Recombinant human collagen-based microspheres mitigate cardiac conduction slowing induced by adipose tissue-derived stromal cells

Background Stem cell therapy to improve cardiac function after myocardial infarction is hampered by poor cell retention, while it may also increase the risk of arrhythmias by providing an arrhythmogenic substrate. We previously showed that porcine adipose tissue-derived-stromal cells (pASC) induce conduction slowing through paracrine actions, whereas rat ASC (rASC) and human ASC (hASC) induce conduction slowing by direct coupling. We postulate that biomaterial microspheres mitigate the conduction slowing influence of pASC by interacting with paracrine signaling. Aim To investigate the modulation of ASC-loaded recombinant human collagen-based microspheres, on the electrophysiological behavior of neonatal rat ventricular myocytes (NRVM). Method Unipolar extracellular electrograms, derived from microelectrode arrays (8x8 electrodes) containing NRVM, co-cultured with ASC or ASC loaded microspheres, were used to determine conduction velocity (CV) and conduction heterogeneity. Conditioned medium (Cme) of (co)cultures was used to assess paracrine mechanisms. Results Microspheres did not affect CV in control (NRVM) monolayers. In co-cultures of NRVM and rASC, hASC or pASC, CV was lower than in controls (14.4+/-1.0, 13.0+/-0.6 and 9.0+/-1.0 vs. 19.5+/-0.5 cm/s respectively, p Conclusion The application of recombinant human collagen-based microspheres mitigates indirect paracrine conduction slowing through interference with a secondary autocrine myocardial factor

Adipose Tissue-Derived Stromal Cells Inhibit TGF-beta 1-Induced Differentiation of Human Dermal Fibroblasts and Keloid Scar-Derived Fibroblasts in a Paracrine Fashion

Background: Adipose tissue-derived stromal cells augment wound healing and skin regeneration. It is unknown whether and how they can also influence dermal scarring. The authors hypothesized that adipose tissue-derived stromal cells inhibit adverse differentiation of dermal fibroblasts induced by the pivotal factor in scarring, namely, transforming growth factor (TGF)-beta. Methods: TGF-beta 1-treated adult human dermal fibroblasts and keloid scar-derived fibroblasts were incubated with adipose tissue-derived stromal cell-conditioned medium and assessed for proliferation and differentiation, particularly the production of collagen, expression of SM22 alpha, and development of hypertrophy and contractility. Results: TGF-beta 1-induced proliferation of adult human dermal fibroblasts was abolished by adipose tissue-derived stromal cell-conditioned medium. Simultaneously, the medium reduced SM22a gene and protein expression of TGF beta 1-treated adult human dermal fibroblasts, and their contractility was reduced also. Furthermore, the medium strongly reduced transcription of collagen I and III genes and their corresponding proteins. In contrast, it tipped the balance of matrix turnover to degradation through stimulating gene expression of matrix metalloproteinase (MMP)-1, MMP-2, and MMP-14, whereas MMP-2 activity was up-regulated also. Even in end-stage myofibroblasts (i.e., keloid scar-derived fibroblasts), adipose tissue-derived stromal cell-conditioned medium suppressed TGF-beta 1-induced myofibroblast contraction and collagen III gene expression. Conclusion: The authors show that adipose tissue-derived stromal cells inhibit TGF-beta 1-induced adverse differentiation and function of adult human dermal fibroblasts and TGF-beta 1-induced contraction in keloid scar-derived fibroblasts, in a paracrine fashion

In vivo behavior of trimethylene carbonate and ε‐caprolactone‐based (co)polymer networks: Degradation and tissue response

The in vivo erosion behavior of crosslinked (co)polymers based on trimethylene carbonate (TMC) and epsilon-caprolactone (CL) was investigated. High molecular weight poly(trimethylene carbonate) (PTMC) homopolymer- and copolymer films were crosslinked by gamma irradiation. To adjust the in vivo erosion rate of the (co)polymer films, both the irradiation dose (25, 50, or 100 kGy) for PTMC and composition (100-70 mol % TMC) at a constant irradiation dose of 25 kGy were varied. After subcutaneous implantation of irradiated films in rats, their in vivo behavior was evaluated qualitatively and quantitatively. When the irradiation dose for PTMC was increased from 25 to 100 kGy, the erosion rate of nonextracted PTMC films (determined at day 5) decreased from 39.7 +/- 16.0 mu m day(-1) to 15.1 +/- 2.5 mu m day(-1), and the number of lymphocytes in the tissue surrounding the films decreased from 235 +/- 114 cells mm(-2) to 64 +/- 33 cells mm(-2). The number of macrophages and giant cells at the tissue-polymer interface also decreased with increasing irradiation dose. All (co)polymer films eroded completely within 28 days of implantation. Variation of the TMC content of gamma irradiated (co)polymer films did not affect the tissue response to the gamma irradiated (co)polymer films and their in vivo erosion behavior much

Polyinosinic acid enhances delivery of adenovirus vectors in vivo by preventing sequestration in liver macrophages

