Colon OCTN2 gene expression is up-regulated by peroxisome proliferator-activated receptor gamma in humans and mice and contributes to local and systemic carnitine homeostasis.

In the large intestine organic cation transporter type-2 (OCTN2) is recognized as a transporter of compounds such as carnitine and colony sporulation factor, promoting health of the colon intestinal epithelium. Recent reports suggest that OCTN2 expression in small intestine is under control of peroxisome proliferator-activated receptor-alpha (PPARalpha). However, PPARalpha contribution to colonic OCTN2 expression remains controversial. Here we examined the transcriptional regulation of colon OCTN2 gene by PPARgamma. To exclude any additional modulation of other PPAR to OCTN2 expression, we used both in vivo and in vitro PPAR-null models and specific PPAR inhibitors. The PPARgamma agonists thiazolidinediones increased both OCTN2 mRNA and protein expression in colonic epithelial cell lines independently by PPARalpha expression. The induction was blocked only by PPARgamma antagonists or by gammaORF4, a PPARgamma isoform with dominant negative activity, suggesting a PPARgamma-dependent mechanism. A conserved noncanonical PPAR-responsive element was found by computational analysis in the first intron of human OCTN2 gene and validated by EMSA assay. Promoter-reporter assays further confirmed transcriptional functionality of the putative PPAR response element, whereas selective mutation caused complete loss of responsiveness to PPARgamma activation. Finally, adenovirus-mediated overexpression of constitutively active PPARgamma mutant increased colon OCTN2 expression in PPARalpha(-/-) mice. Interestingly, animals overexpressing colon PPARgamma showed a significant increase in plasma carnitine, thus demonstrating the functional contribution of large intestine to systemic carnitine homeostasis. This study reveals a PPARgamma-dependent absorption machinery in colon that is likely involved in the health of colon epithelium, in the microbiota-host interactions and in the absorption of nutraceuticals and drugs.

Controlled delivery of the heparan sulfate/FGF-2 complex by a polyelectrolyte scaffold promotes maximal hMSC proliferation and differentiation.

Growth factors and other regulatory molecules are required to direct differentiation of bone marrow-derived human mesenchymal stem cells (hMSC) along specific lineages. However, the therapeutic use of growth factors is limited by their susceptibility to degradation, and the need to maintain prolonged local release of growth factor at levels sufficient to stimulate hMSC. The aim of this study was to investigate whether a device containing heparan sulfate (HS), which is a co-factor in growth factor-mediated cell proliferation and differentiation, could potentiate and prolong the delivery of fibroblast growth factor-2 (FGF-2) and thus enhance hMSC stimulation. To this aim, we synthesized cationic polyelectrolyte polymers covalently and non-covalently anchored to HS and evaluated their effect on hMSC proliferation. Polymers non-covalently bound to HS resulted in the release of an HS/FGF-2 complex rather than FGF-2 alone. The release of this complex significantly restored hMSC proliferation, which was abolished in serum-free medium and only partially restored by the release of FGF-2 alone as occurred with polymer covalently bound to HS. We also demonstrate that exposure to HS/FGF-2 during early growth but not during post-confluence is essential for hMSC differentiation down the fibroblast lineage, which suggests that both factors are required to establish the correct stem cell commitment that is necessary to support subsequent differentiation. In conclusion, the delivery platform described here is a step towards the development of a new class of biomaterial that enables the prolonged, non-covalent binding and controlled delivery of growth factors and cofactors without altering their potency.

Cationic copolymers nanoparticles for nonviral gene vectors: synthesis, characterization, and application in gene delivery.

The major aim of nonviral delivery systems for gene therapy is to mediate high levels of gene expression with low toxicity. Nowadays, one of the most successful synthetic polycations used in gene delivery research is poly(ethylenimine) (PEI) in its high-molecular weight (HMW) branched form. However, PEI is not the ideal transfection agent in vivo because of its overwhelming cytotoxicity. To overcome its toxic effects with a minimal impact on transfection efficiency, PEI has been conjugated with several nonionic biocompatible polymers. Here, we describe the synthesis of nanosized particles consisting of HMW PEI (25 kDa) crosslinked with poly(epsilon-caprolactone) (PCL, 50-60 kDa), a biodegradable aliphatic polyester. PCL was modified by the insertion of glycidyl groups able to condense with the amines of PEI to chemically bind PEI onto PCL. The nanoparticles obtained have been characterized in relation to their physicochemical and biological properties, and the results are extremely promising in terms of low cell toxicity and high transfection efficiency. These biological effects might be related to the peculiar DNA binding to covalently connected polymeric nanoparticles, without the formation of entangled DNA/polymer-soluble aggregates.