Abstract
IntroductionLymphocyte apoptosis has been suggested to play a central role in sepsis pathophysiology, and studies in animal models demonstrated that blocking this pathway improves outcome. However, no routine biomarkers of apoptosis are so far available in patients. Thus, the aim of our study was to assess the different biomarkers of apoptosis putatively usable on a routine basis in septic shock.MethodsThirteen septic shock patients (sampled twice between days 1 to 2 and days 3 to 5 after diagnosis of shock) and 15 sex-matched and age-matched healthy controls were prospectively enrolled. Apoptosis was measured in lymphocyte subpopulations using flow cytometry (Annexin-V binding, activated caspase-3 and Bcl-2 expressions). Representative pro-apoptotic and anti-apoptotic gene expressions were assessed by quantitative reverse-transcription PCR. Monocyte HLA-DR expression and lymphocyte subpopulation cell counts were measured as markers of sepsis-induced immune dysfunctions. To test for statistical significance, the Mann-Whitney U test was used with correction by the number of tests performed.ResultsFlow cytometric measurements of apoptosis in septic shock patients showed an increased Annexin-V binding on CD4+ T cells and an increased active caspase-3 expression on B cells only at days 3 to 5 (sixfold change and twofold change, respectively). Gene expression analysis showed an increased BCL-XL mRNA and an upregulation of the pro-apoptotic genes BID and FAS in septic shock patients (10-fold change and fivefold change, respectively) compared with healthy controls.ConclusionsThe present study highlights the difficulties encountered in monitoring apoptosis on a routine basis in septic patients, whereas in the same sampling conditions and on the same patients, HLA-DR expression and lymphocyte subpopulation cell counts showed characteristics described in the literature. However, pro-apoptotic genes BID and FAS appear to constitute promising apoptosis markers in our hands.
Highlights
Lymphocyte apoptosis has been suggested to play a central role in sepsis pathophysiology, and studies in animal models demonstrated that blocking this pathway improves outcome
The total number of leukocytes was increased in septic patients at days 1 to 2 (D1-2) and days 3 to 5 (D3-5) compared with healthy volunteers (Mann-Whitney U test; Table 3)
Active caspase-3 expression, the central caspase in the apoptosis pathway [12], was increased in CD19+ cells of septic patients at D3-5 compared with healthy volunteers, but no significant changes were observed in CD4+ and CD8+ T cells (Figure 2b)
Summary
Lymphocyte apoptosis has been suggested to play a central role in sepsis pathophysiology, and studies in animal models demonstrated that blocking this pathway improves outcome. The aim of our study was to assess the different biomarkers of apoptosis putatively usable on a routine basis in septic shock. Numerous studies in animal models of sepsis showed that blocking programmed cell death improves outcome after septic challenge [6]. This approach could represent a potential innovative therapeutic strategy in septic shock. Since no clinical signs of apoptosis have been described, there is an urgent need to develop robust biomarkers available on a routine basis for the monitoring of this phenomenon. Weber and colleagues recently observed an extensive apoptosis of circulating lymphocytes after severe sepsis (upregulation of BIM and BID gene expression, and a downregulation of the anti-apoptotic molecules BCL-2 and BCL-XL) [7]
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