Tetrahydrocannabivarin (THCV) Protects Adipose-Derived Mesenchymal Stem Cells (ASC) against Endoplasmic Reticulum Stress Development and Reduces Inflammation during Adipogenesis.

The endoplasmic reticulum (ER) fulfills essential duties in cell physiology, and impairment of this organelle's functions is associated with a wide number of metabolic diseases. When ER stress is generated in the adipose tissue, it is observed that the metabolism and energy homeostasis of the adipocytes are altered, leading to obesity-associated metabolic disorders such as type 2 diabetes (T2D). In the present work, we aimed to evaluate the protective effects of Δ9-tetrahydrocannabivarin (THCV, a cannabinoid compound isolated from Cannabis sativa L.) against ER stress in adipose-derived mesenchymal stem cells. Our results show that pre-treatment with THCV prevents the subcellular alteration of cell components such as nuclei, F-actin, or mitochondria distribution, and restores cell migration, cell proliferation and colony-forming capacity upon ER stress. In addition, THCV partially reverts the effects that ER stress induces regarding the activation of apoptosis and the altered anti- and pro-inflammatory cytokine profile. This indicates the protective characteristics of this cannabinoid compound in the adipose tissue. Most importantly, our data demonstrate that THCV decreases the expression of genes involved in the unfolded protein response (UPR) pathway, which were upregulated upon induction of ER stress. Altogether, our study shows that the cannabinoid THCV is a promising compound that counters the harmful effects triggered by ER stress in the adipose tissue. This work paves the way for the development of new therapeutic means based on THCV and its regenerative properties to create a favorable environment for the development of healthy mature adipocyte tissue and to reduce the incidence and clinical outcome of metabolic diseases such as diabetes.

Astaxanthin and other Nutrients from Haematococcus pluvialis-Multifunctional Applications.

Bioactive compounds of natural origin are gaining increasing popularity. High biological activity and bioavailability, beneficial effects on health and safety of use are some of their most desirable features. Low production and processing costs render them even more attractive. Microorganisms have been used in the food, medicinal, cosmetic and energy industries for years. Among them, microalgae have proved to be an invaluable source of beneficial compounds. Haematococcus pluvialis is known as the richest source of natural carotenoid called astaxanthin. In this paper, we focus on the cultivation methods of this green microalga, its chemical composition, extraction of astaxanthin and analysis of its antioxidant, anti-inflammatory, anti-diabetic and anticancer activities. H. pluvialis, as well as astaxanthin can be used not only for the treatment of human and animal diseases, but also as a valuable component of diet and feed.

Sex hormone binding globulin (SHBG) modulates mitochondrial dynamics in PPARγ-depleted equine adipose derived stromal cells.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a transcription factor that promotes adipogenesis, lipid uptake and storage, insulin sensitivity, and glucose metabolism. Hence, defects in PPARγ have been associated to the development of metabolic disorders. Sex hormone-binding globulin (SHBG) is a glycoprotein primarily produced in the liver that regulates the bioavailability of sex hormones. Alike PPARγ, low SHBG levels have been correlated with insulin resistance and associated endocrine abnormalities. Therefore, this study aimed to verify whether SHBG may restore depleted PPARγ functions and thus serve as a new candidate for the management of metabolic conditions. A model of equine adipose-derived stromal cells (EqASCs) has been used, in which a PPARγ silencing and SHBG treatment have been achieved to determine the changes in cell viability, premature senescence, oxidative stress, and mitochondrial functions. Obtained data demonstrated that loss in PPARγ triggers cell apoptosis which is not reversed by SHBG application. Moreover, PPARγ knockdown cells exhibited premature senescence, which has been substantially alleviated by SHBG concomitantly to increased BAX/BCL2 ratio, suggesting a possible effect on senescence-induced apoptosis resistance. Interestingly, PPARγ silencing induced a significant alteration in mitochondrial membrane potential as well as the expression of dynamics and metabolism-related markers. SHBG treatment enabled to ameliorate the transmembrane potential, to normalize the expression levels of key dynamics and metabolism mediators, and to restore the protein levels of PINK, which is critically involved in mitochondria recycling machinery. Presented data suggest that SHBG may provide new mechanistic insights into the regulation of PPARγ functions, and thus offers a preliminary picture on a possible SHBG-PPARγ metabolic crosstalk. KEY MESSAGES : PPARγ is a transcription factor that tightly regulates cell metabolism. Low SHBG levels correlate with insulin resistance and associated endocrine abnormalities. PPARγ silencing reduces cell viability, triggers premature senescence and profound mitochondrial failure in equine ASCs. SHBG protein reverses senescent phenotype and apoptosis resistance of PPARγ- ASCs. SHBG improves mitochondrial dynamics and metabolism following PPARγ knockdown. SHBG might serve as a PPARγ potential mimicking agent for the modulation of ASCs metabolic processes.

