Ceramide enhanced the hepatic glucagon response through regulation of CREB activity.

BACKGROUND & AIMS: Hyperglycemia is associated with lipid disorders in patients with diabetes. Ceramides are metabolites involved in sphingolipid metabolism that accumulate during lipid disorders and exert deleterious effects on glucose and lipid metabolism. However, the effects of ceramide on glucagon-mediated hepatic gluconeogenesis remain largely unknown. This study was designed to investigate the impact of ceramides on gluconeogenesis in the context of the hepatic glucagon response, with the aim of finding new pharmacological interventions for hyperglycemia in diabetes. METHODS: Liquid chromatography-mass spectrometry was used to quantify ceramide content in the serum of patients with diabetes. Primary hepatocytes were isolated from male C57BL/6J mice to study the effects of ceramide on hepatic glucose production. Immunofluorescence staining was performed to view cAMP-responsive element-binding protein (CREB)- regulated transcription co-activator 2 (CRTC2) nuclear translocation in hepatocytes. Serine palmitoyl-transferase, long chain base subunit 2 (Sptlc2) knockdown mice were generated using an adeno-associated virus containing shRNA, and hepatic glucose production was assessed glucagon tolerance and pyruvate tolerance tests in mice fed a normal chow diet and high-fat diet. RESULTS: Increased ceramide levels were observed in the serum of patients newly diagnosed with type 2 diabetes. De novo ceramide synthesis was activated in mice with metabolic disorders. Ceramide enhanced hepatic glucose production in primary hepatocytes. In contrast, genetic silencing of Sptlc2 prevented this process. Mechanistically, ceramides de-phosphorylate CRTC2 (Ser 171) and facilitate its translocation into the nucleus for CREB activation, thereby augmenting the hepatic glucagon response. Hepatic Sptlc2 silencing blocked ceramide generation in the liver and thus restrained the hepatic glucagon response in mice fed a normal chow diet and high-fat diet. CONCLUSIONS: These data indicate that ceramide serves as an intracellular messenger that augments hepatic glucose production by regulating CRTC2/CREB activity in the context of the hepatic glucagon response, suggesting that CRTC2 phosphorylation might be a potential node for pharmacological interventions to restrain the hyperglycemic response during fasting in diabetes.

S100a16 Deficiency Prevents Alcohol-induced Fatty Liver Injury via Inducing MANF Expression in Mice.

Alcoholic liver disease (ALD) encompasses conditions ranging from simple steatosis to cirrhosis and even liver cancer. It has gained significant global attention in recent years. Despite this, effective pharmacological treatments for ALD remain elusive, and the core mechanisms underlying the disease are not yet fully comprehended. S100A16, a newly identified calcium-binding protein, is linked to lipid metabolism. Our research has discovered elevated levels of the S100A16 protein in both serum and liver tissue of ALD patients. A similar surge in hepatic S100A16 expression was noted in a Gao-binge alcohol feeding mouse model. S100a16 knockdown alleviated ethanol-induced liver injury, steatosis and inflammation. Conversely, S100a16 transgenic mice showed aggravating phenomenon. Mechanistically, we identify mesencephalic astrocyte-derived neurotrophic factor (MANF) as a regulated entity downstream of S100a16 deletion. MANF inhibited ER-stress signal transduction induced by alcohol stimulation. Meanwhile, MANF silencing suppressed the inhibition effect of S100a16 knockout on ethanol-induced lipid droplets accumulation in primary hepatocytes. Our data suggested that S100a16 deletion protects mice against alcoholic liver lipid accumulation and inflammation dependent on upregulating MANF and inhibiting ER stress. This offers a potential therapeutic avenue for ALD treatment.

S100 Calcium Binding Protein A16 Promotes Cell Proliferation by triggering LATS1 ubiquitin degradation mediated by CUL4A ligase to inhibit Hippo pathway in Glioma development.

