Pirfenidone mitigates TGF-ß1-mediated fibrosis in an idiopathic inflammatory myositis-associated interstitial lung disease model.

Idiopathic inflammatory myositis (IIM) is a group of rare diseases of unknown etiology, with a pathognomonic muscular deficiency. Antisynthetase syndrome is a subtype of IIM with an associated interstitial lung disease (ILD), characterized by pulmonary inflammation and fibrosis mediated by TGF-ß. Pirfenidone is a new molecule with anti-inflammatory and anti-fibrotic properties, used for the treatment of idiopathic ILD, but has never been assessed in IIM. The aim of our study is to evaluate the effect of pirfenidone on IIM-associated ILD. Thirty-two BALB/c male mice were divided into three groups: Sham, IIM-untreated (IIM), and IIM pirfenidone-treated (IIM + PIR). IIM was induced by intramuscular injections of guinea pig muscle myosin extract and intraperitoneal injections of Pertussis toxin. Pirfenidone was given orally at a dose of 30 mg kg-1 day-1 for two months. Muscle force, blood and bronchoalveolar lavage fluid samples, as well as muscle and lung tissues, were analyzed. Progressive deterioration of muscle force and infiltration of the muscular tissue by inflammatory cells were observed with IIM. Auto-immune antibodies specific to the antisynthetase syndrome were also increased in IIM mice. Pirfenidone attenuated IIM-associated ILD with anti-inflammatory properties evidenced by decreased peribronchial inflammation and TGF-ß1 in bronchoalveolar lavage fluid. Likewise, pirfenidone attenuated pulmonary fibrosis by fine-tuning TGF-ß1-mediated epithelial-to-mesenchymal and fibrotic signaling pathways; pro-fibrotic SMAD3, ZEB2 and STAT1 expression and activation were decreased, whereas anti-fibrotic SMAD2 activation was increased. This study unravels for the first time that pirfenidone has the potential to fine-tune TGF-ß1 fibrotic signaling in IIM-associated ILD.

New insights in gut-liver axis in wild-type murine imiquimod-induced lupus.

BACKGROUND: Intestinal and hepatic manifestations of lupus seem to be underestimated in comparison to other major organ lesions. Although recent data point to gut-liver axis involvement in lupus, gut permeability dysfunction and liver inflammation need to be more investigated. OBJECTIVE: This study aims to assess fecal calprotectin, intestinal tight junction proteins and liver inflammation pathway in wild-type murine imiquimod- induced lupus. METHODS: C57BL/6 mice were topically treated on their right ears with 1.25 mg of 5% imiquimod cream, three times per week for six weeks. Fecal calprotectin was collected at day 0, 22 and 45. Renal, liver and intestinal pathology, as well as inflammatory markers, intestinal tight junction proteins, and E. coli protein in liver were assessed at sacrifice. RESULTS: At six weeks, lupus nephritis was confirmed on histopathology and NGAL and KIM-1 expression. Calprotectin rise started at day 22 and persists at day 45. Protein expression of Claudine, ZO-1 and occludin was significantly decreased. E. coli protein was significantly increased in liver with necro-inflammation and increased TLR4, TLR7, and pNFκB/NFκB liver expression. CONCLUSION: This study is the first to demonstrate early fecal calprotectin increase and liver activation of TLR4- NFκB pathway in wild-type murine imiquimod-induced lupus.

VEGF-C attenuates renal damage in salt-sensitive hypertension.

Salt-sensitive hypertension is a major risk factor for renal impairment leading to chronic kidney disease. High-salt diet leads to hypertonic skin interstitial volume retention enhancing the activation of the tonicity-responsive enhancer-binding protein (TonEBP) within macrophages leading to vascular endothelial growth factor C (VEGF-C) secretion and NOS3 modulation. This promotes skin lymphangiogenesis and blood pressure regulation. Whether VEGF-C administration enhances renal and skin lymphangiogenesis and attenuates renal damage in salt-sensitive hypertension remains to be elucidated. Hypertension was induced in BALB/c mice by a high-salt diet. VEGF-C was administered subcutaneously to high-salt-treated mice as well as control animals. Analyses of kidney injury, inflammation, fibrosis, and biochemical markers were performed in vivo. VEGF-C reduced plasma inflammatory markers in salt-treated mice. In addition, VEGF-C exhibited a renal anti-inflammatory effect with the induction of macrophage M2 phenotype, followed by reductions in interstitial fibrosis. Antioxidant enzymes within the kidney as well as urinary RNA/DNA damage markers were all revelatory of abolished oxidative stress under VEGF-C. Furthermore, VEGF-C decreased the urinary albumin/creatinine ratio and blood pressure as well as glomerular and tubular damages. These improvements were associated with enhanced TonEBP, NOS3, and lymphangiogenesis within the kidney and skin. Our data show that VEGF-C administration plays a major role in preserving renal histology and reducing blood pressure. VEGF-C might constitute an interesting potential therapeutic target for improving renal remodeling in salt-sensitive hypertension.

