Loss of Phosphatase and Tensin Homologue Increases Transforming Growth Factor β–Mediated Invasion with Enhanced SMAD3 Transcriptional Activity

113

Background: MicroRNA-21 is important in hepatic fibrosis development, but the mechanism is unclear. Results: MicroRNA-21 is predominantly up-regulated in activated hepatic stellate cells and could form a double negative feedback loop that links fibrogenic machinery. Conclusion:The microRNA-21-mediated loop is a main driving force for hepatic fibrosis progression. Significance: It suggests a mechanism for how microRNA-21 contributes to hepatic fibrosis.Sustained activation of hepatic stellate cells (HSCs) leads to hepatic fibrosis, which is characterized by excessive collagen production, and for which there is no available drug clinically. Despite tremendous progress, the cellular activities underlying HSC activation, especially the driving force in the perpetuation stage, are only partially understood. Recently, microRNA-21 (miR-21) has been found to be prevalently up-regulated during fibrogenesis in different tissues, although its detailed role needs to be further elucidated. In the present study, miR-21 expression was examined in human cirrhotic liver samples and in murine fibrotic livers induced by thioacetamide or carbon tetrachloride. A dramatic miR-21 increase was noted in activated HSCs. We further found that miR-21 maintained itself at constant high levels by using a microRNA-21/programmed cell death protein 4/activation protein-1 (miR-21/PDCD4/AP-1) feedback loop. Disrupting this loop with miR-21 antagomir or AP-1 inhibitors significantly suppressed fibrogenic activities in HSCs and ameliorated liver fibrosis. In contrast, reinforcing this loop with small interfering RNA (siRNA) against PDCD4 promoted fibrogenesis in HSCs. Further analysis indicated that the up-regulated miR-21 promoted the central transforming growth factor-␤ (TGF-␤) signaling pathway underlying HSC activation. In summary, we suggest that the miR-21/PDCD4/AP-1 autoregulatory loop is one of the main driving forces for hepatic fibrosis progression. Targeting this aberrantly activated feedback loop may provide a new therapeutic strategy and facilitate drug discovery against hepatic fibrosis.Hepatic fibrosis is a wound healing process in response to chronic liver injuries that leads to unbalanced extracellular matrix (ECM) 3 deposition and resolution. The persistent activation of wound healing responses causes quantitative and qualitative changes in the ECM components and could finally distort liver parenchyma and vascular architecture, which could impair liver function and potentially lead to liver failure and hepatocellular carcinoma. Following liver injury, quiescent hepatic stellate cells (HSCs) transdifferentiate into myofibroblast-like cells that are characterized by the expression of smooth muscle ␣-actin (␣-SMA) and enhanced production of ECM (1). Despite the tremendous progress in understanding the mechanisms during fibrogenesis, the driving force underlying the persistent fibrogenic activities is still only partially understood.Extensive studies have suggested the central role of feedback networks in dictating disease progressi...

Section: Discussionsupporting

confidence: 61%

The Autoregulatory Feedback Loop of MicroRNA-21/Programmed Cell Death Protein 4/Activation Protein-1 (MiR-21/PDCD4/AP-1) as a Driving Force for Hepatic Fibrosis Development

Zhang

Zha

et al. 2013

113

“…Consistent with the previous report, we demonstrate that PTEN forms cytoplasmic complexes with Smad2/3. In contrast to HaCaT cells, in which TGF-␤ stimulation promoted formation of such complexes (33), in fibroblasts, TGF-␤ has an opposite effect by dissociating existing complexes. PTEN did not interfere with nuclear translocation of Smad2/3 in either fibroblasts or HaCaT cells, indicating that the inhibition of TGF-␤ signaling by PTEN does not occur through sequestration of Smad2/3 in the cytoplasm.…”

Section: Discussionmentioning

confidence: 79%

“…Recent studies have suggested that PTEN may also play an important role as a negative regulator of the TGF-␤ signaling pathway through direct interaction with Smad2/3. Specifically, PTEN was shown to inhibit TGF-␤-induced motility and invasion of epithelial cells, including the keratinocyte cell line HaCaT and U87MG glioma cells (33). Another group has reported a role for PTEN as a negative regulator of fibrosis (6).…”

Section: Discussionmentioning

confidence: 99%

Opposite Effects of Dihydrosphingosine 1-Phosphate and Sphingosine 1-Phosphate on Transforming Growth Factor-β/Smad Signaling Are Mediated through the PTEN/PPM1A-dependent Pathway

