Discussion:
Preliminary experiments showed that labeled Porcine EEC and PCF cells
remained viable after being labeled with a cell tracker and transferred
to a single-layer disk. Observation of living cells with a Leica
microscope confirmed the successful labeling and survival of different
cell types. In addition, images of Yokogawa CellVoyager CV8000 at
different z depths show that the cells are widely distributed within a
single-layer disk, indicating their ability to adhere and proliferate on
the disk surface. This finding is consistent with previous studies that
have shown that monolayer cultures are suitable for supporting cell
growth and maintaining viability [47].
The survival of mixed PCF and porcine EEC cells in multilayer disks was
investigated with encouraging results. Yokogawa CellVoyager CV8000
images showed that Porcine fibroblasts - Texas Red and EEC-GFP cells
co-existed and were viable inside the disks after 48 hours. Importantly,
the different cell numbers observed at different Z-depths indicate a
pattern of differential potential distribution within the multilayer.
This finding is consistent with research highlighting the importance of
spatial organization and cell-cell interactions in mixed cell cultures
[48]. Future research should focus on characterizing the spatial
arrangement and intercellular communication between different cell types
within the composite layer.
The simulation of cardiac fluid using cone and plate bioreactors has
provided valuable insights into the mixed cell populations under
conditioning medium. There were significant differences in cell
distribution between the control group and the experimental group after
24 hours of bioreactor treatment. Yokogawa CellVoyager CV8000 images
showed that disks exposed to bioreactor processing showed a wider
distribution of cells compared to controls. This observation suggests
that the dynamic fluid environment promotes the dispersion of cells
within the disk. In addition, a reduction in the number of cells in the
petri dish after bioreactor treatment, particularly PCF labeled with
Texas red, may indicate potential cell detachment or altered cell
survival ratio under the shear stress conditions. Future research could
explore the specific mechanisms of cell detachment under bioreactor
conditions and potential differences in cell viability and function.
In this study, the bioreactor provided the shear stress was used as a
model to study the DSS. The present study investigated the co-cultural
system under bioreactor treatment. The results showed that the mixed
cell population successfully survived, distributed, and reacted on a
monolayer disk, highlighting the behavior of these cells under
bioreactor treatment. The findings of this study provide valuable
insights into cell behavior in DSS.
The use of bioreactors in co-culture systems can significantly affect
cell distribution from the results. The decrease in the number of cells
observed in the bioreactor treated samples suggests that shear stress
caused by the flow dynamics of the bioreactor may lead to cell
detachment. This finding is consistent with previous research and
highlights the importance of optimizing bioreactor conditions to
maintain cell adhesion and prevent cell loss during culture.
Understanding the effects of shear stress on cell behavior is critical
to successfully implementing a co-culture system in DSS.
Also, the successful survival and distribution of a mixed cell
population in a co-culture system including single layer and multiple
layers, which demonstrates the potential for a more holistic approach in
DSS applications. By combining multiple cell types, such as endothelial
cells, and fibroblasts, it is possible to develop more efficient tissue
structures for DSS repair or replacement [49].
The co-culture system is also designed to address the limitations of
single-cell type therapy. By integrating multiple cell types that are
critical to the formation and function of target tissues, researchers
can create tissue structures that more closely resemble natural tissues
[50]. For instance, the addition of endothelial cells helps promote
blood vessel formation, smooth muscle cells support mechanical
properties, and fibroblasts contribute to extracellular matrix
production and tissue stability in DSS [51].
By providing a more comprehensive and versatile approach, co-culture
systems have the potential to create organizational structures that
better mimic the complexity and function of native tissues [52].
This approach allows researchers to study cell-cell interactions,
signaling pathways, and tissue development in a more physiologically
relevant way. Ultimately, it provides more efficient and functional
tissue structures to improve the treatment for DSS patients.