scholarly journals Evaluation of the Therapeutic Effects of Protocatechuic Aldehyde in Diabetic Nephropathy

Toxins ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 560
Author(s):  
Yu-Teng Chang ◽  
Mu-Chi Chung ◽  
Chang-Chi Hsieh ◽  
Jeng-Jer Shieh ◽  
Ming-Ju Wu
Keyword(s):  
Diabetic Nephropathy ◽  
Western Blot ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Kidney Diseases ◽  
Therapeutic Effects ◽  
Mesenchymal Transition ◽  
Protocatechuic Aldehyde ◽  
Kidney Morphology

Diabetic nephropathy (DN) is one of the most severe chronic kidney diseases in diabetes and is the main cause of end-stage renal disease (ESRD). Protocatechuic aldehyde (PCA) is a natural product with a variety of effects on pulmonary fibrosis. In this study, we examined the effects of PCA in C57BL/KS db/db male mice. Kidney morphology, renal function indicators, and Western blot, immunohistochemistry, and hematoxylin and eosin (H&E) staining data were analyzed. The results revealed that treatment with PCA could reduce diabetic-induced renal dysfunction, as indicated by the urine albumin-to-creatinine ratio (db/m: 120.1 ± 46.1μg/mg, db/db: 453.8 ± 78.7 µg/mg, db/db + 30 mg/kg PCA: 196.6 ± 52.9 µg/mg, db/db + 60 mg/kg PCA: 163.3 ± 24.6 μg/mg, p < 0.001). However, PCA did not decrease body weight, fasting plasma glucose, or food and water intake in db/db mice. H&E staining data revealed that PCA reduced glomerular size in db/db mice (db/m: 3506.3 ± 789.3 μm2, db/db: 6538.5 ± 1818.6 μm2, db/db + 30 mg/kg PCA: 4916.9 ± 1149.6 μm2, db/db + 60 mg/kg PCA: 4160.4 ± 1186.5 μm2p < 0.001). Western blot and immunohistochemistry staining indicated that PCA restored the normal levels of diabetes-induced fibrosis markers, such as transforming growth factor-beta (TGF-β) and type IV collagen. Similar results were observed for epithelial–mesenchymal transition-related markers, including fibronectin, E-cadherin, and α-smooth muscle actin (α-SMA). PCA also decreased oxidative stress and inflammation in the kidney of db/db mice. This research provides a foundation for using PCA as an alternative therapy for DN in the future.

Abstract 132: Hypoxia-inducible Factor Prolyl-hydroxylase-2 Mediates Transforming Growth Factor Beta 1-induced Epithelial-mesenchymal Transition In Renal Tubular Cells

Hypertension ◽  
2012 ◽  
Vol 60 (suppl_1) ◽  
Author(s):  
Weiqing Han ◽  
Jun-Pin Hu ◽  
Pin-Lan Li ◽  
Ningjun Li
Keyword(s):  
Signaling Pathway ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Kidney Diseases ◽  
Hypoxia Inducible Factor ◽  
Mesenchymal Transition ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Renal Tubular

Transforming growth factor beta 1 (TGFβ1)-induced epithelial-mesenchymal transition (EMT) in kidney epithelial cells plays a key role in renal tubulointerstitial fibrosis in chronic kidney diseases. As hypoxia-inducible factor (HIF)-1α is found to mediate TGFβ1 signaling pathway, we tested the hypothesis that HIF-1α and its upstream regulator prolyl hydroxylase domain-containing proteins (PHDs) are involved in TGFβ1-induced EMT in renal tubular cells. Our results showed that TGFβ1 treatment for 48 h stimulated EMT in cultured renal tubular cells as indicated by the decrease in epithelial marker P-cadherin from 1.0 ± 0.02 to 0.40 ± 0.05 ( P < 0.05), and the increase in mesenchymal markers α-smooth muscle actin (2.14 ± 0.32 fold, P < 0.05) and fibroblast-specific protein (2.0 ± 0.17 fold, P < 0.05) as shown in Western blot assay. Meanwhile, TGFβ1 time-dependently increased HIF-1α, which reached its maximum value (2.36 ± 0.2 fold, P < 0.05) at 24 h, and that HIF-1α siRNA significantly inhibited TGFβ1-induced EMT, suggesting that HIF-1α mediated TGFβ1 induced-EMT. Real-time PCR showed that PHD1 and PHD2, rather than PHD3, could be detected, with PHD2 as the predominant form of PHDs (PHD1 : PHD2 = 0.21:1.0). Importantly, TGFβ1 time-dependently decreased PHD2 mRNA and protein level, which reached their maximum value from 1.0 ± 0.15 to 0.45 ± 0.08 ( P < 0.05) for mRNA at 16 h and from 1.0 ± 0.08 to 0.26 ± 0.08 ( P < 0.05) for protein at 24 h, respectively. In contrast, TGFβ1 had no effect on PHD1 mRNA and protein levels. Furthermore, over-expression of PHD2 transgene almost fully prevented TGFβ1-induced HIF-1α accumulation and EMT marker changes, indicating that PHD2 is involved in TGFβ1-induced EMT. Finally, Smad2 inhibitor SB431542 prevented TGFβ1-induced PHD2 decrease, suggesting that Smad2 may mediate TGFβ1-induced EMT through PHD2/HIF-1α. It is concluded that TGFβ1 decreased PHD2 expression via a Smad2-dependent signaling pathway, thereby leading to HIF-1α accumulation and EMT in renal tubular cells. The present study suggests that PHD2/HIF-1α is a novel signaling pathway mediating the fibrogenic effect of TGFβ1 and that manipulating PHD2/HIF-1α pathway may be used as a therapeutic strategy in chronic kidney diseases. (support: NIH grant HL89563 and HL106042)