In Vitro Generated Equine Hepatic-Like Progenitor Cells as a Novel Potent Cell Pool for Equine Metabolic Syndrome (EMS) Treatment.

Equine metabolic syndrome (EMS) is recognized as one of the leading cause of health threatening in veterinary medicine worldwide. Recently, PTP1B inhibition has been proposed as an interesting strategy for liver insulin resistance reversion in both equines and humans, however as being a multifactorial disease, proper management of EMS horses further necessities additional interventional approaches aiming at repairing and restoring liver functions. In this study, we hypothesized that in vitro induction of Eq_ASCs hepatogenic differentiation will generate a specialized liver progenitor-like cell population exhibiting similar phenotypic characteristics and regenerative potential as native hepatic progenitor cells. Our obtained data demonstrated that Eq_ASCs-derived liver progenitor cells (Eq_HPCs) displayed typical flattened polygonal morphology with packed fragmented mitochondrial net, lowered mesenchymal CD105 and CD90 surface markers expression, and significant high expression levels of specific hepatic lineage genes including PECAM-1, ALB, AFP and HNF4A. therewith, generated Eq_HPCs exhibited potentiated stemness and pluripotency markers expression (NANOG, SOX-2 and OCT-4). Hence, in vitro generation of hepatic progenitor-like cells retaining high differentiation capacity represents a promising new approach for the establishment of cell-based targeted therapies for the restoration of proper liver functions in EMS affected horses.

Sex Hormone-Binding Globulin (SHBG) Maintains Proper Equine Adipose-Derived Stromal Cells (ASCs)' Metabolic Functions and Negatively Regulates their Basal Adipogenic Potential.

BACKGROUND: Sex hormone binding globulin (SHBG) deteriorated expression has been recently strongly correlated to increased level of circulating pro-inflammatory cytokines and insulin resistance, which are typical manifestations of equine metabolic syndrome (EMS). Despite previous reports demonstrated the potential therapeutic application of SHBG for liver-related dysfunctions, whether SHBG might modulate equine adipose-derived stem/stromal cells (EqASCs) metabolic machinery remains unknown. Therefore, we evaluated for the first time the impact of SHBG protein on metabolic changes in ASCs isolated from healthy horses. METHODS: Beforehand, SHBG protein expression has been experimentally lowered using a predesigned siRNA in EqASCs to verify its metabolic implications and potential therapeutic value. Then, apoptosis profile, oxidative stress, mitochondrial network dynamics and basal adipogenic potential have been evaluated using various molecular and analytical techniques. RESULTS: The SHBG knockdown altered the proliferative and metabolic activity of EqASCs, while dampening basal apoptosis via Bax transcript suppression. Furthermore, the cells treated with siRNA were characterized by senescent phenotype, accumulation of reactive oxygen species (ROS), nitric oxide, as well as decreased mitochondrial potential that was shown by mitochondrial membrane depolarization and lower expression of key mitophagy factors: PINK, PARKIN and MFN. The addition of SHBG protein reversed the impaired and senescent phenotype of EMS-like cells that was proven by enhanced proliferative activity, reduced apoptosis resistance, lower ROS accumulation and greater mitochondrial dynamics, which is proposed to be related to a normalization of Bax expression. Crucially, SHBG silencing enhanced the expression of key pro-adipogenic effectors, while decreased the abundance of anti-adipogenic factors namely HIF1-α and FABP4. The addition of exogenous SHBG further depleted the expression of PPARγ and C/EBPα and restored the levels of FABP4 and HIF1-α evoking a strong inhibitory potential toward ASCs adipogenesis. CONCLUSION: Herein, we provide for the first time the evidence that SHBG protein in importantly involved in various key metabolic pathways governing EqASCs functions, and more importantly we showed that SHBG negatively affect the basal adipogenic potential of tested ASCs through a FABP4-dependant pathway, and provide thus new insights for the development of potential anti-obesity therapeutic approach in both animals and humans.