Circadian Rhythm Disruption Aggravates DSS-Induced Colitis in Mice with Fecal Calprotectin as a Marker of Colitis Severity.

BACKGROUND: Inflammatory bowel disease (IBD) is a chronic immunologically mediated pathology that remains a major health burden. Circadian rhythm disruption leads to a deregulation in the immune system which is a major risk factor for IBD. AIMS: Since fecal calprotectin (FC) has been a useful tool for monitoring IBD, we aimed to evaluate the effect of circadian rhythm alteration on gut inflammation status and whether FC is associated with the severity of colitis. METHODS: C57BL/6J mice were exposed to circadian shifts for 3 months, and then colitis was induced by 2% dextran sulfate sodium (DSS). Colitis was evaluated according to clinical symptoms and histological scoring. Plasma and intestinal inflammatory and permeability markers as well as fecal and intestinal calprotectin were assessed. RESULTS: Circadian shifts aggravated DSS-induced colitis with increased diarrhea, flatulence, and fecal blood associated with decreased colon length. In addition, intestinal cryptic architecture was lost with the presence of increased inflammation, mucosal muscle thickening, and cryptic abscesses. Plasma tumor necrosis factor alpha, interleukin 1 beta, interleukin 6, and C-reactive protein upregulations were paralleled by the deterioration of intestinal permeability. Calprotectin expression and distribution increased in the intestines and feces of shifted animals, and levels highly correlated with the increases in intestinal inflammation and permeability. CONCLUSIONS: Circadian rhythm disruption aggravates DSS-induced colitis, whereas fecal and intestinal calprotectin associates with the severity of disease. Calprotectin might be a useful marker and tool for assessing patients at risk of IBD due to lifestyles with disruptive sleep patterns.

Basic Signaling in Cardiac Fibroblasts.

Cardiac fibroblasts are commonly known as supporting cells of the cardiac network and exert many essential functions that are fundamental for normal cardiac growth as well as for cardiac remodeling process during pathological conditions. This review focuses on the roles of cardiac fibroblasts in the formation and regulation of the extracellular matrix components, and in maintaining structural, biochemical and mechanical properties of the heart. Additionally, though considered as non-excitable cells, we review the functional expression in cardiac fibroblasts of a wide variety of transmembrane ion channels which activity may contribute to key regulation of cardiac physiological processes. All together, cardiac fibroblasts which actively participate to fundamental regulation of cardiac physiology and physiopathology processes may represent pertinent targets for pharmacological approaches of cardiac diseases and lead to new tracks of therapeutic strategies. J. Cell. Physiol. 232: 725-730, 2017. © 2016 Wiley Periodicals, Inc.

An optimized protocol for purification of functional islets of Langerhans.

Islets of Langerhans and ß-cell isolation constitute routinely used cell models for diabetic research, and refining islet isolation protocols and cell quality assessment is a high priority. Numerous protocols have been published describing isolate of islets, but often rigorous and systematic assessment of their integrity is lacking. Herein, we propose a new protocol for optimal generation of islets. Pancreases from mice and rats were excised and digested using a low-activity collagenase solution and islets were then purified by a series of sedimentations and a Percoll gradient. Islets were maintained in culture for 5 days, during which viability, pro/antiapoptotic, and islet-specific genes, glucose-stimulated calcium entry, glucose uptake, and insulin secretion were assessed. The commonly used islet isolation technique by collagenase injection through the common bile duct (CBD) was also performed and compared with the present approach. This new protocol produced islets that retained a healthy status as demonstrated by the yield of stable living cells. Furthermore, calcium oscillation, glucose uptake, and insulin secretion remained intact in the islet cultures. This was reproducible when many rodent species were used, and neither sex nor age affected the cells behavior. When compared with the CBD technique, islet physiology was similar. Finally, this approach was used to uncover new ion channel candidates implicated in insulin secretion. In conclusion, this study outlines an efficient protocol for islet preparation that may support research into new therapeutic targets in diabetes research.

