Nanofiber-based Delivery of Combined Immune-modulating Compounds to Minimize Infection and Enhance Wound Healing

摘要 尽管努力保持手术室无菌，但手术部位感染（SSI）仍然是一个严重的问题， 我们的长期目标是开发新的治疗方法， 有效降低SSI风险并促进伤口愈合。在该奖项的前几个资助周期中， 我们已经证明i）维生素D和其他免疫抑制剂的成功封装和持续释放， 调节化合物诱导更高的凯萨林菌素抗菌肽（CAMP）基因和蛋白（hCAP 18/LL-1）， 37)细胞培养物中免疫细胞和角质形成细胞中的CAMP水平，人CAMP转基因小鼠伤口模型， 人皮肤外植体; ii）局部维生素D增加皮肤伤口感染中金黄色葡萄球菌的杀灭 iii）从用我们的人CAMP转基因小鼠处理的免疫细胞分泌的外来体; 维生素D含有更高水平的hCAP 18/LL-37。此外，我们还开发了一种新型的气体发泡膨胀剂 制造改进的3D支架的技术，洗脱促进免疫细胞浸润的维生素D， 诱导hCAP 18/LL-37，减少炎症，促进新生血管形成和胶原沉积， 人类免疫系统移植小鼠。基于这些发现和我们的初步数据，我们的目标是 该提案的重点是开发纳米纤维敷料，以增强先天免疫力。我们的- 免疫调节假说认为，共包封免疫调节化合物与从细胞分泌的外泌体， 3D支架中的经处理的免疫细胞将协同增强对SSI的保护，并促进 伤口愈合比单独使用任何一种成分都好。为了实现这一目标，有三个具体目标：1）恶魔- 从我们改进的3D层析中基质有效包封和洗脱免疫调节化合物 2）确定来源于引发的或人的外泌体的抗微生物和伤口愈合功效 CAMP转染的免疫细胞;和3）证明免疫调节化合物和外切- 一些-共同纳入3D支架，以促进愈合和预防感染，在我们的人源化的反式， 基因小鼠模型和离体人皮肤外植体。在我们先前赠款工作的基础上， 这一更新目标的完成将为下一代小说的发展奠定坚实的基础 治疗性抗感染伤口敷料，可以大大加快愈合，降低SSI的发生率， 抗生素耐药性的发展。我们还希望这些敷料可以作为有效的治疗方法， 创伤和战斗相关的伤害。

ABSRTACT Despite major efforts to keep operating rooms sterile, surgical site infections (SSIs) remain a serious and stub- born problem, killing up to 8,200 patients a year in the U.S. Our long-term goal is to develop novel therapies that effectively minimize risk of SSIs and promote wound healing. During the previous funding cycles of this award, we have demonstrated that i) successful encapsulation and sustained release of vitamin D and other immuno- modulating compounds induced higher cathelicidin antimicrobial peptide (CAMP) gene and protein (hCAP18/LL- 37) levels in immune cells and keratinocytes in cell culture, a human CAMP transgenic mouse wound model and human skin explants; ii) topical vitamin D increased killing of Staphylococcus aureus in a skin wound-infection model using our human CAMP transgenic mouse; and iii) exosomes secreted from immune cells treated with vitamin D contained higher levels of hCAP18/LL-37. Furthermore, we developed a novel gas-foaming expansion technique to fabricate improved 3D nanofiber scaffolds eluting vitamin D that promoted immune cell infiltration, induced hCAP18/LL-37, decreased inflammation, and promoted neovascularization and collagen deposition in human immune system-engrafted mice. Building on these findings, and our preliminary data, the goal of our proposal focuses on the development of nanofiber-based dressings for enhancing innate immunity. Our over- arching hypothesis is that co-encapsulating immunomodulating compounds with exosomes secreted from treated immune cells in 3D nanofiber scaffolds will synergistically enhance protection against SSIs and promote wound healing better than either component alone. To accomplish this, there are three specific aims: 1) Demon- strate efficient encapsulation and elution of immunomodulating compounds from our improved 3D nanofiber scaffolds; 2) Determine the antimicrobial and wound-healing efficacy of exosomes derived from primed or human CAMP transfected immune cells; and 3) Demonstrate the efficacy of immunomodulating compounds and exo- somes - co-incorporated 3D nanofiber scaffolds to promote healing and prevent infection in our humanized trans- genic mouse model and ex vivo human skin explants. Building on work from our prior grant, we expect successful completion of the aims in this renewal will lay a strong foundation for developing the next generation of novel therapeutic anti-infective wound dressings that could greatly speed healing, reduce rates of SSIs and minimize development of antibiotic resistance. We also expect these dressings could serve as effective treatments for traumatic and combat-related injuries.