A Korean/U.S. collaborative study has shown that increased circulating mitochondrial N-formyl peptide (mtFP) contributes to development of secondary hospital-acquired (nosocomial) infections and increased mortality in septic shock patients who survived the early hyperinflammatory phase.

The study further found that increased susceptibility to secondary infection was at least partly mediated by suppression of formyl peptide receptor-1- (FPR1)-mediated polymorphonuclear leukocyte (PMN) chemotaxis to secondary infection sites.

Incorporating these findings into therapies may improve the clinical outcomes of septic shock patients by preventing PMN chemotactic anergy, or absence of a normal immune response, the authors suggested in the April 27, 2021, edition of Proceedings of the National Academy of Sciences.

"Numerous previous studies have reported the role of mtFPs in PMN chemotaxis in injured patients," said first author Woon Yong Kwon, a professor in the Department of Emergency Medicine at Seoul National University College of Medicine and Hospital.

"However, to my knowledge, there have been no studies investigating the impact of circulating mtFPs on secondary infections in recovering septic shock patients, which prompted the design of the present study."

Kwon designed and conducted the new PNAS study in collaboration with co-leaders Carl Hauser, director of trauma and attending surgeon at Beth Israel Deaconess Medical Center/Harvard Medical School (BIDMC/HMS) in Boston, Massachusetts, and Kiyoshi Itagaki, an assistant professor of surgery at BIDMC/HMS, where Kwon was recently a research fellow.

Septic shock is a life-threatening condition leading to multiorgan injury, both primarily via tissue inflammation and secondarily via hypoperfusion, with secondary nosocomial infections often exacerbating the outcomes of recovering septic shock patients.

"Recent studies have reported that the late death rate attributed to secondary infections in patients recovering from the early hyperinflammatory phase is more than 70%," Kwon told BioWorld Science.

"Our data also showed that development of secondary infection itself increased 90-day mortality by 4.7 times in patients recovering from septic shock," he said.

By promoting migration of PMNs to secondarily inoculated sites, chemotaxis appears to play a key role in inhibiting progression from local bacterial inoculation to secondary infection.

Although both primary and secondary infections can result in mtFP release into the circulation, no studies have assessed the role of circulating mtFPs during septic shock, despite mtFP occupancy of the FPR1 having been shown to suppress PMN chemotaxis.

The new PNAS study consequently investigated the association between circulating mtFPs and development of secondary infection in septic shock patients.

The role of ND6

Kwon and his team collected clinical data and plasma from septic shock patients admitted to intensive care > 72 hours and analyzed the effects of circulating nicotinamide adenine dinucleotide dehydrogenase subunit-6 (ND6) on clinical outcomes.

The ND6 protein is a subunit of NADH dehydrogenase (ubiquinone) involved in redox reactions which is located in the mitochondrial inner membrane, with the new study investigating the role of ND6 in PMN chemotaxis using isolated human PMNs.

"We had previously reported that a subset of five human mtFPs functionally deactivated human PMNs, among which ND6 was the most potent human mtFP," Kwon said.

"Thus, we analyzed the effect of circulating ND6 on clinical outcomes of septic shock patients, with ND6 levels being measured using a commercial ELISA [enzyme-linked immunosorbent assay] kit."

Analysis of plasma samples from 97 Korean septic shock patients revealed that circulating ND6 levels upon admission were independently and significantly associated with secondary infections and increased 90-day mortality.

"An increase in plasma ND6 levels of 1 ng/mL at admission independently increased development of secondary infection by 30-fold, while 90-day mortality was increased by 1.6 times," he said.

In ex vivo experiments, PMN pretreatment with 100 nM ND6 for 45 minutes was shown to suppress FPR1-mediated PMN chemotactic responses to bacterial peptides in the presence of multiple cytokines and chemokines, despite increased non-directional PMN movements.

"In control medium with no added cytokines or chemokines, ND6 pretreatment decreased PMN chemotaxis in response to bacterial peptides by 84%," said Kwon.

Conversely, "in sepsis-mimicking medium containing cytokines and chemokines at concentrations approximating those found in septic shock patients with secondary infection, ND6 pretreatment decreased PMN chemotaxis in response to bacteria peptides by 44%," he said.

"In septic chock patients, ongoing systemic inflammation releases excessive cytokines and chemokines, which activate PMS, into the circulation for several days even after hemodynamic stabilization."

The researchers also found that in sepsis-mimicking medium, PMN movements were increased versus controls, which is important, as PMNs must be chemotactically directed to clear secondary infections.

"However, PMNs activated by cytokines and chemokines do not effectively migrate to secondarily infected sites, with non-directional PMN movements being increased instead," said Kwon.

"This can decrease the number of PMNs moving to secondarily infected sites and killing secondarily inoculated bacteria, resulting in secondary infections, such as ventilator-associated pneumonia (VAP)."

Moreover, circulating mtFPs appeared to contribute to development of secondary infection and increased mortality in septic shock patients who had survived their early hyperinflammatory phase.

This increased susceptibility to secondary infection is probably at least partly mediated by the suppression of FPR1-mediated PMN chemotaxis to secondary infected sites.

"Although we cannot say exactly to what extent circulating mtFPs seemed to contribute to secondary infections and mortality, our data indicate they may suppress PMN chemotaxis by 20-30%," Kwon said.

Collectively, "these findings suggest that immunotherapeutic desensitization and/or elimination of circulating mtFPs before secondary infection occurs, may be a novel therapeutic strategy to reduce secondary infections and mortality in septic shock patients."

"Moreover, five highly potent mtFPs, including ND6, ND3, ND4, ND5 and cytochrome C oxidase subunit 1 (COX1), might be potent candidates for immunotherapy, which may preserve FPR1-mediated PMN chemotaxis in response to bacterial N-formyl peptides, which are more concentrated in secondary infected sites in the presence of cytokines and chemokines."

Looking forward, he said, "we will be investigating whether elimination of circulating mtFPs can rescue PMN chemotaxis. If this is indeed the case, we will look for practical methods with which to eliminate circulating mtFPs, without having adverse effects in septic shock patients."