USP7 alleviates neuronal inflammation and apoptosis in spinal cord injury via deubiquitinating NRF1/KLF7 axis.

BACKGROUND: Ubiquitin-specific protease 7 (USP7) has been found to be associated with motor function recovery after spinal cord injury (SCI). Therefore, its role and mechanism in SCI process need further exploration. METHODS: SCI rat models were established via performing laminectomy at the T9-T11 spinal vertebrae and cutting spinal cord tissues. SCI cell models were constructed by inducing PC12 cells with lipopolysaccharide (LPS). The protein levels of USP7, nuclear respiratory factor 1 (NRF1), Krüppel-like factor 7 (KLF7) and apoptosis-related markers were detected by western blot. Cell viability and apoptosis were tested by cell counting kit-8 assay and flow cytometry. The contents of inflammatory factors were examined using ELISA. The interaction between NRF1 and USP7 or KLF7 was analyzed by co-immunoprecipitation assay, chromatin immunoprecipitation assay and dual-luciferase reporter assay, respectively. RESULTS: USP7 was downregulated in SCI rat models and LPS-induced PC12 cells. Overexpressed USP7 promoted viability, while repressed apoptosis and inflammation in LPS-induced PC12 cells. USP7 could stabilize NRF1 protein expression via deubiquitination, and NRF1 knockdown reversed the protective effect of USP7 against LPS-induced PC12 cell injury. NRF1 is bound to KLF7 promoter to enhance its transcription. NRF1 overexpression inhibited LPS-induced PC12 cell inflammation and apoptosis via increasing KLF7 expression. CONCLUSION: USP7 alleviated inflammation and apoptosis in LPS-induced PC12 cells via NRF1/KLF7 axis, indicating that targeting of USP7/NRF1/KLF7 axis might be a promising treatment strategy for SCI.

KLF7 promotes neuroblastoma differentiation through the GTPase signaling pathway by upregulating neuroblast differentiation-associated protein AHNAKs and glycerophosphodiesterase GDPD5.

The arrest of neural crest-derived sympathoadrenal neuroblast differentiation contributes to neuroblastoma formation, and overriding this blocked differentiation is a clear strategy for treating high-risk neuroblastoma. A better understanding of neuroblast or neuroblastoma differentiation is essential for developing new therapeutic approaches. It has been proposed that Krueppel-like factor 7 (KLF7) is a neuroblastoma super-enhancer-associated transcription factor gene. Moreover, KLF7 was found to be intensely active in postmitotic neuroblasts of the developing nervous system during embryogenesis. However, the role of KLF7 in the differentiation of neuroblast or neuroblastoma is unknown. Here, we find a strong association between high KLF7 expression and favorable clinical outcomes in neuroblastoma. KLF7 induces differentiation of neuroblastoma cells independently of the retinoic acid (RA) pathway and acts cooperatively with RA to induce neuroblastoma differentiation. KLF7 alters the GTPase activity and multiple differentiation-related genes by binding directly to the promoters of neuroblast differentiation-associated protein (AHNAK and AHNAK2) and glycerophosphodiester phosphodiesterase domain-containing protein 5 (GDPD5) and regulating their expression. Furthermore, we also observe that silencing KLF7 in neuroblastoma cells promotes the adrenergic-to-mesenchymal transition accompanied by changes in enhancer-mediated gene expression. Our results reveal that KLF7 is an inducer of neuroblast or neuroblastoma differentiation with prognostic significance and potential therapeutic value.

E2F transcription factor 5, a new regulator in adipogenesis to mediate the role of Krüppel-like factor 7 in chicken preadipocyte differentiation and proliferation.

