Casein Kinase 2 Interacting Protein-1 Research Articles

CKIP-1 inhibits M2 macrophage polarization to suppress the progression of gastric cancer by inactivating JAK/STAT3 signaling.

Gastric cancer (GC) is a frequently occurring malignancy with poor prognosis. Casein kinase 2 interacting protein-1 (CKIP-1) is a PH domain-containing protein implicated in regulating tumorigenesis and macrophage homeostasis. This study aimed to elucidate the role and potential mechanism of CKIP-1 in the progression of GC. CKIP-1 expression in GC tumor and para-carcinoma tissues was detected using RT-qPCR. Then, human monocyte cell line THP-1 was treated with PMA, interleukin (IL)-4 and IL-13 to induce M2-polarized macrophages. CD206, arginase-1 (Arg-1) and transforming growth factorβ1 (TGFβ1) expression in M2-polarized macrophages with or without CKIP-1 overexpression was evaluated. Moreover, GC cell lines (MKN45 and HGC27 cells) were co-cultured with CKIP-1-overexpressed M2-polarized macrophages, and the viability, migration and invasion of GC cells were measured. Additionally, immunoblotting assessed the expression of JAK/STAT3 signaling-related proteins and STAT3 agonist Colivelin was used to treat GC cells to perform the rescue experiments to analyze the changes of malignant phenotypes of GC cells. Results showed that CKIP-1 was downregulated in GC tissues and M2-polarized macrophages. CKIP-1 overexpression inhibited M2 macrophage polarization and decreased TGFβ1 secretion. Besides, elevated CKIP-1 expression in M2-polarized macrophages inhibited the viability, migration and invasion of GC cells. Furthermore, CKIP-1 overexpression inactivated JAK2/STAT3 signaling in GC cells by inhibiting TGFβ1 level. Specifically, Colivelin treatment abrogated the influences of CKIP-1 upregulation on the malignant phenotypes of GC cells. Collectively, CKIP-1 inhibits M2 macrophage polarization to suppress the progression of GC by inactivating JAK/STAT3 signaling pathway.

The Discovery of a Specific CKIP-1 Ligand for the Potential Treatment of Disuse Osteoporosis.

Bone homeostasis relies on the delicate balance between osteoblast-mediated bone formation and osteoclast-mediated bone resorption. The casein kinase 2 interacting protein-1 (CKIP-1), a specific CK2α subunit-interacting protein, has been documented as one of the crucial negative regulators of bone formation. CKIP-1 siRNA therapy has constraints that limit its use in clinical applications. Therefore, it is necessary to explore effective targeting strategies for CKIP-1. In this study, we observed an upregulation of CKIP-1 protein expression in the microgravity environment, while its ubiquitination levels decreased. We further investigated the interaction between CKIP-1 and VHL and found that VHL enhanced CKIP-1 degradation through the ubiquitylation-proteasome system (UPS). Additionally, we discovered a small molecule ligand, named C77, through DNA-encoded library (DEL) screening, which binds to CKIP-1 both in vivo and in vitro, as confirmed by Surface Plasmon Resonance (SPR) and the Cellular Thermal shift assay (CETSA), respectively. Our findings demonstrated the potential of VHL and C77 as guiding factors in the development of CKIP-1-based Proteolysis-Targeting Chimeras (PROTACs), which could be future therapeutic interventions in disuse osteoporosis.

CKIP-1 silencing suppresses OSCC via mitochondrial homeostasis-associated TFAM/cGAS-STING signalling axis.

