Flow cytometric detection of spontaneous apoptosis in human breast cancer using the TUNEL-technique

Area of Science:

• Oncology
• Cell Biology
• Biotechnology

Background:

• Microscopic analysis is reliable for identifying apoptosis but doesn't correlate with cell cycle phases.
• Flow cytometry can identify apoptotic cells in tumors via a hypodiploid sub G0/1-peak due to DNA loss.
• Linking apoptosis induction to specific cell cycle phases requires advanced techniques.

Purpose of the Study:

• To adapt the TUNEL technique for flow cytometry to analyze apoptosis and cell cycle phases simultaneously.
• To investigate the incidence and characteristics of apoptosis in human breast carcinomas.
• To establish correlations between apoptotic cell detection, DNA fragmentation, and tumor proliferation.

Main Methods:

• Multiparametric flow cytometry was employed to analyze DNA content and DNA fragmentation.
• The terminal deoxynucleotidyl transferase nick-end-labeling (TUNEL) assay was integrated into flow cytometry.
• 1700 human breast carcinomas were screened for apoptotic cells and proliferation indices.

Main Results:

• A hypodiploid sub-G0/1 apoptotic peak was detected in 2.3% of the screened breast carcinomas.
• Spontaneous apoptotic fractions varied from 1.5% to 25% within individual tumors.
• Strong correlations were observed between DNA-cytometry, TUNEL-positive cells, and proliferation indices (r²=0.78 and r²=0.807, respectively).

Conclusions:

• Multiparametric flow cytometry with TUNEL enhances apoptosis detection sensitivity and quantifies DNA fragmentation across cell cycle phases.
• This combined approach allows for the correlation of apoptosis with specific cell cycle phases in cancer.
• The findings support the utility of this method for improved understanding and diagnosis of breast cancer proliferation and apoptosis.