E2F transcription factor 5 (E2F5) gene is a transcription factor, plays an important role in the development of a variety of cells. E2F5 is expressed in human and mouse adipocytes, but its specific function in adipogenesis is unclear. Krüppel-like factor 7 (KLF7) facilitates proliferation and inhibits differentiation in chicken preadipocytes. Our previous KLF7 chromatin immunoprecipitation-sequencing analysis revealed a KLF7-binding peak in the 3' flanking region of the E2F5, indicating a regulatory role of KLF7 in this region. In the present study, we investigated E2F5 potential role, the overexpression and knockdown analyses revealed that E2F5 inhibited the differentiation and promoted the proliferation of chicken preadipocytes. Moreover, we identified enhancer activity in the 3' flanking region (nucleotides +22661/+22900) of E2F5 and found that KLF7 overexpression increased E2F5 expression and luciferase activity in this region. Deleting the putative KLF7-binding site eliminated the promoting effect of KLF7 overexpression on E2F5 expression. Further, E2F5 reversed the KLF7-induced decrease in preadipocyte differentiation and increase in preadipocyte proliferation. Taken together, our findings demonstrate that KLF7 inhibits differentiation and promotes proliferation in preadipocytes by enhancing E2F5 transcription.

KLF7 promotes colon adenocarcinoma progression through the PDGFB signaling pathway.

Colon adenocarcinoma (COAD) is the most common malignancy of the digestive tract, which is characterized by a dismal prognosis. No effective treatment has been established presently, thus there is an urgent need to understand the mechanisms driving COAD progression in order to develop effective therapeutic approaches and enhance clinical outcomes. In this study, we found that KLF7 is overexpressed in COAD tissues and correlated with clinicopathological features of COAD. Both gain-of-function and loss-of-function experiments have unequivocally demonstrated that overexpression of KLF7 promotes the growth and metastasis of COAD in vitro and in vivo, while KLF7 knockdown attenuated these effects. Mechanistically, our findings reveal that KLF7 can specifically bind to the promoter region of PDGFB (TGGGTGGAG), thus promoting the transcription of PDGFB and increasing its secretion. Subsequently, secreted PDGFB facilitates the progression of COAD by activating MAPK/ERK, PI3K/AKT, and JAK/STAT3 signaling pathways through PDGFRß. Additionally, we found that sunitinib can block PDGFB signaling and inhibit COAD progression, offering a promising therapeutic strategy for COAD treatment.

Palmitic acid-activated GPRs/KLF7/CCL2 pathway is involved in the crosstalk between bone marrow adipocytes and prostate cancer.

BACKGROUND: Obesity-induced abnormal bone marrow microenvironment is one of the important risk element for bone metastasis in prostate cancer (PCa). The present study aimed to determine whether obesity-induced elevation in palmitic acid (PA), which is the most abundant of the free fatty acids (FFAs), increased CCL2 via the GPRs/KLF7 pathway in bone marrow adipocytes (BMA) to facilitate PCa growth and metastasis. METHODS: We constructed a bone-tumor bearing mouse model with obesity through high-fat diet, and observed the tumor formation ability of PCa cells. In vitro, observe the effect of PA on the expression level of CCL2 in BMA through GPRs/KLF7 signaling pathway. After co-culture of BMA and PCa cells, CCK8 assay and transwell experiment were used to detect the changes in biological behavior of PCa cells stimulated by BMA. RESULTS: The BMA distribution in the bone marrow cavity of BALB/c nude mice fed with the high-fat diet (HFD) was evidently higher than that in the mice fed with the normal diet (ND). Moreover, HFD-induced obesity promoted KLF7/CCL2 expression in BMA and PCa cell growth in the bone marrow cavity of the mice. In the vitro experiment, a conditioned medium with increased CCL2 obtained from the BMA cultured with PA (CM-BMA-PA) was used for culturing the PCa cell lines, which evidently enhanced the proliferation, invasion, and migration ability. KLF7 significantly increased the CCL2 expression and secretion levels in BMA by targeting the promoter region of the CCL2 gene. In addition, GPR40/120 engaged in the PA-induced high KLF7/CCL2 levels in BMA to facilitate the malignant progression of PC-3 cells. CONCLUSIONS: PA-activated GPRs/KLF7/CCL2 pathway in BMA facilitates prostate cancer growth and metastasis.