Limited effective targets have challenged the treatment of oral squamous cell carcinoma (OSCC). Casein kinase 2 interacting protein 1 (CKIP-1) is a scaffold protein involved in various diseases. However, the role of CKIP-1 in OSCC remains unclear. The aim of this study was to explore the regulatory role of CKIP-1 in OSCC, as well as the involved mechanism. First, higher expression of CKIP-1 in OSCC tissues and cell lines were found. Series of gain- and loss-of-function experiments demonstrated suppressed malignant behaviours and enhanced apoptosis of OSCC cells when CKIP-1 was silenced. Also, inhibited tumour growth in CKIP-1-silenced group were proved. Further, mitochondrial transcription factor A (TFAM) downregulation, increased ROS production, decreased mitochondrial membrane potential and cGAS-STING activation in CKIP-1-silenced group were observed. The involvement of mitochondrial homeostasis-related TFAM/cGAS-STING axis in CKIP-1-silenced OSCC cells was finally demonstrated by tetramethylpyrazine (TMP) that inhibits TFAM degradation. Taken together, our study demonstrated that CKIP-1 silencing could significantly antagonize OSCC via TFAM/cGAS-STING axis, which may provide a candidate target for OSCC treatment.

CKIP-1 mediates CK2 translocation to regulate Nav1.5 and Kir2.1 channel complexes in cardiomyocytes.

Sodium and potassium channels, especially Nav1.5 and Kir2.1, play key roles in the formation of action potentials in cardiomyocytes. These channels interact with, and are regulated by, synapse-associated protein 97 (SAP97). However, the regulatory role of SAP97 in myocyte remains incompletely understood. Here, we investigate the function of SAP97 phosphorylation in the regulation of Nav1.5 and Kir2.1 channel complexes and the upstream regulation of SAP97. We found that SAP97 is phosphorylated by casein kinase II (CK2) in vitro. In addition, transfection of casein kinase 2 interacting protein-1 (CKIP-1) into cardiomyocytes to drive CK2 from the nucleus to the cytoplasm, increased SAP97 phosphorylation and Nav1.5 and Kir2.1 current activity. These findings demonstrated that CKIP-1 modulates the subcellular translocation of CK2, which regulates Nav1.5 and Kir2.1 channel complex formation and activity in cardiomyocytes.

CKIP-1-Loaded Cartilage-Affinitive Nanoliposomes Reverse Osteoarthritis by Restoring Chondrocyte Homeostasis.

Osteoarthritis (OA) is a chronic joint disease characterized by cartilage imbalance and disruption of cartilage extracellular matrix secretion. Identifying key genes that regulate cartilage differentiation and developing effective therapeutic strategies to restore their expression is crucial. In a previous study, we observed a significant correlation between the expression of the gene encoding casein kinase-2 interacting protein-1 (CKIP-1) in the cartilage of OA patients and OA severity scores, suggesting its potential involvement in OA development. To test this hypothesis, we synthesized a chondrocyte affinity plasmid, liposomes CKIP-1, to enhance CKIP-1 expression in chondrocytes. Our results demonstrated that injection of CAP-Lipos-CKIP-1 plasmid significantly improved OA joint destruction and restored joint motor function by enhancing cartilage extracellular matrix (ECM) secretion. Histological and cytological analyses confirmed that CKIP-1 maintains altered the phosphorylation of the signal transduction molecule SMAD2/3 of the transforming growth factor-β (TGF-β) pathway by promoting the phosphorylation of the 8T, 416S sit. Taken together, this work highlights a novel approach for the precise modulation of chondrocyte phenotype from an inflammatory to a noninflammatory state for the treatment of OA and may be broadly applicable to patients suffering from other arthritic diseases.

Associations of CKIP-1 and LOX-1 polymorphisms with the risk of type 2 diabetes mellitus with hypertension among Chinese adults.

