Sustained Release of Chondroitiase and Growth Factors from Chitosan Nanoparticles for Repairing the Hemisection Spinal Cord Injured Primate

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Project title

Code/計畫編號

NSC98-2320-B019-001-MY3

Translated Name/計畫中文名

靈長類動物急性暨慢性脊髓修復之合併療法---架接自體周邊神經，植入控制生長因子釋放之生醫材料，自體幹細胞移植與高劑量溶疤脢之療法---從幾丁聚醣奈米粒釋放溶疤脢及生長因子應用於脊髓半斷靈長類動物之研究

Project Coordinator/計畫主持人

Yi-Cheng Huang

Funding Organization/主管機關

National Science and Technology Council

Co-Investigator(s)/共同執行人

黃義侑

Department/Unit

Department of Food Science

Website

https://www.grb.gov.tw/search/planDetail?id=2223666

Year

2011

Start date/計畫起

01-08-2011

Expected Completion/計畫迄

01-07-2012

Bugetid/研究經費

1157千元

ResearchField/研究領域

醫學工程
藥學
生物技術(醫)

"在蛋白質傳輸系統中，可降解性的高分子材料常被用來包覆蛋白質，製備成傳輸系統所需的載體。然而，由於製備過程中有機溶劑的使用及劇烈的溫度、pH 值等反應條件，以致於過去的製備方式無法有效地被廣泛應用。為解決此問題，取而代之的是利用相反電荷離子間的作用力，形成載體攜帶蛋白質。在此研究中，我製備新穎的幾丁聚醣 /肝素及幾丁聚醣/褐藻糖膠奈米粒為載體。肝素及褐藻糖膠都是帶負電荷的聚醣類分子，可在溫和的反應條件下，藉由離子間的靜電作用力和帶正電的幾丁聚醣形成奈米粒。此外，保護蛋白質的活性也是持續釋放之傳輸系統能夠成功的重要因素。實驗中所使用的肝素不但在細胞生長及抗凝血等過程扮演重要的角色，也是纖維母細胞生長因子的鍵結分子。由褐藻中萃取之含有岩藻聚醣的褐藻糖膠，也會與肝素-鍵結生長因子(即纖維母細胞生長因子)結合。換言之，肝素與褐藻糖膠皆能穩定包覆於奈米粒的蛋白質。在此研究中，我們將探討纖維母細胞生長因子及溶疤酶的持續釋放，以促進脊髓損傷之靈長類動物的神經修復。纖維母細胞生長因子家族有多重功能，包括調節細胞的增生、分化、遷移及存活。酸性纖維母細胞生長因子及鹼性纖維母細胞生長因子是此家族中最早被發現的，其性質與功能也廣泛地被研究。已知酸性纖維母細胞生長因子由脊髓中的運動神經元及初級感覺神經元製造，於脊髓或腦部損傷後，已証實此生長因子能促進神經軸突的生長並減少神經元的凋亡。然而，對於此家族其他生長因子的研究卻相對較少。我們將探討由奈米粒中釋放各種纖維母細胞生長因子的效率、觀察這些生長因子在神經修復中所扮演的角色及探討生長因子與奈米粒之間的作用力。此外，我們也將持續釋放纖維母細胞生長因子於幹細胞培養基中，分析幹細胞的生長情形。在此計畫中，不僅是纖維母細胞生長因子的釋放，由奈米粒持續釋放溶疤酶也是研究重點。溶疤酶已被證實可分解疤痕組織中醣蛋白結構的醣鏈，有助於神經細胞與軸突的再生及動物行為的恢復。因此，在神經修復過程中給予溶疤酶治療是重要策略之一。此研究中，我們將完成由奈米粒持續釋放溶疤酶應用於神經修復的體外及體內(包括老鼠及靈長類)實驗，模擬臨床前人類損傷治療的可行性。" "Among the different approaches to achieve protein delivery, the use of biodegradable polymers holds great promise for preparing the macromolecular carrier systems. All the encapsulation methods could not be widely used in peptide and protein delivery because of organic solvents and/or extreme temperature or pH conditions. An alternative approach in these circumstances is the ionic cross-linking of polyelectrolytes or polyiones such as opposite charged polymers. In this study, novel nanoparticles were prepared by chitosan/heparin and chitosan/fucoidan. Heparin and fucoidan, both negative charged polysaccharides, could electrostatically interact with positive charged chitosan to form hydrophilic nanoparticles in mild condition. Furthermore, protecting the proteins from deactivation is important for sustained release successfully. Heparin is not only a pivotal role in processes such as blood coagulation and cell growth but also a well known fibroblast growth factors (FGFs) binging molecule. Fucoidans, vegetal fucose-containing polysaccharides extracted from brown algae, are also known to bind heparin-binding growth factors. In other words, both heparin and fucoidan can stabilize the proteins encapsulated in our nanoparticles. In this research, we will focus on sustained release of FGFs and chondroitinase (Ch ABC) to improve spinal cord recovery of injured primates. The fibroblast growth factors (FGFs) family has multiple functions including being potent modulators of cell proliferation, migration, differentiation, and survival. Acidic fibroblast growth factor (aFGF) and basic fibroblast growth factor (bFGF) were the first members of the family to be discovered, and their properties and functions have been extensively studied. Acidic FGF is normally produced in the spinal cord by motor neurons and primary sensory neurons, was demonstrated to promote axonal regeneration and reduce death of neurons after spinal cord or brain injury. However, there are fewer researches on the other growth factors in this family. We will study the release efficiency of FGFs from our nanoparticles, observing the roles of FGFs in nerve regeneration, and the specific interaction of FGFs and nanoparticles. Moreover, sustained release of FGFs in stem cells culture will also be analyzed in this research. Not only FGFs but also Ch ABC will sustain release from nanoparticles in this project. It has been proved that Ch ABC can digest the polysaccharide of proteoglycan in scar tissue, and promote axonal re-growth, neuron re-growth and animal behavior recovery. Therefore, treating Ch ABC is key strategy for improving nerve repair. Sustained release of Ch ABC from nanoparticles both in vitro and in vivo (for rats and primates) will be done is this project as a preclinical trial before clinical application."

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Sustained Release of Chondroitiase and Growth Factors from Chitosan Nanoparticles for Repairing the Hemisection Spinal Cord Injured Primate

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