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Knockdown of intraislet IKK? by spherical nucleic acid conjugates prevents cytokine-induced injury and enhances graft survival.

ABSTRACT: BACKGROUND:The efficiency of islet graft survival after intraportal implantation is compromised by host innate immune responses and the production of proinflammatory cytokines that cause acute cellular injury. This reaction activates intraislet nuclear factor-?B (NF-?B), causing production of gene products that have detrimental effects on ?-cell survival and function. We hypothesized that small interfering RNA targeting of IKK?, a crucial kinase in the NF-?B activation pathway, in islets before transplantation would ameliorate the detrimental effects of cytokines and improve islet survival after transplantation. METHODS:To test this hypothesis, we prepared small interfering RNA-based spherical nucleic acid nanoparticle conjugates targeting IKK? IKK? SNA-NCs). We treated isolated islets with IKK? SNA-NCs and assessed the functional consequences of IKK? knockdown in vitro and after intraportal transplantation in mice. RESULTS:Treatment of freshly isolated mouse islets with IKK? SNA-NCs reduced constitutive IKK? expression and protected against proinflammatory cytokine-induced NF-?B activation, resulting in improved cell viability and decreased expression of gene products associated with ?-cell dysfunction. Intraportal transplantation of a marginal mass (50 islets) of syngeneic islets treated with nanoparticle conjugates targeting IKK? resulted in reversion to normoglycemia in 50% of streptozotocin-induced diabetic recipients (n=12) compared with 0% of controls (n=12). Histologic analyses showed reduced CD11b(+) cellular infiltration and decreased islet apoptosis. CONCLUSIONS:These results are consistent with the hypothesis that inhibition of intraislet NF-?B activation ameliorates the detrimental effects of host cytokines and demonstrates that preconditioning freshly isolated islets in culture with IKK? SNA-NCs may be a promising therapy to enhance islet graft function and survival after transplantation.

SUBMITTER: Rink JS

PROVIDER: S-EPMC3839058 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Knockdown of intraislet IKKβ by spherical nucleic acid conjugates prevents cytokine-induced injury and enhances graft survival.

Rink Jonathan S JS McMahon Kaylin M KM Zhang Xiaomin X Chen Xiaojuan X Mirkin Chad A CA Thaxton C Shad CS Kaufman Dixon B DB

Transplantation 20131101 10

<h4>Background</h4>The efficiency of islet graft survival after intraportal implantation is compromised by host innate immune responses and the production of proinflammatory cytokines that cause acute cellular injury. This reaction activates intraislet nuclear factor-κB (NF-κB), causing production of gene products that have detrimental effects on β-cell survival and function. We hypothesized that small interfering RNA targeting of IKKβ, a crucial kinase in the NF-κB activation pathway, in islets ...[more]

PMID: 24247900

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Project description:Spherical nucleic acid (SNA) nanoparticle conjugates are a class of bionanomaterials that are extremely potent in many biomedical applications. Their unique ability to enter multiple mammalian cell types as single-entity agents arises from their novel three-dimensional architecture, which consists of a dense shell of highly oriented oligonucleotides chemically attached typically to a gold nanoparticle core. This architecture allows SNAs to engage certain cell surface receptors to facilitate entry. Here, we report studies aimed at determining the intracellular fate of SNAs and the trafficking events that occur inside C166 mouse endothelial cells after cellular entry. We show that SNAs traffic through the endocytic pathway into late endosomes and reside there for up to 24 h after incubation. Disassembly of oligonucleotides from the nanoparticle core is observed 16 h after cellular entry, most likely due to degradation by enzymes such as DNase II localized in late endosomes. Our observations point to these events being likely independent of core composition and treatment conditions, and they do not seem to be particularly dependent upon oligonucleotide sequence. Significantly and surprisingly, the SNAs do not enter the lysosomes under the conditions studied. To independently track the fate of the particle core and the fluorophore-labeled oligonucleotides that comprise its shell, we synthesized a novel class of quantum dot SNAs to determine that as the SNA structures are broken down over the 24 h time course of the experiment, the oligonucleotide fragments are recycled out of the cell while the nanoparticle core is not. This mechanistic insight points to the importance of designing and synthesizing next-generation SNAs that can bypass the degradation bottleneck imposed by their residency in late endosomes, and it also suggests that such structures might be extremely useful for endosomal signaling pathways by engaging receptors that are localized within the endosome.

Dataset Information

Knockdown of intraislet IKK? by spherical nucleic acid conjugates prevents cytokine-induced injury and enhances graft survival.

Publications

Knockdown of intraislet IKKβ by spherical nucleic acid conjugates prevents cytokine-induced injury and enhances graft survival.

Similar Datasets

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