Amplification of tissue plasminogen activator expression from epithelial cell lines

Area of Science:

• Biochemistry
• Cell Biology
• Biotechnology

Background:

• Non-malignant epithelial cell lines (BEB and GPK) secrete tissue plasminogen activator (tPA).
• Epithelial tPA yield is comparable to malignant sources, but specific activity is lower.
• Current production requires a two-step process involving serum and serum-free conditions.

Purpose of the Study:

• To identify methods for potentiating tPA yield from epithelial cells.
• To establish a serum-free culture system for continuous enzyme harvesting.
• To investigate agents that amplify enzyme expression and stimulate stationary-phase cells.

Main Methods:

• Culturing human breast (BEB) and guinea pig ear keratocyte (GPK) cell lines.
• Treating cells with regulatory agents, including the hypomethylating agent 5-azacytidine.
• Assessing the impact of mitogenic lectins, such as Concanavalin A, on tPA secretion.
• Investigating enzyme secretion during cell growth phases using tritiated thymidine.

Main Results:

• 5-azacytidine significantly increased tPA secretion by 3-5 fold.
• Mitogenic lectins resulted in further 15-20 fold increases in tPA yield.
• Concanavalin A extended the period of enzyme secretion, with further investigations underway.

Conclusions:

• 5-azacytidine and specific mitogenic lectins are effective in potentiating tPA yield from epithelial cells.
• These findings suggest a viable strategy for enhancing tPA production in a controlled, serum-free environment.
• Further research is needed to fully elucidate the mechanisms behind lectin-induced secretion and extended culture periods.