Butyrate and propionate induced activated or non-activated neutrophil apoptosis via HDAC inhibitor activity but without activating GPR-41/GPR-43 pathways.
OBJECTIVE
Decreased neutrophil apoptosis is implicated in persistent inflammation resulting in systemic inflammatory response syndrome and multiple organ dysfunctions syndromes. Short-chain fatty acids (SCFAs) may be a candidate to control neutrophil apoptosis because SCFAs are normally produced in the gut and related products have been approved for human use. We investigated the effects of SCFAs on apoptosis of activated and non-activated neutrophils and their mechanisms.


METHODS
Purified neutrophils obtained from healthy volunteers were preincubated for 1 h with or without the G-protein receptor (GPR) inhibitor pertussis toxin (100 ng/mL) or U-73122 (50 ng/mL), extracellular signal-related protein kinase inhibitor PD98059 (10 microM), mitogen-activated protein kinase (MAPK) p38 inhibitor SB203580 (25 microM), Jun kinase inhibitor-I (2 microM), caspase-3 and -7 inhibitor Z-VAD-FMK (100 microM), caspase-8 inhibitor Z-IETD-FMK (50 microM), or caspase-9 inhibitor Z-LEHD-FMK (50 microM). The cells were then cultured with or without SCFAs or trichostatin A, a typical histone deacetylase inhibitor, in the presence or absence of lipopolysaccharide (1 microg/mL) or tumor necrosis factor-alpha (100 ng/mL). Neutrophil apoptosis was assessed by annexin V staining using flow cytometry. The GPR-41 and -43 and apoptosis-related proteins (bax, mcl-1, a1) mRNA were measured by quantitative real-time polymerase chain reaction and the expression of acetylated histone H3 was determined by western blot.


RESULTS
The caspase inhibitors inhibited butyrate- and propionate-induced neutrophil apoptosis treated or untreated with lipopolysaccharide or tumor necrosis factor-alpha, whereas GPR and MAPK inhibitors had no effect. The mRNA expressions of GPR-43 and a1 protein were reduced by butyrate and propionate. The expressions of acetylated histone H3 were induced by butyrate and propionate.


CONCLUSIONS
These results suggest that butyrate and propionate increase apoptosis of neutrophils irrespective of their activation state, by factors other than GPRs and MAPKs, and their mechanisms likely relate to their histone deacetylase inhibition activity, which may control a1 mRNA expression.
Aoyama
Michiko
M
Division of Nutrition and Metabolism, Department of Biophysics, Kobe University Graduate School of Health Sciences, Kobe, Japan.
Kotani
Joji
J
Usami
Makoto
M
D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D002087:Butyrates / Q000494:pharmacology*; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors / Q000378:metabolism; D004963:Estrenes / Q000494:pharmacology; D056572:Histone Deacetylase Inhibitors / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007249:Inflammation / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D037342:Pertussis Toxin; D011422:Propionates / Q000494:pharmacology*; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism*; D012016:Reference Values; D014409:Tumor Necrosis Factor-alpha
D002087:Butyrates; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D004963:Estrenes; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D006877:Hydroxamic Acids; D008070:Lipopolysaccharides; D011422:Propionates; D011506:Proteins; D011760:Pyrrolidinones; D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled; D014409:Tumor Necrosis Factor-alpha; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C012589:trichostatin A; D037342:Pertussis Toxin