Enhanced transduction of colonic cell lines in vitro and the inflamed colon in mice by viral vectors, derived from adeno-associated virus serotype 2, using virus-microbead conjugates bearing lectin

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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In vitro selection of viral vectors with modified tropism: the adeno-associated virus display

Molecular Therapy ◽

10.1016/s1525-0016(03)00123-0 ◽

2003 ◽

Vol 8 (1) ◽

pp. 151-157 ◽

Cited By ~ 128

Author(s):

Luca Perabo ◽

Hildegard Büning ◽

David M Kofler ◽

Martin U Ried ◽

Anne Girod ◽

...

Keyword(s):

Viral Vectors ◽

In Vitro Selection ◽

Adeno Associated Virus ◽

Selection Of

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MiR-200c-3p targets SESN1 and represses the IL-6/AKT loop to prevent cholangiocyte activation and cholestatic liver fibrosis

Laboratory Investigation ◽

10.1038/s41374-021-00710-6 ◽

2021 ◽

Author(s):

Yongfeng Song ◽

Melanie Tran ◽

Li Wang ◽

Dong-Ju Shin ◽

Jianguo Wu

Keyword(s):

Liver Fibrosis ◽

Liver Injury ◽

Feedback Loop ◽

Ductular Reaction ◽

Adeno Associated Virus ◽

Matrix Deposition ◽

Duct Ligation ◽

Virus Serotype ◽

Cholestatic Liver Injury

AbstractCholestasis causes ductular reaction in the liver where the reactive cholangiocytes not only proliferate but also gain a neuroendocrine-like phenotype, leading to inflammatory cell infiltration and extracellular matrix deposition and contributing to the development and progression of cholestatic liver fibrosis. This study aims to elucidate the role of miR-200c in cholestasis-induced biliary liver fibrosis and cholangiocyte activation. We found that miR-200c was extremely abundant in cholangiocytes but was reduced by cholestasis in a bile duct ligation (BDL) mouse model; miR-200c was also decreased by bile acids in vitro. Phenotypically, loss of miR-200c exacerbated cholestatic liver injury, including periductular fibrosis, intrahepatic inflammation, and biliary hyperplasia in both the BDL model and the 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) model. We identified sestrin 1 (SESN1) as a target of miR-200c. Sesn1−/−-BDL mice showed mitigation of cholestatic liver injury. On a molecular level, the pro-proliferative IL-6/AKT feedback loop was activated in Mir200c−/− livers but was inhibited in Sesn1−/− livers upon cholestasis in mice. Furthermore, rescuing expression of miR-200c by the adeno-associated virus serotype 8 ameliorated BDL-induced liver injury in Mir200c−/− mice. Taken together, this study demonstrates that miR-200c restrains the proliferative and neuroendocrine-like activation of cholangiocytes by targeting SESN1 and inhibiting the IL-6/AKT feedback loop to protect against cholestatic liver fibrosis. Our findings provide mechanistic insights regarding biliary liver fibrosis, which may help to reveal novel therapeutic targets for the treatment of cholestatic liver injury and liver fibrosis.

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e0054 Recombinant adeno-associated virus serotype 9 transfection of rats H9C2 cells in vitro

Heart ◽

10.1136/hrt.2010.208967.54 ◽

2010 ◽

Vol 96 (Suppl 3) ◽

pp. A18-A18

Author(s):

G. Xia ◽

M. Yi-Tong ◽

Y. Yi-Ning ◽

X. Yang ◽

C. Bang-Dang ◽

...

Keyword(s):

H9c2 Cells ◽

Adeno Associated Virus ◽

Virus Serotype

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

European Respiratory Journal ◽

10.1183/13993003.03721-2020 ◽

2021 ◽

pp. 2003721

Author(s):

Jiancheng Wang ◽

Xiaofan Lai ◽

Senyu Yao ◽

Hainan Chen ◽

Jianye Cai ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Critical Role ◽

Intermediate Filament Protein ◽

Nestin Expression ◽

Adeno Associated Virus ◽

Intratracheal Administration ◽

Virus Serotype ◽

Fibrotic Lung Disease

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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Impact of capsid modifications by selected peptide ligands on recombinant adeno-associated virus serotype 2-mediated gene transduction

Journal of General Virology ◽

10.1099/vir.0.044735-0 ◽

2012 ◽

Vol 93 (10) ◽

pp. 2131-2141 ◽

Cited By ~ 8

Author(s):

Matthias Naumer ◽

Ruth Popa-Wagner ◽

Jürgen A. Kleinschmidt

Keyword(s):

Gene Therapy ◽

Gene Transfer ◽

Viral Vectors ◽

Peptide Ligands ◽

Gene Transduction ◽

Cell Binding ◽

Adeno Associated Virus ◽

Vector Systems ◽

Virus Serotype ◽

Selection Of

Vectors based on adeno-associated virus serotype 2 (AAV2) belong to today’s most promising and most frequently used viral vectors in human gene therapy. Like in many other vector systems, the broad but non-specific tropism limits their use for certain cell types or tissues. One approach to screen for transduction-improved vectors is the selection of random peptide libraries displayed directly on the AAV2 capsid. Although the AAV2 library system has been widely applied for the successful selection of improved gene therapy vectors, it remains unknown which steps of the transduction process are most affected and therefore critical for the selection of targeting peptides. Attachment to the cell surface is the first essential step of AAV-mediated gene transduction; however, our experiments challenge the conventional belief that enhanced gene transfer is equivalent to more efficient cell binding of recombinant AAV2 vectors. A comparison of the various steps of gene transfer by vectors carrying a wild-type AAV2 capsid or displaying two exemplary peptide ligands selected from AAV2 random libraries on different human tumour cell lines demonstrated strong alterations in cell binding, cellular uptake, as well as intracellular processing of these vectors. Combined, our results suggest that entry and post-entry events are decisive for the selection of the peptides NDVRSAN and GPQGKNS rather than their cell binding efficiency.

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