RETRACTED: One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

Bianchi, Cristina; Torsello, B; Stefano, Vitalba Di; Zipeto, M; Facchetti, Rita; Bombelli, S; Perego, R

doi:10.1016/j.yexcr.2013.05.012

Cited by 8 publications

(13 citation statements)

References 43 publications

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“…Stress fiber density quantification was obtained using NIH ImageJ software (http://www. rsbweb.nih.gor/iJ), as previously described (Bianchi et al, 2013).…”

Section: Human Primary Tubular Cell Cultures and Treatmentsmentioning

confidence: 99%

“…At the indicated time points, primary tubular cell cultures were lysed as described previously (Bianchi et al, 2013). 30 µg, or 60 µg when specified, of protein lysates quantified with a BCA microassay (Sigma-Aldrich) were separated on NuPage 4-12% gels (Invitrogen) and submitted to western blotting (Cifola et al, 2011) with the indicated antibodies (Table S1).…”

Section: Protein Extraction Western Blotting and Immunoprecipitationmentioning

confidence: 99%

“…Remarkably, it has been described that c-Abl and Arg tyrosine kinases associate with and phosphorylate the proteasome PSMA7 subunit with consequent inhibition of proteasome activity (Liu et al, 2006). Arg is also involved in some aspects of the EMT process, like cell migration and cytoskeleton modulation through the RhoA-ROCK pathway (Hernández et al, 2004;Peacock et al, 2007;Bianchi et al, 2013), which is also involved in TGF-β secretion in a high-glucose-treated HK-2 cell line (Gu et al, 2013). Based on these data, in the current study we analyzed the involvement of Arg tyrosine kinase in the production of TGF-β1 induced in well-characterized human primary tubular cell cultures (Perego et al, 2005;Bianchi et al, 2010;Cifola et al, 2011) by treatment with high glucose, which also induced some phenotypical and molecular changes in these tubular cells.…”

Section: Introductionmentioning

confidence: 99%

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Arg tyrosine kinase modulates TGF-β1 production in human renal tubular cells under high-glucose conditions

Torsello

Bianchi

Meregalli

et al. 2016

Journal of Cell Science

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Renal tubular cells are involved in the tubular interstitial fibrosis observed in diabetic nephropathy. It is debated whether epithelialmesenchymal transition (EMT) affects tubular cells, which under highglucose conditions overproduce transforming growth factor-β (TGF-β), a fibrogenic cytokine involved in interstitial fibrosis development. Our study investigated the involvement of non-receptor tyrosine kinase Arg (also called Abl2) in TGF-β production. Human primary tubular cell cultures exposed to high-glucose conditions were used. These cells showed an elongated morphology, stress fibers and vimentin increment but maintained most of the epithelial marker expression and distribution. In these cells exposed to high glucose, which overexpressed and secreted active TGF-β1, Arg protein and activity was downregulated. A further TGF-β1 increase was induced by Arg silencing with siRNA, as with the Arg tyrosine kinase inhibitor Imatinib. In the cells exposed to high glucose, reactive oxygen species (ROS)-dependent Arg kinase downregulation induced both RhoA activation, through p190RhoGAPA (also known as ARHGAP35) modulation, and proteasome activity inhibition. These data evidence a new specific involvement of Arg kinase into the regulation of TGF-β1 expression in tubular cells under high-glucose conditions and provide cues for new translational approaches in diabetic nephropathy.

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“…Stress fiber density quantification was obtained using NIH ImageJ software (http://www. rsbweb.nih.gor/iJ), as previously described (Bianchi et al, 2013).…”

Section: Human Primary Tubular Cell Cultures and Treatmentsmentioning

confidence: 99%

Section: Protein Extraction Western Blotting and Immunoprecipitationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Arg tyrosine kinase modulates TGF-β1 production in human renal tubular cells under high-glucose conditions

Torsello

Bianchi

Meregalli

et al. 2016

Journal of Cell Science

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“…Thirty μg of protein lysates obtained from first-confluent primary cell cultures, quantified with BCA microassay (Sigma Aldrich, St. Louis, MO) and separated on NuPage 4 to 12% Bis-Tris pre-cast gels (Thermo Fisher, Waltham, MA) [ 35 ], were submitted to western blotting [ 36 ] with mouse monoclonal antibody against PLIN2 (dilution 1:500; AP125, Progen, Heidelberg, Germany), or rabbit polyclonal antibody against LDHA (dilution 1:1000; Cell Signaling, Boston, MA), or against β-actin (dilution 1:1000; Sigma-Aldrich). Densitometric analysis of specific bands was performed by ImageJ software, and the specific band intensities were normalized with corresponding β-actin for quantification.…”