The active phthalate metabolite, DHEP, induces proliferation and metastasis of prostate cancer cells via upregulation of ß-catenin and downregulation of KLF7.

Phthalates such as DHEP are among the widely used compounds in industry. It has been shown that DHEP can convey various biological consequences in mammalian cells, among them, the carcinogenic effects of DHEP are emphasized. The present study aimed to assess the impact of DHEP exposure on the proliferation and invasiveness of DU145 prostate cancer cells through in vitro and in vivo models. The DU145 cells were treated with increasing concentrations of DHEP and the tumorigenic parameters were analyzed. KLF7 as a probable mediator of the effect of DHEP was either overexpressed or knocked down in DU145 to evaluate the probable impact of KLF7 on the biological effects of DHEP. The effect of DHEP was also studied in a DU145 xenograft tumor model. The moderate doses of DHEP increased the proliferation and migration of DU145 cells. In the case of gene expression patterns, DHEP reduced the levels of p53 and KLF7 while elevated the expression of ß-catenin. The knock-down of KLF7 conveyed comparable effects to that of DHEP to some degree and increased the proliferation of DU145 cells, while the transduction of KLF7 increased the expressions of p53 and p21 along with controlling the tumor size. The present study demonstrated the potential of DHEP in increasing the tumorigenic properties of DU145 cells along with a focus on the underlying mechanisms. Sustained exposure to DHEP can cause a dysregulation in balance between oncogenes and tumor suppressor genes which is the hallmark of malignant transformation. Thus, special considerations seem necessary for the safe exploitation of phthalates.

KLF7 promotes adipocyte inflammation and glucose metabolism disorder by activating the PKCζ/NF-κB pathway.

In the obesity context, inflammatory cytokines secreted by adipocytes lead to insulin resistance and are key to metabolic syndrome development. In our previous study, we found that the transcription factor KLF7 promoted the expression of p-p65 and IL-6 in adipocytes. However, the specific molecular mechanism remained unclear. In the present study, we found that the expression of KLF7, PKCζ, p-IκB, p-p65, and IL-6 in epididymal white adipose tissue (Epi WAT) in mice fed a high-fat diet (HFD) was significantly increased. In contrast, the expression of PKCζ, p-IκB, p-p65, and IL-6 was significantly decreased in Epi WAT of KLF7 fat conditional knockout mice. In 3T3-L1 adipocytes, KLF7 promoted the expression of IL-6 via the PKCζ/NF-κB pathway. In addition, we performed luciferase reporter and chromatin immunoprecipitation assays, which confirmed that KLF7 upregulated the expression of PKCζ transcripts in HEK-293T cells. Collectively, our results show that KLF7 promotes the expression of IL-6 by upregulating PKCζ expression and activating the NF-κB signaling pathway in adipocytes.

Caprylic Acid (FFA C8:0) promotes the progression of prostate cancer by up-regulating G protein-coupled receptor 84/ Krüppel-like factor 7.

BACKGROUND: In previous study, we found that the content of medium-chain fatty acid Caprylic Acid (FFA C8:0) may be an important risk factor of obesity induced prostate cancer (PCa). However, the relationship between FFA C8:0 and PCa has not been reported. In this study, we explored whether the FFA C8:0 can promotes the progression of PCa by up-regulating Krüppel-like factor 7 (KLF7). METHODS: We collected tissues from PCa patients and Benign Prostate Hyperplasia (BPH), constructed a primary-tumor bearing mouse model with obesity through high-fat diet, and observed the tumor formation ability of PCa cells. In vitro, CCK8 assay, plate cloning, Transwell and scratch experiment were used to detect the changes in biological behavior of PCa cells stimulated by FFA C8:0. RESULTS: First, we found that the expression level of KLF7 is higher in PCa tissues of patients, and the expression of KLF7 is positively correlated with tumour-promoting gene IL-6, while it is negative correlated with another tumour-suppressor gene p21. Then, this study found that PCa cells were more likely to form tumors in diet induced obese mice. Compared with the normal diet group (ND), the expression levels of KLF7 in tumor tissues in high-fat diet group (HFD) were higher. Futhermore, we verified that high concentrations of FFA C8:0 can promote the biological behavior of PCa cells by activating KLF7/IL-6/p21 signaling pathway, which is mediated by the GPR84. CONCLUSIONS: Our research may provide a potential target for clinical prevention and treatment of PCa which induced by obesity.

