Autophagy Induction by a Small Molecule Inhibits
            <i>Salmonella</i>
            Survival in Macrophages and Mice

Nagy, Toni A.; Quintana, Joaquin L. J.; Reens, Abigail L.; Crooks, Amy L.; Detweiler, Corrella S.

doi:10.1128/aac.01536-19

Cited by 14 publications

(22 citation statements)

References 62 publications

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“…We used a fluorescence-based cell culture assay to identify compounds with non-traditional antimicrobial capabilities from chemical and drug libraries. This approach uncovered inhibitors of bacterial efflux pumps, a stimulator of macrophage autophagy and an antibiotic used against Mycobacterium species that was unexpectedly potent against Gram-negative bacteria during infection [21,22,25]. Within the current work, we examined the mechanism of action of a compound identified by SAFIRE, JD1, that strongly reduces S. Typhimurium replication in macrophages and disrupts the bacterial cell membrane upon breaching the outer membrane barrier.…”

Section: Discussionmentioning

confidence: 99%

“…Female C57Bl/67–8-week-old mice were intraperitoneally (IP) inoculated with 8 x 10 3 CFU, and the infectious dose was verified by plating for CFU. Mice were IP-treated with 100 μL of vehicle (70% DMSO) or 1 mg/kg of JD1 in 100 μL of vehicle at 10 minutes and 24 hours post-infection based on a previously established protocol [ 22 ]. Dosages were extrapolated from LDH toxicity and SAFIRE IC 50 assays, according to approved IACUC protocols [ 22 ].…”

Section: Methodsmentioning

confidence: 99%

“…This approach identified from the 14,400 compounds in the Maybridge HitFinder v11 library 58 small molecules that enable the killing of S. Typhimurium within macrophages but not in in standard microbiological media. Three compounds from this screen inhibit bacterial efflux pumps, and another stimulates autophagy in macrophages [20][21][22].…”

Section: Introductionmentioning

confidence: 99%

“…Typhimurium within macrophages but not in in standard microbiological media. Three compounds from this screen inhibit bacterial efflux pumps, and another stimulates autophagy in macrophages [ 20 – 22 ].…”

Section: Introductionmentioning

confidence: 99%

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A small molecule that mitigates bacterial infection disrupts Gram-negative cell membranes and is inhibited by cholesterol and neutral lipids

et al. 2020

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Infections caused by Gram-negative bacteria are difficult to fight because these pathogens exclude or expel many clinical antibiotics and host defense molecules. However, mammals have evolved a substantial immune arsenal that weakens pathogen defenses, suggesting the feasibility of developing therapies that work in concert with innate immunity to kill Gram-negative bacteria. Using chemical genetics, we recently identified a small molecule, JD1, that kills Salmonella enterica serovar Typhimurium (S. Typhimurium) residing within macrophages. JD1 is not antibacterial in standard microbiological media, but rapidly inhibits growth and curtails bacterial survival under broth conditions that compromise the outer membrane or reduce efflux pump activity. Using a combination of cellular indicators and super resolution microscopy, we found that JD1 damaged bacterial cytoplasmic membranes by increasing fluidity, disrupting barrier function, and causing the formation of membrane distortions. We quantified macrophage cell membrane integrity and mitochondrial membrane potential and found that disruption of eukaryotic cell membranes required approximately 30-fold more JD1 than was needed to kill bacteria in macrophages. Moreover, JD1 preferentially damaged liposomes with compositions similar to E. coli inner membranes versus mammalian cell membranes. Cholesterol, a component of mammalian cell membranes, was protective in the presence of neutral lipids. In mice, intraperitoneal administration of JD1 reduced tissue colonization by S. Typhimurium. These observations indicate that during infection, JD1 gains access to and disrupts the cytoplasmic membrane of Gram-negative bacteria, and that neutral lipids and cholesterol protect mammalian membranes from JD1-mediated damage. Thus, it may be possible to develop therapeutics that exploit host innate immunity to gain access to Gram-negative bacteria and then preferentially damage the bacterial cell membrane over host membranes.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A small molecule that mitigates bacterial infection disrupts Gram-negative cell membranes and is inhibited by cholesterol and neutral lipids

et al. 2020

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“…AR-12 microcapsules, prepared using a novel degradable acetalated dextran (Ac-Dex) biopolymer, further enhanced the clearance of intracellular Salmonella and significantly reduced drug toxicity (Hoang et al, 2014). In order to identify small molecules capable of interfering with Salmonella survival or replication in macrophages, Nagy et al (2019) screened the 14,400-compound Maybridge HitFinder v11 library in a high-content Screen for Anti-Infectives. They found that one small molecule, D61, reduced Salmonella load both in vitro and in vivo, through its autophagy-stimulating activity.…”

Section: Certain Drugs Regulate Xenophagy To Combat Salmonella Infectionmentioning

confidence: 99%

Salmonella Interacts With Autophagy to Offense or Defense

Shen

Zhang

et al. 2020

Front. Microbiol.

