Expression of apolipoproteins L in neutrophils during sepsis

Introduction

Sepsis is characterized by a strong systemic inflammatory reaction. The pathogenesis is driven by alterations in the immune system and is associated with high neutrophil counts related to a specific delay in apoptosis [1]. The apolipoproteins L (ApoLs) family comprises six members in humans (ApoL1 to ApoL6). In light of their deregulated expression in several pathologies, they are likely to be important molecular players of programmed cell death [2]. We analyzed ApoL expression in cohorts of septic and nonseptic ICU patients and healthy volunteers in order to test whether ApoLs could be involved in the neutrophil apoptotic program.

Methods

By means of magnetic cell sorting, peripheral neutrophils were purified from 20 healthy volunteers and 40 ICU patients with (n = 20) or without sepsis (n = 20). ApoL expression was analyzed at the mRNA and protein levels by real-time PCR and western blot analysis respectively. Apoptosis of purified neutrophils was assessed using flow cytometry following 4 and 24 hours of in vitro incubation. We monitored the expression of C-reactive protein (CRP), an inflammatory marker, and its correlation with ApoL expression in PMNs was studied by linear regression analysis.

Results

Our results showed a significant downregulation in mRNA expression of ApoL1 (P < 0.0001), ApoL2 (P = 0.0009), ApoL3 (P < 0.0001) and ApoL6 (P = 0.0003) in purified PMNs from ICU patients as compared with the healthy individuals. This downregulation was also validated at the protein level for ApoL1 and ApoL2, whereas ApoL 6 was upregulated in septic patients. We could not detect ApoL3 protein in any of the cohorts. This was accompanied by a significant delay in PMN apoptosis in septic patients as compared with healthy volunteers (P < 0.05) at 4 and 24 hours. We also showed a strong negative correlation in the three mixed groups between CRP and ApoL1 (R = -0.607), ApoL2 (R = -0.651), ApoL3 (R = -0.578) and ApoL6 (R = -0.506).

Conclusion

The altered apoptotic fate of neutrophils in sepsis was correlated with the modification of the expression profile of ApoLs, a family of proteins thought to be involved in the apoptotic process. The role of these proteins in the sepsis-associated phenotype of neutrophils remains to be further elucidated.