A simple, lanthanide-based method to enhance the transduction efficiency of adenovirus vectors

Gd, Palmer; Stoddart, Martin J.; Gouze, Elvire; Jn, Gouze; Ghivizzani, S. C.; Porter, Ryan M.; Ch, Evans

doi:10.1038/sj.gt.3303092

Cited by 23 publications

(15 citation statements)

References 26 publications

(24 reference statements)

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“…Because IκB acts intracellularly, comprehensive effects require its efficient delivery to nearly all cells in culture. Thus we first studied a similar adenoviral construct encoding the GFP reporter (Ad.GFP) to determine transduction efficiency under different conditions, using the lanthanum method (15) to overcome the otherwise modest susceptibility of MSCs to infection by adenovirus. As shown in Figure 2A, the three conditions selected produced between 60-100% GFP + cells (outer gates), with 10-100% of those cells fluorescing more intensely (inner gate).…”

Section: Resultsmentioning

confidence: 99%

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Interleukin‐1β and tumor necrosis factor α inhibit chondrogenesis by human mesenchymal stem cells through NF‐κB–dependent pathways

Wehling¹,

Palmer²,

Pilapil³

et al. 2009

Arthritis & Rheumatism

222

239

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Objective-The differentiation of mesenchymal stem cells (MSCs) into chondrocytes provides an attractive basis for the repair and regeneration of articular cartilage. Under clinical conditions, chondrogenesis will often need to occur in the presence of inflammatory mediators produced in response to injury or disease. Here we examine the effect of two important inflammatory cytokines, interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), on the chondrogenic behavior of human MSCs.Methods-Aggregate cultures of MSCs recovered from the femoral intermedullary canal were used. Chondrogenesis was assessed by the expression of relevant transcripts by quantitative RT-PCR and examination of aggregates by histology and immunohistochemistry. The possible involvement of NF-κB in mediating the effects of IL-1β was examined by delivering a luciferase reporter construct and a dominant negative inhibitor of NF-κB (srIκB), with adenovirus vectors.Results-Both IL-1β and TNF-α inhibited chondrogenesis in a dose-dependent manner. This was associated with a marked activation of NF-κB. Delivery of srIκB abrogated the activation of NF-κB and rescued the chondrogenic response. Although expression of type X collagen followed this pattern, other markers of hypertrophic differentiation responded differently. Matrix metalloproteinase-13 was induced by IL-1β in a NF-κB dependent fashion. Alkaline phosphatase activity, in contrast, was inhibited by IL-1β regardless of srIκB delivery.Conclusions-Cell-based repair of lesions in articular cartilage will be compromised in inflamed joints. Strategies for enabling repair under these conditions include the use of specific antagonists of individual pyrogens, such as IL-1 and TNF, or the targeting of important intracellular mediators, such as NF-κB. There are two general strategies to harnessing MSCs for this purpose. In a tissue engineering approach, MSCs are recovered from the patient and used to generate a graft that is subsequently implanted into the site of cartilage damage (4). Although the graft can be developed in a bioreactor into mature cartilage, there is increasing interest in grafting immature tissue, allowing chondrogenesis to occur in situ. The second strategy, which is already in wide clinical use, supplies MSCs to the defect by penetrating the subchondral bone, thereby allowing marrow to enter the lesion. Various related surgical techniques, including microfracture and subchondral drilling, are used for this purpose. These procedures have the convenience of being performed arthroscopically in large joints (1).Repair strategies that rely on the in situ differentiation of MSCs are attractive, but in many instances require chondrogenesis to take place within an inflamed environment. Intraarticular inflammation may result from disease, such as arthritis, or trauma, including the iatrogenic trauma of the cartilage repair surgery itself. Because interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) are major mediators of local inflammatory processes in joints, the present ...