Fish oil attenuates neurologic severity of antiphospholipid syndrome in a mice experimental model.

INTRODUCTION: Murine experimental models of antiphospholipid syndrome (eAPLS) showed neurologic dysfunction and therapeutic effect of the anticoagulant enoxaparin is well established. Omega-3 fatty acids and curcumin, tested in neuroinflammation and auto-immunity diseases, might be interesting therapeutic candidates. The aim of this study was to evaluate the effects of these candidates on neurologic severity in eAPLS. METHODS: One month after immunization of BALB/c mice with beta-2-glycoprotein I, daily treatments were initiated with enoxaparin (1 mg/kg), omega-3 fatty acids (0.5 g/kg), and curcumin (200 mg/kg) for 3 months. RESULTS: Mortality was significantly decreased by enoxaparin and omega-3 treatments. Fish oil and curcumin group exhibited the highest mean of swimming behavior in forced swim test in surviving mice. Mice under omega-3 fatty acids or curcumin presented low anxiety-like behavior in the elevated plus-maze test. Cerebral histopathology revealed heavy inflammatory infiltrates in cortical and subcortical regions with vacuolization, swelling, and degeneration of astrocytes in the control group, with aggravation under curcumin; no infiltrate was retrieved in enoxaparin and omega-3 groups. CONCLUSION: Our study is the first to demonstrate a potential therapeutic effect of omega-3 fatty acids in eAPLS. Enoxaparin and omega-3 fatty acids combination would be interesting for further investigation.

Evidence of a Role for Fibroblast Transient Receptor Potential Canonical 3 Ca2+ Channel in Renal Fibrosis.

Transient receptor potential canonical (TRPC) Ca(2+)-permeant channels, especially TRPC3, are increasingly implicated in cardiorenal diseases. We studied the possible role of fibroblast TRPC3 in the development of renal fibrosis. In vitro, a macromolecular complex formed by TRPC1/TRPC3/TRPC6 existed in isolated cultured rat renal fibroblasts. However, specific blockade of TRPC3 with the pharmacologic inhibitor pyr3 was sufficient to inhibit both angiotensin II- and 1-oleoyl-2-acetyl-sn-glycerol-induced Ca(2+) entry in these cells, which was detected by fura-2 Ca(2+) imaging. TRPC3 blockade or Ca(2+) removal inhibited fibroblast proliferation and myofibroblast differentiation by suppressing the phosphorylation of extracellular signal-regulated kinase (ERK1/2). In addition, pyr3 inhibited fibrosis and inflammation-associated markers in a noncytotoxic manner. Furthermore, TRPC3 knockdown by siRNA confirmed these pharmacologic findings. In adult male Wistar rats or wild-type mice subjected to unilateral ureteral obstruction, TRPC3 expression increased in the fibroblasts of obstructed kidneys and was associated with increased Ca(2+) entry, ERK1/2 phosphorylation, and fibroblast proliferation. Both TRPC3 blockade in rats and TRPC3 knockout in mice inhibited ERK1/2 phosphorylation and fibroblast activation as well as myofibroblast differentiation and extracellular matrix remodeling in obstructed kidneys, thus ameliorating tubulointerstitial damage and renal fibrosis. In conclusion, TRPC3 channels are present in renal fibroblasts and control fibroblast proliferation, differentiation, and activation through Ca(2+)-mediated ERK signaling. TRPC3 channels might constitute important therapeutic targets for improving renal remodeling in kidney disease.

ANO1 contributes to angiotensin-II-activated Ca2+-dependent Cl- current in human atrial fibroblasts.

Cardiac fibroblasts are an integral part of the myocardial tissue and contribute to its remodelling. This study characterises for the first time the calcium-dependent chloride channels (CaCC) in the plasma membrane of primary human atrial cardiac fibroblasts by means of the iodide efflux and the patch clamp methods. The calcium ionophore A23187 and Angiotensin II (Ang II) activate a chloride conductance in cardiac fibroblasts that shares pharmacological similarities with calcium-dependent chloride channels. This chloride conductance is depressed by RNAi-mediated selective Anoctamine 1 (ANO1) but not by Anoctamine 2 (ANO2) which has been revealed as CaCC and is inhibited by the selective ANO1 inhibitor, T16inh-A01. The effect of Ang II on anion efflux is mediated through AT1 receptors (with an EC50 = 13.8 ± 1.3 nM). The decrease of anion efflux by calphostin C and bisindolylmaleimide I (BIM I) suggests that chloride conductance activation is dependent on PKC. We conclude that ANO1 contributes to CaCC current in human cardiac fibroblasts and that this is regulated by Ang II acting via the AT1 receptor pathway.

