Cell culture under perfusion conditions reduces cellular metabolic stress and mimics the in vivo physiological environment in pancreatic cancer


Year:

Session type:

Theme:

Daniel Hughes1,Frances Willenbrock2,Zahir Soonawalla3,Srikanth Reddy3,Michael Silva3,Somnath Mukherjee3,Eric O'Neill2
1University of Oxford, Oxford, UK,2Department of Oncology, University of Oxford, Oxford, UK,3Oxford University Hospitals, Oxford, UK

Abstract

Background

Tumour responsiveness to chemotherapy in PDAC varies significantly across patients. This reinforces the need to establish an accurate model that preserves the tumour microenvironment in the ex-vivo, allowing a personalized drug screen for patients to identify optimal regime of therapy. Current standard of 2D cell culture is that the media is exchanged at defined time points (i.e. static culture). However, this is not a physiologically representative model as cells are maintained in either a nutrient and substrate ‘hyper-replete’ or deplete environment, rather than an epistatic supply. Moreover, episodic media replacement creates significant and abrupt changes to the culture environment and availability of substrates which induces cellular metabolic stress. We evaluated the role of perfusion culture to establish whether this technique reduced metabolic stress and creates a physiologically representative model.

Method

Cells were cultured  either in static culture (serving as a control, with media exchange every 72 hours) or in a nutritionally replete condition with a constant low perfusion rate of media exchange over 7 days. Daily media extraction was performed in order to evaluate the metabolic activity and viability of cells. Tissue slices were  created from core biopsies and then cultured either in static culture (as control) or under perfusion conditions. 

Results

Under conventional static culture, a decrease in the quantity of metabolic substrates was observed over time indicating Metabolic activity of cells in static culture was suppressed (noted by down regulation of mTOR pathway products). Under perfusion conditions, glucose concentration was maintained over 7 days, indicating cells were able to maintain metabolic activity. Tissue sections were similarly evaluated for metabolic stress and will be presented.

Conclusion

This platform highlights the considerable metabolic stress that cells undergo whilst under conventional static culture. Perfusion culture serves as a technique to reduce cellular metabolic stress in addition to creating a physiologically representative model ex vivo.