scholarly journals Estimating Dynamic Cellular Morphological Properties via the Combination of the RTCA System and a Hough-Transform-Based Algorithm

Cells ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 1287 ◽  
Author(s):  
Lejun Zhang ◽  
Yang Ye ◽  
Rana Dhar ◽  
Jinsong Deng ◽  
Huifang Tang
Keyword(s):  
Western Blot ◽  
Real Time ◽  
Transforming Growth Factor Beta ◽  
Hough Transform ◽  
Cellular Proliferation ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Morphological Properties ◽  
Mesenchymal Transition ◽  
Cell Index

The xCELLigence real-time cell analysis (RTCA) system has the potential to detect cellular proliferation, migration, cytotoxicity, adherence, and remodeling. Although the RTCA system is widely recognized as a noninvasive and efficient tool for real-time monitoring of cellular fate, it cannot describe detailed cell morphological parameters, such as length and intensity. Transforming growth factor beta(TGF-β) induced the epithelial–mesenchymal transition (EMT), which produces significant changes in cellular morphology, so we used TGF-β to treat A549 epithelial cells in this study. We compared it with lipopolysaccharide (LPS) and cigarette smoke extract (CSE) as stimulators. We developed an efficient algorithm to quantify the morphological cell changes. This algorithm is comprised of three major parts: image preprocessing, Hough transform (HT), and post-processing. We used the RTCA system to record the A549 cell index. Western blot was used to confirm the EMT. The RTCA system showed that different stimulators produce different cell index curves. The algorithm determined the lengths of the detected lines of cells, and the results were similar to the RTCA system in the TGF-β group. The Western blot results show that TGF-β changed the EMT markers, but the other stimulator remained unchanged. Optics-based computer vision techniques can supply the requisite information for the RTCA system based on good correspondence between the results.

scholarly journals Epithelial–Mesenchymal Transition Induced by SMAD4 Activation in Invasive Growth Hormone-Secreting Adenomas

2018 ◽  
Vol 16 (1) ◽  
pp. 571-582 ◽  
Author(s):  
Xiaosong Shan ◽  
Qian Liu ◽  
Zhenye Li ◽  
Chuzhong Li ◽  
Hua Gao ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Western Blot ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Progression Free Survival ◽  
Gh3 Cells ◽  
Invasive Growth ◽  
Mesenchymal Transition ◽  
E Cadherin

AbstractBackgroundThe detection and treatment of invasive growth hormone-secreting pituitary adenoma (GHPA) remains challenging. Several transcription factors promoting the epithelial–mesenchymal transition (EMT) can act as cofactors for the transforming growth factor-beta (TGF-ß)/SMAD4. The goal of this study was to investigate the association of SMAD4 expression and clinicopathologic features using a tissue microarray analysis (TMA). The levels of SMAD4 and the related genes of EMT in GHPAs were analyzed by q-PCR and western blot. SMAD4 was strongly expressed in 15/19 cases (78.9%) of invasive GHPA and 10/42 cases (23.8%) of noninvasive GHPA (χ2=10.887,p=0.000). In the high SMAD4 group, a headache was reported in 16/25 cases (64%) compared with 13/36 cases (36.1%) in the low SMAD4 group (χ2=4.565,p=0.032). The progression-free survival (PFS) in the high group was lower than that in the low group (p=0.026). qRT-PCR and western blot analysis further revealed a significant downregulation of E-cadherin and upregulation of N-cadherin and vimentin in the invasive GHPA group. SMAD4 was associated with increased levels of invasion of GH3 cells, as determined by a transwell test. SMAD4 downregulated E-cadherin levels and increased the levels of N-cadherin and vimentin. Our data provide evidence that SMAD4 is a potential prognosis biomarker and a therapeutic target for patients with invasive GHPA.

