Flow Cytometry
Observations collected from the confocal microscopy are indicative of that fish oil supplementation assisted bovine luteal cells in maintaining a higher mitochondrial membrane potential compared to vegetable oil supplementation or the control medium when introduced into a hypoxic environment. However, these observations are qualitative in nature and need to be quantified.
Confocal Microscopy
After cells were subjected to the 5% oxygenated environment for 24 hours,, the fish oil treated cells exhibited higher intensity fluorescence , indicating a higher mitochondrial membrane potential.
Additionally, the comparison from Hour 0 to Hour 24 in a 5% oxygenated environment on the flow cytometer showed that Hypoxia had a negative effect on the cells. It was noticeable that the BSA treated cells were healthier at Hour 0 than Hour 24 of hypoxia.
Although the cells treated with Vegetable Oil in a 5% oxygenated environment were still fluorescing at Hour 24, they were noticeably less fluorescent than the fish oil treated cells nor were the vegetable oil treated cells as fluorescent at Hour 24 as they were at Hour 0.
Additionally, the comparison from Hour 0 to Hour 24 in a 5% oxygenated environment on the flow cytometer showed that Hypoxia had a negative effect on the cells. It was noticeable that the BSA treated cells were healthier at Hour 0 than Hour 24 of hypoxia.
Although the cells treated with Vegetable Oil in a 5% oxygenated environment were still fluorescing at Hour 24, they were noticeably less fluorescent than the fish oil treated cells nor were the vegetable oil treated cells as fluorescent at Hour 24 as they were at Hour 0.
Where could these positive effects come from?
The positive effects may be produced by two fatty acids within fish oil: Docosahexaenoic Acid (DHA) and Eicosapentaenoic Acid (EPA). These acids may play an important part in the physiology of the cell. Fish oil contains these fatty acids whereas Vegetable Oil does not.
Potential errors
During the experiment, there were some potential errors, and conflicting results, that could be caused by human error or other conditions.
For example, cyanide has debilitating properties that, when introduced to cells, should devastate the population. However, the cyanide did not succeed in killing the entire population of cells, only 44.175%, refer to Figure 6. This could be caused by a human error, such as:
The mitochondrial fragmentation of the BSA treated cells is also a source of error. This could have been caused by the BSA treatment, or the stress caused by the hypoxic conditions, or the TMRM dye could have caused the fragmentation.
For example, cyanide has debilitating properties that, when introduced to cells, should devastate the population. However, the cyanide did not succeed in killing the entire population of cells, only 44.175%, refer to Figure 6. This could be caused by a human error, such as:
- The cyanide was diluted too heavily to an unusable concentration and therefore did not have as great of an effect.
- The wrong amount of cyanide could have been pipetted on to the cells.
- When the cyanide was added to the cells, the flask might not have been shaken enough for the cyanide to allow for an even distribution.
- The cyanide was incubated with the cells for only five minutes at 37°C, therefore in future studies we would like to increase the incubation time to hopefully increase the effect of the cyanide on the cells.
The mitochondrial fragmentation of the BSA treated cells is also a source of error. This could have been caused by the BSA treatment, or the stress caused by the hypoxic conditions, or the TMRM dye could have caused the fragmentation.
Future studies
Future cellular studies will be completed because of the implications of this study. This study will be replicated in the future to make certain that the results we gathered will be concrete evidence of fish oil’s effects. A future study could be a whole animal study that researches the effects of cellular treatments on a large scale. Specifically retrieving blood samples to observe the amount of DHA and EPA in the bloodstream of fish oil treated bovine. In the future if these cellular studies are positive and proven to work, this could be transferred to humans to help those with troubles establishing pregnancy or struggling with miscarriages. Also, it would be beneficial to study mitochondrial fragmentation, how it is caused, and how it could affect the corpus luteum as a whole.
Research Paper
Below is my research paper where you can go more in depth about this project.
research_paper.pdf | |
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