Demidov V, Sottosanti J S, Demidova N, Bateman L M, Jackson O P, Craig P L, Hazem D, Gunn J R, Gitajn I L, Elliott J T
Dept. of Orthopaedics, Dartmouth Health, 1 Medical Center Dr, Lebanon, NH USA 03766.
Geisel School of Medicine at Dartmouth, 1 Rope Ferry Rd, Hanover, NH USA 03755.
Proc SPIE Int Soc Opt Eng. 2024 Jan-Feb;12822. doi: 10.1117/12.3000416. Epub 2024 Mar 12.
Well-organized ecosystems of bacteria colonize orthopaedic devices causing biofilm infections that are notoriously difficult to manage. Biofilms typically exhibit increased resistance to antibiotics leading to treatment failure, and tools for eradicating biofilms that do not increase antibiotic resistance are greatly needed. Antimicrobial photodynamic therapy (aPDT) is a promising form of treatment to combat clinically relevant biofilms. Exogenous provision of 5-aminolevulinic acid (5-ALA) to biofilm-forming clinical strains of E.coli, E. faecalis and S. aureus was recently shown by several research groups to result in the accumulation of sufficient quantities of endogenous photosensitizers porphyrins (protoporphyrin IX, coproporphyrin III and others), via the heme biosynthetic pathway, to produce a significant phototoxic effect when exposed to activating light. For clinical translation of this extremely promising approach, here we develop a portable light source for 5-ALA-based aPDT of orthopaedic implant biofilms, spectrally shaped for optimal porphyrin light absorption at wavelengths range approved by FDA for clinical use. After phantom calibration, we tested it on E.coli-E.faecalis biofilms grown in soft lithography-fabricated microfluidic chips and on methicillin-resistant S. aureus (MRSA) biofilms grown on titanium and stainless steel orthopaedic hardware in custom-designed macrofluidic devices. Successful experiments allowed us to conduct a proof-of-concept validation study in a preclinical rat model of MRSA-contaminated open fracture. Following tibia fracture and two hours of wound infection development, a one hour incubation with 20% 5-ALA and treatment with either 90J/cm or three fractions of 30J/cm light doses demonstrated 94% and 99% overall reduction of MRSA, respectively, while the temperature of the tissue remained <39°C, below the threshold for thermal damage. The encouraging results suggest further preclinical testing of the developed light source for optimization of aPDT regimen and 5-ALA concentration to reduce the risk of long-term side effects in animal models of contaminated trauma surgery.
组织良好的细菌生态系统会在骨科器械上定殖，引发生物膜感染，而这种感染 notoriously difficult to manage。生物膜通常对抗生素的耐药性增强，导致治疗失败，因此迫切需要能够根除生物膜且不会增加抗生素耐药性的工具。抗菌光动力疗法（aPDT）是一种有前景的治疗临床相关生物膜的方法。最近，几个研究小组发现，向形成生物膜的大肠杆菌、粪肠球菌和金黄色葡萄球菌临床菌株外源性提供5-氨基乙酰丙酸（5-ALA），会通过血红素生物合成途径导致内源性光敏剂卟啉（原卟啉IX、粪卟啉III等）积累到足够量，从而在暴露于激活光时产生显著的光毒性作用。为了将这种极具前景的方法转化为临床应用，我们在此开发了一种用于基于5-ALA的骨科植入物生物膜aPDT的便携式光源，其光谱形状经过优化，可在FDA批准的临床使用波长范围内实现卟啉的最佳光吸收。经过模型校准后，我们在软光刻制造的微流控芯片中生长的大肠杆菌-粪肠球菌生物膜以及定制设计的大流体装置中钛和不锈钢骨科硬件上生长的耐甲氧西林金黄色葡萄球菌（MRSA）生物膜上对其进行了测试。成功的实验使我们能够在MRSA污染的开放性骨折的临床前大鼠模型中进行概念验证性研究。在胫骨骨折并经过两小时伤口感染发展后，用20%的5-ALA孵育一小时，并用90J/cm或三个30J/cm光剂量的分次照射进行治疗，结果显示MRSA总体减少了94%和99%，而组织温度保持在<39°C，低于热损伤阈值。这些令人鼓舞的结果表明，需要对开发的光源进行进一步的临床前测试，以优化aPDT方案和5-ALA浓度，从而降低污染创伤手术动物模型中长期副作用的风险。
