Secreted Protein Acidic and Rich in Cysteine (Sparc) KO Leads to an Accelerated Ageing Phenotype Which Is Improved by Exercise Whereas SPARC Overexpression Mimics Exercise Effects in Mice.

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein implicated in various functions, including metabolism, tissue regeneration, and functional homeostasis. SPARC/Sparc declines with ageing but increases with exercise. We aim to verify two hypotheses: (1) SPARC deficiency leads to an ageing-like phenotype (metabolic decline, muscle loss, etc.), and (2) SPARC overexpression would mimic exercise, counteract ageing, and improve age-related changes. Our mice experiments are divided into two parts. First, we explore the consequences of Sparc knockout (KO) and compare them to the ageing effects. We also observe the effects of exercise. In the second part, we study the effects of SPARC overexpression and compare them to the exercise benefits. At the end, we make an analysis of the results to point out the analogies between Sparc KO and the ageing-like phenotype on the one hand and make comparisons between SPARC overexpression and exercise in the context of exercise counteracting ageing. The measurements were mainly related to tissue weights, adiposity, metabolism, and muscle strength. The main findings are that Sparc KO reduced glucose tolerance, muscle glucose transporter expression, and abdominal adipose tissue weight but increased glycogen content in the muscle. SPARC overexpression increased muscle strength, muscle mass, and expressions of the muscle glucose transporter and mitochondrial oxidative phosphorylation but lowered the glycemia and the adiposity, especially in males. Collectively, these findings, and the data we have previously reported, show that Sparc KO mice manifest an ageing-like phenotype, whereas SPARC overexpression and exercise generate similar benefits. The benefits are towards counteracting both the SPARC deficiency-induced ageing-like phenotype as well as reversing the age-related changes. The potential applications of these findings are to build/optimize Sparc KO-based animal models of various health conditions and, on the other hand, to develop therapies based on introducing SPARC or targeting SPARC-related pathways to mimic exercise against age-related and metabolic disorders.

Exercise and High-Fat Diet in Obesity: Functional Genomics Perspectives of Two Energy Homeostasis Pillars.

The heavy impact of obesity on both the population general health and the economy makes clarifying the underlying mechanisms, identifying pharmacological targets, and developing efficient therapies for obesity of high importance. The main struggle facing obesity research is that the underlying mechanistic pathways are yet to be fully revealed. This limits both our understanding of pathogenesis and therapeutic progress toward treating the obesity epidemic. The current anti-obesity approaches are mainly a controlled diet and exercise which could have limitations. For instance, the "classical" anti-obesity approach of exercise might not be practical for patients suffering from disabilities that prevent them from routine exercise. Therefore, therapeutic alternatives are urgently required. Within this context, pharmacological agents could be relatively efficient in association to an adequate diet that remains the most efficient approach in such situation. Herein, we put a spotlight on potential therapeutic targets for obesity identified following differential genes expression-based studies aiming to find genes that are differentially expressed under diverse conditions depending on physical activity and diet (mainly high-fat), two key factors influencing obesity development and prognosis. Such functional genomics approaches contribute to elucidate the molecular mechanisms that both control obesity development and switch the genetic, biochemical, and metabolic pathways toward a specific energy balance phenotype. It is important to clarify that by "gene-related pathways", we refer to genes, the corresponding proteins and their potential receptors, the enzymes and molecules within both the cells in the intercellular space, that are related to the activation, the regulation, or the inactivation of the gene or its corresponding protein or pathways. We believe that this emerging area of functional genomics-related exploration will not only lead to novel mechanisms but also new applications and implications along with a new generation of treatments for obesity and the related metabolic disorders especially with the modern advances in pharmacological drug targeting and functional genomics techniques.

Extracellular matrix/mitochondria pathway: A novel potential target for sarcopenia.

Sarcopenia is a geriatric syndrome characterized by a progressive and generalized loss of muscle mass, strength, and endurance. Although the mitochondrial hypothesis of sarcopenia has been proposed, it remains a highly debated topic. The age-related declines in skeletal muscle regenerative capacity have been attributed to the changes in the extracellular matrix (ECM) composition. Therefore, mitochondrial deterioration in the muscle may be implicated in the sarcopenic process, and a possible link between ECM remodeling and mitochondrial dysfunction may be suggested. While exercise training improves elderly muscle function, the contradictory results obtained from different studies, the lack of a well-defined training program as well as age-related falls may raise questions regarding the overall benefits of this interventional strategy for sarcopenic elderly who have limited ability to perform physical exercise. Thus, it remains important to be able to offer less cumbersome but equally effective alternatives such as pharmacological approaches to help elderly patients. In this review, we discuss the involvement of mitochondrial dysfunction as well as the ECM remodeling in sarcopenia, and we suggest that the future of sarcopenia drug development may be targeting ECM/mitochondria pathway.

Secreted protein acidic and rich in cysteine and bioenergetics: Extracellular matrix, adipocytes remodeling and skeletal muscle metabolism.