Background: S100 Calcium Binding Protein A16 (S100A16), a novel member of S100 protein family, is linked to tumorigenic processes and abundantly expressed in CNS tissues. Our study aimed to explore the biological function and possible mechanism of S100A16 in the progression of glioma. Methods: Sequence data of S100A16 and survival prognosis of glioma patients were initially analyzed using public databases. Glioma tissues were collected to examine S100A16 expression levels. Glioma cell lines and nude mice were subjected to in vitro and in vivo functional experiments. Western blot, immunofluorescence (IF), immunoprecipitation (IP) and ubiquitination assays were done to further elucidate the underlying mechanism. Results: This study firstly revealed that S100A16 was markedly up-regulated in glioma, and patients with higher S100A16 levels have a shorter survival time. S100A16 overexpression promoted the proliferation, invasion and migration of glioma cells, and the tumor formation of nude mice. Importantly, we identified S100A16 as a negative regulator of the Hippo pathway which could decrease LATS1 expression levels, promote the YAP nuclear import and initiate the downstream target genes CYR61 and CTGF. Moreover, our data showed that S100A16 destabilized LATS1 protein by inducing the CUL4A-mediated LATS1 ubiquitination degradation. Conclusions: This study demonstrated a vital biological role of S100A16 in glioma progression mechanism by promoting CUL4A-mediated LATS1 ubiquitination to inhibit Hippo signaling pathway. S100A16 could be a novel biomarker and treatment option for glioma patients.

S100a16 deficiency prevents hepatic stellate cells activation and liver fibrosis via inhibiting CXCR4 expression.

INTRODUCTION: Liver fibrosis caused by hepatic stellate cells (HSCs) activation is implicated in the pathogenesis of liver diseases. To date, there has been no effective intervention means for this process. S100 proteins are calcium-binding proteins that regulate cell growth and differentiation. This study aimed to investigate whether S100A16 induces HSCs activation and participates in liver fibrosis progression. METHODS: HSCs were isolated, and the relationship between S100A16 expression and HSCs activation was studied. S100a16 knockdown and transgenic mice were generated and subjected to HSCs activation and liver fibrosis stimulated by different models. Clinical samples were collected for further confirmation. Alterations in gene expression in HSCs were investigated, using transcriptome sequencing to determine the underlying mechanisms. RESULTS: We observed increased S100A16 levels during HSCs activation. Genetic silencing of S100a16 prevented HSCs activation in vitro. Furthermore, S100a16 silencing exhibited obvious protective effects against HSCs activation and fibrosis progression in mice. In contrast, S100a16 transgenic mice exhibited spontaneous liver fibrosis. S100A16 was also upregulated in the HSCs of patients with fibrotic liver diseases. RNA sequencing revealed that C-X-C motif chemokine receptor 4 (Cxcr4) gene was a crucial regulator of S100A16 induction during HSCs activation. Mechanistically, S100A16 bound to P53 to induce its degradation; this augmented CXCR4 expression to activate ERK 1/2 and AKT signaling, which then promoted HSCs activation and liver fibrosis. CONCLUSIONS: These data indicate that S100a16 deficiency prevents liver fibrosis by inhibiting Cxcr4 expression. Targeting S100A16 may provide insight into the pathogenesis of liver fibrosis and pave way for the design of novel clinical therapeutic strategies.

A Novel S100 Family-Based Signature Associated with Prognosis and Immune Microenvironment in Glioma.