Stress-inducible IL-6 is regulated by KLF7 in brown adipocytes.

Stress-inducible interleukin 6 (IL-6) is generated in brown adipocytes via beta-3 adrenergic receptor (ADRB3) signaling, which is necessary in stress hyperglycemia, the kind of metabolic adaptation enabling "fight or flight" response by means of liver gluconeogenesis. Nevertheless, the mechanism of ADRB3 signaling mediates IL-6 in brown adipocytes remains unclear. As a result, it is critical to understand how brown adipocytes produce IL-6 via ADRB3 signaling. We found that the ADRB3 agonist and cold stimulation promoted the expression of KLF7 and IL-6 in brown adipocytes of mice. In parallel to these results in vivo, treatment with ADRB3 agonist promoted the expression of KLF7 and the release of IL-6 in primary brown adipocytes of mice. Notably, we discovered that KLF7 positively controls the expression of IL-6 and downregulated KLF7 largely blunted ADRB3 agonist induced IL-6 expressions in brown adipocytes. Our findings suggest that KLF7 is required for the generation of IL-6 when ADRB3 signaling is activated in brown adipocytes.

Promotion of colorectal cancer by transcription factor BHLHE40 involves upregulation of ADAM19 and KLF7.

BHLHE40 is a transcription factor, whose role in colorectal cancer has remained elusive. We demonstrate that the BHLHE40 gene is upregulated in colorectal tumors. Transcription of BHLHE40 was jointly stimulated by the DNA-binding ETV1 protein and two associated histone demethylases, JMJD1A/KDM3A and JMJD2A/KDM4A, which were shown to also form complexes on their own and whose enzymatic activity was required for BHLHE40 upregulation. Chromatin immunoprecipitation assays revealed that ETV1, JMJD1A and JMJD2A interacted with several regions within the BHLHE40 gene promoter, suggesting that these three factors directly control BHLHE40 transcription. BHLHE40 downregulation suppressed both growth and clonogenic activity of human HCT116 colorectal cancer cells, strongly hinting at a pro-tumorigenic role of BHLHE40. Through RNA sequencing, the transcription factor KLF7 and the metalloproteinase ADAM19 were identified as putative BHLHE40 downstream effectors. Bioinformatic analyses showed that both KLF7 and ADAM19 are upregulated in colorectal tumors as well as associated with worse survival and their downregulation impaired HCT116 clonogenic activity. In addition, ADAM19, but not KLF7, downregulation reduced HCT116 cell growth. Overall, these data have revealed a ETV1/JMJD1A/JMJD2A→BHLHE40 axis that may stimulate colorectal tumorigenesis through upregulation of genes such as KLF7 and ADAM19, suggesting that targeting this axis represents a potential novel therapeutic avenue.

PU.1 interacts with KLF7 to suppress differentiation and promote proliferation in chicken preadipocytes.

Krüppel-like factor 7 (KLF7) is a negative regulator of preadipocyte differentiation. Our previous KLF7 ChIP-seq analysis showed that the binding motif of PU.1 was found among the KLF7 binding peaks, indicating that an interaction between KLF7 and PU.1 at preadipocyte gene promoters and other regulatory elements might be common. Here, Co-IP and FRET assays are used to confirm that PU.1 can directly bind to KLF7 and enhance the transcription activity of cyclin-dependent kinase inhibitor 3 ( CDKN3), which is a downstream target gene of KLF7. We show that the PU.1 expression level is decreased during preadipocyte differentiation. Furthermore, PU.1 overexpression and knockdown experiments reveal that PU.1 negatively regulates chicken preadipocyte differentiation, as evidenced by appropriate changes in lipid droplet accumulation and altered expressions of PPARγ, FAS, and PLIN. In addition, PU.1 overexpression promotes preadipocyte proliferation, while knockdown of PU. 1 inhibits preadipocyte proliferation. We further demonstrate that PU.1 inhibits differentiation and promotes proliferation in preadipocytes, in part by directly interacting with KLF7.