The extracellular matrix (ECM) remodeling plays important roles in both adipocytes shape/expansion remodeling and the skeletal muscle (SM) metabolism. Secreted protein acidic and rich in cysteine (SPARC) is expressed in divers tissues including adipose tissue (AT) and SM where it impacts a variety of remodeling as well as metabolic functions. SPARC, also known as osteonectin or BM-40, is a glycoprotein associated with the ECM. Numerous researches attempted to elucidate the implications of SPARC in these two key metabolic tissues under different conditions. Whereas SPARC deficiency tends to shape the remodeling of the adipocytes and the fat distribution, this deficiency decreases SM metabolic properties. On the other hand, SPARC seems to be an enhancer of the metabolism and a mediator of the exercise-induced adaptation in the SM and as well as an adipogenesis inhibitor. Some findings about the SPARC effects on AT and SM seem "contradictory" in terms of tissue development and energy profile therefore highlighting the mechanistic role of SPARC in both is a priority. Yet, within this review, we expose selected researches and compare the results. We conclude with explanations to "reconcile" the different observations, hypothesize the feedback and regulatory character of SPARC and put its roles within the energetic and structural maps of both adipocytes and myocytes in homeostasis and in situations such as obesity or exercise. These properties explain the modifications and the remodeling seen in AT and SM undergoing adaptive changes (obesity, exercise, etc.) and represent a starting point for precise therapeutic targeting of SPARC-related pathways is conditions such as obesity, sarcopenia and diabetes.

Functional genomics applications and therapeutic implications in sarcopenia.

The human genome contains around 20,000-25,000 genes coding for 30,000 proteins. Some proteins and genes represent therapeutic targets for human diseases. RNA and protein expression profiling tools allow the study of the molecular basis of aging and drug discovery validation. Throughout the life, there is an age-related and disease-related muscle decline. Sarcopenia is defined as a loss of muscle mass and a decrease in functional properties such as muscle strength and physical performance. Yet, there is still no consensus on the evaluation methods of sarcopenia prognosis. The main challenge of this complex biological phenomena is its multifactorial etiology. Thus, functional genomics methods attempt to shape the related scientific approaches via an innovative in-depth view on sarcopenia. Gene and drug high throughput screening combined with functional genomics allow the generation and the interpretation of a large amount of data related to sarcopenia and therapeutic progress. This review focuses on the application of selected functional genomics techniques such as RNA interference, RNA silencing, proteomics, transgenic mice, metabolomics, genomics, and epigenomics to better understand sarcopenia mechanisms.

Sparc, an EPS-induced gene, modulates the extracellular matrix and mitochondrial function via ILK/AMPK pathways in C2C12 cells.

AIMS: Secreted protein acidic and rich in cysteine, (SPARC), is a matricellular protein implicated in the modulation of the extracellular matrix (ECM) and mitochondrial proteins expression. MAIN METHODS: To study the mechanism through which SPARC is involved in the possible link between ECM and mitochondria, C2C12 myoblasts were cultured with/without the exogenous addition/inhibition of SPARC as well as activation/inhibition of adenosine monophosphate-activated protein kinase (AMPK). Electrical pulse stimulation (EPS), was applied for 2 days in myotubes. KEY FINDINGS: The expressions of ECM-related (integrin-linked kinase (ILK), glycogen synthase kinase-3 beta (GSK-3ß), phosphorylated-GSK-3ß (p-GSK-3ß) and collagen 1a1), mitochondrial-related (AMPK, phosphorylated-AMPK (p-AMPK), succinate dehydrogenase (SDHB) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (Pgc1α)) and SPARC proteins and/or genes were measured after modulation of SPARC and/or AMPK as well as with or without EPS. The addition of SPARC in C2C12 myoblast increased the expression of ILK, p-GSK-3ß and p-AMPK whereas anti-SPARC antibody decreased them at different incubation times (0, 10, and 30 min, and 6 h). The AMPK activation increased SPARC, collagen 1a1, p-AMPK and SDHB proteins level, however, AMPK inhibition blunted the effects. EPS induced Sparc and Pgc1a genes expression. SIGNIFICANCE: Sparc, an EPS-induced gene, may be involved in the link between ECM remodeling and mitochondrial function in muscle via its interaction with ILK/AMPK.

Energy and metabolic pathways in trefoil factor family member 2 (Tff2) KO mice beyond the protection from high-fat diet-induced obesity.

