Nestin promotes pulmonary fibrosis via facilitating recycling of TGF-β receptor I

Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic lung disease that is characterised by aberrant proliferation of activated myofibroblasts and pathological remodelling of the extracellular matrix. Previous studies have revealed that the intermediate filament protein Nestin plays key roles in tissue regeneration and wound healing in different organs. Whether Nestin plays a critical role in the pathogenesis of IPF needs to be clarified.Nestin expression in lung tissues from bleomycin-treated mice and IPF patients was determined. Transfection with Nestin shRNA vectors in vitro that regulated TGF-β/Smad signalling was conducted. Biotinylation assays to observe plasma membrane TβRI, TβRI endocytosis and TβRI recycling after Nestin knockdown were performed. Adeno-associated virus serotype 6 (AAV6)-mediated Nestin knockdown was assessed in vivo.We found that Nestin expression was increased in a murine pulmonary fibrosis model and IPF patients, and that the upregulated protein primarily localised in lung α-SMA+ myofibroblasts. Mechanistically, we determined that Nestin knockdown inhibited TGF-β signalling by suppressing recycling of TβRI to the cell surface and that Rab11 was required for the ability of Nestin to promote TβRI recycling. In vivo, we found that intratracheal administration of adeno-associated virus serotype 6 (AAV6)-mediated Nestin knockdown significantly alleviated pulmonary fibrosis in multiple experimental mice models.In conclusion, our findings reveal a pro-fibrotic function of Nestin partially through facilitating Rab11-dependent recycling of TβRI and shed new light on pulmonary fibrosis treatment.

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The 37/67-Kilodalton Laminin Receptor Is a Receptor for Adeno-Associated Virus Serotypes 8, 2, 3, and 9

Journal of Virology ◽

10.1128/jvi.00878-06 ◽

2006 ◽

Vol 80 (19) ◽

pp. 9831-9836 ◽

Cited By ~ 264

Author(s):

Bassel Akache ◽

Dirk Grimm ◽

Kusum Pandey ◽

Stephen R. Yant ◽

Hui Xu ◽

...

Keyword(s):

Gene Therapy ◽

Cultured Cells ◽

Human Gene ◽

Laminin Receptor ◽

Transduction Efficiency ◽

Adeno Associated Virus ◽

Virus Serotype ◽

Tumor Gene

ABSTRACT Adeno-associated virus serotype 8 (AAV8) is currently emerging as a powerful gene transfer vector, owing to its capability to efficiently transduce many different tissues in vivo. While this is believed to be in part due to its ability to uncoat more readily than other AAV serotypes such as AAV2, understanding all the processes behind AAV8 transduction is important for its application and optimal use in human gene therapy. Here, we provide the first report of a cellular receptor for AAV8, the 37/67-kDa laminin receptor (LamR). We document binding of LamR to AAV8 capsid proteins and intact virions in vitro and demonstrate its contribution to AAV8 transduction of cultured cells and mouse liver in vivo. We also show that LamR plays a role in transduction by three other closely related serotypes (AAV2, -3, and -9). Sequence and deletion analysis allowed us to map LamR binding to two protein subdomains predicted to be exposed on the AAV capsid exterior. Use of LamR, which is constitutively expressed in many clinically relevant tissues and is overexpressed in numerous cancers, provides a molecular explanation for AAV8's broad tissue tropism. Along with its robust transduction efficiency, our findings support the continued development of AAV8-based vectors for clinical applications in humans, especially for tumor gene therapy.

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314. Pristimerin Enhances Recombinant Adeno-Associated Virus Serotype 2 Vector-Mediated Transgene Expression Both In Vitro and In Vivo

Molecular Therapy ◽

10.1016/s1525-0016(16)34649-4 ◽

2013 ◽

Vol 21 ◽

pp. S121

Keyword(s):

Transgene Expression ◽

Adeno Associated Virus ◽

Virus Serotype

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115. Bio-Engineering of Adeno-Associated Virus Serotype (AAV)-2 Capsid at Serine/Threonine Residues Improves Its Transduction Efficiency Both In Vitro and In Vivo

Molecular Therapy ◽

10.1016/s1525-0016(16)35919-6 ◽

2012 ◽

Vol 20 ◽

pp. S47-S48

Keyword(s):

