A Non-invasive and Technically Non-intensive Method for Induction and Phenotyping of Experimental Bacterial Pneumonia in Mice

Abstract

Although community-acquired pneumonia remains a major public health problem, murine models of bacterial pneumonia have recently facilitated significant preclinical advances in our understanding of the underlying cellular and molecular pathogenesis. In vivo mouse models capture the integrated physiology and resilience of the host defense response in a manner not revealed by alternative, simplified ex vivo approaches. Several methods have been described in the literature for intrapulmonary inoculation of bacteria in mice, including aerosolization, intranasal delivery, peroral endotracheal cannulation under 'blind' and visualized conditions, and transcutaneous endotracheal cannulation. All methods have relative merits and limitations. Herein, we describe in detail a non-invasive, technically non-intensive, inexpensive, and rapid method for intratracheal delivery of bacteria that involves aspiration (i.e., inhalation) by the mouse of an infectious inoculum pipetted into the oropharynx while under general anesthesia. This method can be used for pulmonary delivery of a wide variety of non-caustic biological and chemical agents, and is relatively easy to learn, even for laboratories with minimal prior experience with pulmonary procedures. In addition to describing the aspiration pneumonia method, we also provide step-by-step procedures for assaying the subsequent in vivo pulmonary innate immune response of the mouse, in particular, methods for quantifying bacterial clearance and the cellular immune response of the infected airway. This integrated and simple approach to pneumonia assessment allows for rapid and robust evaluation of the effect of genetic and environmental manipulations upon pulmonary innate immunity.

Effects of Orally Ingested Arsenic on Respiratory Epithelial Permeability to Bacteria and Small Molecules in Mice

“…Despite this, mice from both arsenic groups had markedly increased bacterial counts in blood and spleen ( Figure 1f ), indicating increased extrapulmonary bacterial dissemination. Consistent with our prior findings ( Madenspacher and Fessler 2016 ), a substantial number of mice had no detectable extrapulmonary bacterial dissemination 24 h post-infection with K. pneumoniae , consistent with an as-yet unbreached alveolocapillary barrier. Consistent with the accelerated escape of bacteria from the arsenic-exposed lung, bacteremia was detected in 8/29 (0), 18/29 ( ), and 23/29 ( ) mice, and splenic bacteria were detected in 18/29 (0), 24/29 ( ), and 25/27 ( ) mice.…”

“…Klebsiella pneumoniae [ATCC 43816; 2,000 CFU (colony-forming units)] or Streptococcus pneumoniae (ATCC 6303; CFU) were delivered to the lung via oropharyngeal aspiration while the mice were under 4% isoflurane anesthesia, as previously reported ( Madenspacher and Fessler 2016 ). In other experiments, K. pneumoniae (960, 6,200, or 92,000 CFU) was injected intravenously (i.v.…”

Tristetraprolin Overexpression in Non-hematopoietic Cells Protects Against Acute Lung Injury in Mice

“…Both male and female adult (8–10-week-old) mice were used for experiments. Mice were anesthetized with isoflurane/oxygen followed by administration of 10 μg Lipopolysaccharide (LPS) from Escherichia coli O111:B4 (L4391, Sigma-Aldrich) per mouse dissolved in sterile endotoxin-free saline (50 μl total volume), or an equivalent volume of sterile endotoxin-free saline as a vehicle control, via oropharyngeal aspiration ( 18 ). Mice were observed for signs of distress including anorexia, weight loss, hunched posture, ruffled haircoat, labored breathing, and dehydration every 8–12 h post LPS challenge.…”

Intrapulmonary Treatment With a Novel Tlr4 Agonist Confers Protection Against Klebsiella Pneumonia

“…Mice were anesthetized using 4% isoflurane for 3 min and suspended by front incisors onto a treatment platform, tongue withdrawn with forceps, and the treatment solution or inoculum was deposited above the vocal cords (14). Aspiration of vehicle, study drug, or bacterial inoculum was confirmed by direct visualization.…”