Type 2 diabetes mellitus (T2DM) and hypertension are common high-incidence diseases, closely related, and have common pathogenic basis such as oxidative stress. Casein kinase 2 interacting protein-1 (CKIP-1) and low-density lipoprotein receptor (LOX-1) are considered to be important factors affect the level of oxidative stress in the body. The main purpose of this study was to explore the relationship between CKIP-1 (rs6693817 A > T, rs2306235 C > G) and LOX-1 (rs1050283 G > A, rs11053646 C > G) polymorphisms and the risk of hypertension and diabetes, and try to find new candidate genes for diabetes and diabetes with hypertension etiology in Chinese population. 574 T2DM patients and 597 controlsfrequently matched by age and sex were selected for genotyping of CKIP-1 (rs6693817 A > T, rs2306235 C > G) and LOX-1 gene (rs1050283 G > A, rs11053646 C > G). Logistic regression was used to analyze the correlation between different genotypes and the risk of T2DM and T2DM with hypertension, and the results were expressed as odds ratio (OR) and 95% confidence interval (95% CI). We found that the risk of T2DM in the AA + AT genotype of rs6693817 was higher than that in the TT genotype in Chinese population (OR = 1.318, 95%CI: 1.011-1.717, P = 0.041), and the difference was still significant after adjustment (OR = 1.370, 95%CI: 1.043-1.799, Padjusted = 0.024), the difference of heterozygotes (AT vs TT: OR = 1.374, 95%CI: 1.026-1.840, Padjusted = 0.033) was statistically significant. But after Bonferroni correction, the significance of the above sites disappeared. And rs6693817 was associated with the risk of T2DM combined with hypertension before and after adjustment in dominant model (OR = 1.424, 95% CI: 1.038-1.954, P = 0.028; OR = 1.460, 95% CI: 1.057-2.015, Padjusted = 0.021, respectively) and in heterozygote model (OR = 1.499, 95% CI: 1.069-2.102, P = 0.019; OR = 1.562, 95% CI: 1.106-2.207, Padjusted = 0.011, respectively). However, only the statistical significance of the heterozygous model remained after Bonferroni correction. rs2306235, rs1050283 and rs11053646 were not significantly correlated with T2DM and T2DM combined with hypertension risk (P > 0.05). The results suggest that CKIP-1 rs6693817 is related to the susceptibility of Chinese people to T2DM with hypertension, providing a new genetic target for the treatment of diabetes with hypertension with in the future.

CKIP-1 mediates P. gingivalis-suppressed osteogenic/cementogenic differentiation of periodontal ligament cells partially via p38 signaling pathway

ABSTRACT Objectives Casein kinase 2 interacting protein-1 (CKIP-1) is a versatile player involved in various biological processes. However, whether CKIP-1 mediates the osteogenic/cementogenic differentiation of periodontal ligament cells (PDLCs) under Porphyromonas gingivalis (Pg) stimulation remains unknown. Material and Methods The effect of Pg on PDLC differentiation was first verified. CKIP-1 expression in Pg-infected PDLCs or in PDL of apical periodontitis (AP) mice was detected. The changes of CKIP-1 during PDLC differentiation was also determined. PDLC differentiation capacity in CKIP-1 knockout (KO) mice and CKIP-1-silenced PDLCs with or without Pg stimulation were further studied. Inhibitor was finally applied to verify the involvement of p38 signaling pathway in PDLC differentiation. Results The suppression effect of Pg on PDLC differentiation was demonstrated. CKIP-1 increased in the PDL of AP mice and Pg-induced PDLCs, and decreased gradually during PDLC differentiation. Increased OSX and RUNX2 expression in PDL were observed in CKIP-1 KO mice. Also, CKIP-1 silencing facilitated and rescued Pg-inhibited PDLC differentiation. Inhibitor for p38 signaling pathway blocked CKIP-1 silencing-facilitated PDLC differentiation. Conclusions CKIP-1 mediated the osteogenic/cementogenic differentiation of PDLCs partially through p38 signaling pathway, which may provide evidence for the regeneration of periodontal hard tissues damaged by Pg.

CKIP-1 Promotes P. gingivalis-Induced Inflammation of Periodontal Soft Tissues by Inhibiting Autophagy.

