Synthesis and Formulation of Four-Arm PolyDMAEA-siRNA Polyplex for Transient Downregulation of Collagen Type III Gene Expression in TGF-β1 Stimulated Tenocyte Culture
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The natural healing process for tendon repair is associated with high upregulation of collagen type III, leading to scar tissue and tendon adhesions with functionally deficient tendons. Gene delivery systems are widely reported as potential nanotherapeutics to treat diseases, providing a promising approach to modulate collagen type III synthesis. This work investigates a proof-of-concept four-arm cationic polymer-siRNA polyplex to mediate a transient downregulation of collagen type III expression in a tendon cell culture system. The tendon culture system was first supplemented with TGF-β1 to stimulate the upregulation of collagen type III prior to silencing experiments. The four-arm poly[2-(dimethylamino) ethyl acrylate] (PDMAEA) polymer was successfully synthesized via RAFT polymerization and then mixed with siRNA to formulate the PDMAEA-siRNA polyplexes. The formation of the polyplex was optimized for the N:P ratio (10:1) and confirmed by agarose gel electrophoresis. The size and solution behavior of the polyplex were analyzed by dynamic light scattering and zeta potential, showing a hydrodynamic diameter of 155 ± 21 nm and overall positive charge of +30 mV at physiological pH. All the polyplex concentrations used had a minimal effect on the metabolic activity of cultured cells, indicating good biocompatibility. The dose and time effects of the TGF-β1 on collagen type III gene expressions were analyzed by qPCR, showing an optimal dose of 10 ng mL-1 TGF-β1 and 3-fold increase of COL3α1 expression at 48 h in cultured tenocytes. The PDMAEA-siRNA polyplex concept observed a limited yet successful and promising efficiency in silencing collagen type III at 48 h compared to PEI-siRNA. Therefore, this concept is a promising approach to reduce tissue scarring and adhesion following injuries.Keywords:
Type I collagen
Biocompatibility
Biocompatibility is a very common word that is used within biomaterial science and used for description of the interactions between the foreign material and the body. However, the meaning of biocompatibility as well as the mechanisms that collectively constitutes is still unclear. With the advance of nanotechnology, new concerns have been observed related to biocompatibility of these biomaterials. Due to their small size and variability of their physical and chemical properties, nanoparticles' (NP) distribution within the body and interactions with the target cells and tissues are highly variable. Here, we tried to provide an overview about NPs, the concept of biocompatibility and biocompatibility-related issues in nanomedicine and several different NPs.
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Fibrosis caused by the host response to long-term transplanted microcapsules and the limitation of traditional L929 cell model for biocompatibility testing inspire the development of an assay of biocompatibility based on macrophage behavior. In this paper, the human monocytic cell line THP-1 was utilized for biocompatibility evaluation of microcapsule materials. The cell viability and secretion of nitric oxide (NO) and cytokines served as index of biocompatibility were assayed. It was found that the evaluated microcapsule materials had no effect on the stimulation of NO and cytokines secretion, which meant that these materials were biocompatible. Furthermore, it suggests the THP-1 cell a convenient in vitro experimental model that might be useful for long-term predictions of material biocompatibility.
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This study investigated the effect of transcriptional gene silencing (TGS) of the heparanase gene on hepatoma SMCC-7721 cells.SiRNAs targeting the promoter region and coding region of the heparanase gene were designed and synthesized. Then the siRNAs were transfected into hepatoma SMCC-7721 cells by nuclear transfection or cytoplasmic transfection. The expression of heparanase was detected by RT-PCR and Western blotting 48 h, 72 h and 96 h post-transfection. In addition, wound healing and invasion assays were performed to estimate the effect of TGS of the heparanase gene on the migration and invasion of hepatoma SMCC-7721 cells.Protein and mRNA expression of the heparanase gene were interfered with by TGS or post-transcriptional gene silencing (PTGS) 48 h after transfection. At 72 h post-transfection, the expression of the PTGS group of genes had recovered unlike the TGS group. At 96 h post-transfection, the expression of the heparanase gene had recovered in both the TGS group and PTGS group. Invasion and wound healing assays showed that both TGS and PTGS of the heparanase gene could inhibit invasion and migration of hepatoma SMCC-7721 cells, especially the TGS group.TGS can effectively interfere with the heparanase gene to reduce the invasion and migration of hepatoma SMCC-7721 cells.
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HeLa
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Polyurethane(PU) has been widely used biomedical material with good stability,excellent mechanical properties and reasonable biocompatibility due to its specific micro-phase separation structure.However,its unsatisfactory blood compatibility results in limitation of application in the biomaterial fields.As far as the dependence of biocompatibility on surface properties of materials is concerned,surface modification has been recognized a preferable way to improve the biocompatibility for biomaterials.In view of many methods applied to modify the surface of polyurethane,the bioactive molecules modified surface through chemical modification has attracted a great of interest.In the present article the approaches of modification to improve the biocompatibility of polyurethane were briefly summarized,and the relationship between polyurethane surface properties and biocompatibility was also discussed.
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The mechanical properties of the human supraspinatus tendon (SST) are highly heterogeneous and may reflect an important adaptive response to its complex, multiaxial loading environment. However, these functional properties are associated with a location-dependent structure and composition that have not been fully elucidated. Therefore, the objective of this study was to determine the concentrations of types I, II and III collagen in six distinct regions of the SST and compare changes in collagen concentration across regions with local changes in mechanical properties. We hypothesized that type I collagen content would be high throughout the tendon, type II collagen would be restricted to regions of compressive loading and type III collagen content would be high in regions associated with damage. We further hypothesized that regions of high type III collagen content would correspond to regions with low tensile modulus and a low degree of collagen alignment. Although type III collagen content was not significantly higher in regions that are frequently damaged, all other hypotheses were supported by our results. In particular, type II collagen content was highest near the insertion while type III collagen was inversely correlated with tendon modulus and collagen alignment. The measured increase in type II collagen under the coracoacromial arch provides evidence of adaptation to compressive loading in the SST. Moreover, the structure-function relationship between type III collagen content and tendon mechanics established in this study demonstrates a mechanism for altered mechanical properties in pathological tendons and provides a guideline for identifying therapeutic targets and pathology-specific biomarkers.
Type I collagen
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