BACKGROUND: Glioma is the most common central nervous system (CNS) cancer with a short survival period and a poor prognosis. The S100 family gene, comprising 25 members, relates to diverse biological processes of human malignancies. Nonetheless, the significance of S100 genes in predicting the prognosis of glioma remains largely unclear. We aimed to build an S100 family-based signature for glioma prognosis. METHODS: We downloaded 665 and 313 glioma patients, respectively, from The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) database with RNAseq data and clinical information. This study established a prognostic signature based on the S100 family genes through multivariate COX and LASSO regression. The Kaplan-Meier curve was plotted to compare overall survival (OS) among groups, whereas Receiver Operating Characteristic (ROC) analysis was performed to evaluate model accuracy. A representative gene S100B was further verified by in vitro experiments. RESULTS: An S100 family-based signature comprising 5 genes was constructed to predict the glioma that stratified TCGA-derived cases as a low- or high-risk group, whereas the significance of prognosis was verified based on CGGA-derived cases. Kaplan-Meier analysis revealed that the high-risk group was associated with the dismal prognosis. Furthermore, the S100 family-based signature was proved to be closely related to immune microenvironment. In vitro analysis showed S100B gene in the signature promoted glioblastoma (GBM) cell proliferation and migration. CONCLUSIONS: We constructed and verified a novel S100 family-based signature associated with tumor immune microenvironment (TIME), which may shed novel light on the glioma diagnosis and treatment.

Declined expressions of vast mitochondria-related genes represented by CYCS and transcription factor ESRRA in skeletal muscle aging.

Age-related skeletal muscle deterioration (sarcopenia) has a significant effect on the elderly's health and quality of life, but the molecular and gene regulatory mechanisms remain largely unknown. It is necessary to identify the candidate genes related to skeletal muscle aging and prospective therapeutic targets for effective treatments. The age-line-related genes (ALRGs) and age-line-related transcripts (ALRTs) were investigated using the gene expression profiles of GSE47881 and GSE118825 from the Gene Expression Omnibus (GEO) database. The protein-protein interaction (PPI) networks were performed to identify the key molecules with Cytoscape, and Gene Set Enrichment Analysis (GSEA) was used to clarify the potential molecular functions. Two hub molecules were finally obtained and verified with quantitative real-time PCR (qRT-PCR). The results showed that the expression of mitochondria genes involved in mitochondrial electron transport, complex assembly of the respiratory chain, tricarboxylic acid cycle, oxidative phosphorylation, and ATP synthesis were down-regulated in skeletal muscle with aging. We further identified a primary hub gene of CYCS (Cytochrome C) and a key transcription factor of ESRRA (Estrogen-related Receptor Alpha) to be associated closely with skeletal muscle aging. PCR analysis confirmed the expressions of CYCS and ESRRA in gastrocnemius muscles of mice of different ages were significantly different, and decreased gradually with age. In conclusion, the main cause of skeletal muscle aging may be the systematically reduced expression of mitochondrial functional genes. The CYCS and ESRRA may play significant roles in the progression of skeletal muscle aging and serve as potential biomarkers for future diagnosis and treatment.

S100A16-induced adipogenesis is associated with up-regulation of 11 ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1).

The steadily increasing epidemic of obesity continues at alarming rates, is an important public health problem, and expression changes of S100A16 and 11 ß-hydroxysteroid dehydrogenase type 1(11ß-HSD1) is attributable to the adipocyte differentiation. In our previous study, we found that 11ß-HSD1 protein expression increased in S100A16-overexpressed 3T3-L1 cell model. In order to further investigate the relationship between S100A16 and 11ß-HSD1, and the molecular mechanisms of S100A16-induced adipogenesis, we constructed S100A16 transgenic and knockout mouse, and S100A16-overexpressed 3T3-L1 preadipocyte cell. Using S100A16 transgenic (S100A16Tg/+) mice fed with normal fat diet (NFD) and high fat diet (HFD) diet model, we evaluated the effect of S100A16 on adipogenesis, expression of 11ß-HSD1, and RNA sequencing and quantification of gene expression. Using the 3T3-L1 cell model, we examined the effect of S100A16 and 11ß-HSD1 on pre-adipocyte differentiation, and cell signaling events of 11ß-HSD1 overexpression induced by S100A16. We found that when compared with C57BL/6 mice, overexpression of S100A16 under the condition of HFD increased lipid content in WAT and fat infiltration in hepatocytes, 11ß-HSD1 protein expression increased along with S100A16. Elevated S100A16 and 11ß-HSD1 expression promoted adipogenesis in 3T3-L1 cells. Overexpression of S100A16 inhibited the degradation of 11ß-HSD1. We conclude that S100A16-induced adipogenesis is associated with up-regulation of 11ß-HSD1.