Kapanadze

Hsu

et al. 2008

103

Transforming growth factor-␤ (TGF-␤) is an important regulator of physiological connective tissue biosynthesis and plays a central role in pathological tissue fibrosis. Previous studies have established that a biologically active lipid mediator, sphingosine 1-phosphate (S1P), mimics some of the profibrotic functions of TGF-␤ through cross-activation of Smad signaling. Here we report that another product of sphingosine kinase, dihydrosphingosine 1-phosphate (dhS1P), has an opposite role in the regulation of TGF-␤ signaling. In contrast to S1P, dhS1P inhibits TGF-␤-induced Smad2/3 phosphorylation and up-regulation of collagen synthesis. The effects of dhS1P require a lipid phosphatase, PTEN, a key modulator of cell growth and survival. dhS1P stimulates phosphorylation of the C-terminal domain of PTEN and its subsequent translocation into the nucleus. We demonstrate a novel function of nuclear PTEN as a co-factor of the Smad2/3 phosphatase, PPM1A. Complex formation of PTEN with PPM1A does not require the lipid phosphatase activity but depends on phosphorylation of the serine/threonine residues located in the C-terminal domain of PTEN. Upon complex formation with PTEN, PPM1A is protected from degradation induced by the TGF-␤ signaling. Consequently, overexpression of PTEN abrogates TGF-␤-induced Smad2/3 phosphorylation. This study establishes a novel role for nuclear PTEN in the stabilization of PPM1A. PTEN-mediated cross-talk between the sphingolipid and TGF-␤ signaling pathways may play an important role in physiological and pathological TGF-␤ signaling. TGF-␤2 is a multifunctional polypeptide growth factor that regulates cell proliferation, functional differentiation, extracellular matrix (ECM) production, cell motility, and apoptosis (1). TGF-␤ signaling is initiated by ligand binding to a heteromeric complex of transmembrane serine/threonine kinases (type I and type II) and subsequent activation of transcriptional coregulators, Smad2 and Smad3 (1). Deregulated TGF-␤ signaling has been implicated in various pathological conditions, including fibrosis and cancer. Depending on the cancer stage, the TGF-␤ signaling pathway may either suppress or promote tumorigenesis (2). Recent studies suggest that TGF-␤ may also influence tumor growth through modulation of the tumor microenvironment (3). TGF-␤ is a potent inducer of the ECM and is required under physiologic conditions, such as wound repair, to induce fibroblasts to produce and contract ECM (4). On the other hand, deregulated TGF-␤ signaling has long been postulated to underlie pathologic fibrosis, but the specific mechanisms involved in this process have not been fully delineated (5). Recent in vivo and in vitro studies also suggest a novel role for the lipid and protein phosphatase, PTEN, as an inhibitor of fibrosis. Reduced levels of PTEN were found in myofibroblasts in human lung specimens of patients with idiopatic pulmonary fibrosis (6). Additional studies using animal models of fibrosis further supported the antifibrotic role of PTEN (6 -8). The specific ro...

“…6250) and were used according to the manufacturer's instructions. Confirmatory experiments were performed in IMR-90 cells stably transfected with retrovirus encoding PTEN short hairpin RNA that targets human PTEN (16), resulting in functional abrogation of PTEN activity (Fig. S1).…”

Section: Methodsmentioning

confidence: 99%

Phosphatase and Tensin Homologue on Chromosome 10 (PTEN) Directs Prostaglandin E2-mediated Fibroblast Responses via Regulation of E Prostanoid 2 Receptor Expression

Sagana

Yan

Cornett

et al. 2009

Self Cite

Prostaglandin E 2 (PGE 2 ) is an arachidonic acid metabolite that counters transforming growth factor-␤-induced fibroblast activation via E prostanoid 2 (EP2) receptor binding. Phosphatase and tensin homologue on chromosome 10 (PTEN) is a lipid phosphatase that, by antagonizing the phosphoinositol 3-kinase (PI3K) pathway, also inhibits fibroblast activation. Here, we show that PTEN directly regulates PGE 2 inhibition of fibroblast activation by augmenting EP2 receptor expression. The increase in collagen production and ␣-smooth muscle actin expression observed in fibroblasts in which PTEN is deficient was resistant to the usual suppressive effects of PGE 2 . This was due to marked down-regulation of EP2, a G s protein-coupled receptor (GPCR) that mediates the inhibitory actions of this prostanoid via cAMP. pten ؊/؊ or PTEN-inhibited fibroblasts in which the PI3K pathway was blocked demonstrated a restoration of EP2 receptor expression, due to augmented gene transcription and mRNA instability. Importantly, restoration of the balance between PI3K and PTEN reestablished the inhibitory effect of PGE 2 on fibroblast activation. No such influence of PTEN was observed on alternative E prostanoid GPCRs. Moreover, our studies identified a positive feedback loop in which cAMP signaling enhanced EP2 receptor expression, independent of PTEN. Therefore, our findings indicate that PTEN regulates the antifibrotic effects of PGE 2 by a specific and permissive effect on EP2 receptor expression. Further, our data imply that cAMP signaling circumvents EP2 down-regulation in pten-deficient cells to restore EP2 receptor expression. This is the first description, to our knowledge, of PI3K/PTEN balance directing GPCR expression, and provides a novel mechanism for cellular effects of PTEN.Prostaglandin E 2 (PGE 2 ) 2 is a cyclooxygenase-derived metabolite of arachidonic acid that influences a spectrum of important biological processes including pain, inflammation, and fibrosis. PGE 2 effects may be cell-specific; for example, PGE 2 promotes migration of epithelial cells (1, 2), but it inhibits migration of fibroblasts (3, 4). These seemingly paradoxical actions of PGE 2 reflect the distinct responses resulting from ligation of a diverse repertoire of E prostanoid (EP) receptors on the cell surface. The four G protein-coupled (GPCR) EP receptors, termed EP1-4, signal by mobilizing intracellular calcium (EP1), increasing intracellular cAMP (EP2 and 4), or decreasing intracellular cAMP (EP3) (5). We have shown previously that PGE 2 is capable, in lung fibroblasts, of tempering the profibrotic, activating effects induced by TGF-␤ (e.g. ␣-smooth muscle actin (␣-SMA) and collagen I expression) (6) as well as the migratory effects of fibroblast growth factor (3). These effects are mediated by intracellular cAMP subsequent to binding of EP2, the most abundant of the fibroblast EP receptors.Phosphatase and tensin homologue on chromosome 10 (PTEN) is a dual specificity lipid and protein phosphatase, but its primary action involves dephosphorylat...