HIF1A-dependent induction of alveolar epithelial PFKFB3 dampens acute lung injury

Abstract

Acute lung injury (ALI) is a severe form of lung inflammation causing acute respiratory distress syndrome in patients. ALI pathogenesis is closely linked to uncontrolled alveolar inflammation.We hypothesize that specific enzymes of the glycolytic pathway could function as key regulators of alveolar inflammation. Therefore, we screened isolated alveolar epithelia from mice exposed to ALI induced by injurious ventilation to assess their metabolic responses. These studies pointed us towards a selective role for isoform 3 of the 6-phosphofructo-2-kinase/fructose-2,6bisphosphatase (PFKFB3). Pharmacologic inhibition or genetic deletion of Pfkfb3 in alveolar epithelia (Pfkfb3 loxp/loxp SPC-ER-Cre+ mice) was associated with profound increases in ALI during injurious mechanical ventilation or acid installation. Studies in genetic models linked Pfkfb3 expression and function to hypoxia-inducible factor Hif1a. Intra-tracheal pyruvate instillation not only reconstituted Pfkfb3 loxp/loxp or Hif1a loxp/loxp SPC ER Cre+ mice, but pyruvate was also effective in ALI treatment of wild-type mice. Finally, proof-of-principle studies in human lung biopsies confirmed increased PFKFB3 staining in injured lungs and co-localized PFKFB3 to alveolar epithelia. These studies reveal a specific role for PFKFB3 in counterbalancing alveolar inflammation and lay the groundwork for novel metabolic therapeutic approaches during ALI.

Lactate produced by alveolar type II cells suppresses inflammatory alveolar macrophages in acute lung injury

“…The acid aspiration model of ALI was described previously 3,4,14,15 . Mice were anesthetized with isoflurane and suspended by their incisors at 45° angle.…”

“…We recently showed that the glycolytic shift of ATII cells in response to injurious stimuli protects against experimental ALI. 3 We hypothesized that the metabolic activity of the alveolar epithelium impacts the alveolar macrophage function. To study the interactions between ATII cells and macrophages, we utilized a non-contact co-culture system.…”

“…The transcription factor HIF1A is a critical regulator of glycolysis and hence lactate production in ATII cells. 3 Mice with ATII-specific genetic suppression of Hif1a (Hif1a loxp/loxp SPC-Cre-ER+) have a more severe phenotype in several models of acute lung injury. 3,4 Therefore, we investigated whether, in Hif1a loxp/loxp SPC-Cre-ER+ mice, the decreased lactate production by ATII cells leads to impaired lactate-mediated paracrine actions between ATII cells and alveolar macrophages.…”

“…1 Our previous work showed that in response to injury, alveolar epithelial type II cells (ATII) demonstrate enhanced glycolysis via the hypoxia-inducible factor (HIF)-1Aphosphofructokinase-2/fructose-2,6-bisphosphatase (PFKFB) 3 axis, which was associated with attenuation of inflammation in the acute phase of acute lung injury in several models of murine ALI. [2][3][4] Alveolar macrophages (AMs) reside in the alveolar space near ATII cells 5 and in this niche, the phenotype of the AMs is affected by the local microenvironment consisting of metabolites, cytokines, and surrounding cells that shape their inflammatory or anti-inflammatory properties. 6 The critical role of alveolar macrophages (AM) in the pathogenesis of ALI/ARDS, especially in the inflammatory phase, has been well-established in animal models and human disease.…”

HIF-dependent induction of alveolar miR-147 dampens SARS-CoV-2 immune evasion

“…For mRNA gene expression assay, cDNA was synthesized using High-Capacity cDNA Reverse Isolation murine alveolar epithelial cells. The protocol to isolate alveolar epithelial cells is adapted from previous publication 53 . For alveolar epithelial cell isolation, mice were euthanized by pentobarbital overdose.…”