S100A16, a novel lipogenesis promoting factor in livers of mice and hepatocytes in vitro.

To investigate the role of S100 calcium-binding protein A16 (S100A16) in hepatic lipid metabolism, S100a16 transgenic, S100a16 knockdown, and wildtype C57BL/6 mice were fed either a high-fat diet (HFD) or normal-fat diet (NFD) for 16 weeks. The results showed that for HFD-fed mice, S100a16 transgenic mice showed significantly more severe fatty liver than other HFD-fed mice, with a significant increase in serum triglyceride (TG) concentration, with more and larger lipid droplets in the liver, whereas S100a16 knockdown mice were completely opposite, with liver fat lesions and TG serological changes being the mildest; for NFD-fed mice, liver fat accumulation and serum TG concentrations were significantly lower than those fed HFD, and no significant lipid droplets were found in the liver. Further, we found that calmodulin (CaM) interacts with S100A16, a member of the AMP-activated protein kinase (AMPK) pathway. Our research found that S100A16 regulates the AMPK pathway-associated protein by interacting with CaM to regulate liver lipid synthesis. S100A16 regulates liver lipid metabolism through the CaM/CAMKK2/AMPK pathway. Overexpression of S100A16 promotes the deterioration of fatty liver induced by HFD, and low expression of S100A16 can attenuate fatty liver.

[S100 calcium binding protein A16 promotes fat synthesis through endoplasmic reticulum stress in HepG2 cells].

The aim of this study was to investigate the role of S100 calcium binding protein A16 (S100A16) in lipid metabolism in hepatocytes and its possible biological mechanism. HepG2 cells (human hepatoma cell line) were cultured with fatty acid to establish fatty acid culture model. The control model was cultured without fatty acid. Each model was divided into three groups and transfected with S100a16 over-expression, shRNA and vector plasmids, respectively. The concentration of triglyceride (TG) in the cells was measured by kit, and the lipid droplets was observed by oil red O staining. Immunoprecipitation and mass spectrometry were used to find the interesting proteins interacting with S100A16, and the interaction was verified by immunoprecipitation. The further mechanism was studied by Western blot and qRT-PCR. The results showed that the intracellular lipid droplet and TG concentrations in the fatty acid culture model were significantly higher than those in the control model. The accumulation of intracellular fat in the S100a16 over-expression group was significantly higher than that in the vector plasmid transfection group. There was an interaction between heat shock protein A5 (HSPA5) and S100A16. Over-expression of S100A16 up-regulated protein expression levels of HSPA5, inositol-requiring enzyme 1α (IRE1α) and pIREα1, which belong to endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway. Meanwhile, over-expression of S100A16 up-regulated the mRNA expression levels of adipose synthesis-related gene Srebp1c, Acc and Fas. In the S100a16 shRNA plasmid transfection group, the above-mentioned protein and mRNA levels were lower than those of vector plasmid transfection group. These results suggest that S100A16 may promote lipid synthesis in HepG2 cells through endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway.

S100B suppresses the differentiation of C3H/10T1/2 murine embryonic mesenchymal cells into osteoblasts.