As a chronic inflammatory disease, periodontitis involves many biological processes including autophagy. At the same time, casein kinase 2 interacting protein-1 (CKIP-1) was reported to play a role in regulation of inflammation. But whether CKIP-1 and autophagy interact in periodontitis remains unclear. In this paper, our research team verified the levels of CKIP-1 expression and autophagy increase in the periodontal tissues of a ligature-induced periodontitis mouse model. And this result was also confirmed in Porphyromonas gingivalis (Pg)-induced human gingival fibroblasts (HGF) and human periodontal ligament cells (PDLC). We also showed the autophagy level in periodontal tissues is higher in Ckip-1 knockout (KO) mice than wild type (WT). At the same time, CKIP-1 knockdown lentivirus was used in PDLC and HGF, and it was found that silencing CKIP-1 significantly activated autophagy. Unfortunately, the regulatory role of autophagy in periodontitis is still unclear. Then, the autophagy agonist Rapamycin and inhibitor 3-MA were used in a periodontitis mouse model to investigate periodontal tissue destruction. We found the inflammation in periodontal tissue was reduced when autophagy activated. All these conclusions have been verified both in vivo and in vitro experiments. Finally, our research proved that silencing CKIP-1 reduces the expression of inflammatory cytokines in Pg-induced PDLC and HGF by regulating autophagy. Overall, a new role for CKIP-1 in regulating periodontal tissue inflammation was demonstrated in our study, and it is possible to treat periodontitis by targeting the CKIP-1 gene.

Effect of CKIP-1 on hepatocyte apoptosis in nonalcoholic fatty liver disease

Objective: To explore the effect and underlying mechanism of casein kinase 2 interacting protein-1 (CKIP-1) on hepatocyte apoptosis in nonalcoholic fatty liver disease (NAFLD). Methods: Experimental study. An NAFLD cell model was established by inducing human hepatoma cell line, HepG2 cells, with oleic acid (OA). Flag-CKIP-1 expression vector and shRNA-CKIP-1 were transfected into HepG2 cells. Flow cytometry was used to detect the effect of CKIP-1 on the activity and apoptosis of NAFLD hepatocytes. The levels of apoptosis-related proteins were detected by Western blot. CKIP-1 knockout mice in C57BL/6 back-ground were fed with either standard or high-fat diet for 8 weeks. Apoptosis-related signal proteins in NAFLD hepatocytes were detected by immunohistochemistry. Results: After CKIP-1 was transfected into HepG2 cells, the degree of OA induced cell liposis was significantly reduced (P<0.05). Annexin V-FITC/PI flow cytometry showed that CKIP-1 reduced the apoptosis of steatotic hepatocytes. Overexpression of CKIP-1 could significantly inhibit the expression of caspase-3 and caspase-9 and increase the expression of Bcl-2/Bax (P<0.05). Knockdown of CKIP-1 could increase the expression of caspase-3 and caspase-9 (P<0.05). CKIP-1 knockout could further increase the expression of caspase-3 and caspase-9 in NAFLD mice (P<0.01,P<0.05), and further decrease the expression of Bcl-2/Bax (P<0.05). Conclusion: CKIP-1 inhibited the apoptosis of steatotic hepatocytes by up-regulating the expression of apoptosis inhibitor gene, Bcl-2/Bax, and affecting the proteases, caspase-3 and caspase-9.

