PT dysfunction, and eventual interstitial fibrosis in a male murine model of DKD. Utilizing combination of snRNA-seq, snATAC-seq, and tandem mass spectrometry, we demonstrate that podocyte-specific KLF6 triggers the release of secretory ApoJ to activate calcium/calmodulin dependent protein kinase 1D (CaMK1D) signaling in neighboring PT cells. CaMK1D is enriched in the first segment of the PT, proximal to the podocytes, and is critical to attenuating mitochondrial fission and restoring mitochondrial function under diabetic conditions. Targeting podocyte-PT signaling by enhancing ApoJ-CaMK1D might be a key therapeutic strategy in attenuating the progression of DKD., (© 2024. The Author(s).)

Aims: Heart failure (HF) is one of the most devastating consequences of cardiovascular diseases. Regardless of etiology, cardiac fibrosis is present and promotes the loss of heart function in HF patients. Cardiac resident fibroblasts, in response to a host of pro-fibrogenic stimuli, trans-differentiate into myofibroblasts to mediate cardiac fibrosis, the underlying mechanism of which remains incompletely understood., Methods: Fibroblast-myofibroblast transition was induced in vitro by exposure to transforming growth factor (TGF-β). Cardiac fibrosis was induced in mice by either transverse aortic constriction (TAC) or by chronic infusion with angiotensin II (Ang II)., Results: Through bioinformatic screening, we identified Kruppel-like factor 6 (KLF6) as a transcription factor preferentially up-regulated in cardiac fibroblasts from individuals with non-ischemic cardiomyopathy (NICM) compared to the healthy donors. Further analysis showed that nuclear factor kappa B (NF-κB) bound to the KLF6 promoter and mediated KLF6 trans-activation by pro-fibrogenic stimuli. KLF6 knockdown attenuated whereas KLF6 over-expression enhanced TGF-β induced fibroblast-myofibroblast transition in vitro. More importantly, myofibroblast-specific KLF6 depletion ameliorated cardiac fibrosis and rescued heart function in mice subjected to the TAC procedure or chronic Ang II infusion., Significance: In conclusion, our data support a role for KLF6 in cardiac fibrosis., Competing Interests: Declaration of competing interest None., (Copyright © 2024. Published by Elsevier Inc.)

The definition of molecular and cellular mechanisms contributing to brain ontogenetic trajectories is essential to investigate the evolution of our species. Yet their functional dissection at an appropriate level of granularity remains challenging. Capitalizing on recent efforts that have extensively profiled neural stem cells from the developing human cortex, we develop an integrative computational framework to perform trajectory inference and gene regulatory network reconstruction, (pseudo)time-informed non-negative matrix factorization for learning the dynamics of gene expression programs, and paleogenomic analysis for a higher-resolution mapping of derived regulatory variants in our species in comparison with our closest relatives. We provide evidence for cell type-specific regulation of gene expression programs during indirect neurogenesis. In particular, our analysis uncovers a key role for a cholesterol program in outer radial glia, regulated by zinc-finger transcription factor KLF6. A cartography of the regulatory landscape impacted by Homo sapiens-derived variants reveals signals of selection clustering around regulatory regions associated with GLI3, a well-known regulator of radial glial cell cycle, and impacting KLF6 regulation. Our study contributes to the evidence of significant changes in metabolic pathways in recent human brain evolution., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Abstract: Background: Myocardial infarction (MI) is a severe condition that typically results from the ischemia and necrosis of heart muscle. Kruppel-like factor 6 (KLF6) can aggravate myocardial ischemia/reperfusion injury. This work aims to reveal its role and mechanism in hypoxia/reoxygenation (H/R)-induced cardiomyocyte injury. Methods: Human cardiomyocyte (AC16) was exposed to hypoxic treatment to mimic MI-like cell injury. mRNA expression levels of KLF6 and WT1-associated protein (WTAP) were detected by quantitative real-time polymerase chain reaction. Protein expression was detected by western blotting assay. Cell viability was assessed by CCK-8 assay. Cell apoptosis and cell cycle were investigated by flow cytometry. Enzyme-linked immunosorbent assays were conducted to detect IL-1β, TNF-α and IL-6 levels. Fe 2+ colorimetric assay kit was used to detect Fe 2+ level. MDA Content Assay Kit was used to detect MDA level. Cellular ROS Assay kit was applied to assess ROS level. The association of KLF6 and WTAP was identified by RNA immunoprecipitation assay and dual-luciferase reporter assay. Results: KLF6 and WTAP expression at mRNA and protein levels were significantly upregulated in serum samples of MI patients and H/R-induced AC16 cells when compared with control groups. KLF6 silencing attenuated H/R-induced AC16 cell apoptosis, inflammatory response, oxidative stress, and ferroptosis. Additionally, WTAP stabilized KLF6 mRNA by regulating its m6A modification. Furthermore, WTAP knockdown rescued H/R-induced AC16 cell apoptosis, inflammatory response, oxidative stress, and ferroptosis by decreasing KLF6 expression. Conclusion: WTAP-mediated m6A modification of KLF6 aggravated hypoxia/reoxygenation-induced apoptosis, inflammatory response, oxidative stress, and ferroptosis of human cardiomyocytes, providing a therapeutic strategy for MI., Competing Interests: The authors report no conflicts of interest., (Copyright © 2024 by the Shock Society.)

Despite decades of research, endometriosis remains a mysterious gynecological disease with unknown etiology and pathogenesis. Krüppel-like Factor 6 (KLF6), a transcription factor, has a wide expression profile and regulates a variety of biological processes. Here, we investigated the expression and function of KLF6 and its possible regulatory mechanisms in endometriosis. To determine the function of KLF6, knockdown and overexpression experiments were performed in eutopic endometrial stromal cells (EU-ESCs) and ectopic endometrial stromal cells (EC-ESCs), respectively. Cell viability, apoptosis, migration, invasion, and angiogenesis assays were conducted in ESCs. ChIP-sequencing and mRNA-sequencing were performed to investigate the functional mechanism of KLF6 in regulating ESCs. We found that KLF6 was highly expressed in eutopic endometrium of endometriosis patients, compared with ectopic endometrium. Similarly, the same was true in EU-ESCs, which was compared with EC-ESCs. Overexpression of KLF6 significantly suppressed EC-ESC proliferation, migration and invasion and induced cell apoptosis, while knockdown of KLF6 resulted in the opposite effects on EU-ESCs. Overexpression of KLF6 significantly inhibited EC-ESC angiogenesis. Mechanistically, the results of ChIP sequencing and mRNA sequencing revealed that CTNNB1 may be a transcriptional target regulated by KLF6. Reintroduction of KLF6 reversed the effects of KLF6 knockdown on EU-ESCs. KLF6 inhibited the proliferation, migration and angiogenesis of EC-ESCs by inhibiting the expression of CTNNB1. Our findings provided a new perspective on the role of KLF6 in endometriosis progression and inspire potential targeted therapeutic strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Background: Intracranial aneurysm (IA) is a cerebrovascular disease with a high mortality rate due to ruptured subarachnoid hemorrhage. While Krüppel-like factor 6 (KLF6) dysregulation has been implicated in cancer and cardiovascular diseases, its role in IA remains unclear., Materials and Methods: The GSE122897 and GSE15629 datasets were downloaded from the Gene Expression Omnibus database. Immune cell infiltration and hypoxia analysis were performed to explore the effects of KLF6 on IA. Weighted gene co-expression network analysis was used to identify hub genes related to KLF6 expression for subsequent analyses. Hypoxia-related genes were identified. Drug prediction was performed for IA. Samples from healthy individuals and patients with IA were collected to detect the expression of endothelin-1 (ET-1), vascular hematoma factor (vWF), and KLF6. A model of H 2 O 2 -induced human brain vascular smooth muscle cells (HBVSMC) injury was constructed to explore the effects of KLF6 and melatonin to treat IA., Results: T cells CD4 memory resting and monocytes were significantly different in the KLF6 high and low expression groups. Four hypoxia-related gene sets were significantly enriched in the KLF6 high-expression group. Six hypoxia-related hub genes were obtained, which were significantly associated with KLF6. Drug prediction showed that melatonin may be a potential drug for IA. The levels of ET-1, vWF, and KLF6 were significantly upregulated in patients with IA. KLF6 exacerbates H 2 O 2 -induced injury in HBVSMC, ameliorated by melatonin., Conclusion: KLF6 may be a potential target for IA treatment, with melatonin-mediated KLF6 effects playing a crucial role in the development of IA.

Resistance to immunity is associated with the selection of cancer cells with superior capacities to survive inflammatory reactions. Here, we tailored an ex vivo immune selection model for acute myeloid leukemia (AML) and isolated the residual subpopulations as "immune-experienced" AML (ieAML) cells. We confirmed that upon surviving the immune reactions, the malignant blasts frequently decelerated proliferation, displayed features of myeloid differentiation and activation, and lost immunogenicity. Transcriptomic analyses revealed a limited number of commonly altered pathways and differentially expressed genes in all ieAML cells derived from distinct parental cell lines. Molecular signatures predominantly associated with interferon and inflammatory cytokine signaling were enriched in the AML cells resisting the T-cell-mediated immune reactions. Moreover, the expression and nuclear localization of the transcription factors c-MYB and KLF6 were noted as the putative markers for immune resistance and identified in subpopulations of AML blasts in the patients' bone marrow aspirates. The immune modulatory capacities of ieAML cells lasted for a restricted period when the immune selection pressure was omitted. In conclusion, myeloid leukemia cells harbor subpopulations that can adapt to the harsh conditions established by immune reactions, and a previous "immune experience" is marked with IFN signature and may pave the way for susceptibility to immune intervention therapies., (© 2024 The Authors. European Journal of Immunology published by Wiley‐VCH GmbH.)