The PTP1B inhibitor MSI-1436 ameliorates liver insulin sensitivity by modulating autophagy, ER stress and systemic inflammation in Equine metabolic syndrome affected horses.

Background: Equine metabolic syndrome (EMS) is a multifactorial pathology gathering insulin resistance, low-grade inflammation and past or chronic laminitis. Among the several molecular mechanisms underlying EMS pathogenesis, increased negative insulin signalling regulation mediated by protein tyrosine phosphatase 1 B (PTP1B) has emerged as a critical axis in the development of liver insulin resistance and general metabolic distress associated to increased ER stress, inflammation and disrupted autophagy. Thus, the use of PTP1B selective inhibitors such as MSI-1436 might be considered as a golden therapeutic tool for the proper management of EMS and associated conditions. Therefore, the present investigation aimed at verifying the clinical efficacy of MSI-1436 systemic administration on liver metabolic balance, insulin sensitivity and inflammatory status in EMS affected horses. Moreover, the impact of MSI-1436 treatment on liver autophagy machinery and associated ER stress in liver tissue has been analysed. Methods: Liver explants isolated from healthy and EMS horses have been treated with MSI-1436 prior to gene and protein expression analysis of main markers mediating ER stress, mitophagy and autophagy. Furthermore, EMS horses have been intravenously treated with a single dose of MSI-1436, and evaluated for their metabolic and inflammatory status. Results: Clinical application of MSI-1436 to EMS horses restored proper adiponectin levels and attenuated the typical hyperinsulinemia and hyperglycemia. Moreover, administration of MSI-1436 further reduced the circulating levels of key pro-inflammatory mediators including IL-1ß, TNF-α and TGF-ß and triggered the Tregs cells activation. At the molecular level, PTP1B inhibition resulted in a noticeable mitigation of liver ER stress, improvement of mitochondrial dynamics and consequently, a regulation of autophagic response. Similarly, short-term ex vivo treatment of EMS liver explants with trodusquemine (MSI-1436) substantially enhanced autophagy by upregulating the levels of HSC70 and Beclin-1 at both mRNA and protein level. Moreover, the PTP1B inhibitor potentiated mitophagy and associated expression of MFN2 and PINK1. Interestingly, inhibition of PTP1B resulted in potent attenuation of ER stress key mediators' expression namely, CHOP, ATF6, HSPA5 and XBP1. Conclusion: Presented findings shed for the first time promising new insights in the development of an MSI-1436-based therapy for proper equine metabolic syndrome intervention and may additionally find potential translational application to human metabolic syndrome treatment.

Sex hormone-binding globulin improves lipid metabolism and reduces inflammation in subcutaneous adipose tissue of metabolic syndrome-affected horses.

Equine metabolic syndrome (EMS) is a steadily growing endocrine disorder representing a real challenge in veterinary practice. As a multifactorial condition, EMS is characterized by three main metabolic abnormalities including insulin resistance, increased adiposity or obesity and hoof laminitis. Adipose tissue dysfunction is recognized as a core pathophysiological determinant of EMS, as it strongly participates to lipotoxicity and systemic metaflammation, both of which have been closely linked to the development of generalized insulin resistance. Besides, sex hormone binding globulin (SHBG) is an important sex steroids transporters that has been recently proposed as an important metabolic mediator. Therefore, the aim of this study was to verify whether SHBG treatment may ameliorate subcutaneous adipose tissue metabolic failure under EMS condition in terms of lipidome homeostasis, lipid metabolism programs, insulin signalling and local inflammation. Subcutaneous adipose tissue (SAT) biopsies were collected post-mortem from healthy (n = 3) and EMS (n = 3) slaughtered horses. SHBG protein has been applied to SAT samples from EMS horses for 24 h at a final concentration of 50 nM, while control groups (healthy and untreated EMS) were cultured in the presence of SHBG-vehicle only. Tissues from all groups were afterwards secured for downstream analysis of gene expression using RT-qPCR, protein levels by Western blot and ELISA assay and lipidomics through GC-MS technique. Obtained results showcased that SHBG intervention efficiently normalized the altered fatty acids (FAs) profiles by lowering the accumulation of saturated and trans FAs, as well as the pro-inflammatory arachidonic and linoleic acids. Moreover, SHBG showed promising value for the regulation of adipocyte lipolysis and engorgement by lowering the levels of perilipin-1. SHBG exerted moderated effect toward SCD1 and FASN enzymes expression, but increased the LPL abundance. Interestingly, SHBG exhibited a negative regulatory effect on pro-adipogenic stimulators and induced higher expression of KLF3, IRF3 and ß-catenin, known as strong adipogenesis repressors. Finally, SHBG protein showed remarkable ability in restoring the insulin signal transduction, IR/IRS/Pi3K/AKT phosphorylation events and GLUT4 transporter abundance, and further attenuate pro-inflammatory response by lowering IL-6 tissue levels and targeting the PDIA3/ERK axis. Overall, the obtained data clearly demonstrate the benefice of SHBG treatment in the regulation of adipose tissue metabolism in the course of EMS and provide new insights for the development of molecular therapies with potential translational application to human metabolic disorders.