Balancing the mind: Toward a complete picture of the interplay between gut microbiota, inflammation and major depressive disorder.

The intricate interplay existing between gut microbiota and homeostasis extends to the realm of the brain, where emerging research underscores the significant impact of the microbiota on mood regulation and overall neurological well-being and vice-versa, with inflammation playing a pivotal role in mediating these complex interactions. This comprehensive review explores the complex interplay between inflammation, alterations in gut microbiota, and their impact on major depressive disorder (MDD). It provides a cohesive framework for the puzzle pieces of this triad, emphasizing recent advancements in understanding the gut microbiota and inflammatory states' contribution to the depressive features. Two directions of communication between the gut and the brain in depression are discussed, with inflammation serving as a potential modulator. Therapeutic implications were discussed as well, drawing insights from interventional studies on the effects of probiotics on gut bacterial composition and depressive symptoms. Ultimately, this review will attempt to provide a complete and valuable framework for future research and therapeutic interventions in MDD.

Antioxidant and Anti-Apoptotic Neuroprotective Effects of Cinnamon in Imiquimod-Induced Lupus.

BACKGROUND: Despite accumulating evidence correlating oxidative stress with lupus disease activity, the brain redox pathways are still poorly investigated. Cinnamomum cassia, a widely used spice with powerful antioxidant properties, could be a novel therapeutic candidate in lupus. METHODS: C57BL/6J female mice were divided into five groups: sham, sham-cinnamon, lupus, lupus-cinnamon starting from induction, and lupus-cinnamon starting two weeks before induction. Lupus was induced by skin application on the right ear with 1.25 mg of 5% imiquimod cream three times per week for six weeks. Cinnamomum cassia was given orally, five days per week, at 200 mg/kg. RESULTS: Concomitant to TLR7-MYD88 pathway activation, the p-NRF2/NRF2 and p-FOXO3/FOXO3 ratios were increased in the hippocampus and alleviated by cinnamon treatment. BCL-2 positivity was enhanced in hippocampal neurons and reversed only by preventive cinnamon administration. In vitro, exposure of hippocampal cells to the plasma of different groups induced a surge in oxidative stress. This was associated with an increased t-BID/BID ratio. Cinnamon treatment, particularly in the preventive arm, normalized these modifications. CONCLUSIONS: Our study shows a neuroprotective effect of cinnamon by rescuing brain redox and apoptosis homeostasis in lupus, paving the way for its use as a natural therapeutic compound in the clinical management of lupus.

Circadian Rhythm Perturbation Aggravates Gut Microbiota Dysbiosis in Dextran Sulfate Sodium-Induced Colitis in Mice.

Circadian rhythm disruption is increasingly considered an environmental risk factor for the development and exacerbation of inflammatory bowel disease. We have reported in a previous study that nychthemeral dysregulation is associated with an increase in intestinal barrier permeability and inflammation in mice with dextran sulfate sodium (DSS)-induced colitis. To investigate the effect of circadian rhythm disruption on the composition and diversity of the gut microbiota (GM), sixty male C57BL/6J mice were initially divided to two groups, with the shifted group (n = 30) exposed to circadian shifts for three months and the non-shifted group (n = 30) kept under a normal light-dark cycle. The mice of the shifted group were cyclically housed for five days under the normal 12:12 h light-dark cycle, followed by another five days under a reversed light-dark cycle. At the end of the three months, a colitis was induced by 2% DSS given in the drinking water of 30 mice. Animals were then divided into four groups (n = 15 per group): sham group non-shifted (Sham-NS), sham group shifted (Sham-S), DSS non-shifted (DSS-NS) and DSS shifted (DSS-S). Fecal samples were collected from rectal content to investigate changes in GM composition via DNA extraction, followed by high-throughput sequencing of the bacterial 16S rRNA gene. The mouse GM was dominated by three phyla: Firmicutes, Bacteroidetes and Actinobacteria. The Firmicutes/Bacteroidetes ratio decreased in mice with induced colitis. The richness and diversity of the GM were reduced in the colitis group, especially in the group with inverted circadian rhythm. Moreover, the GM composition was modified in the inverted circadian rhythm group, with an increase in Alloprevotella, Turicibacter, Bacteroides and Streptococcus genera. Circadian rhythm inversion exacerbates GM dysbiosis to a less rich and diversified extent in a DSS-induced colitis model. These findings show possible interplay between circadian rhythm disruption, GM dynamics and colitis pathogenesis.