miR-132-3p and KLF7 as novel regulators of aortic stiffening-associated EndMT in type 2 diabetes mellitus.

BACKGROUND: The prevalence of diabetes mellitus has risen considerably and currently affects more than 422 million people worldwide. Cardiovascular diseases including myocardial infarction and heart failure represent the major cause of death in type 2 diabetes (T2D). Diabetes patients exhibit accelerated aortic stiffening which is an independent predictor of cardiovascular disease and mortality. We recently showed that aortic stiffness precedes hypertension in a mouse model of diabetes (db/db mice), making aortic stiffness an early contributor to cardiovascular disease development. Elucidating how aortic stiffening develops is a pressing need in order to halt the pathophysiological process at an early time point. METHODS: To assess EndMT occurrence, we performed co-immunofluorescence staining of an endothelial marker (CD31) with mesenchymal markers (α-SMA/S100A4) in aortic sections from db/db mice. Moreover, we performed qRT-PCR to analyze mRNA expression of EndMT transcription factors in aortic sections of db/db mice and diabetic patients. To identify the underlying mechanism by which EndMT contributes to aortic stiffening, we used aortas from db/db mice and diabetic patients in combination with high glucose-treated human umbilical vein endothelial cells (HUVECs) as an in vitro model of diabetes-associated EndMT. RESULTS: We demonstrate robust CD31/α-SMA and CD31/S100A4 co-localization in aortic sections of db/db mice which was almost absent in control mice. Moreover, we demonstrate a significant upregulation of EndMT transcription factors in aortic sections of db/db mice and diabetic patients. As underlying regulator, we identified miR-132-3p as the most significantly downregulated miR in the micronome of db/db mice and high glucose-treated HUVECs. Indeed, miR-132-3p was also significantly downregulated in aortic tissue from diabetic patients. We identified Kruppel-like factor 7 (KLF7) as a target of miR-132-3p and show a significant upregulation of KLF7 in aortic sections of db/db mice and diabetic patients as well as in high glucose-treated HUVECs. We further demonstrate that miR-132-3p overexpression and KLF7 downregulation ameliorates EndMT in high glucose-treated HUVECs. CONCLUSIONS: We demonstrate for the first time that EndMT contributes to aortic stiffening in T2D. We identified miR-132-3p and KLF7 as novel EndMT regulators in this context. Altogether, this gives us new insights in the development of aortic stiffening in T2D.

Circular hsa_circ_0020377 regulates KLF7 by targeting miR-194-5p to facilitate tumor cell malignant behaviors and glycolysis in oral squamous cell carcinoma progression.