S100 calcium-binding protein B (S100B) is expressed and released by adipocytes, and is positively correlated with body mass index, however, the direct effects of S100B on adipocytes remain unclear. Bone marrow­derived mesenchymal stem cells have the capacity to differentiate into osteoblasts and adipocytes, which is important for bone metabolism. The current study aimed to determine the effect of S100B on adipogenesis and osteogenesis. The mouse embryo cell line C3H/10T1/2 was used to build cell models with varying levels of S100B protein expression. Western blot analysis was performed to assess the expression of various marker proteins. Oil red O staining and alizarin red S staining were used to detect adipogenesis and osteogenesis, respectively. S100B overexpression was associated with a significant increase in oil red O staining and a significant reduction in alizarin red S staining. Runt­related transcription factor­2 and bone morphogenetic protein 2 expression levels were significantly increased in the S100B underexpression group, however not in the S100B overexpression group. By contrast, the expression levels of the adipogenesis markers peroxisome proliferator­activated receptor Î³ and CCAAT­enhancer­binding protein α was significantly increased in the S100B overexpression group, however not in the S100B underexpression group. Osteogenesis stimulation increased extracellular signal­regulated kinase (ERK) phosphorylation, and adipogenesis stimulation increased c­Jun N­terminal kinase (JNK) phosphorylation. The results suggest that S100B inhibits osteogenesis, however stimulates adipogenesis. The ERK pathway is involved in the regulation of osteogenesis, whereas the JNK pathway is involved in the regulation of adipogenesis.

S100A16 promotes cell proliferation and metastasis via AKT and ERK cell signaling pathways in human prostate cancer.

S100A16 is a member of the S100 calcium-binding protein family. It is overexpressed in many types of tumors and associated with proliferation, migration, and invasion; however, its function in human prostate cancer is unresolved. Our objective was to determine its effects and the underlying pathways of S100A16 in prostate cancer tissues and cells. We measured S100A16 expression by quantitative real-time polymerase and Western blotting in eight matched prostate cancer and adjacent normal tissues, and in three prostate cancer cell lines, DU-145, LNCaP, and PC-3, compared to a normal prostate epithelial cell line PrEC. DU-145 cells stably overexpressing S100A16 and PC-3 cells with S100A16 knockdown were established by transfection with S100A16 overexpression plasmid or shRNAs. Invasion, migration, and proliferation were analyzed by transwell assay, wound healing, and colony formation assays, respectively. Western blotting and invasion assays were performed to determine expressions and activation of AKT, ERK, p21, and p27. S100A16 was significantly overexpressed in both prostate cancer tissues and cells lines compared to normal controls (P < 0.05). Overexpression of S100A16 significantly promoted invasion, migration, and proliferation in prostate cancer cells in vitro, whereas silencing S100A16 showed the converse effects (P < 0.05). Furthermore, overexpression of S100A16 activated cell signaling proteins AKT and ERK and downregulated tumor suppressors p21 and p27. Specific inhibitors, LY294002 and PD98059, suppressed activation of AKT and ERK, which attenuated DU-145 cell clone formation and invasion induced by S100A16 overexpression. S100A16 may promote human prostate cancer progression via signaling pathways involving AKT, ERK, p21, and p27 downstream effectors. Our findings suggest that S100A16 may serve as a novel therapeutic or diagnostic target in human prostate cancer.

Protective effects of astragaloside IV on db/db mice with diabetic retinopathy.

OBJECTIVES: Diabetic retinopathy (DR) is a common diabetic eye disease which is well-known as the result of microvascular retinal changes. Although the potential biological functions of astragaloside IV (AS IV) have long been described in traditional system of medicine, its protective effect on DR remains unclear. This study aims to investigate the function and mechanism of AS IV on type 2 diabetic db/db mice. METHODS: Db/db mice were treated with AS IV (4.5 mg/kg or 9 mg/kg) or physiological saline by oral gavage for 20 weeks along with db/m mice. In each group, retinal ganglion cell (RGC) function was measured by pattern electroretinogram (ERG) and apoptosis was determined by Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Blood and retina aldose reductase (AR) activity were quantified by chemiluminescence analysis. The expressions of phosporylated-ERK1/2, NF-κB were determined by Western blot analysis. Furthermore, the expression of related downstream proteins were quantified by Label-based Mouse Antibody Array. RESULTS: Administration of AS IV significantly improved the amplitude in pattern ERG and reduced the apoptosis of RGCs.in db/db mice. Furthermore, downregulation of AR activity, ERK1/2 phosphorylation, NF-κB and related cytokine were observed in AS IV treatment group. CONCLUSIONS: Our study indicated that AS IV, as an inhibitor of AR, could prevent the activation of ERK1/2 phosporylation and NF-kB and further relieve the RGCs disfunction in db/db mice with DR. It has provided a basis for investigating the clinical efficacy of AR inhibitors in preventing DR.

