Baculovirus‐mediated vascular endothelial growth factor‐D<sup>ΔNΔC</sup> gene transfer induces angiogenesis in rabbit skeletal muscle

Heikura, Tommi; Nieminen, Tiina; Roschier, Miia M.; Karvinen, Henna; Lesch, Hanna P.; Rissanen, Tuomas T.; Laitinen, Olli H.; Airenne, Kari J.; Ylä‐Herttuala, Seppo

doi:10.1002/jgm.1637

Cited by 14 publications

(11 citation statements)

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“…Cloning and production of 2A pro and 3C pro CVB3 Nancy (UniProtKB/Swiss-Prot: P03313.4) polyprotein sequence was used as a template to construct recombinant His-tagged 2A pro and 3C pro proteases. Artificial genes encoding the proteases were ordered from Life Technologies (Germany) and were cloned into the pBVboostFGII expression vector as described in (Heikura et al, 2012;Laitinen et al, 2005). Proteases were expressed in E. coli, strain BL21-AI (Life Technologies), and then purified with immobilized metal affinity chromatography (IMAC) using the ÄKTA P-100 liquid chromatography system and HisTrap FF crude IMAC columns (GE Healthcare Bio-Science AB).…”

Section: Methodsmentioning

confidence: 99%

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

Laitinen

Svedin

Kapell

et al. 2018

Journal of Virological Methods

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Enteroviruses (EVs), such as the Coxsackie B-viruses (CVBs), are common human pathogens, which can cause severe diseases including meningitis, myocarditis and neonatal sepsis. EVs encode two proteases (2A and 3C), which perform the proteolytic cleavage of the CVB polyprotein and also cleave host cell proteins to facilitate viral replication. The 2A cause direct damage to the infected heart and tools to investigate 2A and 3C expression may contribute new knowledge on virus-induced pathologies. Here, we developed new antibodies to CVB-encoded 2A and 3C; Two monoclonal 2A antibodies and one 3C antibody were produced. Using cells infected with selected viruses belonging to the EV A, B and C species and immunocytochemistry, we demonstrate that the 3C antibody detects all of the EV species B (EV-B) viruses tested and that the 2A antibody detects all EV-B viruses apart from Echovirus 9. We furthermore show that the new antibodies work in Western blotting, immunocyto- and immunohistochemistry, and flow cytometry to detect CVBs. Confocal microscopy demonstrated the expression kinetics of 2A and 3C, and revealed a preferential cytosolic localization of the proteases in CVB3 infected cells. In summary, the new antibodies detect proteases that belong to EV species B in cells and tissue using multiple applications.

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Section: Methodsmentioning

confidence: 99%

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

Laitinen

Svedin

Kapell

et al. 2018

Journal of Virological Methods

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“…The VP1 antigen (CVB3, strain Nancy) were prepared as described in [20]. Briefly, attL-flanked His-tagged VP1 protein coding synthetic genes were ordered from Life technologies and Gateway cloned into pBVboostFGII-vector [31,32] for antigen production. After sequence verification, the VP1 containing plasmids were transferred to E. coli BL21 DE3 Star cells, the proteins were expressed by inducing the cells with Isopropyl beta-D-1-thiogalactopyranoside and the produced recombinant proteins were purified from clarified bacterial cell lysate using immobilized metal anion chromatography (Ni-NTA IMAC).…”

Section: Appendix A2 Histologymentioning

confidence: 99%

Antibody Responses against Enterovirus Proteases are Potential Markers for an Acute Infection

Saarinen

Stone

Hankaniemi

et al. 2020

Viruses

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Background: Enteroviruses are a group of common non-enveloped RNA viruses that cause symptoms ranging from mild respiratory infections to paralysis. Due to the abundance of enterovirus infections it is hard to distinguish between on-going and previous infections using immunological assays unless the IgM fraction is studied. Methods: In this study we show using Indirect ELISA and capture IgM ELISA that an IgG antibody response against the nonstructural enteroviral proteins 2A and 3C can be used to distinguish between IgM positive (n = 22) and IgM negative (n = 20) human patients with 83% accuracy and a diagnostic odds ratio of 30. Using a mouse model, we establish that the antibody response to the proteases is short-lived compared to the antibody response to the structural proteins in. As such, the protease antibody response serves as a potential marker for an acute infection. Conclusions: Antibody responses against enterovirus proteases are shorter-lived than against structural proteins and can differentiate between IgM positive and negative patients, and therefore they are a potential marker for acute infections.

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“… 24 ). For example, expression of VEGF-D-induced vascularization in rabbit skeletal muscle suggesting that baculovirus-driven VEGF-D expression might be an option to cure lymphatic disorders 40 . Nevertheless as a non-integrative virus it is a priori limited to transient expression, which can be an advantage or a disadvantage depending on the application.…”

Section: Discussionmentioning

confidence: 99%

Highly efficient baculovirus-mediated multigene delivery in primary cells

et al. 2016

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Multigene delivery and subsequent cellular expression is emerging as a key technology required in diverse research fields including, synthetic and structural biology, cellular reprogramming and functional pharmaceutical screening. Current viral delivery systems such as retro- and adenoviruses suffer from limited DNA cargo capacity, thus impeding unrestricted multigene expression. We developed MultiPrime, a modular, non-cytotoxic, non-integrating, baculovirus-based vector system expediting highly efficient transient multigene expression from a variety of promoters. MultiPrime viruses efficiently transduce a wide range of cell types, including non-dividing primary neurons and induced-pluripotent stem cells (iPS). We show that MultiPrime can be used for reprogramming, and for genome editing and engineering by CRISPR/Cas9. Moreover, we implemented dual-host-specific cassettes enabling multiprotein expression in insect and mammalian cells using a single reagent. Our experiments establish MultiPrime as a powerful and highly efficient tool, to deliver multiple genes for a wide range of applications in primary and established mammalian cells.

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Baculovirus‐mediated vascular endothelial growth factor‐D^ΔNΔC gene transfer induces angiogenesis in rabbit skeletal muscle

Abstract: We show the first evidence of biologically significant baculoviral gene transfer in skeletal muscle of NZW rabbits using VEGF-D(ΔNΔC) as a therapeutic transgene.

Cited by 14 publications

References 51 publications

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

Antibody Responses against Enterovirus Proteases are Potential Markers for an Acute Infection

Highly efficient baculovirus-mediated multigene delivery in primary cells

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Baculovirus‐mediated vascular endothelial growth factor‐DΔNΔC gene transfer induces angiogenesis in rabbit skeletal muscle

Abstract: We show the first evidence of biologically significant baculoviral gene transfer in skeletal muscle of NZW rabbits using VEGF-D(ΔNΔC) as a therapeutic transgene.

Cited by 14 publications

References 51 publications

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

New Coxsackievirus 2Apro and 3Cpro protease antibodies for virus detection and discovery of pathogenic mechanisms

Antibody Responses against Enterovirus Proteases are Potential Markers for an Acute Infection

Highly efficient baculovirus-mediated multigene delivery in primary cells

Contact Info

Product

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Baculovirus‐mediated vascular endothelial growth factor‐D^ΔNΔC gene transfer induces angiogenesis in rabbit skeletal muscle