From in vivo to in vitro: Major metabolic alterations take place in hepatocytes during and following isolation
Bibliographic record
Abstract
The liver plays a key role in maintaining physiological homeostasis and hepatocytes are largely responsible for this. The use of isolated primary hepatocytes has become an essential tool for the study of nutrient physiology, xenobiotic metabolism and several liver pathologies. Since hepatocytes are removed from their normal environment, the isolation procedure and in vitro culture of primary hepatocytes is partially known to induce undesired metabolic changes. We aimed to perform a thorough metabolic profiling of primary cells before, during and after isolation using state-of-the-art techniques. Extensive metabolite measurements using HPLC were performed in situ in the liver, during hepatocyte isolation using the two-step collagenase perfusion method and during in vitro cell culture for up to 48 hours. Assessment of mitochondrial respiratory capacity and ATP-linked respiration of isolated primary hepatocytes was performed using extracellular flux analysis. Primary hepatocytes displayed a drastic decrease in antioxidative-related metabolites (NADPH, NADP, GSH and GSSG) during the isolation procedure when compared to the in situ liver (P<0.001). Parallel assessment of citric acid cycle activity showed a significant decrease of up to 95% in Acetyl-CoA, Isocitrate/Citrate ratio, Succinate, Fumarate and Malate in comparison to the in situ liver (P<0.001). While the levels of several cellular energetic metabolites such as Adenosine, AMP, ADP and ATP were found to be progressively reduced during the isolation procedure and cell culture (P<0.001), higher ATP/ADP ratio and energy charge level were observed when primary cells were cultured in vitro compared to the in situ liver (P<0.05). In addition, a significant decrease in the respiratory capacity occurred after 24 hours in culture. Interestingly, this was not associated with a significant modification of ATP-linked respiration. In conclusion, major metabolic alterations occur immediately after hepatocytes are removed from the liver. These changes persist or increase during in vitro culture. These observations need to be taken into account when using primary hepatocytes for the study of metabolism or liver physiopathology.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".