Chemo-enzymatic synthesis and biological activity evaluation of propenylbenzene derivatives.

Propenylbenzenes, including isosafrole, anethole, isoeugenol, and their derivatives, are natural compounds found in essential oils from various plants. Compounds of this group are important and valuable, and are used in the flavour and fragrance industries as well as the pharmaceutical and cosmetic industries. The aim of this study was to develop an efficient process for synthesising oxygenated derivatives of these compounds and evaluate their potential biological activities. In this paper, we propose a two-step chemo-enzymatic method. The first step involves the synthesis of corresponding diols 1b-5b from propenylbenzenes 1a-5avia lipase catalysed epoxidation followed by epoxide hydrolysis. The second step involves the microbial oxidation of a diasteroisomeric mixture of diols 1b-5b to yield the corresponding hydroxy ketones 1c-4c, which in this study was performed on a preparative scale using Dietzia sp. DSM44016, Rhodococcus erythropolis DSM44534, R. erythropolis PCM2150, and Rhodococcus ruber PCM2166. Application of scaled-up processes allowed to obtain hydroxy ketones 1-4c with the following yield range 36-62.5%. The propenylbenzene derivatives thus obtained and the starting compounds were tested for various biological activities, including antimicrobial, antioxidant, haemolytic, and anticancer activities, and their impact on membrane fluidity. Fungistatic activity assay against selected strains of Candida albicans results in MIC50 value varied from 37 to 124 µg/mL for compounds 1a, 3a-c, 4a,b, and 5a,b. The highest antiradical activity was shown by propenylbenzenes 1-5a with a double bond in their structure with EC50 value ranged from 19 to 31 µg/mL. Haemolytic activity assay showed no cytotoxicity of the tested compounds on human RBCs whereas, compounds 2b-4b and 2c-4c affected the fluidity of the RBCs membrane. The tested compounds depending on their concentration showed different antiproliferative activity against HepG2, Caco-2, and MG63. The results indicate the potential utility of these compounds as fungistatics, antioxidants, and proliferation inhibitors of selected cell lines.

Postbiotics as potential new therapeutic agents for metabolic disorders management.

The prevalence of obesity, diabetes, non-alcoholic fatty liver disease, and related metabolic disorders has been steadily increasing in the past few decades. Apart from the establishment of caloric restrictions in combination with improved physical activity, there are no effective pharmacological treatments for most metabolic disorders. Many scientific-studies have described various beneficial effects of probiotics in regulating metabolism but others questioned their effectiveness and safety. Postbiotics are defined as preparation of inanimate microorganisms, and/or their components, which determine their safety of use and confers a health benefit to the host. Additionally, unlike probiotics postbiotics do not require stringent production/storage conditions. Recently, many lines of evidence demonstrated that postbiotics may be beneficial in metabolic disorders management via several potential effects including anti-inflammatory, antibacterial, immunomodulatory, anti-carcinogenic, antioxidant, antihypertensive, anti-proliferative, and hypocholesterolaemia properties that enhance both the immune system and intestinal barrier functions by acting directly on specific tissues of the intestinal epithelium, but also on various organs or tissues. In view of the many reports that demonstrated the high biological activity and safety of postbiotics, we summarized in the present review the current findings reporting the beneficial effects of various probiotics derivatives for the management of metabolic disorders and related alterations.

Astaxanthin Carotenoid Modulates Oxidative Stress in Adipose-Derived Stromal Cells Isolated from Equine Metabolic Syndrome Affected Horses by Targeting Mitochondrial Biogenesis.

