3D-bioprinting of sustained- and phased-release antibiotic and probiotic scaffolds to treat bacterial vaginosis

项目总结 细菌性阴道病(BV)是一种阴道微生物群失调，影响29%的育龄妇女。 并与不良妊娠结局、术后感染和性传播的高风险有关。 感染。虽然乳酸菌通常在健康的阴道中占主导地位，但BV的特点是乳酸菌水平较低。 以及各种厌氧细菌的过度生长，最常见的包括加德纳氏菌和普雷沃泰氏菌。当前口述 或局部抗生素治疗可以缓解80%的女性的症状，至少是暂时的。然而，大多数女性 会在治疗后一年内复发。最近的一项临床研究表明，一个阴道 阴道抗生素疗程后使用的益生菌治疗方案(含脆乳杆菌) (甲硝唑)，显著提高远期疗效。不幸的是，电流是局部应用的 配方需要重复给药(每天一到两次)，这会阻碍女性的便利性和 坚持治疗，特别是在接受数周治疗时(需要同时进行两种治疗 抗生素和益生菌)。 在这个项目中，我们将使用3D打印和计算建模，通过功能研究反复启用 体外和体内的原型支架，以设计维持治疗输送的长效产品，而 使抗生素和益生菌能够分阶段输送到女性生殖道。我们的团队聚集在一起 女性生殖道--在接生工具设计、计算建模和临床前方面的专业知识 BV车型。最终目标是使用3D生物打印来整合不同的设备隔间，这 依次释放针对厌氧过度生长的抗生素，然后释放恢复平衡的活益生菌 回到了对阴道乳杆菌的支持。在目标1中，我们将设计和表征3D打印硅胶支架， 维持仅限抗生素的给药。Aim 2将设计和评估3D打印硅胶和明胶海藻酸盐 依次释放抗生素和益生菌的复合体，提供了杀死BV的“1-2拳”策略 细菌，并提供一种“健康的”乳酸菌替代品。每个目标首先将重点放在以材料为基础的 含甲硫氨酸的有机硅(1A)或甲硫氨酸-有机硅益生菌-明胶海藻酸盐复合材料(2A)的表征。 来自体外释放实验的测量结果将用于开发和测试计算模型， 预测抗生素(1B)或双重制剂(2B)在“虚拟女性生殖道”中的输送，并最终在 一种小鼠混合感染模型。我们将评估甲硅氧烷(1C)和甲硅氧烷益生菌-明胶海藻酸盐 组合物(2C)对阴道上皮的细胞毒性和刺激可溶性促炎的能力 中介物和下游组织病理学。在我们的小鼠模型中，我们将测量存活的加德纳氏菌的水平 普雷沃泰尔菌在用空白或活化剂处理后从阴道和子宫组织中恢复- 包含3D打印的脚手架。如果成功，该项目将支持多用途平台的开发 预防和治疗BV以及其他女性生殖道应用。

PROJECT SUMMARY Bacterial vaginosis (BV) is a dysbiosis of the vaginal microbiome that affects ~29% of reproductive age women and is linked with higher risks of adverse pregnancy outcomes, postsurgical infections, and sexually transmitted infections. While lactobacilli typically dominate the healthy vagina, BV is characterized by low lactobacilli levels and an overgrowth of diverse anaerobic bacteria, most often including Gardnerella and Prevotella. Current oral or topical antibiotic treatments alleviate symptoms in 80% of women, at least temporarily. However, most women will experience a recurrence of BV within one-year post-treatment. A recent clinical study showed that a vaginal probiotic treatment regimen (with Lactobacillus crispatus), used following a vaginal course of antibiotics (metronidazole), significantly improved long-term treatment efficacy. Unfortunately, current topically-applied formulations require repeated administrations (once to twice daily), which can hinder female convenience and adherence to treatment, particularly when undergoing multiple weeks of treatment (required to administer both antibiotic and probiotics). In this project we will use 3D printing and computational modeling, iteratively enabled by functional investigation of prototype scaffolds in vitro and in vivo, to design long-acting products that sustain therapeutic delivery, while enabling phased-delivery of antibiotics and probiotics to the female reproductive tract. Our team brings together female reproductive tract-specific expertise in delivery vehicle design, computational modeling, and preclinical BV models. The ultimate goal is to use 3D-bioprinting to incorporate different device compartments, which sequentially release antibiotics that target anaerobic overgrowth, followed by live probiotics that restore balance back in favor of vaginal lactobacilli. In Aim 1, we will design and characterize 3D-printed silicone scaffolds that sustain antibiotic-only delivery. Aim 2 will design and evaluate 3D-printed silicone and gelatin alginate composites that sequentially release antibiotics followed by probiotics, providing a “1-2 punch” strategy to kill BV bacteria and provide a ‘healthy’ Lactobacillus alternative. Each aim will focus first on a materials-based characterization of Met-containing silicone (1A) or Met-silicone probiotic-gelatin alginate composites (2A). Measurements from in vitro release experiments will be used to develop and test computational models that predict the delivery of antibiotic (1B) or dual agents (2B) in a “virtual female reproductive tract” and ultimately in a mouse co-infection model. We will evaluate Met-silicone (1C) and Met-silicone probiotic-gelatin alginate composites (2C) for cytotoxicity to the vaginal epithelium and ability to stimulate soluble proinflammatory mediators and downstream histopathology. In our mouse model, we will measure levels of viable Gardnerella and Prevotella recovered from vaginal and uterine tissues following treatment with blank or active agent- containing 3D-printed scaffolds. If successful, this project will support the development of multipurpose platforms to prevent and treat BV as well as other female reproductive tract applications.