Unmasking unique immune altering aspects of the microbiome as a tool to correct sepsis induced immune dysfunction
Author(s):
Isabella Heffernan; Tatiana Johnston; Alfred Ayala; Daithi Heffernan
Background:
Sepsis mortality is driven by lymphocyte dysfunction and anergy. Although PD-1 induces T-cell anergy, PD-1-high-T-cells prevent immune mediated tissue injury from infections. Bidirectional microorganism– immune cell cross talks exists. Gut Bacteroides fragilis-T-cell cross talk maintains innate immune cell/pathogen homeostasis. Commensal gut Clostridia spp suppress inflammation and induce gut tolerance. However, microbiota therapies for sepsis-induced microbiome disruptions are rarely tailored to induce specific, or required, immune responses.
Hypothesis:
Among options commonly employed as probiotic therapy, Bacteroides fragilis (BF) will induce the least anergic and most restorative (IL-33 and PD-1-high) lymphocyte phenotype.
Methods:
1×106 T-lymphocytes [primary T-cell(ATCC-HB-11052)] were cultured with monomicrobial (MMicr) B. fragilis (BF), C. perfringnes (CP) or L. acidophilus [LA] or PBS for control. Polymicrobial environments (PMicr) were dominant in either BF(B50), CP(C50) or LA(LA50). Cytokines included IL-22 & IL-33 (mediators of gut epithelium–bacteria interactions and epithelial repair). Flow cytometry was used for phenotying. T-cell DNA was extracted for pyrosequencing (CpG islands) for epigenetic changes. For translation, blood was obtained from septic SICU patients with either PMicr or MMicr infections.
Results:
In reviewing T-cell cytokine responses, LA & LA50 consistently induced both IL-22 (42.6 vs 24.5 vs 24.2pg/ml;p<0.05) and IL-33. MMicrCP only induced IL-22, and BF only induced IL-33 under PMicr(B50) conditions(65 vs 30 vs 24 pg/ml;p<0.05). Within SICU patients IL-33 levels were higher in PMicr vs MMicr patients. Phenotype: All exposures uniformly elevated CD69. PD-1+ expression was lowest with either MMicr BF exposure (65.2% vs 85.7% vs 82.3%;p<0.05) or PMicr B50(79% vs 86% vs 95.3%;p<0.05). Conversely BF exposure induced a distinct PD-1-high subpopulation [BF (20.6%) & B50 (16.3%)]. BTLA expression did not differ following individual MMicr. Among PMicr, B50 and C50, but not LA50 increased BTLA+ expression(23.1% vs 20.6% vs 5.5%;p<0.05). Epigenetic changes: No methylation variation for cytokines. Only B50 induced methylation of HLA-DR4. BF & PMicr B50 induced demethylation of CTLA.
Conclusions:
L.acidophilus induced the potential for short term resolution (IL-22 & IL-33) whereas B. fragilis induced a T-cell phenotype consistent with potential long term immune recovery. Microbiota therapies tailored to specific sepsis induced immune dysfunctions may obviate the need for prolonged antimicrobial therapy in immune paralyzed septic patients.