TRPC3 Regulates Islet Beta-Cell Insulin Secretion.

Insulin release is tightly controlled by glucose-stimulated calcium (GSCa) through hitherto equivocal pathways. This study investigates TRPC3, a non-selective cation channel, as a critical regulator of insulin secretion and glucose control. TRPC3's involvement in glucose-stimulated insulin secretion (GSIS) is studied in human and animal islets. TRPC3-dependent in vivo insulin secretion is investigated using pharmacological tools and Trpc3-/- mice. TRPC3's involvement in islet glucose uptake and GSCa is explored using fluorescent glucose analogue 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose and calcium imaging. TRPC3 modulation by a small-molecule activator, GSK1702934A, is evaluated in type 2 diabetic mice. TRPC3 is functionally expressed in human and mouse islet beta cells. TRPC3-controlled insulin secretion is KATP -independent and primarily mediated by diacylglycerol channel regulation of the cytosolic calcium oscillations following glucose stimulation. Conversely, glucose uptake in islets is independent of TRPC3. TRPC3 pharmacologic inhibition and knockout in mice lead to defective insulin secretion and glucose intolerance. Subsequently, TRPC3 activation through targeted small-molecule enhances insulin secretion and alleviates diabetes hallmarks in animals. This study imputes a function for TRPC3 at the onset of GSIS. These insights strengthen one's knowledge of insulin secretion physiology and set forth the TRPC3 channel as an appealing candidate for drug development in the treatment of diabetes.

A new method of ultrasonic nonviral gene delivery to the adult myocardium.

Cardiac gene transfer is a powerful molecular tool to improve our understanding of the role of new proteins and mutants in cardiac pathophysiology. There is a need for a simple efficient myocardial gene delivery technique in order to study the physiological role of proteins in their native environment. Here we tested a new method of myocardial nonviral gene delivery, by using the combination of ultrasound energy (USE), liposomes and high pressure injections to the rat heart. Wistar rats were subjected to intra-myocardial injections of liposomes-DNA or siRNA mix. The heart was exposed after an inter-costal incision, and then injections were conducted between two sets of USE heart exposure. Ultrasound application resulted in much higher transfection efficiency (2% of left ventricle) than the liposomes-DNA alone (0.12% of left ventricle) as shown by the beta-galactosidase staining. The ultrasonic based liposomes-DNA delivery resulted in low inflammatory response, as well as in low cardiac fibrosis as shown by total collagen staining. Quantitative real time polymerase chain reaction (PCR) showed that the ultrasonic delivery resulted in cardiac specific transduction. Moreover, 23,906±2197 and 71,883±4065 calcium tolerant transfected cardiac myocytes were isolated following the delivery of a GFP plasmid or tagged siRNA, respectively. This was sufficient to perform single cell physiological measurements and biochemical experiments on homogenates. We developed an interesting safe method for local gene transfer in the heart using ultrasound and liposomes gene delivery. This method is particularly useful to study the effect of gene transfer on cardiac myocytes maintained in their normal environment in animal models.

Calcitriol Supplementation Protects Against Apoptosis and Alleviates the Severity of Abdominal Aortic Aneurysm Induced by Angiotensin II and Anti-TGFß.

The present study aims to assess the effect of vitamin D deficiency (VDD) and its supplementation on the severity of AAA in mice. AAA was induced by AngII and anti-TGF-ß administration. Animals were divided into four groups: Sham, mice with AAA, mice with AAA, and VDD, and mice with AAA supplemented with calcitriol. Blood pressure, echocardiography, abdominal aortic tissues, and plasma samples were monitored for all groups. VDD was associated with enhanced activity of cleaved MMP-9 and elastin degradation and positively correlated with the severity of AAA. Calcitriol supplementation decreased the INFγ/IL-10 ratio and enhanced the Nrf2 pathway. Moreover, Cu/Zn-superoxide dismutase expression and catalase and neutral sphingomyelinase activity were exacerbated in AAA and VDD groups. Furthermore, calcitriol supplementation showed a significantly lower protein expression of caspase-8, caspase-3, Bid, and t-Bid, and prevented the apoptosis of VSMCs treated by AngII and anti-TGF-ß. Calcitriol supplementation may alleviate AAA severity and could be of great interest in the clinical management of AAA. VDD enhances antioxidant enzymes activity and expression, whereas calcitriol supplementation alleviates AAA severity by re-activating Nrf2 and inhibiting apoptotic pathways.

