[PMC free article] [PubMed] [CrossRef] [Google Scholar] 73. that mirrors the TG 100572 HCl spectrum of pneumococcal lung disease in patients, from moderate self-resolving contamination to severe TG 100572 HCl pneumonia refractory to antibiotics. A delay in antibiotic treatment resulted in ongoing inflammation and renal and hepatic dysfunction despite effective bacterial killing. The addition of fluid resuscitation to the model improved renal function but worsened the severity of lung injury based on direct measurements of pulmonary edema and lung compliance, analogous to patients with pneumonia and sepsis hSNFS who develop ARDS following fluid administration. (56). Patients recognized to have pneumonia are uniformly treated with broad-spectrum antibiotics within 1C2 h of presenting for medical care (25). However, much of the work done with bacterial pathogens in animal models has not included the use of antibiotics. Notably, treatment of severe pneumococcal infections with effective antibiotics releases large quantities of bacterial cell wall products over a short time and has been shown to produce a wave of inflammation that can worsen organ injury (41, 76, 77). As part of our research designed to model ARDS in mice and to study the effects of tobacco product exposure on acute infection-related lung injury, our goal was to develop a clinically relevant mouse TG 100572 HCl model of pneumococcal pneumonia. After critiquing the substantial literature on animal models of bacterial pneumonia, we found few manuscripts that provided methodological insights into model development to meet the specific needs of investigators. Herein we present our results in developing a mouse pneumococcal model incorporating antibiotic and fluid therapy that we hope will be helpful to other investigators studying sepsis, pneumonia, and ARDS in preclinical models. MATERIALS AND METHODS Animals Adult 8C10-wk-old female C57BL/6 mice were ordered from your National Malignancy Institute (Frederick, MD) and housed in pathogen-free housing. Female mice were used for regularity with prior experiments involving tobacco smoke, in which female mice were found to fight less during smoke exposure (21, 23). Furthermore, it was our goal to develop a reliable model that was strong to possible hormonal effects that have generally biased researchers away from the use of female animals. Animals were cared for in accordance with NIH guidelines by the Laboratory Animal Resource Center of the University or college of California, San Francisco, and all experiments were conducted under protocols approved by the University or college of California, San Francisco (UCSF) Institutional Animal Care and Make use of Committee. Group size was established to ensure sufficient statistical power predicated on our intensive encounter with the types of severe lung damage (16, 21, 38). INFECTION, Antibiotic Administration, and Microbiology serotype 19F [American Type Tradition Collection (ATCC) 49619, Manassas, VA] was expanded in brain-heart broth (Becton Dickinson 237500, Sparks, MD) and gathered in the mid-log stage [optical denseness (OD) 0.50 at 600 nm]. To boost reliability across tests, all cultures had been produced from aliquots of an individual bacterial expansion freezing at ?80C in 30% glycerol. Once OD 0.5 was reached, the bacterial tradition was spun at 4,000 revolutions/min (2,700 check or MannCWhitney check (when data weren’t normally distributed). Evaluations greater than two organizations were made out of ANOVA (accompanied by Tukeys or Dunnetts multiple evaluations testing) or KruskalCWallis (accompanied by Dunns multiple evaluations check). In instances of lacking data, a mixed-effects model was utilized. Survival evaluation was finished with log rank check with check for craze where suitable. 0.05 was considered to be significant statistically. Statistical evaluation and graph creation were finished with Prism (GraphPad, La Jolla, CA)..