Hydrocortisone fails to abolish NF-κB1 protein nuclear translocation in deletion allele carriers of the NFKB1 promoter polymorphism (-94ins/delATTG) and is associated with increased 30-day mortality in septic shock

Abstract

Background: Previous investigations and meta-analyses on the effect of glucocorticoids on mortality in septic shock revealed mixed results. This heterogeneity might be evoked by genetic variations. Such candidate is a promoter polymorphism (-94ins/delATTG) of the gene encoding the ubiquitous transcription-factor nuclear-factor-κB (NF-κB) which binds to recognition elements in the promoter of several genes encoding for the innate immune-system. In turn, hydrocortisone inhibits NF-κB nuclear translocation and thus transcription of key immune-response regulators. Accordingly, we tested the hypotheses that hydrocortisone has a NFKB1 genotype dependent effect on 1) NF-κB1 nuclear translocation evoked by lipopolysaccharide (LPS) in monocytes in vitro, and 2) mortality in septic shock.

Methods: Monocytes of volunteers with the homozygous insertion (II; n = 5) or deletion (DD; n = 6) NFKB1 genotype were incubated with 10 µgml-1 LPS ± hydrocortisone (10-5M), and NF-κB1 nuclear translocation was assessed (immunofluorescence). Furthermore, we analyzed 30-day-mortality in 160 patients with septic shock stratified for both genotype and hydrocortisone therapy.

Results: Hydrocortisone inhibited LPS induced nuclear translocation of NF-κB1 in II (25%±11;p = 0.0001) but not in DD genotypes (51%±15;p = n.s.). Onehundredandfour of 160 patients with septic shock received hydrocortisone, at the discretion of the intensivist. NFKB1 deletion allele carriers (ID/DD) receiving hydrocortisone had a much greater 30-day-mortality (57.6%) than II genotypes (24.4%; HR:3.18, 95%-CI:1.61-6.28;p = 0.001). In contrast, 30-day mortality was 22.2% in ID/DD and 25.0% in II genotypes without hydrocortisone therapy. Results were similar when using propensity score matching to account for possible bias in the intensivists' decision to administer hydrocortisone.

Conclusion: Hydrocortisone fails to inhibit LPS induced nuclear NF-κB1 translocation in deletion allele carriers of the NFKB1 promoter polymorphism (-94ins/delATTG). In septic shock, hydrocortisone treatment is associated with markedly increased 30-day-mortality only in such carriers. Accordingly, previous heterogeneous results regarding the benefit of hydrocortisone in septic shock may be reconciled by genetic variation of the NFKB1 promoter polymorphism.

Figure 1. Percentage of NF-κB1 positive monocytes according to the NFKB1 promoter polymorphism (-94ins/delATTG) following lipopolysaccharide (LPS) incubation with and without hydrocortisone (HC).

Hydrocortisone significantly decreased the percentage of NF-κB1 positive cell nuclei in LPS stimulated monocytes of II genotype individuals, but not in those of the DD genotype individuals, which showed an unchanged high percentage of NF-κB1 positive cell nuclei. There was no difference in the percentage of NF-κB1 positive cells both in unstimulated and hydrocortisone (HC) treated cells. * p<0.0001 vs. control; # p<0.01 vs. II genotype. II  =  homozygous insertion genotype; DD  =  homozygous deletion genotype.

Figure 2. Kaplan–Meier plot of 30-day mortality of patients with septic shock stratified both by NFKB1 promoter polymorphism (−94ins/delATTG) and hydrocortisone therapy.

Kaplan-Meier estimators for the four subgroups for all 160 septic shock patients. DD  =  homozygous deletion genotype. ID  =  heterozygous deletion genotype. II  =  homozygous insertion genotype.

Figure 3. Kaplan–Meier plot of 30-day mortality of patients with septic shock stratified both by NFKB1 promoter polymorphism (−94ins/delATTG) and hydrocortisone therapy.

Kaplan-Meier estimators for the four subgroups in 2x24 matched patients of the propensity score analysis. DD  =  homozygous deletion genotype. ID  =  heterozygous deletion genotype. II  =  homozygous insertion genotype.