Electrophysiological and functional effects of sphingosine-1-phosphate in mouse ventricular fibroblasts.

The aim of this study was to characterize the effects of sphingosine-1-phosphate (S1P) on cardiac ventricular fibroblasts. Impacts of S1P on fibroblast excitability, cell migration, proliferation and secretion were characterized. The patch-clamp technique in the whole-cell configuration was used to study the S1P-induced current from mouse ventricular fibroblasts. The expression level of the S1P receptor during cell culture duration was evaluated by western-blot. Fibroblast proliferation and migration were quantified using the methylene blue assay and the Boyden chamber technique, respectively. Finally, fibroblast secretion properties were estimated by quantification of the IL-6 and collagen levels using ELISA and SIRCOL collagen assays, respectively. We found that S1P activated SUR2/Kir6.1 channel and that this effect was sensitive to specific inhibition of the S1P receptor of type 3 (S1P3R). In contrast, S1P1R receptor inhibition had no effect. Moreover, the S1P-induced current increased with cell culture duration whereas S1P3R expression level remained constant. The activation of SUR2/Kir6.1 channel by S1P via S1P3R stimulated cell proliferation and decreased IL-6 and collagen secretions. S1P also stimulated fibroblast migration via S1P3R but independently from SUR2/Kir6.1 channel activation. This study demonstrates that S1P, via S1P3R, affects cardiac ventricular fibroblasts function independently or through activation of SUR2/Kir6.1 channel. The latter effect occurs after fibroblasts differentiate into myofibroblasts, opening a new potential therapeutic strategy to modulate fibrosis after cardiac physiopathological injury.

Pargyline reduces renal damage associated with ischaemia-reperfusion and cyclosporin.

BACKGROUND: The slow deterioration of the kidney graft is characterized histologically by interstitial fibrosis and tubular atrophy (IFTA). Immunological and non-immunological stress is the main cause of progression towards IFTA. Our study focused on the non-immunological injuries induced by ischaemia-reperfusion (IR) and cyclosporin (CsA) toxicity, which remain the two stress factors putting a damper on the outcome of the renal graft. Endogenous reactive oxygen species (ROS) are essentially produced by mitochondria, and we have previously shown that the blockage of the mitochondrial enzymes monoamine oxidases (MAOs) prevents H2O2 production in the early reperfusion stage following IR. METHODS: We used a rat model of IFTA consisting in unilateral nephrectomy followed by IR and daily CsA administration. Four weeks after IR, we analysed renal function, histological alterations and a number of inflammatory and fibrotic genes. RESULTS: We observed, 28 days after pargyline-mediated blockade of MAO (before or after IR), improved renal function as well as a net decrease in renal inflammation associated to lower IL-1ß and TNF-α gene expression. However, significant reduction in apoptosis, necrosis and fibrosis was only observed when pargyline was administrated before IR. This protective effect was associated to decreased expression of TGF-ß1, collagen types I, III and IV and also to the normalization of antioxidant (SOD1, catalase) and inflammatory (COX2, LOX5) gene expression. CONCLUSION: It appears that the blockage of ROS produced by MAO and subsequent cell death might be an effective protective strategy against IFTA progression.

Ranolazine: An Old Drug with Emerging Potential; Lessons from Pre-Clinical and Clinical Investigations for Possible Repositioning.