The ubiquitination of CKIP-1 mediated by Src aggravates diabetic renal fibrosis (original article)

Renal chronic inflammation is an important hallmark of diabetic renal fibrosis. Casein kinase 2 interacting protein 1 (CKIP-1) performs a nephroprotective role in the pathogenesis of diabetic nephropathy (DN), which is dramatically decreased in diabetic kidneys. However, whether CKIP-1 regulates inflammation to ameliorate renal fibrosis remains unclear and it is interesting to clarify the degradation mechanism of CKIP-1. Here, we identified CKIP-1 expression was down-regulated in diabetic kidneys and knockout (KO) of CKIP-1 increased c-Jun expression and extra cellular matrix (ECM) in kidneys of normal mice, and knockout (KO) of CKIP-1 further exacerbated renal inflammatory fibrosis in diabetic mice. Moreover, the activated Src kinase interacted with CKIP-1 at Lys252 and increased K48 linked polyubiquitination and proteasome degradation of CKIP-1 in HG induced GMCs and diabetic kidneys. Mechanistically, Src facilitating the binding of c-Cbl with CKIP-1 by promoting the phosphorylation of c-Cbl, thereby increasing Cbl-mediated ubiquitination of CKIP-1 to down-regulate CKIP-1 protein expression. Thus, our study highlighted the anti-inflammation role of CKIP-1 and clarified the mechanism of CKIP-1 degradation in DN.

Corrigendum: Inflammation-mediated age-dependent effects of casein kinase 2-interacting protein-1 on osteogenesis in mesenchymal stem cells

Corrigendum: Inflammation-mediated age-dependent effects of casein kinase 2-interacting protein-1 on osteogenesis in mesenchymal stem cells

A novel CKIP-1 SiRNA slow-release coating on porous titanium implants for enhanced osseointegration.

Osseointegration between implants and bone tissue lays the foundation for the long-term stability of implants. The incorporation of a porous structure and local slow release of siRNA to silence casein kinase-2 interacting protein-1 (CKIP-1), a downregulator of bone formation, is expected to promote osseointegration. Here, porous implants with a porous outer layer and dense inner core were prepared by metal coinjection molding (MIM). Mg-doped calcium phosphate nanoparticles (CaPNPs)-grafted arginine-glycine-aspartate cell adhesion sequence (RGD) and transcribed activator (TAT) (MCPRT)/CKIP-1 siRNA complex and polylysine (PLL) were coated onto the surface of the porous implants by layer-by-layer (LBL) self-deposition. The in vitro results showed that the MCPRT-siRNA coating promoted MG63 cell adhesion and proliferation, enhanced the protein expressions (ALP and OC) and bone formation-related gene expression (OPN, OC and COL-1α) in vitro. The in vivo results demonstrated that the porous structure enhanced bone ingrowth and that the local slow release of MCPRT-siRNA accelerated new bone formation at the early stage. The porous structure coupled with local CKIP-1 siRNA delivery constitutes a promising approach to achieve faster and stronger osseointegration for dental implants.

IFP35 Is a Relevant Factor in Innate Immunity, Multiple Sclerosis, and Other Chronic Inflammatory Diseases: A Review.

Simple SummaryIn this review, we focused on the emerging role of IFP35, a highly conserved leucine zipper protein from fish to humans, with a still unknown biological function. The considered literature indicates this protein as a key-pleiotropic factor reflecting JAK-STAT and DAMPs pathways activation in innate immunity-dependent inflammation, as well as in the physiology and general pathology of a wide range of phylogenetically distant organisms. These findings also indicate IFP35 as a biologically relevant molecule in human demyelinating diseases of the central nervous system, including Multiple Sclerosis, and other organ-specific chronic inflammatory disorders.Discovered in 1993 by Bange et al., the 35-kDa interferon-induced protein (IFP35) is a highly conserved cytosolic interferon-induced leucine zipper protein with a 17q12-21 coding gene and unknown function. Belonging to interferon stimulated genes (ISG), the IFP35 reflects the type I interferon (IFN) activity induced through the JAK-STAT phosphorylation, and it can homodimerize with N-myc-interactor (NMI) and basic leucine zipper transcription factor (BATF), resulting in nuclear translocation and a functional expression. Casein kinase 2-interacting protein-1 (CKIP-1), retinoic acid-inducible gene I (RIG-I), and laboratory of genetics and physiology 2 Epinephelus coioides (EcLGP2) are thought to regulate IFP35, via the innate immunity pathway. Several in vitro and in vivo studies on fish and mammals have confirmed the IFP35 as an ISG factor with antiviral and antiproliferative functions. However, in a mice model of sepsis, IFP35 was found working as a damage associated molecular pattern (DAMP) molecule, which enhances inflammation by acting in the innate immune-mediated way. In human pathology, the IFP35 expression level predicts disease outcome and response to therapy in Multiple Sclerosis (MS), reflecting IFN activity. Specifically, IFP35 was upregulated in Lupus Nephritis (LN), Rheumatoid Arthritis (RA), and untreated MS. However, it normalized in the MS patients undergoing therapy. The considered data indicate IFP35 as a pleiotropic factor, suggesting it as biologically relevant in the innate immunity, general pathology, and human demyelinating diseases of the central nervous system.