AIMS: Trefoil factor family member 2 (TFF2) is a small gut peptide. We have previously shown that Tff2 knock out (KO) mice are protected from high-fat (HF) diet-induced obesity (De Giorgio et al., 2013a). Thus, exploring Tff2 KO-related pathways of mice at the genomic, proteinic and biochemical levels would allow us to elucidate the processes behind this protection from obesity. MAIN METHODS: To explore the metabolic and energetic effects related to Tff2 deficiency, we used sampled blood from the previous study to measure levels of free fatty acids, glucose, glycerol and triglycerides in serum. Expression levels of selected genes and proteins related to energy metabolism in the skeletal muscle, liver and adipose tissue were also studied. KEY FINDINGS: Following the 12-wk challenging of Tff2 KO and WT mice with both HF and low-fat diet, Tff2 KO mice had lower levels of serum glucose, triglycerides and glycerol. Importantly, western blotting and Q_RT-PCR revealed that the expression levels of selected genes and proteins are toward less fat storage and increased energy expenditure by enhancing lipid and glucose utilization via oxidative phosphorylation. SIGNIFICANCE: We mapped a part of the metabolic and biochemical pathways of lipids and glucose involving the adipose tissue, liver, skeletal muscle and sympathetic nervous system that protect Tff2 KO mice from the HF diet-induced obesity. Our data highlight Tff2-related pathways as potential targets for obesity therapies.

Implication of SPARC in the modulation of the extracellular matrix and mitochondrial function in muscle cells.

Secreted protein, acidic and rich in cysteine (SPARC) is differentially associated with cell proliferation and extracellular matrix (ECM) assembly. We show here the effect of exogenous SPARC inhibition/induction on ECM and mitochondrial proteins expression and on the differentiation of C2C12 cells. The cells were cultured in growth medium (GM) supplemented with different experimental conditions. The differentiation of myoblasts was studied for 5 days, the expressions of ECM and mitochondrial proteins were measured and the formation of the myotubes was quantified after exogenous induction/inhibition of SPARC. The results indicate that the addition of recombinant SPARC protein (rSPARC) in cell culture medium increased the differentiation of C2C12 myoblasts and myogenin expression during the myotube formation. However, the treatment with antibody specific for SPARC (anti-SPARC) prevented the differentiation and decreased myogenin expression. The induction of SPARC in the proliferating and differentiating C2C12 cells increased collagen 1a1 protein expression, whereas the inhibition decreased it. The effects on fibronectin protein expression were opposite. Furthermore, the addition of rSPARC in C2C12 myoblast increased the expression of mitochondrial proteins, ubiquinol-cytochrome c reductase core protein II (UQCRC2) and succinate dehydrogenase iron-sulfur subunit (SDHB), whereas the anti-SPARC decreased them. During the differentiation, only the anti-SPARC had the effects on mitochondrial proteins, NADH dehydrogenase ubiquinone 1 beta subcomplex subunit 8 (NADHB8), SDHB and cytochrome c oxidase 1 (MTCO1). Thus, SPARC plays a crucial role in the proliferation and differentiation of C2C12 and may be involved in the link between the ECM remodeling and mitochondrial function.

Differential gene expression analysis in ageing muscle and drug discovery perspectives.

Identifying therapeutic target genes represents the key step in functional genomics-based therapies. Within this context, the disease heterogeneity, the exogenous factors and the complexity of genomic structure and function represent important challenges. The functional genomics aims to overcome such obstacles via identifying the gene functions and therefore highlight disease-causing genes as therapeutic targets. Genomic technologies promise to reshape the research on ageing muscle, exercise response and drug discovery. Herein, we describe the functional genomics strategies, mainly differential gene expression methods microarray, serial analysis of gene expression (SAGE), massively parallel signature sequence (MPSS), RNA sequencing (RNA seq), representational difference analysis (RDA), and suppression subtractive hybridization (SSH). Furthermore, we review these illustrative approaches that have been used to discover new therapeutic targets for some complex diseases along with the application of these tools to study the modulation of the skeletal muscle transcriptome.

Identification of the principal transcriptional regulators for low-fat and high-fat meal responsive genes in small intestine.

BACKGROUND: High-fat (HF) diet is a well-known cause of obesity. To identify principle transcriptional regulators that could be therapeutic targets of obesity, we investigated transcriptomic modulation in the duodenal mucosa following low-fat (LF) and HF meal ingestion. METHODS: Whereas one group of mice was sacrificed after fasting, the others were fed ad libitum with LF or HF meal, and sacrificed 30 min, 1 h and 3 h after the beginning of the meal. A transcriptome analysis of the duodenal mucosa of the 7 groups was conducted using both microarray and serial analysis of gene expression (SAGE) method followed by an Ingenuity Pathways Analysis (IPA). RESULTS: SAGE and microarray showed that the modulation of a total of 896 transcripts in the duodenal mucosa after LF and/or HF meal, compared to the fasting condition. The IPA identified lipid metabolism, molecular transport, and small molecule biochemistry as top three molecular and cellular functions for the HF-responsive, HF-specific, HF-delay, and LF-HF different genes. Moreover, the top transcriptional regulator for the HF-responsive and HF-specific genes was peroxisome proliferator-activated receptor alpha (PPARα). On the other hand, the LF-responsive and LF-specific genes were related to carbohydrate metabolism, cellular function and maintenance, and cell death/cellular growth and proliferation, and the top transcriptional regulators were forkhead box protein O1 (FOXO1) and cAMP response element binding protein 1 (CREB1), respectively. CONCLUSIONS: These results will help to understand the molecular mechanisms of intestinal response after LF and HF ingestions, and contribute to identify therapeutic targets for obesity and obesity-related diseases.