Ischemic heart disease is a significant public health problem with high mortality and morbidity. Extensive scientific investigations from basic sciences to clinics revealed multilevel alterations from metabolic imbalance, altered electrophysiology, and defective Ca2+/Na+ homeostasis leading to lethal arrhythmias. Despite the recent identification of numerous molecular targets with potential therapeutic interest, a pragmatic observation on the current pharmacological R&D output confirms the lack of new therapeutic offers to patients. By contrast, from recent trials, molecules initially developed for other fields of application have shown cardiovascular benefits, as illustrated with some anti-diabetic agents, regardless of the presence or absence of diabetes, emphasizing the clear advantage of "old" drug repositioning. Ranolazine is approved as an antianginal agent and has a favorable overall safety profile. This drug, developed initially as a metabolic modulator, was also identified as an inhibitor of the cardiac late Na+ current, although it also blocks other ionic currents, including the hERG/Ikr K+ current. The latter actions have been involved in this drug's antiarrhythmic effects, both on supraventricular and ventricular arrhythmias (VA). However, despite initial enthusiasm and promising development in the cardiovascular field, ranolazine is only authorized as a second-line treatment in patients with chronic angina pectoris, notwithstanding its antiarrhythmic properties. A plausible reason for this is the apparent difficulty in linking the clinical benefits to the multiple molecular actions of this drug. Here, we review ranolazine's experimental and clinical knowledge on cardiac metabolism and arrhythmias. We also highlight advances in understanding novel effects on neurons, the vascular system, skeletal muscles, blood sugar control, and cancer, which may open the way to reposition this "old" drug alone or in combination with other medications.

Reno-Protective Effect of GLP-1 Receptor Agonists in Type1 Diabetes: Dual Action on TRPC6 and NADPH Oxidases.

Diabetic kidney disease (DKD), a serious diabetic complication, results in podocyte loss and proteinuria through NADPH oxidases (NOX)-mediated ROS production. DUOX1 and 2 are NOX enzymes that require calcium for their activation which enters renal cells through the pivotal TRPC channels. Hypoglycemic drugs such as liraglutide can interfere with this deleterious mechanism imparting reno-protection. Herein, we aim to investigate the reno-protective effect of GLP1 receptor agonist (GLP1-RA), via its effect on TRPC6 and NADPH oxidases. To achieve our aim, control or STZ-induced T1DM Sprague-Dawley rats were used. Rats were treated with liraglutide, metformin, or their combination. Functional, histological, and molecular parameters of the kidneys were assessed. Our results show that treatment with liraglutide, metformin or their combination ameliorates DKD by rectifying renal function tests and protecting against fibrosis paralleled by restored mRNA levels of nephrin, DUOX1 and 2, and reduced ROS production. Treatment with liraglutide reduces TRPC6 expression, while metformin treatment shows no effect. Furthermore, TRPC6 was found to be directly interacting with nephrin, and indirectly interacting with DUOX1, DUOX2 and GLP1-R. Our findings suggest that treatment with liraglutide may prevent the progression of diabetic nephropathy by modulating the crosstalk between TRPC6 and NADPH oxidases.

Cardiac Phosphodiesterases Are Differentially Increased in Diabetic Cardiomyopathy.

AIM: Diabetic cardiomyopathy (DCM) accomodates a spectrum of cardiac abnormalities. This study aims to investigate whether DCM is associated with changes in cyclic adenosine 3'-5' monophosphate (cAMP) signaling, particularly cyclic nucleotide phosphodiesterases (PDEs). MAIN METHODS: Type 1 diabetes (T1D) was induced in rats by streptozotocin (STZ, 65 mg/kg) injection. Myocardial remodeling, structure and function were evaluated by histology and echocardiography, respectively. We delineated the sequential changes affecting cAMP signaling and characterized the expression pattern of the predominant cardiac PDE isoforms (PDE 1-5) and ß-adrenergic (ß-AR) receptors at 4, 8 and 12 weeks following diabetes induction, by real-time quantitative PCR and Western blot. cAMP levels were measured by immunoassays. KEY FINDINGS: T1D-induced DCM was associated with cardiac remodeling, steatosis and fibrosis. Upregulation of ß1-AR receptor transcripts was noted in diabetic hearts at 4 weeks along with an increase in cAMP levels and an upregulation in the ejection fraction and fraction shortening. However, ß2-AR receptors expression remained unchanged regardless of the disease stage. Moreover, we noted an early and specific upregulation of cardiac PDE1A, PDE2A, PDE4B, PDE4D and PDE5A expression at week 4, followed by increases in PDE3A levels in diabetic hearts at week 8. However, DCM was not associated with changes in PDE4A gene expression irrespective of the disease stage. SIGNIFICANCE: We show for the first time differential and time-specific regulations in cardiac PDEs, data that may prove useful in proposing new therapeutic approaches in T1D-induced DCM.