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Updated: May 10, 2026
Quantifying the Modulation of Elastase Enzyme Activity Through Colorimetric Analysis
Published on: January 17, 2025
[Neutrophil elastase inhibitor on proliferation and apoptosis of U937 cells]
Peng-peng Ma1, Dan Zhu, Bei-zhong Liu
1Central Laboratory of Yong-chuan Hospital, Chongqing Medical University, Chongqing 402160, China.
Objective:
To study and compare the effect of neutrophil elastase inhibitors (GW311616A and sivelestat) on the proliferation and apoptosis of U937 cells.
Methods:
Inhibitory effects of GW311616A and sivelestat on the proliferation of U937 cells were assayed by MTT assay. The morphologic changes of U937 cells were detected by transmission electron microscope, and apoptosis was observed by AnnexinV-FITC/PI staining. The changes of cell cycle and apoptosis were detected by flow cytometry. The expression of NE in U937 cells was observed by indirect immunofluorescence, the variations of content and activity of NE in U937 cells were measured through ELISA assay and colorimetric method.
Results:
MTT showed that both NE inhibitors could inhibit the proliferation of U937 cells in a dose dependent manner. The IC50 of GW311616A and sivelestat were 150 and 214 μmol/L respectively. The inhibition effect of GW311616A was significantly higher than of sivelestat (P<0.01). Typical apoptosis morphological changes of U937 cells was observed through electron microscope. AnnexinV-FITC/PI staining showed that U937 cells could be induced to undergo apoptosis by the two inhibitors, the apoptosis ratio of 150μmol/L GW311616A group (13.60%) was significantly higher than that of 150μmol/L sivelestat group (3.69%)(P<0.01). The result of flow cytometry indicated that the apoptosis ratio of 150 μmol/L GW311616A group was 14.61%, U937 cell cycle was mainly blocked in G2/M phase; meanwhile 150 μmol/L sivelestat group as 4.25% with cell cycle in S phase. The fluorescence intensity of GW311616A group obviously decreased than of sivelestat group. And the two inhibitors could reduce the content and activity of NE in U937 cells, but the effect of GW311616A was significantly higher than of sivelestat (P<0.01).
Conclusion:
GW311616A and sivelestat could inhibit the proliferation and cause apoptosis of U937 cells. Furthermore, GW311616A was more effective and harmful to cells than sivelestat.
Insights
Neutrophil elastase inhibitors GW311616A and sivelestat inhibit U937 cell proliferation and induce apoptosis. GW311616A demonstrated greater efficacy and cellular toxicity compared to sivelestat in this study.
Area of Science:
- Pharmacology
- Cell Biology
- Biochemistry
Background:
- Neutrophil elastase (NE) plays a role in inflammatory diseases.
- Investigating NE inhibitors offers potential therapeutic strategies.
- U937 cells serve as a model for studying cellular responses to NE inhibition.
Purpose of the Study:
- To compare the effects of GW311616A and sivelestat on U937 cell proliferation and apoptosis.
- To evaluate the dose-dependent inhibitory effects of these neutrophil elastase inhibitors.
- To assess the impact of these compounds on NE expression and activity in U937 cells.
Main Methods:
- MTT assay for proliferation inhibition.
- Transmission electron microscopy and AnnexinV-FITC/PI staining for apoptosis analysis.
- Flow cytometry for cell cycle and apoptosis assessment.
- ELISA and colorimetric methods for NE content and activity measurement.
Main Results:
- Both GW311616A and sivelestat inhibited U937 cell proliferation in a dose-dependent manner.
- GW311616A exhibited a significantly higher inhibitory effect (IC50: 150 μmol/L) than sivelestat (IC50: 214 μmol/L).
- Both compounds induced apoptosis, with GW311616A showing a markedly higher apoptosis ratio (13.60%) compared to sivelestat (3.69%) at 150 μmol/L.
- GW311616A primarily blocked the cell cycle at the G2/M phase, while sivelestat induced S phase arrest.
- GW311616A significantly reduced NE content and activity more than sivelestat.
Conclusions:
- GW311616A and sivelestat effectively inhibit U937 cell proliferation and induce apoptosis.
- GW311616A demonstrates superior efficacy and greater cytotoxic effects on U937 cells compared to sivelestat.
- These findings highlight GW311616A as a potentially more potent neutrophil elastase inhibitor for further investigation.
