Emerging antibiotic resistance is a major concern when treating microbial infections, and development of alternative therapies remains an urgent need.
9,10 In this regard, extracellular HMGB1, a member of a family of molecules referred to as DAMPS or alarmins, contributes to the pathogenesis of Pseudomonas keratitis,
15 as well as other infectious
13,38 and noninfectious diseases.
38,39 Extracellular HMGB1 is a late mediator of the inflammatory response,
12 in contrast to other proinflammatory molecules
12 (e.g., TNF-α, IL-1β, and IFN-α) that are released by activated immune cells early in the disease response. For example, in sepsis and endotoxemia, HMGB1 levels plateaued between 24 to 36 hours after infection. This wide therapeutic window
12 and the strong correlation between HMGB1 and the pathogenesis of various infectious diseases
13 suggest that it may provide an optimum target for treatment and clinical use. In fact, previous preclinical animal studies have reported various strategies to do precisely that. Unfortunately, many of those approaches have drawbacks that limit their use in a clinical setting.
38 Case in point, recently, this laboratory demonstrated the use of a small interfering RNA (siHMGB1) to knock down HMGB1 in a mouse model of
P. aeruginosa keratitis.
15 Treatment with siHMGB1 led to improved disease outcomes along with reduction in proinflammatory cytokines, an increase in anti-inflammatory cytokines and reduced neutrophil infiltration. For the current study, we used a higher bacterial concentration for the clinical isolate than was used in the above paper
15 where 1 × 10
6 CFU/μL was used for both the cytotoxic and the clinical isolate strains and silencing (scrambled treatment for controls siRNA for HMGB1) was our experimental approach to decrease HMGB1 levels. This proof of principle study established HMGB1 as a target for treatment. In the GLY studies reported herein, we used 1 × 10
6 CFU/μL of the clinical isolate in two separate preliminary experiments (data not shown) and found no significant difference between the PBS control (light opacity/infection) and GLY-treated mice. This led us to test an inoculum of 1 × 10
7 CFU/μL, which provided consistent statistically significant data for the clinical isolate with GLY treatment compared with PBS controls as shown in the manuscript.