A metal stent that could elute plasmid DNA (pDNA) in a controlled manner for substrate-mediated gene transfection was fabricated by first coating with hyaluronic acid (HA) and subsequent deposition of pDNA. To create robust HA coating layer on stainless steel (SS316L) surface, HA was derivatized with dopamine which is a well-known adsorptive molecule involving mussel adhesion process. The HA-coated surface was verified by various analytical techniques and proved to be very hydrophilic and stable, also showing superior biocompatibility in terms of suppressed plasma protein adsorption. For surface loading of pDNA, cationic pDNA/polyethylenimine (PEI) polyplexes were prepared and ionically adsorbed onto the HA-coated SS316L surface. The adsorbed surface exhibited evenly distributed nano-granular topography while the polyplexes maintained the nano-particular morphology. The pDNA was released out in a controlled manner for a period of 10 days with maintaining structural integrity. The dual coated substrate with HA and pDNA/PEI polyplexes exhibited greatly enhanced gene transfection efficiency, when compared to both bare substrate adsorbed with the polyplexes and PEI/pDNA polyelectrolyte multilayers. Dually functionalized stent with HA and pDNA exhibited effective biocompatibility and gene transfection.
Drug-eluting stent (DES) has been widely used for the effective treatment of obstructive coronary artery disease, preventing the occurrence of restenosis which was mainly caused by hyper-proliferation of smooth muscle cells. Here, we demonstrate the immobilization of heparin on the metal surface via a bio-inspired manner and subsequent build-up of therapeutic lay-by-layer multilayer assembled from paclitaxel (PTX) encapsulated poly(lactic-co-glycolic acid) grafted hyaluronic acid (HA-g-PLGA) micelles and poly-L-lysine (PLL). It was hypothesized that the heparinized metallic surface would create a non-thrombogenic environment, while controlled release of PTX from the surface could induce anti-proliferation of smooth muscle cells. For the surface immobilization of heparin on the surface of cobalt-chromium alloy (L605), dopamine-derivatized heparin was synthesized and anchored on the surface by a mussel-inspired adhesion mechanism. An amphiphilic graft copolymer of HA-g-PLGA was synthesized and utilized for the formation of anionic PTX loaded micelles. A PTX eluting multilayer composed of anionic HA-g-PLGA micelles and PLL was self-assembled on the metal surface by a layer-by-layer (LbL) fashion. Loading amount of PTX on the metal surface could be readily controlled with concomitantly achieving sustained release profiles of PTX over an extended period. The proliferation of human coronary artery smooth muscle cells (CASMCs) were successfully arrested by the resulting PTX releasing multilayer coating on the L605 substrate.
Biodegradable polymeric nanocylinders were fabricated by segmental degradation of electro-spun nanofibers. Poly(L-lactic acid) (PLA) was electro-spun to produce non-crystalline nanofibers, which were immediately treated with amino group-containing strong bases to fabricate semi-crystalline PLA nanocylinders with tunable aspect ratio. The formation of PLA nanocylinders was attributed to two concurrent events occurring during the aminolysis reaction: i) development of stacked transverse lamellae and ii) transversely oriented degradation and fragmentation of the amorphous gaps between the lamellae, both responsible for the periodic fragmentation of PLA nanofibers into uniformly shaped nanocylinders. The aspect ratio of PLA nanocylinders was readily tunable by varying aminolysis time and controlling nanofiber diameter. This new class of biocompatible and biodegradable PLA nanocylinders can be potentially utilized for many biomedical applications.
The critical essentials of soft tissue engineering are to overcome the cell source shortage and to achieve long-term maintenance of regenerated tissue contour. Thus, the utilization of stem cells and the vascularization of regenerated tissues can be the potential solution for clinical success of soft tissue reconstruction. We prepared the multicellular spheroids comprised of mesenchymal stem cells (MSCs) and synthetic bio-mimicking nanofilaments. The nanofilaments were proved to facilitate the adipogenesis of MSCs in the spheroid culture system. This bio-functionally optimized hybrid spheroid was used as a building block for three dimensional bio-hybrid construction with assistance of microstructured-framework fabricated by direct polymer melt deposition process. At the same time, the angiogenic growth factor was delivered from the surface of the framework. As a result, the multi-scaled and multi-functional hybrid construct with each scaled structure providing a different function enabled the successful formation of vascularized adipose tissue in vivo.
약물전달용 스텐트 (drug eluting stent) 는 심근세포의 지나친 증식으로 발생되는 restenosis를 방지하기 위하여 효과적인 치료수단으로 많이 사용되어 왔다. 이 연구에서는 restenosis 억제를 목적으로 파클리탁셀 (paclitaxel) 또는 플라스미드 DNA (pDNA) 를 금속 스텐트 표면으로부터 제어 방출할 수 있도록 하는 표면 디자인에 대하여 설명한다. 첫째로, pDNA를 전달하기 위하여 생체 고분자인 하이알루로닉 산으로 표면 개질을 시도하였는데, 이때 안정된 표면 고정을 위하여 도파민으로 수식된 하이알루로닉 산을 사용하였다. 도파민은 홍합의 접착 단백질로부터 유도된 성분으로 홍합의 접착력과 같은 강한 표면 부착력을 제공한다. 이렇게 개질된 표면에 pDNA/PEI polyplexes를 정전기적 인력으로 표면 흡착시켰을 때 세포내부로 뛰어난 pDNA 전달능력을 보여주었다. 또한, paclitaxel의 표면 고정을 위하여 먼저 도파민으로 수식된 헤파린을 금속 스텐트 표면에 고정하고 마이셀 형태의paclitaxel을 layer-by-layer 방식으로 표면에 흡착시켰다. Layer-by-layer로 적층하는 방식은 paclitaxel의 용량과 그 방출 패턴을 쉽게 제어할 수 있을 뿐 아니라, 헤파린과 같은 생체고분자를 동시에 방출시킬 수 있는 장점을 가진다. 결과적으로 이렇게 표면 개질된 스텐트는 심근세포의 성장을 효과적으로 저해시킬 수 있음이 관찰되었다.
생체 적합 고분자 구조체의 표면 개질은 세포의 접착, 증식, 분화 등의 세포거동을 제어할 수 있는 효과적인 수단을 제공한다. 특히 나노구조의 고분자 구조체는 세포의 거동을 제어함에 있어 더 뛰어난 특징을 보여주며, 물리적 구조와 화학적 표면특성의 적절한 조합으로 세포의 최종적 운명을 결정할 수 있다. 이에 본 연구에서는 전기방사된 나노섬유를 분해하여 일차원 나노필라멘트를 제작하고 이를 mesenchymal stem cell (MSC)과 결합시켜 스페로이드 형태의 마이크로 구조체를 만들었다. 이런 하이브리드 구조체는 내부의 나노필라멘트의 양에 따라 세포 분화의 제어를 가능하게 하며 뛰어난 분화 능력을 가짐이 확인되었다. 분화능력에 있어 최적화된 하이브리드 스페로이드를 쾌속 조형방식으로 제작된 마이크로 구조체를 이용하여 3차원적으로 조립하였다. 동시에 마이크로 구조체 표면을 신생혈관재생을 위한 bFGF로 수식하였다. 최종적으로 만들어진 3차원 다기능성 하이브리드 구조체는 in vivo실험을 통하여 혈관이 재생된 지방조직을 형성시킬 수 있음을 확인하였다.