Oral squamous cell carcinoma (OSCC) is a common malignant tumor with high recurrence, metastasis rates, and poor prognosis. Numerous studies discover that circular RNA (circRNA) is closely associated with OSCC progression. Hsa_circ_0020377 has been aberrantly expressed in OSCC, but its role in tumor growth and metastasis remains largely unclear. Hsa_circ_0020377, microRNA-194-5p (miR-194-5p), and Krüppel-like factor 7 (KLF7) contents were determined by real-time quantitative polymerase chain reaction (RT-qPCR). Cell proliferative, cycle progression migration, and invasion were measured using 5-ethynyl-2'-deoxyuridine (EdU), Cell Counting Kit-8 (CCK-8), flow cytometry, wound healing, and Transwell assays. The glycolysis level was detected via specific kits. Cyclin D1, E-cadherin, hexokinase 2 (HK2), and KLF7 protein levels were detected via western blot. Using predicting bioinformatics software, the binding between miR-194-5p and hsa_circ_0020377 or KLF7 was verified using a dual-luciferase reporter and RNA Immunoprecipitation (RIP). Beyond that, a xenograft tumor model was used to analyze the role of hsa_circ_0020377 on tumor cell growth in vivo. Increased hsa_circ_0020377 and KLF7 and reduced miR-194-5p were found in OSCC tissues and cell lines. Loss-of-function experiments proved that hsa_circ_0020377 depletion might block OSCC cell proliferation, cycle progression, migration, invasion, and glycolysis in vitro. In xenograft mouse models, hsa_circ_0020377 silencing might suppress tumor growth. In addition, mechanism research suggested that hsa_circ_0020377 could bind with miR-194-5p and enhance its target gene (KLF7), thereby affecting OSCC development. These results broaden our insights regarding the regulation of OSCC progression via circRNA and act as a reference for future clinical studies in OSCC diagnosis and treatment.

Knockdown of KLF7 inhibits the differentiation of both intramuscular and subcutaneous preadipocytes in goat.

KLF7 belongs to the Krüppel-like factors (KLFs) family, which function as transcriptional regulators controlling a number of basic cellular processes, involving proliferation, differentiation, and migration. Here, we reveal insights into the differentiated expression of KLF7 in different goat tissues and different stages of growth, and the inhibition role of KLF7 knockdown to differentiation by using goat intramuscular and subcutaneous preadipocytes. We demonstrate that KLF7 expression is obviously changed during the differentiation of preadipocytes into mature adipocytes. Knockdown of KLF7 inhibited lipid droplet accumulation, reduced the expression of adipogenic markers both in intramuscular and subcutaneous preadipocytes in goats, suggesting that KLF7 is a novel regulator of adipogenesis. KLF7 expression changed also up or down-regulation the other KLF family members, but there were differences between these two types of cells. Investigation into the mechanism that KLF7 regulates preadipocyte differentiation revealed that KLF family members KLF1, KLF5, KLF6, KLF8, KLF11, KLF12, KLF16, KLF17 and adipogenic markers C/EBPα and SREBP1 promoter region present KLF7 transcriptional binding sites. Altogether, the data here identify KLF7 as a novel regulator of adipogenesis.

Krüppel-like Factor 7 inhibits proliferation and migration of pulmonary smooth muscle cells via p21 activation.

The aberrant proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) are critical contributors to the pulmonary vascular remodeling that occurs during the development of Pulmonary arterial hypertension (PAH). Krüppel-like Factor 7 (KLF7) has been reported to be involved in the development of certain cardiovascular diseases. However, the role of KLF7 in PAH remains unknown. Here, we aimed to explore whether KLF7 mediates the proliferation and migration of PASMCs and its underlying mechanism. In this study, Sprague Dawley rats were exposed to 60 mg/kg monocrotaline (MCT) for 3 weeks to induce PAH and human PASMCs were stimulated with 20 ng/ml platelet-derived growth factor-BB (PDGF-BB) for 24 h to induce proliferation and migration. The mRNA and protein expression of KLF7 were significantly down-regulated in MCT-induced PAH rats and PDGF-BB-treated PASMCs. Under normal conditions, KLF7 knockdown obviously promoted PASMCs proliferation and migration, whereas KLF7 overexpression exhibited the opposite effects. Furthermore, PDGF-BB promoted the PASMCs proliferation and migration, increased the cell proportion in S phase, which was significantly attenuated by overexpression of KLF7. Mechanistic investigation indicated that KLF7 through activation its target protein, the cell cycle inhibitor p21, which finally leading to the inhibition of PASMCs growth. Consistently, UC2288, a specific inhibitor of p21, partially reversed the PASMCs proliferation inhibited by KLF7 overexpression. Taken collectively, the data suggested that KLF7 inhibits PASMCs proliferation and migration via p21 pathway and it may be used as a new therapeutic target for the PAH.

