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Published on: January 7, 2012
Cellular Respiration in Thymic Fragments from Mice
Mariam Alshamsi1, Maya Hassane1, Farida Almarzooqi2
1Department of Microbiology and Immunology, College of Medicine and Health Sciences, UAE University, 15551 Al Ain, Abu Dhabi, United Arab Emirates.
Background:
This report aims to detail the use of the phosphorescence oxygen analyzer for in vitro investigation of thymic responses to pharmaceutical agents, in particular immunosuppressants and immunomodulators. Sirolimus (a highly specific inhibitor of the 'molecular target of rapamycin', mTOR) and ozanimod (an agonist of the sphingosine 1-phosphate receptor, recently approved for treatment of multiple sclerosis and ulcerative colitis) are used for this purpose.
Methods:
Thymic fragments from mice were placed in glass vials containing phosphate-buffered saline, bovine albumin, and Pd(II) meso-tetra (sulfophenyl) tetrabenzoporphyrin. The vials were sealed from air, and the cellular oxygen consumption was monitored as function of time.
Results:
The decline of dissolved oxygen concentration with time (d[O2]/dt) was linear; thus, its rate (thymocyte respiration) was expressed as μM O2 min-1. Cyanide inhibited respiration, confirming the oxygen consumption was in cytochrome oxidase. In age-matched mice, the rate of thymocyte respiration (mean ± SD, in μM O2 min-1 mg-1) was 0.046 ± 0.011 (median = 0.043, range = 0.028 to 0.062, n = 10). In thymic fragments from littermates, this rate was inhibited in the presence of sirolimus (16% lower) or ozanimod (29% lower).
Conclusions:
Thymocyte respiration can serve as a surrogate biomarker for studying the mode-of-action and the cytotoxicity of immunotoxins and immunosuppressants.
Insights
Phosphorescence oxygen analysis measures thymocyte respiration in vitro. Sirolimus and ozanimod, common immunosuppressants, significantly reduced oxygen consumption in thymic fragments, indicating their impact on cellular metabolism.
Area of Science:
- Immunology
- Pharmacology
- Biochemistry
Background:
- Investigating in vitro thymic responses to pharmaceutical agents is crucial for understanding immune modulation.
- Sirolimus (mTOR inhibitor) and ozanimod (S1P receptor agonist) are key immunomodulatory drugs with distinct mechanisms of action.
Purpose of the Study:
- To detail the use of phosphorescence oxygen analyzer for in vitro thymic response assessment.
- To evaluate the impact of sirolimus and ozanimod on thymocyte respiration.
Main Methods:
- In vitro incubation of mouse thymic fragments in oxygen-sensing medium.
- Monitoring cellular oxygen consumption over time using phosphorescence oxygen analyzer.
- Quantifying thymocyte respiration rate (μM O2 min-1 mg-1) and assessing inhibition by pharmaceutical agents.
Main Results:
- Thymocyte respiration rate was linear and confirmed to be cytochrome oxidase-dependent.
- Baseline thymocyte respiration rate in control mice was established (0.046 ± 0.011 μM O2 min-1 mg-1).
- Sirolimus inhibited respiration by 16%, and ozanimod by 29% in thymic fragments.
Conclusions:
- Thymocyte respiration serves as a valid surrogate biomarker for evaluating drug effects.
- This method aids in studying the mode-of-action and cytotoxicity of immunosuppressants and immunomodulators.
- Phosphorescence oxygen analysis offers a sensitive tool for in vitro drug screening in immunology.
