thapsigargin and Fibrosis

Fibrosis is a common final pathway of chronic kidney disease, which is a major incurable disease. Although fibrosis has an irreversible pathophysiology, the molecular and cellular mechanisms responsible remain unclear and no specific treatment is available to halt the progress of renal fibrosis. Thus, an improved understanding of the cellular mechanism involved and a novel therapeutic approach are urgently required for end-stage renal disease (ESRD). We investigated the role played by interleukin-10 (IL-10, a potent anti-inflammatory cytokine) in kidney fibrosis and the mechanisms involved using

Topics: Animals; Apoptosis; Brefeldin A; Disease Models, Animal; Endoplasmic Reticulum Stress; Fibrosis; Interleukin-10; Kidney; Kidney Diseases; Mice; Renal Insufficiency, Chronic; RNA, Small Interfering; Thapsigargin; Tunicamycin; Ureteral Obstruction

Cardiac fibrosis plays a vital role in the pathogenesis of heart failure. Fibroblast activity is enhanced by increases in store-operated Ca

Topics: Actins; Boron Compounds; Calcium; Cell Movement; Cell Proliferation; Cells, Cultured; Collagen; Fibroblasts; Fibrosis; Homeostasis; Humans; Lithium Chloride; Myocardium; ORAI1 Protein; Phosphorylation; RNA, Small Interfering; Thapsigargin

Restoration of kidney tubular epithelium following sublethal injury sequentially involves partial epithelial-mesenchymal transition (pEMT), proliferation, and further redifferentiation into specialized tubule epithelial cells (TECs). Because the immunosuppressant cyclosporine-A produces pEMT in TECs and inhibits the peptidyl-prolyl isomerase (PPIase) activity of cyclophilin (Cyp) proteins, we hypothesized that cyclophilins could regulate TEC phenotype. Here we demonstrate that in cultured TECs, CypA silencing triggers loss of epithelial features and enhances transforming growth factor β (TGFβ)-induced EMT in association with upregulation of epithelial repressors Slug and Snail. This pro-epithelial action of CypA relies on its PPIase activity. By contrast, CypB emerges as an epithelial repressor, because CypB silencing promotes epithelial differentiation, prevents TGFβ-induced EMT, and induces tubular structures in 3D cultures. In addition, in the kidneys of CypB knockout mice subjected to unilateral ureteral obstruction, inflammatory and pro-fibrotic events were attenuated. CypB silencing/knockout leads to Slug, but not Snail, downregulation. CypB support of Slug expression depends on its endoplasmic reticulum location, where it interacts with calreticulin, a calcium-buffering chaperone related to Slug expression. As CypB silencing reduces ionomycin-induced calcium release and Slug upregulation, we suggest that Slug expression may rely on CypB modulation of calreticulin-dependent calcium signaling. In conclusion, this work uncovers new roles for CypA and CypB in modulating TEC plasticity and identifies CypB as a druggable target potentially relevant in promoting kidney repair.

Topics: Animals; Basigin; Calcium; Cell Line; Cyclophilins; Endoplasmic Reticulum; Epithelial Cells; Fibrosis; Gene Silencing; Humans; Inflammation; Ionomycin; Kidney Tubules; Mice; Phenotype; Protein Transport; Smad Proteins; Snail Family Transcription Factors; Thapsigargin; Transforming Growth Factor beta; Ureteral Obstruction

We investigated the effect of saturated fatty acids on chronic pancreatitis pathogenesis by elucidating the endoplasmic reticulum (ER) stress response in pancreatic stellate cells (PSCs), which are major effector cells in pancreatic fibrosis.. Wistar Bonn/Kobori rats were fed either control diet or high-fat diet (HFD) for 4 weeks. Meanwhile, cultured rat PSCs were stimulated with thapsigargin, an ER stress inducer, or palmitic acid (PA). Pancreatic fibrosis, expressions of fibrosis-related and ER stress-related proteins and mRNA, cell viability, and apoptosis were examined.. The HFD reduced fibrosis and α-smooth muscle actin expression (ie, activated PSCs) but upregulated ER stress-related mRNA expression in the pancreas of young HFD-fed Wistar Bonn/Kobori rats. Induction of ER stress response in PSCs with thapsigargin or PA induced apoptosis, activated the protein kinase-like ER kinase (PERK) pathway, inhibited cell viability, and downregulated fibrosis-related protein and mRNA expression. The PERK inhibitor negated PA-induced ER stress response.. Saturated fatty acids can inhibit but may not promote the fibrogenesis of chronic pancreatitis, at least in the early stage, via an ER stress response (ie, the PERK pathway) in PSCs. Moreover, induction of an apoptotic ER stress response in PSCs might be a novel therapeutic strategy for pancreatic fibrosis.