Astaxanthin is gaining recognition as a natural bioactive component. This study aimed to test whether astaxanthin could protect adipose-derived stromal stem cells (ASCs) from apoptosis, mitochondrial dysfunction and oxidative stress. Phaffia rhodozyma was used to extract astaxanthin, whose biocompatibility was tested after 24, 48 and 72 h of incubation with the cells; no harmful impact was found. ASCs were treated with optimal concentrations of astaxanthin. Several parameters were examined: cell viability, apoptosis, reactive oxygen levels, mitochondrial dynamics and metabolism, superoxide dismutase activity, and astaxanthin's antioxidant capacity. A RT PCR analysis was performed after each test. The astaxanthin treatment significantly reduced apoptosis by modifying the normalized caspase activity of pro-apoptotic pathways (p21, p53, and Bax). Furthermore, by regulating the expression of related master factors SOD1, SOD2, PARKIN, PINK 1, and MFN 1, astaxanthin alleviated the oxidative stress and mitochondrial dynamics failure caused by EMS. Astaxanthin restored mitochondrial oxidative phosphorylation by stimulating markers associated with the OXPHOS machinery: COX4I1, COX4I2, UQCRC2, NDUFA9, and TFAM. Our results suggest that astaxanthin has the potential to open new possibilities for potential bio-drugs to control and suppress oxidative stress, thereby improving the overall metabolic status of equine ASCs suffering from metabolic syndrome.

Catestatin peptide of chromogranin A as a potential new target for several risk factors management in the course of metabolic syndrome.

Obesity, lipodystrophy, diabetes, and hypertension collectively constitute the main features of Metabolic Syndrome (MetS), together with insulin resistance (IR), which is considered as a defining element. MetS generally leads to the development of cardiovascular disease (CVD), which is a determinant cause of mortality and morbidity in humans and animals. Therefore, it is essential to implement and put in place adequate management strategies for the treatment of this disease. Catestatin is a bioactive peptide with 21 amino acids, which is derived through cleaving of the prohormone chromogranin A (CHGA/CgA) that is co-released with catecholamines from secretory vesicles and, which is responsible for hepatic/plasma lipids and insulin levels regulation, improves insulin sensitivity, reduces hypertension and attenuates obesity in murine models. In humans, there were few published studies, which showed that low levels of catestatin are significant risk factors for hypertension in adult patients. These accumulating evidence documents clearly that catestatin peptide (CST) is linked to inflammatory and metabolic syndrome diseases and can be a novel regulator of insulin and lipid levels, blood pressure, and cardiac function. The goal of this review is to provide an overview of the CST effects in metabolic syndrome given its role in metabolic regulation and thus, provide new insights into the use of CST as a diagnostic marker and therapeutic target.

Probiotics-rich emulsion improves insulin signalling in Palmitate/Oleate-challenged human hepatocarcinoma cells through the modulation of Fetuin-A/TLR4-JNK-NF-κB pathway.

BACKGROUND: Fetuin-A, also known as α2-Heremans-Schmid glycoprotein (AHSG), is an abundant plasmatic serum protein synthesized predominantly in liver and adipose tissue. This glycoprotein is known to negatively regulate insulin signaling through the inhibition of insulin receptor (IR) autophosphorylation and tyrosine kinase activity, which participates in insulin resistance (IR) and metabolic syndrome development. Recent studies demonstrated that IR and associated metabolic disorders, are closely related to the gut microbiota and modulating it by probiotics could be effective in metabolic diseases management. OBJECTIVE: In this present work we aimed to evaluate the effects of a probiotics-rich emulsion on reducing the IR induced by free fatty acids accumulation in human hepatocarcinoma cell line, and to elucidate the implicated molecular pathways, with a specific emphasis on the hepatokin Fetuin-A-related axis. RESULTS: Here we showed, that probiotics improve HepG2 viability, protect against apoptosis under normal and IR conditions. Moreover, the emulsion was successful in attenuating oxidative stress as well as improving mitochondrial metabolism and dynamics. Interestingly, application of the probiotics to lipotoxic HepG2 cells resulted in significant reduction of Fetuin-A/TLR4/JNK/NF-κB pathway activation, which suggests a protective effect against inflammation, obesity as well as liver related insulin resistant. CONCLUSION: Overall, the presented data reports clearly on the potent potential of probiotics formulated in an emulsion vehicle to enhance metabolic functions of affected IR HepG2 cells, and suggest the possibility of using such preparations as insulin sensitizing therapy, playing at the same time protective role for the development of liver related insulin resistant.