Combination therapy of iPSC‐derived conditioned medium with ceftriaxone alleviates bacteria‐induced lung injury by targeting the NLRP3 inflammasome

Abstract

The lung is the first and most frequent organ to fail among sepsis patients. The mortality rate of sepsis-related acute lung injury (ALI) is high. Despite appropriate antimicrobial therapy, no treatment strategies are available for sepsis-induced ALI. Stem cell-mediated paracrine signaling is a potential treatment method for various diseases. This study aimed to examine the effects of induced pluripotent stem cell-derived conditioned medium (iPSC-CM) combined with antibiotics on ALI in a rat model of Escherichia coli-induced sepsis. Rats were administered either iPSC-CM or the vehicle (saline) with antibiotics (ceftriaxone). After 72 h, liquid biopsy, bronchoalveolar lavage fluid (BALF), and tissues were harvested for analysis. Survival rates were observed for up to 3 days. Furthermore, we examined the effects of iPSC-CM on cytokine production, metalloproteinase 9 (MMP-9) expression, and NLRP3-ASC interaction in RAW264.7 cells stimulated with lipopolysaccharide/interferon-γ (LPS/IFN-γ). Combined treatment of iPSC-CM with antibiotics significantly improved survival in E. coli-infected rats (p = 0.0006). iPSC-CM ameliorated E. coli-induced infiltration of macrophages, reducing the number of cells in BALF, and suppressing interleukin (IL)-1β, MIP-2, IL-6, and MMP-9 messenger RNA in lung sections. iPSC-CM treatment attenuated NLRP3 expression and inhibited NLRP3 inflammasome activation by disrupting NLRP3-mediated ASC complex formation in LPS/IFN-γ-primed RAW264.7 cells. This study reveals the mechanisms underlying iPSC-CMconferred anti-inflammatory activity in ALI through the attenuation of macrophage recruitment to the lung, thus inactivating NLRP3 inflammasomes in macrophages. iPSC-CM therapy may be a useful adjuvant treatment to reduce sepsis-related mortality by ameliorating ALI.

Salvianolic acid A alleviates lipopolysaccharide-induced disseminated intravascular coagulation by inhibiting complement activation

“…Therefore, the possible reason is that LPS is only a challenge for 3 h, and DIC is in the early stage, which can detect the biochemical indicators of renal and liver damage, but it has not yet reached the level of histopathological damage. The lung is the earliest and most easily damaged organ in the pathological process of sepsis, and patients with sepsis can have lung damage in the early stage of the disease course [46][47][48]. We investigated the effect of SAA on the lung histopathologic change in rats with LPS challenges.…”

“…The lung is the earliest and most easily damaged organ in the pathological process of sepsis, and patients with sepsis can have lung damage in the early stage of the disease course [46][47][48]. We investigated the effect of SAA on the lung histopathologic change in rats with LPS challenges.…”

P2X7 receptor blockade reduces pyroptotic inflammation and promotes phagocytosis in Vibrio vulnificus infection

“…High throughput imaging and analysis platform for quantification of LC3 + dots RAW264.7 cells were seeded in a 96 well black/clear bottom plate and then infected with V. vulnificus for 30 or 60 min. The cells were fixed, permeabilized, stained using an anti-LC3II antibody, and counterstained with 4',6-diamidino-2-phenylindole (DAPI), as previously described (Wann, Chi, Huang, Cheng, & Chang, 2022). Images were captured using an ImageXpress pico system (Molecular Devices).…”

“…Growth-arrested RAW264.7 cells were then infected with V. vulnificus at 37 degC for 1 and 2 h. The cells were washed, scraped, collected, and centrifuged at 45,000 x g at 4 degC for 1 h to prepare the whole-cell extract, as previously described (Wann et al, 2022). Membranes were incubated for 24 h with the primary antibody (1:1000) and then for 1 h with horseradish peroxidase (HRP)-conjugated anti-mouse or anti-rabbit secondary antibodies.…”

“…The following primer sets were used: mouse NLRP3 forward, 5'-TCA CAA CTC GCC CAA GGA A-3' and reverse, 5'-AAG AGA CCA CGG CAG AAG CTA G-3'; ASC forward, 5'-CTG CTC AGA GTA CAG CCA GAA C-3' and reverse, 5'-CTG TCC TTC AGT CAG CAC ACT G-3'; and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) forward, 5'-AAG AGG GAT GCT GCC CTT AC-3' and reverse, 5'-CCA TTT TGT CTA CGG GAC GA-3'. The relative expression levels of the different genes were determined according to the 2 -C t method (Wann et al, 2022).…”