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Published on: March 25, 2016
Mcl-1 correlates with reduced apoptosis in neutrophils from patients with sepsis
Luc Härter1, Ladislav Mica, Reto Stocker
1Division of Trauma Surgery, University Hospital Zurich, Switzerland.
Background:
Reduction of PMN apoptosis during sepsis contributes to the pathogenesis of multiple organ failure. Differential expression of Bcl-2 proteins, which participate in apoptosis regulation, may be responsible for the dysbalanced apoptosis seen in neutrophils from septic patients. In this study, expression of Mcl-1, Bid, Bcl-2, and Bax were investigated in septic neutrophils.
Study Design:
PMN (1 x 10(6)/mL) from septic patients (n = 16) or healthy volunteers (n = 10) were incubated with either lipopolysaccharide (1 microg/mL), agonistic CD95 antibody (100 ng/mL), or medium for 16 hours. Apoptosis was quantified in FACS after propidium iodine staining. Mcl-1, Bid, Bcl-2, and Bax mRNA expression was detected by reverse transcriptase-polymerase chain reaction and protein determined by Western blot analysis.
Results:
Spontaneous apoptosis was significantly reduced in PMN from septic patients (28.8% versus 64.0% in controls). Mcl-1 protein levels decreased in patients after 16 hours but remained stable in controls. Mcl-1 mRNA was found in freshly isolated PMN from controls and patients but remained elevated only in patients. Bid protein level decreased significantly in control PMN undergoing apoptosis but differences were less prominent in septic patients. Bid mRNA was detected only in freshly isolated PMN. No Bcl-2 mRNA or protein was detected in neutrophils from patients or controls, and detectable Bax protein and mRNA levels remained unchanged in all samples.
Conclusions:
Alterations of Bid and Mcl-1 protein in neutrophils may reflect the level of apoptosis. The upregulation of Mcl-1 mRNA in patients with sepsis suggests an active role for Mcl-1 in regulation of apoptosis during sepsis; Bax remains unchanged.
Insights
Reduced neutrophil apoptosis in sepsis is linked to Mcl-1 mRNA upregulation. Septic neutrophils show altered Bid and Mcl-1 protein levels, impacting apoptosis regulation and potentially contributing to organ failure.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Sepsis-induced reduction in polymorphonuclear neutrophil (PMN) apoptosis contributes to multiple organ failure.
- Dysregulated apoptosis in septic neutrophils may stem from altered expression of Bcl-2 family proteins.
- This study investigates Mcl-1, Bid, Bcl-2, and Bax expression in neutrophils from septic patients.
Purpose of the Study:
- To investigate the expression levels of Mcl-1, Bid, Bcl-2, and Bax in neutrophils during sepsis.
- To determine the role of these proteins in the altered apoptosis observed in septic neutrophils.
- To elucidate the mechanisms underlying reduced PMN apoptosis in sepsis.
Main Methods:
- Polymorphonuclear neutrophils (PMNs) from septic patients and healthy controls were stimulated with lipopolysaccharide or CD95 antibody.
- Apoptosis was quantified using flow cytometry (FACS) with propidium iodide staining.
- Messenger RNA (mRNA) and protein expression of Mcl-1, Bid, Bcl-2, and Bax were analyzed via reverse transcriptase-polymerase chain reaction and Western blot.
Main Results:
- Spontaneous apoptosis was significantly lower in PMNs from septic patients (28.8%) compared to controls (64.0%).
- Mcl-1 protein decreased in septic PMNs, but Mcl-1 mRNA remained elevated, unlike in controls.
- Bid protein changes were less pronounced in septic PMNs compared to controls, while Bax levels were unchanged.
Conclusions:
- Altered Bid and Mcl-1 protein levels in neutrophils correlate with apoptosis levels during sepsis.
- Upregulated Mcl-1 mRNA in septic neutrophils suggests an active role in regulating apoptosis.
- Bax expression remains unchanged, indicating it is not a primary factor in sepsis-related apoptosis dysregulation.