Section: Methodsmentioning

confidence: 99%

The glucose and lipid metabolism reprogramming is grade-dependent in clear cell renal cell carcinoma primary cultures and is targetable to modulate cell viability and proliferation

et al. 2017

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Clear cell renal cell carcinoma (ccRCC) has a poor prognosis despite novel biological targeted therapies. Tumor aggressiveness and poor survival may correlate with tumor grade at diagnosis and with complex metabolic alterations, also involving glucose and lipid metabolism. However, currently no grade-specific metabolic therapy addresses these alterations. Here we used primary cell cultures from ccRCC of low- and high-grade to investigate the effect on energy state and reduced pyridine nucleotide level, and on viability and proliferation, of specific inhibition of glycolysis with 2-deoxy-D-glucose (2DG), or fatty acid oxidation with Etomoxir. Our primary cultures retained the tissue grade-dependent modulation of lipid and glycogen storage and aerobic glycolysis (Warburg effect). 2DG affected lactate production, energy state and reduced pyridine nucleotide level in high-grade ccRCC cultures, but the energy state only in low-grade. Rather, Etomoxir affected energy state in high-grade and reduced pyridine nucleotide level in low-grade cultures. Energy state and reduced pyridine nucleotide level were evaluated by ATP and reduced 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) dye quantification, respectively. 2DG treatment impaired cell proliferation and viability of low-grade ccRCC and normal cortex cultures, whereas Etomoxir showed a cytostatic and cytotoxic effect only in high-grade ccRCC cultures. Our data indicate that in ccRCC the Warburg effect is a grade-dependent feature, and fatty acid oxidation can be activated for different grade-dependent metabolic needs. A possible grade-dependent metabolic therapeutic approach in ccRCC is also highlighted.

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“…Immunofluorescence was performed on primary cell cultures as we previously described 35 using antibody A (dilution, 1:200), antibody B (dilution, 1:200), antibody against Paxillin (dilution, 1:50; clone 349, Becton Dickinson, San Jose, CA), and the Alexa Fluor 488 or 594 conjugated anti-rabbit or anti-mouse antibodies (dilution, 1:100; Invitrogen, Carlsberg, CA). Nuclei were counterstained with mounting DAPI (Invitrogen).…”

Section: Immunohistochemistry and Immunofluorescencementioning

confidence: 99%

Major Action of Endogenous Lysyl Oxidase in Clear Cell Renal Cell Carcinoma Progression and Collagen Stiffness Revealed by Primary Cell Cultures

Stefano

Torsello

Bianchi

et al. 2016

The American Journal of Pathology

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Human clear cell renal cell carcinoma (ccRCC) is therapy resistant; therefore, it is worthwhile studying in depth the molecular aspects of its progression. In ccRCC the biallelic inactivation of the VHL gene leads to stabilization of hypoxia-inducible factors (HIFs). Among the targets of HIF-1α transcriptional activity is the LOX gene, which codes for the inactive proenzyme (Pro-Lox) from which, after extracellular secretion and proteolysis, derives the active enzyme (Lox) and the propeptide (Lox-PP). By increasing stiffness of extracellular matrix by collagen crosslinking, Lox promotes tumor progression and metastasis. Lox and Lox-PP can reenter the cells where Lox promotes cell proliferation and invasion, whereas Lox-PP acts as tumor suppressor because of its Ras recision and apoptotic activity. Few data are available concerning LOX in ccRCC. Using an in vitro model of ccRCC primary cell cultures, we performed, for the first time in ccRCC, a detailed study of endogenous LOX and also investigated their transcriptomic profile. We found that endogenous LOX is overexpressed in ccRCC, is involved in a positive-regulative loop with HIF-1α, and has a major action on ccRCC progression through cellular adhesion, migration, and collagen matrix stiffness increment; however, the oncosuppressive action of Lox-PP was not found to prevail. These findings may suggest translational approaches for new therapeutic strategies in ccRCC.

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RETRACTED: One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

Cited by 8 publications

References 43 publications

Arg tyrosine kinase modulates TGF-β1 production in human renal tubular cells under high-glucose conditions

Arg tyrosine kinase modulates TGF-β1 production in human renal tubular cells under high-glucose conditions

The glucose and lipid metabolism reprogramming is grade-dependent in clear cell renal cell carcinoma primary cultures and is targetable to modulate cell viability and proliferation

Major Action of Endogenous Lysyl Oxidase in Clear Cell Renal Cell Carcinoma Progression and Collagen Stiffness Revealed by Primary Cell Cultures

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