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Section: Resultsmentioning

confidence: 99%

“…For experiments requiring high transduction efficiency, viral transduction was enhanced by co-precipitation with lanthanum (La) phosphates (15). Briefly, lanthanum chloride anhydrous salt (Sigma, St. Louis, MO) was dissolved in deionized (DI) water to a stock concentration of 0.4 M (pH 5.5-6.0).…”

Section: Methodsmentioning

confidence: 99%

Interleukin‐1β and tumor necrosis factor α inhibit chondrogenesis by human mesenchymal stem cells through NF‐κB–dependent pathways

Wehling¹,

Palmer²,

Pilapil³

et al. 2009

Arthritis & Rheumatism

222

239

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“…45 La 3+ was found to be superior to Gd 3+ , Y 3+ , Lu 3+ , and even Ca 2+ . Lanthanide oxide nanostructures are promising drug delivery materials due to their ease of synthesis and exceptionally low dimensionality.…”

Section: Drug/gene Deliverymentioning

confidence: 91%

Lanthanides: Applications in Cancer Diagnosis and Therapy

2016

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Lanthanide complexes are of increasing importance in cancer diagnosis and therapy, owing to the versatile chemical and magnetic properties of the lanthanide-ion 4f electronic configuration. Following the first implementation of gadolinium(III)-based contrast agents in magnetic resonance imaging in the 1980s, lanthanide-based small molecules and nanomaterials have been investigated as cytotoxic agents and inhibitors, in photodynamic therapy, radiation therapy, drug/gene delivery, biosensing, and bioimaging. As the potential utility of lanthanides in these areas continues to increase, this timely perspective of current applications will be useful to medicinal chemists and other investigators interested in the latest developments and trends in this emerging field.

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“…21,22 The lanthofection procedure was described previously by Palmer et al 23 Briefly, virus was diluted in 8 mL DMEM per 300 cm 2 flask to about 10,000 viral particles=cell (which is approximately 100 MOI), and LaCl 3 (Sigma-Aldrich GmbH, Buchs, Switzerland) was added dropwise to a final concentration of 100 mM. After 30 min of incubation the solution was added to the flasks containing 20 mL DMEM with 10% fetal bovine serum and incubated overnight.…”

Section: Msc Transductionmentioning

confidence: 99%

Improving Chondrogenesis: Potential and Limitations ofSOX9Gene Transfer and Mechanical Stimulation for Cartilage Tissue Engineering

Kupcsik

Stoddart

et al. 2010

Tissue Engineering Part A

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Articular cartilage injuries and degeneration affect a large proportion of the population in developed countries world wide. Stem cells can be differentiated into chondrocytes by adding transforming growth factor-beta1 and dexamethasone to a pellet culture, which are unfeasible for tissue engineering purposes. We attempted to achieve stable chondrogenesis without any requirement for exogenous growth factors. Human mesenchymal stem cells were transduced with an adenoviral vector containing the SRY-related HMG-box gene 9 (SOX9), and were cultured in a three-dimensional (3D) hydrogel scaffold composite. As an additional treatment, mechanical stimulation was applied in a custom-made bioreactor. SOX9 increased the expression level of its known target genes, as well as its cofactors: the long form of SOX5 and SOX6. However, it was unable to increase the synthesis of sulfated glycosaminoglycans (GAGs). Mechanical stimulation slightly enhanced collagen type X and increased lubricin expression. The combination of SOX9 and mechanical load boosted GAG synthesis as shown by (35)S incorporation. GAG production rate corresponded well with the amount of (endogenous) transforming growth factor-beta1. Finally, cartilage oligomeric matrix protein expression was increased by both treatments. These findings provide insight into the mechanotransduction of mesenchymal stem cells and demonstrate the potential of a transcription factor in stem cell therapy.

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A simple, lanthanide-based method to enhance the transduction efficiency of adenovirus vectors

Cited by 23 publications

References 26 publications

Interleukin‐1β and tumor necrosis factor α inhibit chondrogenesis by human mesenchymal stem cells through NF‐κB–dependent pathways

Interleukin‐1β and tumor necrosis factor α inhibit chondrogenesis by human mesenchymal stem cells through NF‐κB–dependent pathways

Lanthanides: Applications in Cancer Diagnosis and Therapy

Improving Chondrogenesis: Potential and Limitations ofSOX9Gene Transfer and Mechanical Stimulation for Cartilage Tissue Engineering

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