Analysis of health and lung injury markers in mice treated with TLR4-interacting SPA4 peptide

Abstract

Abstract The SPA4 peptide (GDFRYSDGTPVNYTNWYRGE) interacts with TLR4, affects intracellular signaling, improves host defense, and alleviates tissue injury in mouse models of bacterial lung infection and lipopolysaccharide-induced lung inflammation. In this study, we investigated the effect of different doses of SPA4 peptide on health, and systemic and lung injury parameters in outbred CD-1 mice. Mice were intratracheally administered with 0.625, 1.25, 2.5, 5, and 10 μg SPA4 peptide per g body weight every day for three days. Mice left untreated or those treated with equivalent volume of vehicle were included as controls. Signs of sickness and body weights were monitored every 24 h; food and water were provided ad libitum. After 24 h of last intratracheal instillation, mice were euthanized, and blood and major organs were collected. Blood gas and electrolytes (pCO2, total CO2, pO2, pH, anion gap, base excess, total hemoglobin, Na+, K+, Cl−, HCO3−) were analyzed, and markers of lung injury (lung wet/dry weight ratio, and levels of albumin, lactate, and total protein) were determined in lung tissue homogenates. Major organs (stomach, intestines, lung, liver, spleen, kidney, heart, and thymus) were fixed, sectioned, stained with hematoxylin and eosin, and examined for tissue damage or inflammatory response. No signs of sickness or changes in body weight were noted. Blood gas and electrolyte parameters were not significantly altered in SPA4 peptide-treated mice. The lung injury parameters were also not significantly different in SPA4 peptide-treated mice. Histological examination revealed no sign of tissue injury or inflammation in any of the major organs. Together, these results provide an initial safety assessment of the SPA4 peptide.