Acute liver failure (ALF) is a complex syndrome characterized by overactivation of innate immunity, and the recruitment and differentiation of immune cells at inflammatory sites. The present study aimed to explore the role of microRNA (miRNA/miR)‑21 and its potential mechanisms underlying inflammatory responses in ALF. Baseline serum miR‑21 was analyzed in patients with ALF and healthy controls. In addition, miR‑21 antagomir was injected via the tail vein into C57BL/6 mice, and lipopolysaccharide/D‑galactosamine (LPS/GalN) was injected into mice after 48 h. The expression levels of miR‑21, Krüppel‑like‑factor‑6 (KLF6), autophagy‑related proteins and interleukin (IL)‑23, and hepatic pathology were then assessed in the liver tissue. Furthermore, THP‑1‑derived macrophages were transfected with a miRNA negative control, miR‑21 inhibitor, miR‑21 mimics or KLF6 overexpression plasmid, followed by treatment with or without rapamycin, and the expression levels of miR‑21, KLF6, autophagy‑related proteins and IL‑23 were evaluated. The results revealed that baseline serum miR‑21 levels were significantly upregulated in patients with ALF. In addition, LPS/GalN‑induced ALF was attenuated in the antagomir‑21 mouse group. KLF6 was identified as a target of miR‑21‑5p with one putative seed match site identified by TargetScan. A subsequent luciferase activity assay demonstrated a direct interaction between miR‑21‑5p and the 3'‑UTR of KLF6 mRNA. Further experiments suggested that miR‑21 promoted the expression of IL‑23 via inhibiting KLF6, which regulated autophagy. In conclusion, in the present study, baseline serum miR‑21 levels were highly upregulated in patients with ALF, antagomir‑21 attenuated LPS/GalN‑induced ALF in a mouse model, and miR‑21 could promote the expression of IL‑23 via inhibiting KLF6.

Intramuscular fat is a crucial determinant of carcass quality traits like tenderness and taste, which in turn is influenced by the proliferation of intramuscular preadipocytes. This study aimed to investigate the Krüppel-like factor 6 (KLF6)-mediated proliferation of bovine preadipocytes and identify underlying molecular mechanisms. Down-regulation of KLF6 by siKLF6 resulted in a significant ( p < 0.01) suppression of cell cycle-related genes including CDK1, MCM6, ZNF4, PCNA, CDK2, CCNB1, and CDK6. Conversely, the expression level of p27 was significantly ( p < 0.01) increased. Moreover, EdU (5-ethynyl-20-deoxyuridine) staining revealed a significant decrease in EdU-labeled cells due to KLF6 down-regulation. Collectively, these findings indicate that KLF6 down-regulation inhibits adipocyte proliferation. Furthermore, RNA sequencing of preadipocytes transfected with siKLF6 and NC, followed by differential gene expression analysis, identified 100 up-regulated and 70 down-regulated genes. Additionally, the differentially expressed genes also significantly influenced various Gene Ontology (GO) terms related to cell cycle, nuclear chromosomes, and catalytic activity on DNA. Furthermore, the top 20 pathways enriched in these DEGs included cell cycle, DNA replication, cellular senescence, and homologous recombination. These GO terms and KEGG pathways play key roles in bovine preadipocyte proliferation. In conclusion, the results of this study suggest that KLF6 positively regulates the proliferation of bovine preadipocytes.

Background: The incidence of chronic obstructive pulmonary disease (COPD) is increasing year by year. Kruppel-like factor 6 (KLF6) plays an important role in inflammatory diseases. However, the regulatory role of KLF6 in COPD has not been reported so far., Methods: The viability of human bronchial epithelial cells BEAS-2B induced by cigarette smoke extract (CSE) was detected by CCK-8 assay. The protein expression of KLF6 and sirtuin 4 (SIRT4) was appraised with Western blot. RT-qPCR and Western blot were applied to examine the transfection efficacy of sh-KLF6 and Oe-KLF6. Cell apoptosis was detected using flow cytometry. The levels of inflammatory factors IL-6, TNF-α and IL-1β were assessed with ELISA assay. DCFH-DA staining was employed for the detection of ROS activity and the levels of oxidative stress markers SOD, CAT and MDA were estimated with corresponding assay kits. The mitochondrial membrane potential (MMP), adenosine triphosphate (ATP) content and Complex I activity were evaluated with JC-1 staining, ATP colorimetric/fluorometric assay kit and Complex I enzyme activity microplate assay kit. With the application of mitochondrial permeability transition pore detection kit, mPTP opening was measured. Luciferase report assay was employed to evaluate the activity of SIRT4 promoter and chromatin immunoprecipitation (ChIP) to verify the binding ability of KLF6 and SIRT4 promoter., Results: KLF6 expression was significantly elevated in CSE-induced cells. KLF6 was confirmed to suppress SIRT4 transcription. Interference with KLF6 expression significantly inhibited cell viability damage, cell apoptosis, inflammatory response, oxidative stress and mitochondrial dysfunction in CSE-induced BEAS-2B cells, which were all reversed by SIRT4 overexpression., Conclusion: Silencing KLF6 alleviated CSE-induced mitochondrial dysfunction in bronchial epithelial cells by SIRT4 upregulation., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Wan et al.)