Role of p53 in preadipocyte differentiation.

Insulin resistance, diabetes and many kinds of cancers are common in overweight and obese individuals. The tumor suppressor p53 is important in securing genetic stability, but its role in the regulation of metabolic processes and cell differentiation remains unclear. We have investigated the role of p53 in adipocyte differentiation. Using 3T3-L1 cells, a mouse embryonic fibroblast preadipocyte model and DIO rat model, p53 expression and function during adipocyte differentiation were investigated. p53 expression increased on the second and fourth day of adipocyte differentiation and decreased thereafter. Its overexpression in 3T3-L1 preadipocytes markedly reduced adipogenesis and marker gene expression. p53 activity was weakened in DIO rat abdominal adipose tissue because of an decreased expression of its activated phosphorylated form. In contrast, p53 knockout enhanced adipogenesis and the expression of marker genes, but significantly reduced insulin-stimulated Akt phosphorylation. These results indicate that p53 partly suppresses preadipocyte differentiation and adipogenesis by regulating adipocyte gene expression and Akt signaling.

Estrogen suppresses adipogenesis by inhibiting S100A16 expression.

The aim of this study is to determine the effects of E2 on metabolic syndrome and the molecular mechanisms involving S100A16. Ovariectomized (OVX) rat models and mouse embryonic fibroblasts cell models were used. E2 loss in OVX rats induced body weight gain and central abdominal fat accumulation, which were ameliorated by E2 treatment under chow and high-fat diet (HFD) conditions. E2 decreased the expression of the adipocyte marker genes PPARγ, aP2, C/EBPα, and S100A16. E2 inhibited adipogenesis. Overexpression of S100A16 reversed the E2-induced adipogenesis effect. A luciferase assay showed that E2 inhibited the expression of S100A16. E2 treatment decreased body weight gain and central abdominal fat accumulation under both chow and HFD conditions. Also, E2 suppressed adipogenesis by inhibiting S100A16 expression.

S100A16 inhibits osteogenesis but stimulates adipogenesis.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) have the capacity to differentiate into osteoblasts and adipocytes. Bone marrow adipogenesis exerts an inhibitory effect on osteogenesis, which leads to osteoporosis. S100A16, a novel member of the S100 family, is ubiquitously expressed, and markedly enhances adipogenesis. The aim of this study was to demonstrate, in the mouse BM-MSC model, whether S100A16 significantly stimulates adipogenic, rather than osteogenic differentiation. The overexpression of S100A16 led to a significant increase in Oil Red O staining (a marker of adipocyte differentiation) but a decrease in Alizarin Red S staining (a marker of osteoblast differentiation). In contrast, reducing the expression of S100A16 resulted in minimal Oil Red O staining but increased Alizarin Red S staining. During differentiation into osteoblasts, RUNX2 expression increased fourfold in the S100A16(KO+/-) BM-MSCs, but only increased by approximately 1.5-fold in the S100A16(TG+/+) BM-MSCs. And BMP2 occurred in the same changes. Upon induction of BM-MSC differentiation into adipocytes, peroxisome proliferator-activated receptor-γ (PPARγ) and CCAAT/enhancer binding protein-α expression were significantly higher in the cells overexpressing S100A16 protein but lower in the cells with reduced expression of S100A16 protein, compared with the control cells, which were BM-MSCs derived from C57/BL6. S100A16 increased PPARγ promoter luciferase activity and decreased RUNX2 promoter luciferase activity. ERK1/2 phosphorylation was stimulated during osteogenesis, whereas p-JNK phosphorylation was increased by stimulation of adipogenesis. Our results suggest that S100A16 inhibits osteogenesis but stimulates adipogenesis by increasing the transcription of PPARγ and decreasing the transcription of RUNX2. The ERK1/2 pathway is involved in the regulation of osteogenesis whereas the JNK pathway is involved in adipogenesis.

