Phages as immunomodulators and their promising use as anti-inflammatory agents in a cftr loss-of-function zebrafish model

Abstract

Cystic Fibrosis (CF), one of the most frequent hereditary disease due to mutations in the CFTR gene, causes mortality in humans mainly due to infection in the respiratory system. However, besides the massive inflammatory response triggered by chronic bacterial infections, a constitutive pro-inflammatory state associated with the most common CFTR mutations has been reported in paediatric cases before the onset of bacterial colonization. In a previous works we isolated and characterized a mix of virulent bacteriophages (phage cocktail) able to efficiently counteract Pseudomonas aeruginosa infection in a zebrafish model of CF, but also showing anti-inflammatory effects in CF zebrafish embryos not infected by bacteria.On these premises, in this work we demonstrated the anti-inflammatory role of the phage cocktail both in the wild-type (WT) and hyper-inflamed CF zebrafish embryos in terms of reduction of pro-inflammatory markers. We also dissect that only the virion proteinaceous components but not the phage DNA, are responsible for the immune-modulatory effect and that this action is elicited through the activation of the Toll-like Receptor (TLR) pathway. In the CF context, we demonstrated that phages injection significantly reduces neutrophil migration following acute inflammatory induction. The elucidation of the molecular interaction between phages and the cells of vertebrate immune system might open new possibility in their manipulation for therapeutic benefits especially in diseases such as cystic fibrosis, characterized by chronic infection and inflammation.

Prospects of Inhaled Phage Therapy for Combatting Pulmonary Infections

“…Two-way cooperation exists between phages and the immune system called "Immunophage Synergy", whereby both complement each other towards faster resolution of infections and minimal tissue damage, as seen in neutrophil-phage cooperation, reported by Roach et al (2017) against P. aeruginosa. In a similar study, Cafora et al (2020) showed the anti-inflammatory role of the phage cocktail in terms of reduction of pro-inflammatory markers in P. aeruginosa infection using the zebrafish model. Phage cocktail injections significantly reduced neutrophil migration and heightened proinflammatory cytokine levels highlighting the molecular interaction between phages and the cells of the vertebrate immune system in CF disease and the anti-inflammatory role of phages.…”

Zebrafish Embryo Infection Model to Investigate Pseudomonas aeruginosa Interaction With Innate Immunity and Validate New Therapeutics

“…Phage therapy, done by injecting a phage cocktail in the yolk sac of PAO1-infected embryos, was shown to reduce lethality, bacterial burden, and the proinflammatory response caused by PAO1 infection at 20 hpi both in wild-type and CF zebrafish ( Cafora et al., 2019 ). In addition, phages alone mitigate inflammation in wild-type and CF zebrafish by reducing the expression levels of proinflammatory cytokines and the neutrophilic recruitment to the infection site ( Cafora et al., 2020a ).…”

Phage Therapy for Multi-Drug Resistant Respiratory Tract Infections

“…To the authors’ knowledge, the only publication measuring the effects of phage on neutrophilic migration was conducted using Wild-type and hyper-inflamed cftr loss-of-function zebrafish embryo models [ 58 ]. Using these, it was determined that a four-phage cocktail had immunomodulatory effects via the downregulation of pro-inflammatory cytokines and reduced neutrophil migration in response to acute inflammatory induction by tailfin amputation [ 58 ].…”

“…To the authors’ knowledge, the only publication measuring the effects of phage on neutrophilic migration was conducted using Wild-type and hyper-inflamed cftr loss-of-function zebrafish embryo models [ 58 ]. Using these, it was determined that a four-phage cocktail had immunomodulatory effects via the downregulation of pro-inflammatory cytokines and reduced neutrophil migration in response to acute inflammatory induction by tailfin amputation [ 58 ]. Recently, the development of a transepithelial migration model has enabled researchers to measure polymorphonuclear neutrophilic migration through epithelial monolayers at ALI in response to a stimulus such as purified airway supernatants from CF patients [ 59 ].…”

“…For example, some antibiotics can induce a large release of bacterial cell wall components such as lipopolysaccharide (LPS) from Gram-negative bacteria that can result in septic shock [ 61 ]. There are data suggesting phages have an immunomodulatory rather than inflammatory effect; however, this remains pertinent, especially in situations where inflammation is a primary concern for the patient [ 58 , 62 ]. Concerns surrounding bacterial lysis could be addressed by using in vitro models to perform biofilm dispersal assays on pAECs in response to phage-mediated dispersal.…”