Prolidase (PEPD) is the only hydrolase that cleaves the dipeptides containing C-terminal proline or hydroxyproline-the rate-limiting step in collagen biosynthesis. However, the molecular regulation of prolidase expression remains largely unknown. In this study, we have identified overlapping binding sites for the transcription factors Krüppel-like factor 6 (KLF6) and Specificity protein 1 (Sp1) in the PEPD promoter and demonstrate that KLF6/Sp1 transcriptionally regulate prolidase expression. By cloning the PEPD promoter into a luciferase reporter and through site-directed deletion, we pinpointed the minimal sequences required for KLF6 and Sp1-mediated PEPD promoter-driven transcription. Interestingly, Sp1 inhibition abrogated KLF6-mediated PEPD promoter activity, suggesting that Sp1 is required for the basal expression of prolidase. We further studied the regulation of PEPD by KLF6 and Sp1 during transforming growth factor β 1 (TGF-β 1 ) signaling, since both KLF6 and Sp1 are key players in TGF-β1 mediated collagen biosynthesis. Mouse and human fibroblasts exposed to TGF-β1 resulted in the induction of PEPD transcription and prolidase expression. Inhibition of TGF-β1 signaling abrogated PEPD promoter-driven transcriptional activity of KLF6 and Sp1. Knock-down of KLF6 as well as Sp1 inhibition also reduced prolidase expression. Chromatin immunoprecipitation assay supported direct binding of KLF6 and Sp1 to the PEPD promoter and this binding was enriched by TGF-β1 treatment. Finally, immunofluorescence studies showed that KLF6 co-operates with Sp1 in the nucleus to activate prolidase expression and enhance collagen biosynthesis. Collectively, our results identify functional elements of the PEPD promoter for KLF6 and Sp1-mediated transcriptional activation and describe the molecular mechanism of prolidase expression., Competing Interests: Conflict of interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: We aim to accurately distinguish ubiquitin-specific proteases (USPs) from other members within the deubiquitinating enzyme families based on protein sequences. Additionally, we seek to elucidate the specific regulatory mechanisms through which USP26 modulates Krüppel-like factor 6 (KLF6) and assess the subsequent effects of this regulation on both the proliferation and migration of cervical cancer cells., Methods: All the deubiquitinase (DUB) sequences were classified into USPs and non-USPs. Feature vectors, including 188D, n-gram, and 400D dimensions, were extracted from these sequences and subjected to binary classification via the Weka software. Next, thirty human USPs were also analyzed to identify conserved motifs and ascertained evolutionary relationships. Experimentally, more than 90 unique DUB-encoding plasmids were transfected into HeLa cell lines to assess alterations in KLF6 protein levels and to isolate a specific DUB involved in KLF6 regulation. Subsequent experiments utilized both wild-type (WT) USP26 overexpression and shRNA-mediated USP26 knockdown to examine changes in KLF6 protein levels. The half-life experiment was performed to assess the influence of USP26 on KLF6 protein stability. Immunoprecipitation was applied to confirm the USP26-KLF6 interaction, and ubiquitination assays to explore the role of USP26 in KLF6 deubiquitination. Additional cellular assays were conducted to evaluate the effects of USP26 on HeLa cell proliferation and migration., Results: 1. Among the extracted feature vectors of 188D, 400D, and n-gram, all 12 classifiers demonstrated excellent performance. The RandomForest classifier demonstrated superior performance in this assessment. Phylogenetic analysis of 30 human USPs revealed the presence of nine unique motifs, comprising zinc finger and ubiquitin-specific protease domains. 2. Through a systematic screening of the deubiquitinase library, USP26 was identified as the sole DUB associated with KLF6. 3. USP26 positively regulated the protein level of KLF6, as evidenced by the decrease in KLF6 protein expression upon shUSP26 knockdown in both 293T and Hela cell lines. Additionally, half-life experiments demonstrated that USP26 prolonged the stability of KLF6. 4. Immunoprecipitation experiments revealed a strong interaction between USP26 and KLF6. Notably, the functional interaction domain was mapped to amino acids 285-913 of USP26, as opposed to the 1-295 region. 5. WT USP26 was found to attenuate the ubiquitination levels of KLF6. However, the mutant USP26 abrogated its deubiquitination activity. 6. Functional biological assays demonstrated that overexpression of USP26 inhibited both proliferation and migration of HeLa cells. Conversely, knockdown of USP26 was shown to promote these oncogenic properties., Conclusions: 1. At the protein sequence level, members of the USP family can be effectively differentiated from non-USP proteins. Furthermore, specific functional motifs have been identified within the sequences of human USPs. 2. The deubiquitinating enzyme USP26 has been shown to target KLF6 for deubiquitination, thereby modulating its stability. Importantly, USP26 plays a pivotal role in the modulation of proliferation and migration in cervical cancer cells., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Nicotine contributes to the causation of atherosclerosis, which the prominent cellular components are macrophages. Long non-coding RNAs (lncRNAs) play an important role in regulating cell functions such as cell proliferation, differentiation and programmed death. However, the function and mechanism of lncRNAs in nicotine-induced macrophage pyroptosis has not been reported. We screened the deferentially expressed lncRNAs of human carotid artery plaque (GSE97210) and verified them in nicotine-induced pyroptosis of macrophages. Results showed only LINC01272 was up-regulated in a dose-dependent manner in macrophages. The immunofluorescence staining result confirmed that interfering LINC01272 inhibited nicotine-induced macrophage pyroptosis. Through bioinformatics analysis, dual luciferase reporter gene assay and qPCR, we identified miR-515 was significantly negatively correlated with the expression of LINC01272, and KLF6 is the target gene of miR-515. Furthermore, our results demonstrated that LINC01272/miR-515/KLF6 axis meditated nicotine-induced macrophage pyroptosis. In addition, in human peripheral blood mononuclear cells of smoking populations, the expression of GSDMD-N, NLRP3, LINC01272 and KLF6 was significantly increased, while the level of miR-515 was reduced. This study confirmed that nicotine increases the expression of LINC01272 to competitively bind with miR-515 in macrophages, reducing the inhibitory effect of miR-515 on its target gene KLF6, which ultimately induces macrophage pyroptosis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Skin color, one of the most diverse human traits, is determined by the quantity, type, and distribution of melanin. In this study, we leveraged the light-scattering properties of melanin to conduct a genome-wide screen for regulators of melanogenesis. We identified 169 functionally diverse genes that converge on melanosome biogenesis, endosomal transport, and gene regulation, of which 135 represented previously unknown associations with pigmentation. In agreement with their melanin-promoting function, the majority of screen hits were up-regulated in melanocytes from darkly pigmented individuals. We further unraveled functions of KLF6 as a transcription factor that regulates melanosome maturation and pigmentation in vivo, and of the endosomal trafficking protein COMMD3 in modulating melanosomal pH. Our study reveals a plethora of melanin-promoting genes, with broad implications for human variation, cell biology, and medicine.

The transcription factor SREBP2 is the main regulator of cholesterol homeostasis and is central to the mechanism of action of lipid-lowering drugs, such as statins, which are responsible for the largest overall reduction in cardiovascular risk and mortality in humans with atherosclerotic disease. Recently, SREBP2 has been implicated in leukocyte innate and adaptive immune responses by upregulation of cholesterol flux or direct transcriptional activation of pro-inflammatory genes. Here, we investigate the role of SREBP2 in endothelial cells (ECs), since ECs are at the interface of circulating lipids with tissues and crucial to the pathogenesis of cardiovascular disease. Loss of SREBF2 inhibits the production of pro-inflammatory chemokines but amplifies type I interferon response genes in response to inflammatory stimulus. Furthermore, SREBP2 regulates chemokine expression not through enhancement of endogenous cholesterol synthesis or lipoprotein uptake but partially through direct transcriptional activation. Chromatin immunoprecipitation sequencing of endogenous SREBP2 reveals that SREBP2 bound to the promoter regions of two nonclassical sterol responsive genes involved in immune modulation, BHLHE40 and KLF6. SREBP2 upregulation of KLF6 was responsible for the downstream amplification of chemokine expression, highlighting a novel relationship between cholesterol homeostasis and inflammatory phenotypes in ECs., Competing Interests: Conflict of interest The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: The purpose of this study was to identify novel abnormally expressed microRNAs (miRNAs) and their downstream target in diabetic cataract (DC)., Methods: General feature, fasting blood glucose, glycosylated hemoglobin, and type A1c (HbA1c) expression level of patients were collected. DC capsular tissues were obtained from patients and the lens cells (HLE-B3) exposed to different concentrations of glucose were used to simulate the model in vitro. Both mimic and inhibitor of miR-22-3p were transferred into HLE-B3 to up- and downregulate miR-22-3p expression, respectively. The cellular apoptosis was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR), Western blot, and immunofluorescence. The downstream target gene of miR-22-3p was identified by dual luciferase reporter., Results: In DC capsules and HLE-B3 under hyperglycemia, miR-22-3p showed a significant downward trend. The expression of BAX was upregulated and the BCL-2 was downregulated following high glucose. The expression of BAX was significantly down- or upregulated in HLE-B3 cells following transfection of mimic or inhibitor of miR-22-3p, respectively. Conversely, BCL-2 was significantly increased or decreased. Dual luciferase reporter assay showed that miR-22-3p directly targeted Krüppel Like Factor 6 (KLF6) to regulate cell apoptosis. In addition, the expression of KLF6 were significantly up- or downregulated following transfection of inhibitor or mimic of miR-22-3p., Conclusions: This study suggested that miR-22-3p could inhibit lens apoptosis by targeting KLF6 directly under high glucose condition. The miR-22-3p/KLF6 signal axis may provide novel insights into the pathogenesis of DC., Translational Relevance: Differential expression of miR-22-3p may account for the pathogenesis of DC and lead to a new therapeutic strategy for DC.

The incidence of laryngeal carcinoma accounts for 1 to 5% of systemic malignancies and ranks second among head and neck malignancies. Screening more effective targets are meaningful for the treatment of laryngeal carcinoma. The purpose was to research the action of miR-21-5p in the occurrence of laryngeal carcinoma. Genecards combined with g:profiler was used for cluster analysis to predict gene-related miRNAs. Q-PCR assay was performed for measuring the level of miR-21-5p and Kruppel-like factor 6 (KLF6). miR-21-5p-mimic, miR-21-5p-inhibitor and sh-KLF6 were transfected using LipofectamineTM 2000. Both CCK-8 and EdU experiments were undertaken to detect cell proliferation ability. Western blot was used to detect apoptosis and epithelial-mesenchymal transition (EMT) related proteins. Wound healing assay and transwell assay were undertaken for migration and invasion, respectively. Three online software (ENCORI, miRWalk, and miRDB) were applied to screen the downstream of miR-21-5p. At the same time, a dual-luciferase reporter experiment was processed to verify the binding. Finally, a rescue experiment was applied to reveal the mediating role of miR-21-5p and KLF6. MiR-21-5p expressed highly in laryngeal carcinoma tissues and cell lines. Knockdown of miR-21-5p reduced the EMT, while enhancing apoptosis of laryngeal carcinoma cell lines. MiR-21-5p targeted KLF6 with negative relationships. The rescue assay results confirmed that sh-KLF6 rescued the action of miR-21-5p knockdown in developing laryngeal carcinoma cells. MiR-21-5p promotes the occurrence and development of laryngeal cancer by targeting KLF6. This finding may provide new insights into miRNA as a biomarker for diagnosing and treating laryngeal carcinoma in the future., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: MicroRNAs (miRNAs) exert certain functions in the development of several cancers and can be a potential hallmark for cancer diagnosis and prognosis. MiR-191-5p has been proven to have high expression in breast cancer (BC), while its biological role and potential regulatory mechanisms in BC remain an open issue., Objective: Bioinformatics was utilized to assay miR-191-5p level in BC tissues and predict its downstream target gene as well as the enriched signaling pathways of the target gene., Methods: qRT-PCR was carried out to assay miR-191-5p and KLF6 levels in BC cells as well as miR-191-5p level in blood-derived exosomes from BC patients. Western blot was to examine the expression of proteins linked with cell adhesion, epithelial-mesenchymal transition (EMT), and exosome markers. A dual luciferase reporter assay was utilized to verify the interaction between miR-191-5p and KLF6. Abilities of cell phenotypes of BC cells were detected by CCK8, Transwell, and cell adhesion assay, separately., Results: Upregulated miR-191-5p expression and downregulated KLF6 expression were observed in BC cells. There was a targeting relationship between miR-191-5p and KLF6. MiR-191-5p negatively regulated KLF6 to promote EMT and malignant progression of BC cells. Additionally, we described a dramatically high level of miR-191-5p in the blood exosomes of BC patients., Conclusion: MiR-191-5p advances the EMT of BC by targeting KLF6, indicating that miR-191-5p and KLF6 may be new biomarkers for BC.