Insulin glargine and risk of cancer: a meta-analysis.

AIMS: Recently, more and more attention has been drawn on the long-term effects of insulin glargine. Here we strived to estimate the association of cancer occurrence with the use of insulin glargine. METHODS: We searched all the publications regarding the association between cancer occurrence and the use of insulin glargine using the US National Library of Medicine's PubMed database. Data were independently extracted and analyzed using random or fixed effects meta-analysis depending upon the degree of heterogeneity. RESULTS: Seven cohort studies were included in the meta-analysis. Cancer occurrence had no significant difference in glargine-treated patients compared to patients treated with other insulins (RR=0.86, 95% CI=0.69-1.07, p=0.17, P(heterogeneity)<0.00001). In our subgroup analysis, glargine, compared to other insulins, did not increase the risk of breast cancer (RR=1.14, 95% CI=0.65-2.02, p=0.65, P(heterogeneity)=0.002), prostate cancer (RR=1.00, 95% CI=0.79-1.26, p=0.99, P(heterogeneity)=0.78), pancreatic cancer (RR=0.57, 95% CI=0.14-2.35, p=0.44, P(heterogeneity)=0.0002) and gastrointestinal cancer (RR=0.80, 95% CI=0.62-1.02, p=0.07, P(heterogeneity)=0.86). CONCLUSIONS: This meta-analysis of open-label studies does not support an increased cancer risk in patients treated with insulin glargine. The result provides confidence for the development of insulin glargine, but needs confirmation by further clinical studies.

S100A16 mediation of weight gain attenuation induced by dietary calcium.

Dietary calcium influences the regulation of energy metabolism, and weight gain is attenuated by a high-calcium diet. S100A16 is a novel calcium-binding signaling protein of the EF-hand superfamily that promotes adipogenesis. This study aimed to investigate the effect of S100A16 on weight gain attenuation with a calcium-rich diet. An obese rat model was produced after feeding with a high-fat diet. Animals were randomly divided into 4 groups according to the diet provided over 8 weeks: normal diet group; high-fat, normal-calcium diet group; high-fat, high-calcium diet (HH) group; and high-fat, low-calcium diet group. Serum biochemistry was analyzed, and body weight and visceral fat pads were measured. Expression of S100A16 was assayed by Western blotting. Adipogenesis was detected by oil red O staining. Increases in body weight and visceral fat weight were attenuated in the HH group. High-calcium diets decreased the concentrations of serum total cholesterol and triglyceride. Expression of S100A16 decreased in the HH group. Using the 3T3-L1 preadipocyte model, it was observed that elevation of intracellular Ca(2+) via calcium ionophores led to the exclusion of S100A16 from the nucleus. Overexpression of S100A16 in 3T3-L1 preadipocytes enhanced adipogenesis, although a significant reduction in Akt phosphorylation was also detected. High-calcium diets were associated with a significant reduction in body weight gain. High-calcium diets may lead to nuclear exclusion of S100A16, which results in the inhibition of adipogenesis and enhanced insulin sensitivity.

Osteopontin enhances the expression and activity of MMP-2 via the SDF-1/CXCR4 axis in hepatocellular carcinoma cell lines.