3' untranslated region of Ckip-1 inhibits cardiac hypertrophy independently of its cognate protein.

3' untranslated region (3' UTR) of mRNA is more conserved than other non-coding sequences in vertebrate genomes, and its sequence space has substantially expanded during the evolution of higher organisms, which substantiates their significance in biological regulation. However, the independent role of 3' UTR in cardiovascular disease was largely unknown. Using bioinformatics, RNA fluorescent in situ hybridization and quantitative real-time polymerase chain reaction, we found that 3' UTR and coding sequence regions of Ckip-1 mRNA exhibited diverse expression and localization in cardiomyocytes. We generated cardiac-specific Ckip-1 3' UTR overexpression mice under wild type and casein kinase 2 interacting protein-1 (CKIP-1) knockout background. Cardiac remodelling was assessed by histological, echocardiography, and molecular analyses at 4 weeks after transverse aortic constriction (TAC) surgery. The results showed that cardiac Ckip-1 3' UTR significantly inhibited TAC-induced cardiac hypertrophy independent of CKIP-1 protein. To determine the mechanism of Ckip-1 3' UTR in cardiac hypertrophy, we performed transcriptome and metabolomics analyses, RNA immunoprecipitation, biotin-based RNA pull-down, and reporter gene assays. We found that Ckip-1 3' UTR promoted fatty acid metabolism through AMPK-PPARα-CPT1b axis, leading to its protection against pathological cardiac hypertrophy. Moreover, Ckip-1 3' UTR RNA therapy using adeno-associated virus obviously alleviates cardiac hypertrophy and improves heart function. These findings disclose that Ckip-1 3' UTR inhibits cardiac hypertrophy independently of its cognate protein. Ckip-1 3' UTR is an effective RNA-based therapy tool for treating cardiac hypertrophy and heart failure.

An activatable near-infrared fluorescent probe targeting CKIP-1 for monitoring osteoporosis in vivo

Osteoporosis (OP) is characterized by decreased bone mass and strength. Patients with OP have a higher risk of bone fragility and fracture. Currently, bone mineral density (BMD) determined by radioactive scanning is one of the most critical parameters applied in the clinical diagnosis and monitoring of therapeutic response in osteoporosis. However, radioactive scanning exposes the potential risks of long-time radiation damage. This calls for development of less toxic and highly sensitive molecular fluorescence imaging techniques. Currently, this is not possible because there are no effective indicators. Here, we found a negative correlation between osteoporosis and casein kinase 2-interacting protein-1 (CKIP-1, also known as PLEKHO1). This kinase was previously found to negatively regulate bone formation. Using this new biomarker, we designed an activatable near-infrared (NIR) fluorescent probe targeting CKIP-1 mRNA (NPC) with the toe-hold mediated strand displacement reaction and fuel amplification strategy. NIR fluorophore Cy5 and its quencher part BHQ3 were incorporated in the probe. The ratio of NPC probe and its fuel strand were also optimized.The designed detection system effectively visualized CKIP-1 at the cell level and in three different osteoporosis models. Results revealed that the NPC probe targeting CKIP-1 can be used to monitor changes in bone mass and therapy responses in vivo.