Cutaneous carcinoma is one of the most common neoplasm tumors in the West. Its incidence rate is one of the fastest growing tumors in China. The Krüppel-like factor 6 (KLF6) is a latent tumor suppressor. Decreased KLF6 is related to the occurrence and progression of many cancers in human. Our previous studies have demonstrated that KLF6 was down-regulation in cutaneous malignant melanoma (CMM), and was significant correlated with ulcer, lymph node metastasis and clinical stage, suggesting that KLF6 loss is expected to become a biological indicator of poor prognosis in CMM patients. In this research, we would further study the features of KLF6 in the malignant progression of CMM. The expression of KLF6 was up-regulated by lentivirus infection containing KLF6, and short hairpin RNA (shRNA) was used for knockdown of KLF6 in CMM cells. Western blot, RT-qpcr, CCK8 assay, transwell migration assays, wound healing assay and flow cytometry were used to test the role of KLF6 in the CMM. We found that reduced expression of KLF6 significantly enhanced proliferation, migration and invasion. Moreover, KLF6 induced CMM cell apoptosis and G1 cycle arrest. The decreased KLF6 expression is expected to be a biological indicator of poor prognosis in CMM patients.

Members of the Krüppel-like family of transcription factors are widely expressed, including in the kidney. Expression of some KLFs changes in acute kidney injury, and this may be adaptive or maladaptive, and result in effects on various cellular pathways. This mini-review will highlight the roles of KLF6 and KLF15 in control of proximal tubular cell metabolism., (© 2023 S. Karger AG, Basel.)

Background: Colorectal cancer (CRC) is one of the most important cancers in the world, and its prevalence varies depending on the geographical area. Genetically, tumor regeneration in CRC as a multi-step process involves activating mutations in protocogenes and losing the function of tumor suppressor genes as well as DNA repair and recovery genes. Occur in this way, our goal was to investigate the expression of KLF6 genes as a tumor suppressor and MUTYH involved in the DNA repair process in colorectal cancer., Methods: This research was conducted during the years 2019-2018 in Razi Hospital, Rasht. The subjects included 30 tumoral and 30 non-tumoral tissues of colorectal cancer and 20 healthy controls. The real-time PCR method was used to investigate the gene expression. For data analysis by SPSS, parametric statistical tests ANOVA and T test and regression analysis were used and p value values less than 0.05 were considered significant., Results: The expression of KLF6 gene in tumoral tissues showed a significant decrease compared to non-tumoral tissues (P = 0.04). Also, the expression of MUTYH gene in tumor tissue showed a significant decrease compared to non-tumoral (P = 0.02) and this decrease in MUTYH gene expression had a significant relationship with increasing tumor stage (P = 0.01)., Conclusion: These findings suggest that decreased expression of KLF6 and MUTYH genes in the study population has a significant relationship with colorectal cancer and can be considered as tumor marker in diagnostic purpose., (© 2022. The Author(s).)

Purpose: The phenotypic transformation of human brain vascular smooth muscle cells (HBVSMC) is widely involved in the appearance and progression of intracranial aneurysms (IA). Aneurysm (IA) Circular RNA circ&#95;FOXO3 functions pivotally in vascular diseases and tumors, but its regulatory role as well as its molecular mechanism in IA is still uncertain. This research was to explore how circ&#95;FOXO3 works and its mechanism in vitro model of HBVSMC IA induced by H2O2., Methods: Thirty-eight patients with IA and their normal tissues were clinically collected. Examination of endothelin-1, vascular hematoma factor, circ&#95;FOXO3, microRNA (miR)-122-5p and KLF6 and the correlation of circ&#95;FOXO3 with clinical case information were ensured. Establishment of an in vitro IA model was through HBVSMC induced by H2O2 and transfection with circ&#95;FOXO3, miR-122-5p and KLF6 related plasmids was to figure out their roles in cell growth. The relationship among circ&#95;FOXO3, miR-122-5p with KLF6 was detected., Results: Up-regulated circ&#95;FOXO3 and KLF6 and reduced miR-122-5p were in IA tissues; Circ&#95;FOXO3 was associated with smoking history, Hunt-Hess grading and endothelial injury degree. Repressive circ&#95;FOXO3 or KLF6 and strengthening miR-122-5p facilitated H2O2-induced proliferation and repressed HBVSMC apoptosis, while elevation of circ&#95;FOXO3 or depressive miR-122-5p was opposite. circ&#95;FOXO3 bound to miR-122-5p, whose target was KLF6, which participated in controlling IA by mediating the circ&#95;FOXO3/miR-122-5p axis., Conclusion: In summary, the findings suggest that circ&#95;FOXO3 suppresses H2O2-induced proliferation of HBVSMC but promotes apoptosis via modulation of miR-122-5p/KLF6 axis. Targeted therapy of circ&#95;FOXO3/miR-122-5p/KLF6 axis is supposed to be a promising treatment approach for IA patients.

MicroRNAs (miRNAs) are non-coding RNAs that regulate the expression of their target genes involved in many cellular functions at the post-transcriptional level. Previously, bta-miR-148a showed significantly high expression in bovine mammary epithelial cells (BMECs) of Chinese Holstein cows producing high milk fat compared to those with low milk fat content. Here, we investigated the role of bta-miR-148a through targeting Krüppel-like factor 6 (KLF6) and further analyzed the role of KLF6 in regulating fat metabolism through targeting PPARA, AMPK/mTOR/PPARG, and other fat marker genes in BMECs of Chinese Holstein. The bioinformatics analysis showed that the 3' UTR of KLF6 mRNA possesses the binding sites for bta-miR-148a, which was further verified through dual-luciferase reporter assay. The BMECs were transfected with bta-miR-148a-mimic, inhibitor, and shNC, and the expression of KLF6 was found to be negatively regulated by bta-miR-148a. Moreover, the contents of triglyceride (TG), and cholesterol (CHO) in BMECs transfected with bta-miR-148a-mimic were significantly lower than the contents in BMECs transfected with bta-miR-148a-shNC. Meanwhile, the TG and CHO contents were significantly increased in BMECs transfected with bta-miR-148a-inhibitor than in BMECs transfected with bta-miR-148a-shNC. In addition, the TG and CHO contents were significantly decreased in BMECs upon the down-regulation of KLF6 through transfection with pb7sk- KLF6 -siRNA1 compared to the control group. Contrarily, when KLF6 was overexpressed in BMECs through transfection with pBI-CMV3- KLF6 , the TG and CHO contents were significantly increased compared to the control group. Whereas, the qPCR and Western blot evaluation of PPARA, AMPK/mTOR/PPARG, and other fat marker genes revealed that all of the genes were considerably down-regulated in the KLF6 -KO-BMECs compared to the normal BMECs. Taking advantage of deploying new molecular markers and regulators for increasing the production of better-quality milk with tailored fat contents would be the hallmark in dairy sector. Hence, bta-miR-148a and KLF6 are potential candidates for increased milk synthesis and the production of valuable milk components in dairy cattle through marker-assisted selection in molecular breeding. Furthermore, this study hints at the extrapolation of a myriad of functions of other KLF family members in milk fat synthesis.

Background: The development of acute lung injury (ALI) into a severe stage leads to acute respiratory distress syndrome (ARDS). The morbidity and mortality of ALI and ARDS are very high. Objective : This study is aimed to explore the effect of Krüppel-like factor 6 (KLF6) on lipopolysaccharide (LPS)-induced type II alveolar epithelial cells in ALI by interacting with cysteine-rich angiogenic inducer 61 (CYR61)., Material and Methods: ALI mice model and LPS-induced type II alveolar epithelial cells were conducted to simulate ALI in vivo and in vitro . The messenger RNA (mRNA) and protein expression of KLF6 in lung tissues were detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis. Pathological changes in lung tissues were observed by hematoxylin and eosin (H&E) staining. The viability and KLF6 expression of A549 cells treated with different concentrations of LPS were detected by cell counting kit-8 (CCK-8) assay, RT-qPCR, and Western blot analysis. After indicated treatment, the viability and apoptosis of A549 cells were analyzed by CCK-8 and TUNEL assays, and the inflammation factors of A549 cells were detected by Enzyme-linked-immunosorbent serologic assay, RT-qPCR, and Western blot analysis. The combination of KLF6 and CYR61 was determined by chromatin immunoprecipitation (ChIP)-PCR and dual-luciferase reporter assay., Results: KLF6 expression was increased in lung tissues of ALI mice and LPS-induced A549 cells. Interference with KLF6 improved the viability, reduced the inflammatory damage, and promoted the apoptosis of LPS-induced A549 cells. In addition, KLF6 could bind to CYR61. Interference with KLF6 could decrease CYR61 expression in LPS-induced A549 cells. LPS also enhanced the TLR4/MYD88 signaling pathway, which was reversed by KLF6 interference. The above phenomena in LPS-induced A549 cells transfected with Si-KLF6 could be reversed by overexpression of CYR61., Conclusion: Inhibition of KLF6 promoted the viability and reduced the inflammation and apoptosis of LPS-induced A549 cells, which was reversed by CYR61., Competing Interests: The authors had no competing interests to declare.

