Background:The 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 un...Background:The 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.Methods:Bone 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)-1b 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.Results:ALP 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 withWTmice(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 themRNA level,P=0.04).Finally,the expression levels of CKIP-1 in bone marrow(3.2-fold increase at themRNA level,P=0.03)and cultured MSCs were up-regulated on chronic inflammatory stimulation by IL-1b.Conclusions:CKIP-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.展开更多
Background:Gastric cancer(GC)is one of the most common malignancies,and intestinal-type GC is the main histopathologic type of GC in China.We previously reported that casein kinase 2 interacting protein 1(CKIP-1)acts ...Background:Gastric cancer(GC)is one of the most common malignancies,and intestinal-type GC is the main histopathologic type of GC in China.We previously reported that casein kinase 2 interacting protein 1(CKIP-1)acts as a candidate tumor suppressor in intestinal-type GC.CKIP-1 participates in the regulation of multiple signaling pathways,including the Wnt/b-catenin pathway,of which caudal-related homeobox 1(CDX1)may be a downstream target gene.The purpose of this study was to investigate the relationship between CKIP-1 and CDX1 in intestinal-type GC.Methods:Sixty-seven gastroscopy biopsy specimens and surgically resected gastric specimens were divided into four groups:gastric mucosa group,intestinal metaplasia(IM)group,dysplasia group,and intestinal-type GC group.The expression levels of CKIP-1 and CDX1 were detected in these groups and GC cell lines,and the correlations between these expression levels were analyzed.SGC7901 and BGC823 cells were divided into CKIP-1 shRNA groups and CKIP-1 over-expression groups,and CDX1 expression was detected.b-Catenin expression was detected in intestinal-type GC tissue samples and CKIP-1 shRNA and CKIP-1 overexpression SGC7901 cells,and its correlation with CKIP-1 expression in intestinal-type GC tissue was analyzed.The Wnt/b-catenin pathway inhibitor DKK-1 and activator LiCl were incubated with SGC7901 cells,BGC823 cells,and CKIP-1 shRNA and CKIP-1 over-expression SGC7901 and BGC823 cells,following which CDX1 and Ki-67 expression were detected.Results:The expression levels of CKIP-1 and CDX1 were lower in patients with intestinal-type GC than in patients with IM and dysplasia(both P<0.05).CKIP-1 and CDX1 expression levels were positively correlated in IM,dysplasia,and intestinal-type GC tissue and cell lines(r=0.771,P<0.01;r=0.597,P<0.01;r=0.654,P<0.01;r=0.811,P<0.01,respectively).CDX1 expression was decreased in the CKIP-1 shRNA groups and increased in the CKIP-1 over-expression groups of SGC7901 and BGC823 cells compared to that in the corresponding control groups(both P<0.05).CKIP-1 expression was negatively correlated with b-catenin expression in intestinal-typeGCpatients(r=0.458,P<0.01).Compared to the control group,b-catenin expression was increased in the CKIP-1 shRNA SGC7901 cell group and decreased in the CKIP-1 over-expression SGC7901 cell group(P<0.05).CDX1 expression was increased inSGC7901 andBGC823 cells treatedwithDKK-1,DKK-1 increasedCDX1 expression and decreased Ki-67 expression in the CKIP-1 shRNA group;the opposite result was observed in SGC7901 and BGC823 cells treated with LiCl,and LiCl decreased CDX1 expression and increased Ki-67 expression in the CKIP-1 over-expression group(both P<0.05).Conclusions:Through the Wnt/b-catenin signaling pathway,CKIP-1 may positively regulate CDX1 in intestinal-type GC.展开更多
基金Supported by the grants from the National Key Research and Development Project(No.2017YFB1304300)Conversion Fund of PLA General Hospital(No.2017tm-018)the Clinical Research Support Fund of PLA General Hospital(No.2017fc-tsys-2013).
文摘Background:The 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.Methods:Bone 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)-1b 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.Results:ALP 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 withWTmice(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 themRNA level,P=0.04).Finally,the expression levels of CKIP-1 in bone marrow(3.2-fold increase at themRNA level,P=0.03)and cultured MSCs were up-regulated on chronic inflammatory stimulation by IL-1b.Conclusions:CKIP-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.
基金This work was supported by the grants from the National Natural Science Foundation of China(No.81560088)Guizhou Provincial Science and Technology Foundation(No.[2019]1209).
文摘Background:Gastric cancer(GC)is one of the most common malignancies,and intestinal-type GC is the main histopathologic type of GC in China.We previously reported that casein kinase 2 interacting protein 1(CKIP-1)acts as a candidate tumor suppressor in intestinal-type GC.CKIP-1 participates in the regulation of multiple signaling pathways,including the Wnt/b-catenin pathway,of which caudal-related homeobox 1(CDX1)may be a downstream target gene.The purpose of this study was to investigate the relationship between CKIP-1 and CDX1 in intestinal-type GC.Methods:Sixty-seven gastroscopy biopsy specimens and surgically resected gastric specimens were divided into four groups:gastric mucosa group,intestinal metaplasia(IM)group,dysplasia group,and intestinal-type GC group.The expression levels of CKIP-1 and CDX1 were detected in these groups and GC cell lines,and the correlations between these expression levels were analyzed.SGC7901 and BGC823 cells were divided into CKIP-1 shRNA groups and CKIP-1 over-expression groups,and CDX1 expression was detected.b-Catenin expression was detected in intestinal-type GC tissue samples and CKIP-1 shRNA and CKIP-1 overexpression SGC7901 cells,and its correlation with CKIP-1 expression in intestinal-type GC tissue was analyzed.The Wnt/b-catenin pathway inhibitor DKK-1 and activator LiCl were incubated with SGC7901 cells,BGC823 cells,and CKIP-1 shRNA and CKIP-1 over-expression SGC7901 and BGC823 cells,following which CDX1 and Ki-67 expression were detected.Results:The expression levels of CKIP-1 and CDX1 were lower in patients with intestinal-type GC than in patients with IM and dysplasia(both P<0.05).CKIP-1 and CDX1 expression levels were positively correlated in IM,dysplasia,and intestinal-type GC tissue and cell lines(r=0.771,P<0.01;r=0.597,P<0.01;r=0.654,P<0.01;r=0.811,P<0.01,respectively).CDX1 expression was decreased in the CKIP-1 shRNA groups and increased in the CKIP-1 over-expression groups of SGC7901 and BGC823 cells compared to that in the corresponding control groups(both P<0.05).CKIP-1 expression was negatively correlated with b-catenin expression in intestinal-typeGCpatients(r=0.458,P<0.01).Compared to the control group,b-catenin expression was increased in the CKIP-1 shRNA SGC7901 cell group and decreased in the CKIP-1 over-expression SGC7901 cell group(P<0.05).CDX1 expression was increased inSGC7901 andBGC823 cells treatedwithDKK-1,DKK-1 increasedCDX1 expression and decreased Ki-67 expression in the CKIP-1 shRNA group;the opposite result was observed in SGC7901 and BGC823 cells treated with LiCl,and LiCl decreased CDX1 expression and increased Ki-67 expression in the CKIP-1 over-expression group(both P<0.05).Conclusions:Through the Wnt/b-catenin signaling pathway,CKIP-1 may positively regulate CDX1 in intestinal-type GC.