Casein Kinase-2 Interacting Protein-1 Regulates Physiological Cardiac Hypertrophy via Inhibition of Histone Deacetylase 4 Phosphorylation.

Different kinds of mechanical stimuli acting on the heart lead to different myocardial phenotypes. Physiological stress, such as exercise, leads to adaptive cardiac hypertrophy, which is characterized by a normal cardiac structure and improved cardiac function. Pathological stress, such as sustained cardiac pressure overload, causes maladaptive cardiac remodeling and, eventually, heart failure. Casein kinase-2 interacting protein-1 (CKIP-1) is an important regulator of pathological cardiac remodeling. However, the role of CKIP-1 in physiological cardiac hypertrophy is unknown. We subjected wild-type (WT) mice to a swimming exercise program for 21 days, which caused an increase in myocardial CKIP-1 protein and mRNA expression. We then subjected CKIP-1 knockout (KO) mice and myocardial-specific CKIP-1-overexpressing mice to the 21-day swimming exercise program. Histological and echocardiography analyses revealed that CKIP-1 KO mice underwent pathological cardiac remodeling after swimming, whereas the CKIP-1-overexpressing mice had a similar cardiac phenotype to the WT controls. Histone deacetylase 4 (HDAC4) is a key molecule in the signaling cascade associated with pathological hypertrophy; the phosphorylation levels of HDAC4 were markedly higher in CKIP-1 KO mouse hearts after the swimming exercise program. The phosphorylation levels of HDAC4 did not change after swimming in the hearts of CKIP-1-overexpressing or WT mice. Our results indicate that swimming, a mechanical stress that leads to physiological hypertrophy, triggers pathological cardiac remodeling in CKIP-1 KO mice. CKIP-1 is necessary for physiological cardiac hypertrophy in vivo, and for modulating the phosphorylation level of HDAC4 after physiological stress. Genetically engineering CKIP-1 expression affected heart health in response to exercise.

Casein Kinase 2-Interacting Protein-1 Alleviates High Glucose-Reduced Autophagy, Oxidative Stress, and Apoptosis in Retinal Pigment Epithelial Cells via Activating the p62/KEAP1/NRF2 Signaling Pathway.

Background Casein kinase 2-interacting protein-1 (CKIP-1) has been proved to be associated with complications of diabetes. Diabetic retinopathy is a main diabetic complication which usually leads to blindness. The current study aims to investigate the role of CKIP-1 in high glucose-treated retinal pigment epithelial (RPE) cells which is a component of blood-retinal barriers. Methods The RPE cells, ARPE-19, are treated with high glucose to mimic the diabetic stimulation. CKIP-1 was overexpressed in ARPE-19 cells to evaluate its effects on autophagy, oxidative stress, and apoptosis induced by high glucose treatment, using Western blot, immunofluorescence, and flow cytometry assays, respectively. Results CKIP-1 was expressed at a lower level in high glucose-treated cells than in normal glucose cells. Overexpression of CKIP-1 enhanced the Nrf2 translocation to the nucleus. Furthermore, high glucose-induced autophagy, oxidative stress, and apoptosis were inhibited after overexpression of CKIP-1. Also, CKIP-1 regulates the p62/Keap1/Nrf2 signaling, which might be the potential mechanism in this model. Conclusion In conclusion, CKIP-1 may be a potential therapeutic target that protects RPE cells from injury and subsequent diabetic retinopathy induced by high glucose.

Let-7i-5p functions as a putative osteogenic differentiation promoter by targeting CKIP-1.

The online version of this article (10.1007/s10616-020-00444-1) contains supplementary material, which is available to authorized users.