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Autophagy is an important component of the innate immune system in mammals. Low levels of basic autophagy are sustained in normal cells, to help with the clearance of aging organelles and misfolded proteins, thus maintaining their structural and functional stability. However, when cells are faced with challenges, such as starvation or pathogenic infection, their level of autophagy increases significantly. Salmonella is a facultative intracellular pathogen, which imposes an economic burden on the poultry farming industry and human public health. Previous studies have shown that Salmonella can induce the autophagy of cells following invasion, which to a certain extent helps to protect the cells from bacterial colonization. This review summarizes the latest research in the field of Salmonella-induced autophagy, including: (i) the autophagy induction and escape mechanisms employed by Salmonella during the infection of host cells; (ii) the effect of autophagy on intracellular Salmonella; (iii) the important autophagy adaptors that recognize intracellular Salmonella in host cells; and (iv) the effect of autophagy-modulating drugs on Salmonella infection.

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Lactobacillus plantarum postbiotics trigger AMPK‐dependent autophagy to suppress Salmonella intracellular infection and NLRP3 inflammasome activation

Wu¹,

Hu²,

Shu³

et al. 2023

Journal Cellular Physiology

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We previously found that Lactobacillus plantarum (LP)‐derived postbiotics protected animals against Salmonella infection, but the molecular mechanism remains obscure. This study clarified the mechanisms from the perspective of autophagy. Intestinal porcine epithelial cells (IPEC‐J2) were pretreated with LP‐derived postbiotics (the culture supernatant, LPC; or heat‐killed bacteria, LPB), and then challenged with Salmonella enterica Typhimurium (ST). Results showed that LP postbiotics markedly triggered autophagy under ST infection, as indicated by the increased LC3 and Beclin1 and the decreased p62 levels. Meanwhile, LP postbiotics (particularly LPC) exhibited a strong capacity of inhibiting ST adhesion, invasion and replication. Pretreatment with the autophagy inhibitor 3‐methyladenine (3‐MA) led to a significant decrease of autophagy and the aggravated infection, indicating the importance of autophagy in LP postbiotics‐mediated Salmonella elimination. LP postbiotics (especially LPB) significantly suppressed ST‐induced inflammation by modulating inflammatory cytokines (the increased interleukin (IL)‐4 and IL‐10, and decreased tumor necrosis factor‐α (TNF), IL‐1β, IL‐6 and IL‐18). Furthermore, LP postbiotics inhibited NOD‐like receptor protein 3 (NLRP3) inflammasome activation, as evidenced by the decreased levels of NLRP3, Caspase‐1 and apoptosis‐associated speck‐like protein containing a caspase recruitment domain (ASC). Deficits in autophagy resulted in an increase of inflammatory response and inflammasome activation. Finally, we found that both LPC and LPB triggered AMP‐activated protein kinase (AMPK) signaling pathway to induce autophagy, and this was further confirmed by AMPK RNA interference. The intracellular infection and NLRP3 inflammasome were aggravated after AMPK knockdown. In summary, LP postbiotics trigger AMPK‐mediated autophagy to suppress Salmonella intracellular infection and NLRP3 inflammasome in IPEC‐J2 cells. Our findings highlight the effectiveness of postbiotics, and provide a new strategy for preventing Salmonella infection.

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Autophagy Induction by a Small Molecule Inhibits Salmonella Survival in Macrophages and Mice

Cited by 14 publications

References 62 publications

A small molecule that mitigates bacterial infection disrupts Gram-negative cell membranes and is inhibited by cholesterol and neutral lipids

A small molecule that mitigates bacterial infection disrupts Gram-negative cell membranes and is inhibited by cholesterol and neutral lipids

Salmonella Interacts With Autophagy to Offense or Defense

Lactobacillus plantarum postbiotics trigger AMPK‐dependent autophagy to suppress Salmonella intracellular infection and NLRP3 inflammasome activation

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