scholarly journals Microparticles as Potential Mediators of High Glucose-Induced Renal Cell Injury

Biomolecules ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 348 ◽  
Author(s):  
Ravindran ◽  
Pasha ◽  
Agouni ◽  
Munusamy
Keyword(s):  
Er Stress ◽  
Signaling Pathways ◽  
Transforming Growth Factor Beta ◽  
High Glucose ◽  
Cell Injury ◽  
Transforming Growth Factor ◽  
Nuclear Translocation ◽  
Epithelial Mesenchymal Transition ◽  
Smooth Muscle Actin ◽  
Mesenchymal Transition

Diabetic nephropathy (DN) is the most common cause of chronic kidney disease worldwide. Activation of signaling pathways such as the mammalian target of rapamycin (mTOR), extracellular signal-regulated kinases (ERK), endoplasmic reticulum (ER) stress, transforming growth factor-beta (TGF-β), and epithelial-mesenchymal transition (EMT), are thought to play a significant role in the etiology of DN. Microparticles (MPs), the small membrane vesicles containing bioactive signals shed by cells upon activation or during apoptosis, are elevated in diabetes and were identified as biomarkers in DN. However, their exact role in the pathophysiology of DN remains unclear. Here, we examined the effect of MPs shed from renal proximal tubular cells (RPTCs) exposed to high glucose conditions on naïve RPTCs in vitro. Our results showed significant increases in the levels of phosphorylated forms of 4E-binding protein 1 and ERK1/2 (the downstream targets of mTOR and ERK pathways), phosphorylated-eIF2α (an ER stress marker), alpha smooth muscle actin (an EMT marker), and phosphorylated-SMAD2 and nuclear translocation of SMAD4 (markers of TGF-β signaling). Together, our findings indicate that MPs activate key signaling pathways in RPTCs under high glucose conditions. Pharmacological interventions to inhibit shedding of MPs from RPTCs might serve as an effective strategy to prevent the progression of DN.

scholarly journals The Synergistic Anti-Cancer Effects of NVP-BEZ235 and Regorafenib in Hepatocellular Carcinoma

Molecules ◽  
2020 ◽  
Vol 25 (10) ◽  
pp. 2454
Author(s):  
Cheng-Chan Yu ◽  
Sung-Ying Huang ◽  
Shu-Fang Chang ◽  
Kuan-Fu Liao ◽  
Sheng-Chun Chiu
Keyword(s):  
Hepatocellular Carcinoma ◽  
Western Blot ◽  
Kinase Inhibitor ◽  
Epithelial Mesenchymal Transition ◽  
Combined Treatment ◽  
Gelatin Zymography ◽  
Migration And Invasion ◽  
Therapeutic Effects ◽  
Mesenchymal Transition ◽  
Anti Cancer

Hepatocellular carcinoma (HCC) is the most common type of liver cancer worldwide. Regorafenib is a multi-kinase inhibitor and the second-line treatment for HCC. Since the PI3K/Akt/mTOR signaling pathway is dysregulated in HCC, we evaluated the therapeutic effects of regorafenib combined with a dual PI3K/mTOR inhibitor BEZ235 in the human HCC cell lines (n = 3). The combined treatment with BEZ235 and regorafenib enhanced the inhibition of cell proliferation and increased the expression of cleaved caspase-3 and cleaved PARP in HCC cells. Moreover, the combined treatment suppressed HCC cell migration and invasion in the transwell assay. Further, the Western blot analyses confirmed the involvement of epithelial-mesenchymal transition (EMT)-related genes such as slug, vimentin, and matrix metalloproteinase (MMP)-9/-2. Additionally, the proteinase activity of MMP-9/-2 was analyzed using gelatin zymography. Furthermore, the inhibition of phosphorylation of the Akt, mTOR, p70S6K, and 4EBP1 after combined treatment was validated using Western blot analysis. Therefore, these results suggest that the combined treatment with BEZ235 and regorafenib benefits patients with HCC.