Genome-wide survey identifies TNNI2 as a target of KLF7 that inhibits chicken adipogenesis via downregulating FABP4.

Krüppel-like factor 7 (KLF7) negatively regulates adipocyte differentiation; however, the mechanism underlying its activity in mammals and birds remains poorly understood. To identify genome-wide KLF7-binding motifs in preadipocytes, we conducted a chromatin immunoprecipitation-sequencing analysis of immortalized chicken preadipocytes (ICP2), which revealed 11,063 specific binding sites. Intergenic binding site analysis showed that KLF7 regulates several novel factors whose functions in chicken and mammal adipogenesis are underexplored. We identified a novel regulator, troponin I2 (TNNI2), which is positively regulated by KLF7. TNNI2 is downregulated during preadipocyte differentiation and acts as an adipogenic repressor at least in part by repressing FABP4 promoter activity. In conclusion, we demonstrated that KLF7 functions through cis-regulation of TNNI2, which inhibits adipogenesis. Our findings not only provide the first genome-wide picture of KLF7 associations in preadipocytes but also identify a novel function of TNNI2.

Inhibition of microRNA-19a-3p alleviates the neuropathic pain (NP) in rats after chronic constriction injury (CCI) via targeting KLF7.

BACKGROUND/PURPOSE: Neuropathic pain(NP) is derived from the dysfunctions of nerve system. The current research is to explore the impact and mechanism of miR-19a-3p in neuropathic pain in rats. METHODS: The NP was induced through the chronic constriction injury (CCI) surgery in rats. The pro-inflammatory factors (IL-1ß, IL-6, TNF-α) in spinal cord tissues from rats were measured using Elisa kits. Moreover, the different levels of thermal hyperalgesia and mechanical allodynia in rats were examined through paw withdrawal latency (PWL) and paw withdrawal threshold (PWT). To investigate into the role of miR-19a-3p and KLF7 in NP of rats, the knockdown of miR-19a-3p alone or along with KLF7 downregulation in rats were achieved through lentivirus injection. The miR-19a-3p and KLF7 expression in spinal cord of rats on Day 3,7,14 after CCI were detected using RT-qPCR. The protein expression of KLF7 were measured by Western blot. Bioinformatics and luciferase assays were used for the prediction and verification of bindings between KLF7 and miR-19a-3p. RESULTS: CCI surgery caused neuropathic pain in rats with the levels of inflammatory cytokines increased and PWL and PWT decreased. Moreover, miR-19a-3p expression was increased while the protein and mRNA levels were decreased in spinal cord tissues in rats after CCI surgery. In rat microglial cells, miR-19a-3p downregulation could promote the KLF7 in both mRNA and protein expression. In spinal cord tissues of rats, the inhibition of miR-19a-3p enhanced the KLF7 expression. Furthermore, miR-19a-3p downregulation suppressed the IL-1ß, IL-6 and TNF-α concentrations, and could decrease the NP but inhibition of KLF7 could partially reverse this in CCI rats. CONCLUSION: miR-19a-3p inhibition may alleviate NP via KLF7 in CCI rats.

Krüppel-like factor 7 deficiency causes autistic-like behavior in mice via regulating Clock gene.

BACKGROUND: Krüppel-like factor 7 (klf7), a transcription factor in the nervous system to regulate cell proliferation and differentiation, has been recently identified as a causal gene for autism spectrum disorder (ASD), but the mechanism behind remains unknown. RESULT: To uncover this mechanism, in this study we characterized the involvement of klf7 in circadian rhythm by knocking down klf7 in N2A cells and examining the rhythmic expression of circadian genes, especially Clock gene. We constructed klf7-/- mice and then investigated into klf7 regulation on the expression of rhythm genes in vivo as well as the use of melatonin to rescue the autism behavior. Our results illustrated that circadian rhythm was disrupted in klf7 knockdown cells and that klf7-/- mice showed autism-like behavior. Also, we found that Clock gene was downregulated in the brain of these klf7-/- mice and that the downstream rhythm genes of Clock were disturbed. Melatonin, as a circadian regulation drug, could regulate the expression level and amplitude of rhythm genes in klf7 knockout cells and further rescue the autistic behavior of klf7-/- mice. CONCLUSION: Klf7 deficiency causes ASD by disrupting circadian rhythm related genes to trigger rhythm oscillations. To treat ASD, maintaining circadian homeostasis is promising with the use of melatonin.