Tumor-derived extracellular vesicles (EVs) promote ovarian cancer (OC) metastasis by carrying microRNAs (miRs). This study investigated the mechanism of miR-106a-5p carried by OC cell-derived EVs in OC. miR-106a-5p expression in OC tissues and cells was measured. EVs were extracted from SKOV3 cells and normal cells. The internalization of EVs in OC cells was observed. OC cells were treated with SKOV3-EVs or SKOV3-EVs overexpressing miR-106a-5p to detect the proliferation, migration, and invasion. The expression levels of miR-106a-5p, KLF6, and PTTG1 were detected and their binding relationships were identified. Combined experiments were designed to detect the effects of KLF6 and PTTG1 on OC cells. A xenograft tumor experiment was performed to verify the mechanism of EVs-miR-106a-5p and KLF6 in OC metastasis. Consequently, miR-106a-5p was enhanced in OC and correlated with OC metastasis. SKOV3-EVs promoted the proliferation, migration, and invasion of OC cells. Mechanistically, EVs carried miR-106a-5p into other OC cells, inhibited KLF6, reduced the binding of KLF6 to the PTTG1 promoter, and upregulated PTTG1 transcription. Overexpression of KLF6 or silencing of PTTG1 attenuated the promoting effect of EVs-miR-106a-5p on OC cells. EVs-miR-106a-5p facilitated OC metastasis via the KLF6/PTTG1 axis. To conclude, OC cell-derived EVs facilitated the progression and metastasis of OC via the miR-106a-5p/KLF6/PTTG1 axis., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Trophoblast cell differentiation is of paramount importance for successful pregnancy. Krüppel-like factor 6 (KLF6), a transcription factor with diverse roles in cell physiology and tumor biology, is required for trophoblast differentiation through the syncytial pathway. Herein, we demonstrate that extravillous trophoblast (EVT) cell migration and mesenchymal phenotype are increased upon KLF6 downregulation or the expression of a deletion mutant lacking its transcriptional regulatory domain (KΔac). Raman spectroscopy revealed molecular modifications compatible with increased differentiation in cells stably expressing the KΔac mutant. Moreover, abnormally invasive placenta showed lower KLF6 immunostaining compared with the normal placenta. Thus, impaired KLF6 expression or function stimulates EVT migration and differentiation in vitro and may contribute to the physiopathology of the abnormally invasive placenta., (© 2022 Federation of European Biochemical Societies.)

Objective: To explore the effect of circular RNA circ-ABCB10 on the proliferation and apoptosis of laryngeal carcinoma via inhibiting KLF6., Methods: RT-qPCR assay was adopted to detect the expression of circ-ABCB10 and KFL6 in laryngeal carcinoma tissues and cell lines. Cell counting kit-8 (CCK-8) and clone formation assay were employed to detect laryngeal cancer cell viability and proliferation when circ-ABCB10 was silenced or upregulated. In this study, the apoptosis rate was detected by flow cytometry and the protein expression was detected by Western blotting. Wound healing and cross-hole invasion were used to study the migration and invasion of laryngeal cancer cells when circ-ABCB10 was silenced or upregulated., Results: The results of RT-qPCR detection indicated that the expression of circ-ABCB10 in all three laryngeal carcinoma cells was downregulated by 3.2 times compared with that of HaCat cells. There is low expression of circ-ABCB10 in most laryngeal carcinoma tissues, the diagnostic cutoff value of circ-ABCB10 is 0.0008, the area under the curve is 0.718, the sensitivity is 0.981, and the specificity is 0.556. The expression level of KLF6 in laryngeal carcinoma is on the rise, which is significantly higher compared to healthy tissues ( P < 0.05); 48 hours after transfection, RT-qPCR analysis confirmed the transfection efficiency, and upregulation of circ-ABCB10 could significantly promote cell proliferation. Compared with the control group, silencing circ-MTCL1 could inhibit cell proliferation, overexpression of circ-ABCB10 promoted cell migration, and downregulation of circ-ABCB10 significantly inhibited cell movement ( P < 0.001). Upregulation of circ-ABCB10 significantly enhanced the invasiveness and motility of laryngeal cancer cells, while downregulation of circ-ABCB10 was the opposite. Compared with the KLF6 NC group, KLF6 level increased significantly in the KLF6 group, while cell viability, colony formation, scratch healing rate, invasive cell number, and Bcl-2 expression level decreased significantly in the KLF6 group, while apoptosis rate and Bax expression level increased significantly ( P < 0.05). KLF6 level in the si-circ-ABCB10+anti-KLF6 group was significantly lower than that in the si-circ-ABCB10+anti-KLF6-NC group ( P < 0.05). Meanwhile, the cell activity, colony formation number, cell scratch healing rate, number of invaded cells, and Bcl-2 all indicated an upward trend, while the cell apoptosis rate and Bax expression indicated a downward trend ( P < 0.05)., Conclusion: The expression of circ-ABCB10 in laryngeal carcinoma was significantly higher compared to that in paracancerous tissues. Silencing circ-ABCB10 could significantly inhibit the growth and proliferation of laryngeal adenocarcinoma cells, while overexpression of circ-ABCB10 could significantly promote the growth of laryngeal adenocarcinoma cells, probably by inhibiting KLF6 to enhance the proliferation of laryngeal carcinoma and inhibit apoptosis., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Zhuxiang Chen et al.)

Purpose: We aimed to uncover the role of microRNA-181 (miR-181) in the disease onset of diabetic retinopathy (DR) and its underlying mechanism., Methods: MiR-181 levels in plasma and aqueous humor samples of non-proliferative diabetic retinopathy (NPDR), proliferative diabetic retinopathy (PDR) and healthy subjects were analyzed by microarray and quantitative real-time polymerase chain reaction (qRT-PCR). Proliferative and migrative capacities of human retinal endothelial cells (hRECs) regulated by miR-181 were assessed. The binding between miR-181 and Kruppel-like factor 6 (KLF6) was verified by dual-luciferase reporter assay., Results: MiR-181 was upregulated in plasma and aqueous humor samples of NPDR and PDR patients. Overexpression of miR-181 stimulated hRECs to proliferate and migrate. KLF6 was the downstream gene binding miR-181, which was involved in the regulation of hRECs by miR-181., Conclusions: MiR-181 is upregulated in plasma and aqueous humor of DR patients. It enhances proliferative and migratory potentials of retinal endothelial cells by targeting KLF6.

Super-enhancer consists of a large cluster of transcription enhancers that regulates the expression of genes playing an important role in the growth and development of malignant tumors. Recently, several attempts for the identification of super-enhancers have been made, but their functional role in tumor cells remains unclear. This paper aims at elucidating the functional properties of KLF6 super-enhancer related to the growth regulation of HepG2 cells, in relation to transcription factors (TFs). First, some TFs specifying KLF6 super enhancer were identified using CRISPR/Cas9 system and siRNA. Then, their effects on the expression of the target gene KLF6 were assessed. Last, their influence on the proliferation of tumor cells was considered using the MTT method. The study shows that the active enhancers of KLF6 super-enhancer recruit GATA2 and SOX10 TFs to control the expression of the target gene, KLF6. Our findings suggest that the activity of KLF6 super-enhancer is regulated by two TFs (GATA2 and SOX10), and its targeting may be a potential therapeutic strategy for the liver cancer therapy.

Background and Aim: Based on the emerging role of Kruppel-like factor 6 (KLF6) in lipid metabolism, we examined whether there is a relationship between the KLF6 rs3750861 genetic variant and plasma ceramide levels in people with type 2 diabetes mellitus (T2DM)., Methods and Result: We measured six previously identified plasma ceramides, which have been associated with increased cardiovascular risk [Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:0) and Cer(d18:1/24:1)] amongst 101 Caucasian post-menopausal women with T2DM, who consecutively attended our diabetes outpatient service during a 3-month period. Plasma ceramides were measured by targeted liquid chromatography-tandem mass spectrometry assay. Genotyping of the KLF6 rs3750861 polymorphism was performed by TaqMan-Based RT-PCR system. Overall, 87 (86.1%) patients had KLF6 rs3750861 C/C genotype and 14 (13.9%) had C/T or T/T genotypes. After adjustment for age, diabetes-related variables, use of lipid-lowering drugs and other potential confounders, patients with C/T or T/T genotypes had higher plasma Cer(d18:1/18:0) (0.159 ± 0.05 vs. 0.120 ± 0.04 μmol/L, p = 0.012), Cer(d18:1/20:0) (0.129 ± 0.04 vs. 0.098 ± 0.03 μmol/L, p = 0.008), and Cer(d18:1/24:1) (1.236 ± 0.38 vs. 0.978 ± 0.36 μmol/L, p = 0.032) compared with those with C/C genotype., Conclusions: The C/T or T/T genotypes of rs3750861 in the KLF6 gene were closely associated with higher levels of specific plasma ceramides in post-menopausal women with T2DM., Competing Interests: Declaration of competing interest All authors declare no conflicts of interest., (Copyright © 2022 The Italian Diabetes Society, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.)