scholarly journals Dysregulated YAP1/TAZ and TGF-β signaling mediate hepatocarcinogenesis in Mob1a/1b-deficient mice

2015 ◽  
Vol 113 (1) ◽  
pp. E71-E80 ◽  
Author(s):  
Miki Nishio ◽  
Keishi Sugimachi ◽  
Hiroki Goto ◽  
Jia Wang ◽  
Takumi Morikawa ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Tissue Growth ◽  
Mutant Mice ◽  
Binding Motif ◽  
Hippo Signaling ◽  
Mesenchymal Transition ◽  
Liver Cancers

Mps One Binder Kinase Activator (MOB)1A/1B are core components of the Hippo pathway that coactivate large tumor suppressor homolog (LATS) kinases. Mob1a/1b double deficiency in mouse liver (LMob1DKO) results in hyperplasia of oval cells and immature cholangiocytes accompanied by inflammatory cell infiltration and fibrosis. More than half of mutant mice die within 3 wk of birth. All survivors eventually develop liver cancers, particularly combined hepatocellular and cholangiocarcinomas (cHC-CCs) and intrahepatic cholangiocellular carcinomas (ICCs), and die by age 60 wk. Because this phenotype is the most severe among mutant mice lacking a Hippo signaling component, MOB1A/1B constitute the critical hub of Hippo signaling in mammalian liver. LMob1DKO liver cells show hyperproliferation, increased cell saturation density, hepatocyte dedifferentiation, enhanced epithelial–mesenchymal transition and cell migration, and elevated transforming growth factor beta(TGF-β)2/3 production. These changes are strongly dependent on Yes-Associated Protein-1 (Yap1) and partially dependent on PDZ-binding motif (Taz) and Tgfbr2, but independent of connective tissue growth factor (Ctgf). In human liver cancers, YAP1 activation is frequent in cHC-CCs and ICCs and correlates with SMAD family member 2 activation. Drug screening revealed that antiparasitic macrocyclic lactones inhibit YAP1 activation in vitro and in vivo. Targeting YAP1/TAZ with these drugs in combination with inhibition of the TGF-β pathway may be effective treatment for cHC-CCs and ICCs.

scholarly journals SMAD4 mutations do not preclude epithelial–mesenchymal transition in colorectal cancer

Oncogene ◽  
2021 ◽  
Author(s):  
Patrick Frey ◽  
Antoine Devisme ◽  
Katja Rose ◽  
Monika Schrempp ◽  
Vivien Freihen ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Morphological Changes ◽  
Expression Patterns ◽  
Human Tumor ◽  
Marker Genes ◽  
Mesenchymal Transition ◽  
Gene Regulatory

AbstractTransforming growth factor beta (TGFβ) superfamily signaling is a prime inducer of epithelial-mesenchymal transitions (EMT) that foster cancer cell invasion and metastasis, a major cause of cancer-related deaths. Yet, TGFβ signaling is frequently inactivated in human tumor entities including colorectal cancer (CRC) and pancreatic adenocarcinoma (PAAD) with a high proportion of mutations incapacitating SMAD4, which codes for a transcription factor (TF) central to canonical TGFβ and bone morphogenetic protein (BMP) signaling. Beyond its role in initiating EMT, SMAD4 was reported to crucially contribute to subsequent gene regulatory events during EMT execution. It is therefore widely assumed that SMAD4-mutant (SMAD4mut) cancer cells are unable to undergo EMT. Here, we scrutinized this notion and probed for potential SMAD4-independent EMT execution using SMAD4mut CRC cell lines. We show that SMAD4mut cells exhibit morphological changes, become invasive, and regulate EMT marker genes upon induction of the EMT-TF SNAIL1. Furthermore, SNAIL1-induced EMT in SMAD4mut cells was found to be entirely independent of TGFβ/BMP receptor activity. Global assessment of the SNAIL1-dependent transcriptome confirmed the manifestation of an EMT gene regulatory program in SMAD4mut cells highly related to established EMT signatures. Finally, analyses of human tumor transcriptomes showed that SMAD4 mutations are not underrepresented in mesenchymal tumor samples and that expression patterns of EMT-associated genes are similar in SMAD4mut and SMAD4 wild-type (SMAD4wt) cases. Altogether, our findings suggest that alternative TFs take over the gene regulatory functions of SMAD4 downstream of EMT-TFs, arguing for considerable plasticity of gene regulatory networks operating in EMT execution. Further, they establish that EMT is not categorically precluded in SMAD4mut tumors, which is relevant for their diagnostic and therapeutic evaluation.

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