CKIP-1 augments autophagy in steatotic hepatocytes by inhibiting Akt/mTOR signal pathway

Nonalcoholic fatty liver disease (NAFLD), which is characterized by aberrant accumulation of intrahepatic triglycerides and lipid droplets (LDs) in the liver cells, is becoming increasingly prevalent at an alarming rate worldwide. LDs can be consumed by either hydrolysis or autophagy, which is shown to be of importance in the regulation of hepatic lipid metabolism. We have shown that deficiency of pleckstrin homology domain-containing casein kinase 2 interacting protein-1 (CKIP-1), a scaffold protein that interacts with various proteins in multiple signal pathways, in mice aggravates high-fat diet induced fatty liver. However, its underlying mechanisms remain largely unknown. In this study, we found that the mRNA and protein levels of CKIP-1 decreased dramatically in steatotic HepG2 cells induced by oleic acid (OA) treatment. Coincidently, hepatic autophagy was also dynamically regulated in steatotic HepG2 cells. In addition, overexpression of CKIP-1 activated autophagy by suppression of Akt/mTOR signaling, which in turn reduced lipid accumulation. Moreover, these phenomena were reversed in CKIP-1-shRNA transfected steatotic hepatocytes. To further evaluate the potential role of CKIP-1 in autophagy, we determined the level of autophagy related proteins in CKIP-1 knockout mice. These results supported our findings in vitro. In summary, we found CKIP-1 to be a positive regulator of hepatic autophagy and a promising therapeutic target for treatment of NAFLD.

Inflammation-mediated age-dependent effects of casein kinase 2-interacting protein-1 on osteogenesis in mesenchymal stem cells.

BackgroundThe casein kinase 2-interacting protein-1 (CKIP-1) is important in the development of osteoblasts and cardiomyocytes. However, the effects of CKIP-1 on osteoblast precursor mesenchymal stem cells (MSCs) remain unclear. This study aimed to determine whether CKIP-1 affects osteogenic differentiation in MSCs and explore the relationship of CKIP-1 and inflammation.MethodsBone marrow MSCs of CKIP-1 wild type (WT) and knockout (KO) mice were cultivated in vitro. Cell phenotype was analyzed by flow cytometry, colony formation was detected to study the proliferative ability. Osteogenic and adipogenic induction were performed. The osteogenic ability was explored by alizarin red staining, alkaline phosphatase (ALP) staining and ALP activity detection. Quantitative real-time polymerase chain reaction (qRT-PCR) was carried out to determine the mRNA expression levels of osteoblast marker genes. The adipogenic ability was detected by oil red O staining. Content of the bone was analyzed to observe the differences of bone imaging parameters including trabecular bone volume/tissue volume (BV/TV), bone surface area fraction/trabecular BV, trabecular number (Tb.N), and trabecular spacing (Tb.sp). Interleukin (IL)-1β was injected on WT mice of 2 months old and 18 months old, respectively. Difference in CKIP-1 expression was detected by RT-PCR and western blot. The relationship between CKIP-1 and inflammation was explored by RT-PCR and western blot.ResultsALP assays, alizarin red staining, and qRT-PCR showed that MSCs derived from CKIP-1 KO mice exhibited a stronger capability for osteogenesis. Micro-computed tomography detection showed that among 18-month-old mice, CKIP-1 KO mice presented significantly higher bone mass compared with WT mice (P = 0.02). No significant difference was observed in 2-month-old mice. In vivo data showed that expression of CKIP-1 was higher in the bone marrow of aging mice than in young mice (4.3-fold increase at the mRNA level, P = 0.04). Finally, the expression levels of CKIP-1 in bone marrow (3.2-fold increase at the mRNA level, P = 0.03) and cultured MSCs were up-regulated on chronic inflammatory stimulation by IL-1β.ConclusionsCKIP-1 is responsible for negative regulation of MSC osteogenesis with age-dependent effects. Increasing levels of inflammation with aging may be the primary factor responsible for higher expression levels of CKIP-1 but may not necessarily affect MSC aging.