Long non-coding RNA (lncRNA) ZFAS1 (zinc finger antisense 1) was demonstrated to play critical roles in various cancer progression. However, the functions of ZFAS in cervical cancers (CC) are unclear. Human CC cell lines were used for in vitro experiments. RT-qPCR (Real Time Quantitative PCR) was performed to detect the expression of ZFAS1, microRNA-190a-3p (miR-190a-3p) and Kruppel-like factor 6 (KLF6). Cell proliferation, invasion and migration assays were used to investigate biological behaviors of CC cells related to CC progression. The relationship of KLF6 to ZFAS1 and miR-190a-3p was analyzed by circRIP and luciferase reporter assay. In addition, in vivo experiment was carried out to explore the function of ZFAS1 in tumor growth of CC. The expression levels of ZFAS1 and KLF6 were both significantly elevated, while the expression of miR-190a-3p was inhibited in CC tumor tissues. In addition, ZFAS1 influenced CC tumor growth through miR-190a-3p. KLF6 was a target of miR-190a-3p and inhibited miR-190a-3p-induced CC tumor growth. Furthermore, KLF6 was negatively regulated by miR-190a-3p, but positively regulated by ZFAS1. Overexpression of ZFAS1 and inhibition of miR-190a-3p significantly increased the expression levels of KLF6. Finally, in vitro assays demonstrated that inhibition of ZFAS1 reduced CC tumor growth and the expression levels of KLF6, but increased the expression levels of miR-190a-3p. ZFAS1 could regulate CC pathogenesis via regulating the miR-190a-3p/KLF6 axis, which might be considered as new CC therapeutic targets.

Purpose: Research in this article was performed to explore the biological role and clinical significance of Krüppel-like transcription factor 6 (KLF6) in non-small-cell lung cancer (NSCLC)., Methods: KLF6 expression in NSCLC cell lines was analyzed using reverse transcription PCR and Western blot. The expressed KLF6 protein was examined in 50 surgical NSCLC tissues using immunohistochemistry. Statistical analyses were employed for clinical association examinations. CCK8 assay and Annexin V/PI analysis were used to execute cell proliferation and apoptosis in KLF6-overexpression cell lines and the control groups. Cleaved caspase-3 expression was also detected in KLF6-overexpression cells and NSCLC tissues. KLF6 expression correlation with cleaved caspase-3 was also examined., Results: It was discovered that downregulation of KLF6 was seen in human NSCLC cell lines. Low KLF6 expression in NSCLC tissues was correlated with poor patient prognosis ( P < 0.005); patients with less KLF6 expression possessed a lower cumulative 5-year survival rate. Multivariate analysis showed KLF6 expression as an independent prognostic indicator for NSCLC individuals. Expression levels of KLF6 were associated with NSCLC tumor size ( P = 0.041). Overexpression of KLF6 inhibited cell proliferation and stimulated A549 and H322 cell line apoptosis. Cleaved caspase-3 protein had higher expression levels in KLF6-overexpressed cells than in the control group. The KLF6 expression levels were positively related to the cleaved caspase-3 protein expression in NSCLC tissues ( r = 0.689, P = 0.001)., Conclusions: The results indicate that downregulation of KLF6 is a significant NSCLC progression marker. KLF6 prevents cell growth and promotes cell apoptosis, possibly caspase-3 activations., Competing Interests: The authors declare that they have no conflict of interest., (Copyright © 2022 Binbin Zeng et al.)

Marbling influences the taste and tenderness of meat and is the main determinant of carcass quality in many countries. This study aims to investigate the influence of KLF6 (Kruppel Like Factor 6) and associated molecular mechanisms on lipid metabolism in bovine adipocytes. In the current study, KLF6 gene expression was down regulated via siRNA (small interfering RNA) in bovine adipocytes in vitro. Subsequently, adipogenic cells were collected from the culture media after 9 days, and subjected to fluorescent imaging and RNA sequencing. After confirming that KLF6 was down regulated in bovine adipocytes by siRNA, differential gene expression analysis was used to characterize the infuence of KLF6 on gene expression profiles in bovine adipocytes. A total of 10,812 genes were characterized as differentially expressed genes (DEGs) of which, 109 were up-regulated and 62 were down-regulated genes. KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis identified that the DEGs were associated with lipid metabolism, carbohydrate metabolism, cell growth and death, cancer, and the signaling pathways for calcium, AMPK (Adenosine Monophosphate-Activated Protein Kinase), PI3K-Akt (Phosphatidylinositol 3-kinase), PPAR (Peroxisome proliferator-activated receptors), MAPK (mitogen-activated protein kinase), cAMP (Cyclic adenosine monophosphate), and Wnt (Wingless-related integration site). Similarly, gene ontology analysis indicated that down-regulation of KLF6 gene significantly up regulated the genes that regulate adipogenesis, differentiation and regulation of adipocytes and homeostasis of bovine adipocytes, specifically regulating the cell-type specific apoptotic action, negative regulation of apoptotic pathways, programmed cell death, and growth. Results indicate that KLF6 has a role in regulating lipid metabolism in bovine adipocytes. These findings provide evidence that may inform further investigations into molecular mechanisms that underlie the role of bovine KLF6 gene in regulating adipogenesis., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Purpose: To analyse the relationship between the transcriptional expression of Krüppel-like factor-6 (KLF6) and local response to treatment with radiotherapy in patients with head and neck squamous cell carcinoma (HNSCC)., Methods: We determined the transcriptional expression of KLF6 in tumour biopsies obtained before treatment with radiotherapy in 83 HNSCC patients. The KLF6 expression was categorized according to the local control of the disease with a recursive partitioning analysis., Results: During the follow-up period, 27 patients (32.5%) had a local recurrence of the tumour. Patients with local recurrence had significantly higher levels of KLF6 expression than patients in which radiotherapy achieved local control of the disease (P = 0.029). Five-year local recurrence-free survival for patients with a high transcriptional expression of KLF6 (n = 46) was 51.1% (95% CI 36.4-66.2%), and for patients with low expression it was 85.6% (95% CI 73.9-97.3%) (P = 0.0001). The results of a multivariate analysis showed that patients with a high KLF6 expression had a 3.8 times higher risk of local recurrence after treatment with radiotherapy (95% CI 1.4-10.5, P = 0.008)., Conclusion: Transcriptional expression of KLF6 was significantly related to local control in HNSCC patients treated with radiotherapy. Patients with high levels of KLF6 expression had a significantly higher risk of local recurrence after treatment., (© 2021. Federación de Sociedades Españolas de Oncología (FESEO).)

Background: To further understand the development of the spinal cord, an exploration of the patterns and transcriptional features of spinal cord development in newborn mice at the cellular transcriptome level was carried out., Methods: The mouse single-cell sequencing (scRNA-seq) dataset was downloaded from the GSE108788 dataset. Single-cell RNA-Seq (scRNA-Seq) was conducted on cervical and lumbar spinal V2a interneurons from 2 P0 neonates. Single-cell analysis using the Seurat package was completed, and marker mRNAs were identified for each cluster. Then, pseudotemporal analysis was used to analyze the transcription changes of marker mRNAs in different clusters over time. Finally, the functions of these marker mRNAs were assessed by enrichment analysis and protein-protein interaction (PPI) networks. A transcriptional regulatory network was then constructed using the TRRUST dataset., Results: A total of 949 cells were screened. Single-cell analysis was conducted based on marker mRNAs of each cluster, which revealed the heterogeneity of neonatal mouse spinal cord neuronal cells. Functional analysis of pseudotemporal trajectory-related marker mRNAs suggested that pregnancy-specific glycoproteins (PSGs) and carcinoembryonic antigen cell adhesion molecules (CEACAMs) were the core mRNAs in cluster 3. GSVA analysis then demonstrated that the different clusters had differences in pathway activity. By constructing a transcriptional regulatory network, USF2 was identified to be a transcriptional regulator of CEACAM1 and CEACAM5, while KLF6 was identified to be a transcriptional regulator of PSG3 and PSG5. This conclusion was then validated using the Genotype-Tissue Expression (GTEx) spinal cord transcriptome dataset., Conclusions: This study completed an integrated analysis of a single-cell dataset with the utilization of marker mRNAs. USF2/CEACAM1&5 and KLF6/PSG3&5 transcriptional regulatory networks were identified by spinal cord single-cell analysis., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2021 Xinbing Liu et al.)