Topics: Actins; Animals; Apoptosis; Cells, Cultured; Diet, High-Fat; Endoplasmic Reticulum Stress; Enzyme Inhibitors; Fatty Acids; Fibrosis; Gene Expression; Palmitic Acid; Pancreas; Pancreatic Stellate Cells; Rats, Wistar; Thapsigargin

Calcium transient triggered firing (CTTF) is induced by large intracellular calcium (Ca(i)) transient and short action potential duration (APD). We hypothesized that CTTF underlies the mechanisms of early afterdepolarization (EAD) and spontaneous recurrent atrial fibrillation (AF) in transgenic (Tx) mice with overexpression of transforming growth factor β1 (TGF-β1).. MHC-TGFcys(33)ser Tx mice develop atrial fibrosis because of elevated levels of TGF-β1. We studied membrane potential and Ca(i)transients of isolated superfused atria from Tx and wild-type (Wt) littermates. Short APD and persistently elevated Ca(i) transients promoted spontaneous repetitive EADs, triggered activity and spontaneous AF after cessation of burst pacing in Tx but not Wt atria (39% vs. 0%, P=0.008). We were able to map optically 4 episodes of spontaneous AF re-initiation. All first and second beats of spontaneous AF originated from the right atrium (4/4, 100%), which is more severely fibrotic than the left atrium. Ryanodine and thapsigargin inhibited spontaneous re-initiation of AF in all 7 Tx atria tested. Western blotting showed no significant changes of calsequestrin or sarco/endoplasmic reticulum Ca(2+)-ATPase 2a.. Spontaneous AF may occur in the Tx atrium because of CTTF, characterized by APD shortening, prolonged Ca(i) transient, EAD and triggered activity. Inhibition of Ca(2+) release from the sarcoplasmic reticulum suppressed spontaneous AF. Our results indicate that CTTF is an important arrhythmogenic mechanism in TGF-β1 Tx atria.

Topics: Action Potentials; Animals; Atrial Fibrillation; Atrial Function; Blotting, Western; Calcium Signaling; Cardiac Pacing, Artificial; Disease Models, Animal; Electrophysiologic Techniques, Cardiac; Enzyme Inhibitors; Fibrosis; Heart Atria; Heart Conduction System; Mice; Mice, Transgenic; Ryanodine; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Thapsigargin; Time Factors; Transforming Growth Factor beta1; Up-Regulation

Survival of hepatic stellate cells (HSCs) is a hallmark of liver fibrosis, while the induction of HSC apoptosis may induce recovery. Activated HSC are resistant to many pro-apoptotic stimuli. To this issue, the role of Endoplasmic Reticulum (ER) stress in promoting apoptosis of HSCs and consequently fibrosis resolution is still debated.. To evaluate the potential ER stress-mediated apoptosis of HSCs and fibrosis resolution. HSCs were incubated with the ER stress agonists, tunicamycin or thapsigargin. In vivo, HSC were isolated from normal, bile duct-ligated (BDL) and bile duct-diverted (BDD) rats.. In activated HSC, the specific inhibitor of ER stress-induced apoptosis, calpastatin, is significantly increased vs. quiescent HSCs. Calpain is conversely reduced in activated HSCs. This pattern of protein expression provides HSCs resistance to the ER stress signals of apoptosis (apoptosis-resistant phenotype). However, both tunicamycin and thapsigargin are able to induce apoptosis in HSCs in vitro, completely reversing the calpain/calpastatin pattern expression. Furthermore, in vivo, the fibrosis resolution observed in rat livers subjected to bile duct ligation (BDL) and subsequent bile duct diversion (BDD), leads to fibrosis resolution through a mechanism of HSCs apoptosis, potentially associated with ER stress: in fact, BDD rat liver shows an increased number of apoptotic HSCs associated with reduced calapstatin and increased calpain protein expression, leading to an apoptosis-sensible phenotype.. ER stress sensitizes HSC to apoptosis both in vitro and in vivo. Thus, ER stress represents a key target to trigger cell death in activated HSC and promotes fibrosis resolution.

Topics: Animals; Apoptosis; Bile Ducts; Blotting, Western; Calcium-Binding Proteins; Calpain; Caspase 8; Endoplasmic Reticulum Stress; Fibrosis; Hepatic Stellate Cells; Immunohistochemistry; In Situ Nick-End Labeling; Ligation; Liver; Rats; RNA, Small Interfering; Thapsigargin; Tunicamycin