## 2 Results
### Antisepsis Effect of Pro10-1D Peptide on E. coli K1-induced Mouse Model of Septic Shock  
We next examined the potency of Pro10-1D as a candidate antiseptic peptide drug using an effective septic shock mouse model established using _E. coli_ K1. Pro10-1D reduced the number of bacteria in the mouse lung, liver, and kidney by approximately 73%, 87%, and 71%, respectively. This indicated that Pro10-1D can effectively inhibit bacterial growth in visceral organs of _E. coli_ K1-infected mice (Figure 12A). Furthermore, serum endotoxin measurement by the LAL test (Figure 12B) revealed that _E. coli_ K1-infected mice exhibited 100% endotoxin activation, whereas pretreatment with Pro10-1D effectively reduced the endotoxin levels by over 33%. We also quantified the inflammatory cytokine levels (TNF-\(\alpha\) and IL-6) using ELISA to determine the suppressive effect of Pro10-1D. As shown in Figure 12C,D, _E. coli_ K1-challenged mice exhibited significantly upregulated TNF-\(\alpha\) and IL-6 levels compared with the control. Treatment with Pro10-1D evidently suppressed TNF-\(\alpha\) and IL-6 levels by 45% and 56%, respectively, in serum and by 51% and 62%, respectively, in lung tissue of septic mice. Considering the role of sepsis in multiple-organ failure, we analyzed the serum levels of marker enzymes in the liver and kidney. As expected, _E. coli_ K1-infected mice displayed 100% enhanced AST, ALT, and BUN levels compared to the control. However, the mice pretreated with Pro10-1D showed significant reduction of the enzyme levels to 26% for AST, 48% for ALT, and 37% for BUN (Figure 12E). Pro10-1D Treatment Suppresses Polymorphonuclear Lymphocyte (PMN) Infiltration in E. coli K1-Induced Mouse Model  
Histological patterns of _E. coli_ K1-infected lung tissues displayed severe edema, pulmonary congestion, and alveolar hemorrhage, along with infiltrated PMNs in the lungs (Figure 13). These microanatomic features validate that increased PMNs are associated with lung damage, leading to sepsis severity upon _E. coli_ K1 infection. Pro10-1D effectively alleviated the pathological changes induced by _E. coli_ K1 in septic mice. Hence, we suggest that Pro10-1D can restore _E. coli_. Figure 11: Pro10-1D exerts nontoxic effects in vivo. The representative graph indicates (**A**) aspartate aminotransferase (AST), (**B**) alanine aminotransferase (ALT), and (**C**) blood urea nitrogen (BUN) levels of mice treated with Pro10-1D (1 mg/kg and 5 mg/Kg) for 24 h. Control mice only received phosphate-buffered saline. Data are presented as the mean \(\pm\) SEM (5 mice/group). Statistical analysis was performed by _one-way ANOVA ([PERSON]’s post hoc test;_ ns, not significant). Figure 12: Pro10-1D treatment alleviates septic shock in _E. coli_ K1-infected mice: (**A**) Inhibition of bacterial growth by Pro10-1D in the lung, liver, and kidney lysates of the septic mice. Mice were pretreated with 1 mg/kg of Pro10-1D (1 h) and _E. coli_ K1 (\(6\times 10^{6}\) Colony-Forming Unit (CFU)/mice, 16 h). (**B**) Pro10-1D reduces circulating endotoxin (LPS) levels in serum of septic mice. Effect of Pro10-1D on TNF-\(\alpha\) and IL-6 levels in the (**C**) serum, and (**D**) lung lysates of septic mice. (**E**) Pro10-1D normalizes the serum levels of (**E**) aspartate aminotransferase (AST), alanine aminotransferase (ALT), and blood urea nitrogen (BUN) in septic mice infected with _E. coli_ K1. The graphs indicate the mean \(\pm\) SEM (5 mice/group). \({}^{**}\)\(p<0.01\); \({}^{***}\)\(p<0.001\); ns, nonsignificant (by _one-way ANOVA_, _[PERSON]’s post hoc test_). Figure 13: Pro10-1D treatment ameliorates lung microanatomical changes in _E. coli_ K1-infected mice. Representative hematoxylin and eosin staining of lung tissue of (**A**) the control shows normal lung anatomy, while that of (**B**) _E. coli_ K1-infected mice (\(6\times 10^{6}\) CFU/mice, Intra-peritoneal injection (I.P.)) displays severe lung damage associated with neutrophil infiltration. (**C**) No sign of lung damage can be observed in the Pro10-1D-treated control mice (1 mg/Kg, I.P.