Background: Gastric cell carcinoma (GCC) is a common and high-incidence malignant gastrointestinal cancer that seriously threatens human life and safety. Evidences suggest that microRNAs (miRNAs) exhibit an essential role in regulating the occurrence and development of GCC, while the effects and possible mechanisms remain to be further explored., Objective: This study was designed to explore whether miR-200c-3p exerted its functional role in the growth and metastasis of GCC, and investigate the possible mechanisms., Methods: The expression levels of miR-200c-3p in GCC tissues and cell lines were detected by qRT-PCR analysis. The functional role of miR-200c-3p in the viability, proliferation, migration and invasion of GCC cells were evaluated by CCK-8, EdU, wound healing and Transwell assays. In addition, the candidate targets of miR-200c-3p was predicted and confirmed by dual-luciferase reporter assay. Moreover, the relationship between miR-200c-3p and target (Krüppel like factor 6, KLF6) was assessed by qRT-PCR and western blot assays. Besides, the expression levels of KLF6 in GCC cells were determined by qRT-PCR and western blot assays. Furthermore, the role of KLF6 in the viability, proliferation, migration and invasion of GCC cells mediated with miR-200c-3p mimics was evaluated by CCK-8, EdU, wound healing and Transwell assays., Results: In the present study, a new tumor promoting function of miR-200c-3p was disclosed in GCC. We found that the expression of miR-200c-3p was obviously increased in clinic GCC tissues and cell lines. In addition, down-regulation of miR-200c-3p suppressed cell viability, proliferation, migration, and invasion in GCC cells. Moreover, KLF6 was verified as a direct target of miR-200c-3p by binding its 3'-UTR. Additionally, KLF6 was remarkably decreased and was negatively associated with the miR-200c-3p expression in GCC cell lines. Furthermore, over-expression of KLF6 retarded the effects of miR-200c-3p on the growth and metastasis of GCC cell lines., Conclusions: MiR-200c-3p potentially played a tumor-promoting role in the occurrence and development of GCC, which may be achieved by targeting KLF6., (© 2021. The Author(s).)

Ischemic stroke (IS), caused by a permanent or transient local reduction in blood supply to the brain, is one of the most widespread causes of public health problems in modern society. Long non-coding RNA (LncRNA) has been reported to be related to angiogenesis following IS. In this study, we explored the effect and potential molecular mechanism of lncRNA homeobox antisense non-coding RNA (HOTAIR) in IS. Permanent middle cerebral artery occlusion (pMCAO) model and oxygen and glucose deprivation (OGD) model were established. HOTAIR was increased in vivo and in vitro models post-ischemic. HOTAIR knockdown promoted neurological function recovery, manifesting in decreased modified neurological severity score, cerebral infarcted area, apoptosis and inflammation, and improved balance ability, spatial learning and memory ability. Silencing HOTAIR also improved the viability of OGD-induced N2a cells, and attenuated apoptosis and inflammation. HOTAIR can compete with KLF6 to bind to miR-148a-3p. miR-148a-3p knockdown or KLF6 overexpression partially reversed the effect of sh-HOTAIR on OGD-induced N2a cells. HOTAIR suppressed the activation of STAT3 pathway via the miR-148a-3p/KLF6 axis. To summarize, this study demonstrated that lncRNA HOTAIR absorbed miR-148a-3p and up-regulated KLF6 expression through ceRNA mechanism, and inhibited STAT3 pathway, promoted apoptosis and inflammation, and aggravated neurological injury post-IS., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Breast cancer (BC) is the most commonly occurring malignancy in women. This study aimed to investigate the functions of the long noncoding RNA ZBED3-AS1 (ZBED3-AS1) in BC and its molecular mechanisms. qRT-PCR was conducted to access the expression of ZBED3-AS1, microRNA-513a-5p (miR-513a-5p), and Kruppel like factor 6 (KLF6) in BC. Additionally, BC cell viability and proliferative capacity were measured by MTT and 5-Ethynyl-20-deoxyuridine (EdU) assays. A transwell assay was used for evaluating BC cell migration and invasion. The interactions among ZBED3-AS1, miR-513a-5p, and KLF6 in BC were confirmed by dual-luciferase reporter assay. Furthermore, feedback approaches were performed to determine whether ZBED3-AS1 influences BC cell behaviors by regulating the miR-513a-5p/KLF6 axis. The murine xenograft model was established to assess the effect of ZBED3-AS1 on tumor growth. The expression of ZBED3-AS1 and KLF6 was reduced, while miR-513a-5p expression was elevated in BC. ZBED3-AS1 elevation attenuated the malignant behaviors of BC cells, including viability, proliferative capacity, migration, and invasion. Mechanical experiments revealed that ZBED3-AS1 targeted miR-513a-5p, and miR-513a-5p targeted KLF6 in BC. Feedback approaches validated that miR-513a-5p overexpression or KLF6 depletion reversed the inhibitory effects of ZBED3-AS1 upregulation on viability, proliferative capacity, migration, and invasion of BC cells. Furthermore, ZBED3-AS1 elevation attenuated the tumor growth in the murine xenograft model. ZBED3-AS1 hindered the malignant development of BC cells by regulating the miR-513a-5p/KLF6 axis, providing a novel therapeutic target in BC., (© 2021 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Background: The heart is one of the target organs vulnerable to sepsis. About 50% of sepsis patients will suffer from myocardial injury and cardiac dysfunction, which will aggravate the sepsis and affect its prognosis., Objectives: Here, we attempt to investigate the function of long non coding RNA PVT1 in LPS-induced cardiac fibroblasts in vitro, and explore its potential mechanism., Methods: The expression of PVT1 in LPS-induced cardiac fibroblasts was detected by qRT-PCR. CCK-8 assay, cell migration, qRT-PCR and western blotting analysis were applied to evaluating the effect of PVT1 knockdown on LPS-induced cardiac fibroblasts. The bioinformatics analysis and the rescue experiment were devoted to the underlying mechanism., Results: PVT1 expression was up-regulated in LPS-induced cardiac fibroblasts. And knockdown of PVT1 inhibited cell viability and migration, alleviated inflammation cytokines production of LPS-treated cardiac fibroblasts. The bioinformatics analysis predicted PVT1 negatively regulates miR-24 and KLF6 is a direct target of miR-24., Conclusions: In a word, we observed PVT1 expression level was up-regulated in LPS- treated cardiac fibroblasts. PVT1 knockdown could alleviate LPS-induced biological behavior of cardiac fibroblasts through sponging miR-24 in vitro., (© 2021. The Genetics Society of Korea.)

Increasing evidences have confirmed that long non-coding RNA LOXL1-AS1 functions in multiple human diseases. Here, we aim to explore the function and mechanism of LOXL1-AS1 in modulating oxidized low-density lipoprotein (ox-LDL)-induced angiogenesis of endothelial cells (ECs). Presently, we found that LOXL1-AS1 and KLF6 were upregulated in ECs treated by Ox-LDL in a dose- and time-dependent manner while miR-590-5p was downregulated. Overexpression of LOXL1-AS1 aggravated Ox-LDL mediated ECs proliferation and migration, and promoted angiogenesis both in vitro and in vivo . On the contrary, enhancing miR-590-5p or inhibiting LOXL1-AS1 level led to suppressive effects on the proliferation, migration and angiogenesis of ECs. Moreover, LOXL1-AS1 upregulation promoted the expression of vascular endothelial growth factor (VEGF), MMPs (including MMP2, MMP9 and MMP14) and also activated VEGF/VEGFR2/PI3K/Akt/eNOS pathway. Mechanistically, LOXL1-AS1 works as a competitive endogenous RNA (ceRNA) by sponging miR-590-5p, which targeted at the 3'-untranslated region (3'UTR) of KLF6. Additionally, the proliferation, migration and angiogenesis of ECs were elevated following KLF6 upregulation. By detecting the expression of LOXL1-AS1 and miR-590-5p in the serum of healthy donors and atherosclerosis patients, it was found that LOXL1-AS1 was upregulated in atherosclerosis patients (compared with healthy donors) and had a negative relationship with miR-590-5p. Taken together, LOXL1-AS1 promoted Ox-LDL induced angiogenesis via regulating miR-590-5p-modulated KLF6/VEGF signaling pathway. The LOXL1-AS1-miR-590-5p axis exerts a novel role in the progression of atherosclerosis.

Altered cellular metabolism in kidney proximal tubule (PT) cells plays a critical role in acute kidney injury (AKI). The transcription factor Krüppel-like factor 6 (KLF6) is rapidly and robustly induced early in the PT after AKI. We found that PT-specific Klf6 knockdown ( Klf6 PTKD ) is protective against AKI and kidney fibrosis in mice. Combined RNA and chromatin immunoprecipitation sequencing analysis demonstrated that expression of genes encoding branched-chain amino acid (BCAA) catabolic enzymes was preserved in Klf6 PTKD mice, with KLF6 occupying the promoter region of these genes. Conversely, inducible KLF6 overexpression suppressed expression of BCAA genes and exacerbated kidney injury and fibrosis in mice. In vitro , injured cells overexpressing KLF6 had similar decreases in BCAA catabolic gene expression and were less able to utilize BCAA. Furthermore, knockdown of BCKDHB , which encodes one subunit of the rate-limiting enzyme in BCAA catabolism, resulted in reduced ATP production, while treatment with BCAA catabolism enhancer BT2 increased metabolism. Analysis of kidney function, KLF6 , and BCAA gene expression in human chronic kidney disease patients showed significant inverse correlations between KLF6 and both kidney function and BCAA expression. Thus, targeting KLF6-mediated suppression of BCAA catabolism may serve as a key therapeutic target in AKI and kidney fibrosis., Competing Interests: Competing interest statement: J.C.H. declares a Shangpharma research grant of $250,000 per year in 2019 to 2020 and is a Scientific Advisory Board member for RenalytixAI.