Notoginsenoside R1 alleviates spinal cord injury through the miR-301a/KLF7 axis to activate Wnt/ß-catenin pathway.

Spinal cord injury (SCI) is a devastating incident that induces neuronal loss and dysfunction. Notoginsenoside R1 (NGR1) has been reported to exhibit a neuroprotective role after SCI. In this study, the effect and molecular mechanisms of NGR1 in models of SCI were further investigated. Rat adrenal pheochromocytoma cell line (PC-12) were stimulated with lipopolysaccharide (LPS) to establish a cell model of SCI-like condition. The changes of proinflammatory cytokines and associated proteins were analyzed using enzyme linked immunosorbent assay (ELISA) and western blotting. A rat model of SCI was established. Nissl staining were used to observe the morphological characteristics of spinal cord tissues. reverse transcription-quantitative PCR (RT-qPCR) was used to measure the expression of miR-301a andKrüppel-like factor 7 (KLF7). Our results showed that NGR1 alleviated LPS-triggered apoptosis and inflammation in PC-12 cells. MiR-301a was upregulated in LPS-stimulated PC-12 cells and was downregulated by NGR1 treatment. MiR-301a overexpression reversed the effect of NGR1 in LPS-treated PC-12 cells. KLF7 was verified to be targeted by miR-301a. NGR1 activated Wnt/ß-catenin signaling in LPS-treated PC-12 cells by inhibiting miR-301a and upregulating KLF7. Moreover, blocking wingless/integrated (Wnt)/ß-catenin signaling eliminated the protective effect of NGR1 against SCI in vitro and in vivo. Overall, NGR1 could reduce inflammation and apoptosis and promote functional recovery of SCI rats by activating Wnt/ß-catenin pathway.

Cancer-suppressing miR-520-3p gene inhibits proliferation, migration, and invasion of gastric cancer cells through targeted regulation of KLF7.

Gastric cancer (GC) is among the most common malignant tumors. Numerous studies have reported that microRNAs (miRNAs) play significant roles in carcinogenesis and treatment. An miRNA, miR-520-3p, has been identified as a cancer-suppressing gene in several cancers. However, the role and underlying mechanism of miR-520-3p regulation of GC remain unknown. In this study, the expression levels of miR-520-3p in cancer tissues of patients with GC - and in adjacent normal tissues, gastric cancer cell lines, and human normal gastric epithelial cells - were detected by qRT-PCR. RNA interference was performed in GC cell lines. After the corresponding treatment, the cells were characterized in vitro or in vivo to evaluate their molecular function. CCK-8, cell colony formation, and a Transwell assay were used to detect cell proliferation rate, viability, and invasion ability. A dual-luciferase reporter gene experiment was used to explore the potential molecular mechanisms of miR-520-3p. The results showed that the expression of miR-520-3p was significantly downregulated in GC tissues and cells, and upregulation of miR-520-3p could inhibit the proliferation, vitality, and invasion of GC cells both in vivo and in vitro. The expression of Kruppel-like factor 7 (KLF7) was greatly upregulated in GC tissues. MiR-520-3p can adsorb KLF7 in GC cells, and KLF7 can reverse the inhibitory effect of miR-520-3p overexpression on the proliferation of GC cells. This study revealed that miR-520-3p plays a significant role in inhibiting the proliferation, invasion, and migration of GC cells by targeting KLF7. These data demonstrate that miR-520-3p may serve as a novel prognostic biomarker and a potential therapeutic target for GC.