Adenovirus is among the preferred vectors for gene therapy because of its superior in vivo gene-transfer efficiency. However, upon systemic administration, adenovirus is preferentially sequestered by the liver, resulting in reduced adenovirus-mediated transgene expression in targeted tissues. In the liver, Kupffer cells are responsible for adenovirus degradation and contribute to the inflammatory response. As scavenger receptors present on Kupffer cells are responsible for the elimination of blood-borne pathogens, we investigated the possible implication of these receptors in the clearance of the adenovirus vector. Polyinosinic acid [poly(I)], a scavenger receptor A ligand, was analysed for its capability to inhibit adenovirus uptake specifically in macrophages. In in vitro studies, the addition of poly(I) before virus infection resulted in a specific inhibition of adenovirus-induced gene expression in a J774 macrophage cell line and in primary Kupffer cells. In in vivo experiments, pre-administration of poly(I) caused a 10-fold transient increase in the number of adenovirus particles circulating in the blood. As a consequence, transgene expression levels measured in different tissues were enhanced (by 5- to 15-fold) compared with those in animals that did not receive poly(I). Finally, necrosis of Kupffer cells, which normally occurs as a consequence of systemic adenovirus administration, was prevented by the use of poly(I). No toxicity, as measured by liver-enzyme levels, was observed after poly(I) treatment. From our data, we conclude that poly(I) can prevent adenovirus sequestration by liver macrophages. These results imply that, by inhibiting adenovirus uptake by Kupffer cells, it is possible to reduce the dose of the viral vector to diminish the liver-toxicity effect and to improve the level of transgene expression in target tissues. In systemic gene-therapy applications, this will have great impact on the development of targeted adenoviral vectors

In vivo behavior of trimethylene carbonate and epsilon-caprolactone-based (co)polymer networks:Degradation and tissue response

The in vivo erosion behavior of crosslinked (co)polymers based on trimethylene carbonate (TMC) and epsilon-caprolactone (CL) was investigated. High molecular weight poly(trimethylene carbonate) (PTMC) homopolymer- and copolymer films were crosslinked by gamma irradiation. To adjust the in vivo erosion rate of the (co)polymer films, both the irradiation dose (25, 50, or 100 kGy) for PTMC and composition (100-70 mol % TMC) at a constant irradiation dose of 25 kGy were varied. After subcutaneous implantation of irradiated films in rats, their in vivo behavior was evaluated qualitatively and quantitatively. When the irradiation dose for PTMC was increased from 25 to 100 kGy, the erosion rate of nonextracted PTMC films (determined at day 5) decreased from 39.7 +/- 16.0 mu m day(-1) to 15.1 +/- 2.5 mu m day(-1), and the number of lymphocytes in the tissue surrounding the films decreased from 235 +/- 114 cells mm(-2) to 64 +/- 33 cells mm(-2). The number of macrophages and giant cells at the tissue-polymer interface also decreased with increasing irradiation dose. All (co)polymer films eroded completely within 28 days of implantation. Variation of the TMC content of gamma irradiated (co)polymer films did not affect the tissue response to the gamma irradiated (co)polymer films and their in vivo erosion behavior much. (C) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 95A: 940-949, 2010

Endotoxin contamination delays the foreign body reaction

Biomaterials are at continuous risk of bacterial contamination during production and application. In vivo, bacterial contamination of biomaterials delays the foreign body reaction (FBR). Endotoxins such as lipopolysaccharides (LPS), major constituents of the bacterial cell wall, are potent stimulators of the immune system in vitro and in vivo. In vitro, biomaterials contaminated with LPS induce the production of proinflammatory cytokines by adherent macrophages. This suggests that the presence of endotoxins on biomaterials will intensify the FBR. The effects of LPS on the course of the FBR have never been studied in vivo. In this study, the influence of LPS contamination on the FBR to subcutaneously implanted Puramatrix-loaded hexamethylenediisocyanate-crosslinked dermal sheep collagen (HDSC) disks was studied in rats. During the onset phase of the FBR, a massive influx of granulocytes was detected in LPS-contaminated disks, while their presence was prolonged. IL-10 production inside LPS-contaminated disks was increased at days 10 and 21. Macrophage densities were not affected by the presence of LPS. However, macrophage functionality was altered: giant cell formation and biomaterial degradation were delayed by LPS-contamination up to 21 days. On the basis of these results, we conclude that LPS delays the FBR. This finding indicates that endotoxin contamination has significant implications for the in vivo function of biomaterials and medical devices and emphasizes the importance of endotoxin testing. (C) 2011 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 98A: 527-534, 2011

Development of recombinant collagen-peptide-based vehicles for delivery of adipose-derived stromal cells

Stem cell therapy is a promising approach for repair, remodeling and even regenerate tissue of otherwise irreparable damage, such as after myocardial infarction (aMI). A severe limitation of cardiac stem cell therapy is the generally poor retention of administered cells in the target tissue. In tissue repair the main mode of action of adipose tissue-derived stem cells (ADSC) is the production of various growth factors, cytokines, anti-inflammatory and anti-apoptotic factors that together augment repair, remodeling, and regeneration. In this experiment, we used recombinant collagen peptide (RCP) with additional integrin-binding motives and different crosslinkers. Formulated as 50-100 mu m microspheres with bound ADSC, we hypothesized that this would improve ADSC retention and function. Crosslinking was performed with chemical crosslinkers (EDC and HMDIC) at high and low concentrations or by thermal treatment (DHT). ADSC adhesion, proliferation, apoptosis/necrosis, and gene expressions in two-dimensional and three-dimensional were analyzed. In addition, the effect of ADSC conditioned medium (ADSC-CM) on proapoptotic/sprouting HUVEC was examined. Our results show that all materials support cell adhesion in short time point, however, EDC-High crosslinker induced ADSC apoptosis/necrosis. Gene expression results revealed lower expression of proinflammatory genes in chemical crosslinked materials, despite EDC-High the proinflammatory genes expressions were similar or higher than TCPS. In addition, cultured ADSC on DHT crosslinked RCP showed a proinflammatory phenotype compared to TCPS. Sprouting assay results confirmed the protective effect of ADSC-CM derived from TCPS and HMDIC-High crosslinked RCP proapoptotic HUVEC. We conclude that ADSC adhere to the materials and maintain their therapeutic profile. (c) 2015 Wiley Periodicals, Inc