MicroRNA-543-3p (miR-543-3p) has been reported to be involved in many human disease's progression, but its role in inflammation is still unclear. After bacterial infection, innate immune cells are activated to trigger inflammation by recognizing lipopolysaccharide (LPS) on the bacterial outer membrane. In our research, it showed that miR-543-3p was down-regulated in LPS-treated periodontal ligament cells (PDLCs). And it mediated the apoptosis of PDLC induced by LPS, which may be involved in periodontitis development. Besides, up-regulation of miR-543-3p alleviated the inflammatory damage induced by LPS. Furthermore, our research demonstrated Kruppel-like factor 6 (KLF6) served as a direct downstream target of miR-543-3p to play a vital role in periodontitis. Simply put, these findings suggest that miR-543-3p could down-regulate inflammation and inhibit periodontitis by targeting KLF6, and it provides a new insight into the molecular mechanism of periodontitis, which may be helpful for the early diagnosis and treatment of this disease., (© 2021 The Author(s).)

Transcription factors (TFs) act as powerful levers to regulate neural physiology and can be targeted to improve cellular responses to injury or disease. Because TFs often depend on cooperative activity, a major challenge is to identify and deploy optimal sets. Here we developed a bioinformatics pipeline, centered on TF co-occupancy of regulatory DNA, and used it to predict factors that potentiate the effects of pro-regenerative Klf6 in vitro. High content screens of neurite outgrowth identified cooperative activity by 12 candidates, and systematic testing in a mouse model of corticospinal tract (CST) damage substantiated three novel instances of pairwise cooperation. Combined Klf6 and Nr5a2 drove the strongest growth, and transcriptional profiling of CST neurons identified Klf6/Nr5a2-responsive gene networks involved in macromolecule biosynthesis and DNA repair. These data identify TF combinations that promote enhanced CST growth, clarify the transcriptional correlates, and provide a bioinformatics approach to detect TF cooperation.

Krüppel-like factors (KLFs) are a highly conserved family of transcription factors. We analysed expression profile data of KLFs and identified KLF6 as a new potential regulator of erythropoiesis. Knocking down the expression of KLF6 significantly raised γ-globin mRNA and protein levels in the erythroid cell line HUDEP-2 and haematopoietic progenitor (CD34 + ) cells. We found that overexpression of microRNA (miR)-2355-5p in HUDEP-2 and CD34 + cells correlated with increased γ-globin synthesis by suppressing expression of KLF6. Our discovery that the interaction between miR-2355-5p and KLF6 affects the expression of γ-globin may provide more information for the clinical management of β-thalassaemia patients., (© 2020 British Society for Haematology and John Wiley & Sons Ltd.)

Psychiatric disorders are highly genetically correlated, but little research has been conducted on the genetic differences between disorders. We developed a new method (case-case genome-wide association study; CC-GWAS) to test for differences in allele frequency between cases of two disorders using summary statistics from the respective case-control GWAS, transcending current methods that require individual-level data. Simulations and analytical computations confirm that CC-GWAS is well powered with effective control of type I error. We applied CC-GWAS to publicly available summary statistics for schizophrenia, bipolar disorder, major depressive disorder and five other psychiatric disorders. CC-GWAS identified 196 independent case-case loci, including 72 CC-GWAS-specific loci that were not significant at the genome-wide level in the input case-control summary statistics; two of the CC-GWAS-specific loci implicate the genes KLF6 and KLF16 (from the Krüppel-like family of transcription factors), which have been linked to neurite outgrowth and axon regeneration. CC-GWAS loci replicated convincingly in applications to datasets with independent replication data.

Small nucleolar RNA host gene 6 (SNHG6) is a newly discovered long non-coding RNA (lncRNA), while the regulatory mechanism of SNHG6 in chondrosarcoma is largely unknown. Here we found that SNHG6 expression was upregulated and showed positive correlation with the progression of chondrosarcoma. Functional assays demonstrated that SNHG6 was required for the proliferation, migration, and invasion of chondrosarcoma cells. Mechanistic study revealed that SNHG6 could recruit EZH2 and maintain high level of H3K27me3 to repress the transcription of tumor-suppressor genes, including KLF6. KLF6 was found to bind to the promoter region of SP1 and restrained its transcription, while SP1 could be recruited to the promoter region of SNHG6 and promoted its transcription to form a positive loop. In summary, this study reveals that SP1-induced SNHG6 forms a positive loop to facilitate the carcinogenesis of chondrosarcoma through the suppression of KLF6 by recruiting EZH2, which manifests the oncogenic function of SNHG6 in chondrosarcoma.

Oncogenic K-Ras (K-Ras G12V ) promotes senescence in normal cells but fuels transformation of cancer cells after the senescence barrier is bypassed. The mechanisms regulating this pleiotropic function of K-Ras remain to be fully established and bear high pathological significance. We find that K-Ras G12V activates the angiotensinogen (AGT) gene promoter and promotes AGT protein expression in a Kruppel-like factor 6-dependent manner in normal cells. We show that AGT is then converted to angiotensin II (Ang II) in a cell-autonomous manner by cellular proteases. We show that blockade of the Ang II receptor type 1 (AT 1 -R) in normal cells inhibits oncogene-induced senescence. We provide evidence that the oncogenic K-Ras-induced synthesis of Ang II and AT 1 -R activation promote senescence through caveolin-1-dependent and nicotinamide adenine dinucleotide phosphate oxidase 2-mediated oxidative stress. Interestingly, we find that expression of AGT remains elevated in lung cancer cells but in a Kruppel-like factor 6-independent and high-mobility group AT-hook 1-dependent manner. We show that Ang II-mediated activation of the AT 1 -R promotes cell proliferation and anchorage-independent growth of lung cancer cells through a STAT3-dependent pathway. Finally, we find that expression of AGT is elevated in lung tumors of K-Ras LA2-G12D mice, a mouse model of lung cancer, and human lung cancer. Treatment with the AT 1 -R antagonist losartan inhibits lung tumor formation in K-Ras LA2-G12D mice. Together, our data provide evidence of the existence of a novel cell-autonomous and pleiotropic Ang II-dependent signaling pathway through which oncogenic K-Ras promotes oncogene-induced senescence in normal cells while fueling transformation in cancer cells., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Circular RNAs (circRNAs) could regulate gene expression which may induce tumor occurrence and progression. In the current study, we first investigated the expression of circMTO1 in osteosarcoma, and the underlying mechanism was further elucidated., Patients and Methods: Circular RNA microarrays were used to identify the differential expression of circRNAs in osteosarcoma tissues and the corresponding normal tissues. qRT-PCR was used to examine the level of circMTO1 in osteosarcoma tissues and cell lines. In addition, circMTO1 overexpression was constructed using lentiviral transfection in cell lines. Subsequently, the Cell Counting Kit-8 (CCK8), cell migration and invasion, and flow cytometry were used to investigate the effect of circMTO1 on the biological functions of cells. The Western Blot and the recovery experiments were used to explore the potential mechanism., Results: Here, we measured 20 circRNAs which were downregulated in osteosarcoma tissues using circRNA microarray. CircMTO1 expression was decreased in osteosarcoma cell lines. Besides, circMTO1 could inhibit cell proliferation, migration and invasion, and induced apoptosis in osteosarcoma cells. Bioinformatics analysis showed that circMTO1 serves as a sponge for miR-630 and KLF6 is a direct target of miR-630. Furthermore, circMTO1 functions through regulation of miR-630/KLF6 axis., Conclusions: Our study suggests circMTO1 could suppress osteosarcoma progression by regulating miR-630/KLF6 axis, which may highlight the diagnostic and therapeutic potential of these molecules in osteosarcoma treatment.

Background: Krüppel-like factor 6 (KLF6) is a transcription factor that participate in various pathophysiological processes, but its contribution in ischemia acute kidney injury (AKI) is lacking so far. The study aimed to investigate the expression and the role of KLF6 in kidney ischemia-reperfusion (IR) injury., Method: Microarray data were collected from GSE58438 and GSE52004. The rat IR model was established to evaluate the mRNA and protein expression of KLF6 and inflammatory cytokines in serum and kidney tissues. SiRNA-KLF6 was transfected with HK-2 cells, and then a cell-based hypoxia-reoxygenation (HR) model was established., Results: Bioinformatics showed KLF6 mRNA in kidney tissue is up-regulated in 3 h after IR in rat kidney, which involved in cell activation, leukocyte activation, and response to hydrogen peroxide after IR. The rat IR model results showed that KLF6 expression was peaking at 6 h, and the expression of pro-inflammatory cytokines MCP-1 and TNF-α was increased both in serum and kidney tissues, while anti-inflammatory cytokine IL-10 was decreased after IR. Furthermore, in vitro results showed that KLF6 knock-down reduced the pro-inflammatory cytokines expression., Conclusion: These results suggest that (1) KLF6 might be a novel biomarker for early diagnosis of AKI and (2) KLF6 may play a role in promoting inflammation in AKI.