Transduction Efficiency ◽

Adeno Associated Virus ◽

Virus Serotype ◽

Bio Engineering

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Successful Gene Transfer Using Adeno-Associated Virus Vectors into the Kidney: Comparison among Adeno-Associated Virus Serotype 1–5 Vectors in vitro and in vivo

Nephron Experimental Nephrology ◽

10.1159/000077378 ◽

2004 ◽

Vol 96 (4) ◽

pp. e119-e126 ◽

Cited By ~ 37

Author(s):

Shin-ichi Takeda ◽

Masafumi Takahashi ◽

Hiroaki Mizukami ◽

Eiji Kobayashi ◽

Koichi Takeuchi ◽

...

Keyword(s):

Gene Transfer ◽

Adeno Associated Virus ◽

Virus Vectors ◽

Virus Serotype ◽

Serotype 1

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Detectable reporter gene expression following transduction of adenovirus and adeno-associated virus serotype 2 vectors within full-thickness osteoarthritic and unaffected canine cartilage in vitro and unaffected guinea pig cartilage in vivo

Journal of Orthopaedic Research® ◽

10.1002/jor.20975 ◽

2009 ◽

Vol 28 (2) ◽

pp. 149-155 ◽

Cited By ~ 12

Author(s):

Kelly S. Santangelo ◽

Sarah A. Baker ◽

Gerard Nuovo ◽

Jonathan Dyce ◽

Jeffrey S. Bartlett ◽

...

Keyword(s):

Gene Expression ◽

Guinea Pig ◽

Reporter Gene ◽

Reporter Gene Expression ◽

Full Thickness ◽

Adeno Associated Virus ◽

Virus Serotype

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Abstract 254: Profiling Transcriptional Changes and Identifying Novel Molecular Markers During the Dedifferentiation of Cardiomyocyte

Circulation Research ◽

10.1161/res.127.suppl_1.254 ◽

2020 ◽

Vol 127 (Suppl_1) ◽

Author(s):

Yike Zhu ◽

Wilson Tan Lek Wen ◽

Do Dang Vinh ◽

Matthew A Ackers-Johnson ◽

Roger Foo

Keyword(s):

Cell Cycle ◽

Molecular Markers ◽

Cell Division ◽

Cardiac Function ◽

Adeno Associated Virus ◽

Genome Wide ◽

Virus Serotype ◽

Transcriptional Changes

The loss of cardiomyocytes (CMs) during heart failure (HF) cannot be replaced by new CMs due to their limited proliferative capacity. Regenerating the failing heart by promoting CM cell-cycle re-entry has thus become a possible solution rigorously pursued. Some genes have been proven to promote endogenous CM proliferation, nearly always preceded by CM dedifferentiation, wherein terminally-differentiated CMs are reversed back initially to a less-matured status, preceding cell division. However, very little else is known about CM dedifferentiation, including the lack of robust molecular markers, the mechanisms driving it, whether it necessarily leads to CM proliferation, and the effect of CM dedifferentiation alone on cardiac function. Therefore, investigating CM dedifferentiation is of great significance. Here, we profiled genome-wide transcriptional changes and identified novel molecular markers during CM dedifferentiation. We used the established in vitro model where adult mouse CMs dedifferentiated and re-differentiated (DR) upon 2-week long culture, without detectable cell division. DR was evident by breakdown and rebuilding of sarcomeres, and differential regulation of cardiac fetal genes. Tracking the transcriptome change pointed to gene pathways involved in this process and potential markers of CM dedifferentiation. Another assumption is that transient induction of Yamanaka factors ( Oct4, Sox2, Klf4, Myc, OSKM ) induces CM dedifferentiation of CMs in vivo , since the four factors are themselves sufficient to induce pluripotency of human and mouse somatic cells. Our preliminary results show that CM-specific overexpression of OSKM using adeno-associated virus serotype 9 (AAV9) induced fetal-like hallmarks, down-regulation of sarcomeric genes, re-activation of cell cycle, reduced fatty acid utilization, and associated loss of cardiac function . Putative markers identified in the in vitro model was also validated to be upregulated in OSKM -overexpressed CMs. In conclusion, through 2 models of CM dedifferentiation, we have screened out potential novel markers and regulators of CM dedifferentiation which now requires further validation.

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