BACKGROUND AND AIMS: Osteopontin, SDF-1α, and MMP-2 are important secreted molecules involved in the pathophysiology of human hepatocellular carcinoma (HCC). This study investigates the effect of the SDF-1α/CXCR4 axis on expression and activity of MMP-2 induced by osteopontin. METHODS: The expression of CXCR4, SDF-1α, MMP-2 and their associated cellular signaling cascades, involving Akt and MAP Kinases, were determined by Western blotting. The activities of MMP-2 and MMP-9 were assayed by gel zymography. The role of the osteopontin receptors integrin α(v)ß3 and CD44v6 was evaluated using neutralizing antibodies. We also established CXCR4-deficient SMMC7721 cell lines by transfection with miRNA-CXCR4 plasmids and determined cell invasion activity in a transwell assay. RESULTS: In comparison with untreated cells, recombinant human osteopontin (rhOPN) up-regulated CXCR4, SDF-1α, and MMP-2 expression about 5-, 4-, and 6-fold on the protein levels through binding to integrin α(v)ß3 and CD44v6 in hepatocellular carcinoma cells (SMMC7721 and HepG2). Inhibition of the SDF-1α/CXCR4 axis down-regulated the rhOPN-induced MMP-2 expression and activity. rhOPN also activated Akt, p38 and JNK. Down-regulation of CXCR4 decreased the rhOPN-induced invasion in SMMC7721 cells. CONCLUSION: These results indicate that rhOPN up-regulates MMP-2 through the SDF-1α/CXCR4 axis, mediated by binding to integrin α(v)ß3 and CD44v6 and activating the PI-3K/Akt and JNK pathways in HepG2 and SMMC7721 cells. Therefore, the osteopontin-SDF-1α/CXCR4-MMP-2 system may be a new therapeutic target for treating HCC progression.

Requirement of Osteopontin in the migration and protection against Taxol-induced apoptosis via the ATX-LPA axis in SGC7901 cells.

BACKGROUND: Autotaxin (ATX) possesses lysophospholipase D (lyso PLD) activity, which converts lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA). The ATX-LPA signaling axis has been implicated in angiogenesis, chronic inflammation and tumor progression. Osteopontin (OPN) is an important chemokine involved in the survival, proliferation, migration, invasion and metastasis of gastric cancer cells. The focus of the present study was to investigate the relationship between the ATX-LPA axis and OPN. RESULTS: In comparison with non-treated cells, we found that the ATX-LPA axis up-regulated OPN expression by 1.92-fold in protein levels and 1.3-fold in mRNA levels. The ATX-LPA axis activates LPA2, Akt, ERK and ELK-1 and also protects SGC7901 cells from apoptosis induced by Taxol treatment. CONCLUSIONS: This study provides the first evidence that expression of OPN induced by ATX-LPA axis is mediated by the activation of Akt and MAPK/ERK pathways through the LPA2 receptor. In addition, OPN is required for the protective effects of ATX-LPA against Taxol-induced apoptosis and ATX-LPA-induced migration of SGC7901 cells.

ATX-LPA axis induces expression of OPN in hepatic cancer cell SMMC7721.

Osteopontin (OPN) and autotaxin (ATX) are important chemokines involved in the survival, proliferation, migration, invasion, and metastasis of many cancer cells. The focus of the study was to investigate the relationship between OPN and ATX-lysophosphatidic acid (LPA) axis. The expression of OPN and its cellular cascades were determined by western blot and real-time quantitative transcription polymerase chain reaction (real-time PCR) analyses. Cell migration activity was determined by a Transwell-migration assay. In comparison with nontreated cells, we found that the ATX-LPA axis upregulated OPN expression by 2.91-fold in protein levels and 2.52-fold in mRNA levels. The ATX-LPA axis activates Akt and ATX/LPC-induced OPN expression in SMMC7721 cells was largely reduced by the inhibitors of phosphatidylinositol 3-kinase (PI3K)/Akt or LPA receptor. This study provides the first evidence that the induction of the OPN expression by ATX-LPA axis was mediated by the activation of Akt through LPA receptors and OPN was required for migration of SMMC7721 cells induced by ATX-LPA axis.