Coelho 2015 Abstract Thyroid Cancer

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AMPK overactivation in BCPAP tumor cells induces cell death through increased reactive oxygen species (ROS).

Link: Open Access

Coelho R, Cazarin J, Albuquerque JA, Galina A, Andrade B, Carvalho DP (2015)

Event: Thyroid Cancer

AMP-activated protein kinase (AMPK) is a sensor of cellular energy status that acts directly on cell proliferation and on transition from anaerobic to aerobic metabolic state. Our group showed that the activated form of AMPK (pAMPK) is overexpressed in papillary thyroid carcinoma (PTC) cases by immunohistochemistry. AMPK presents anti proliferative effects, so the biological meaning of AMPK activation in thyroid tumor cells and the consequences of its further activation on PTC metabolism is not known.

We used the PTC-derived BCPAP cell line to analyze cellular responses to a further stimulation of AMPK with the pharmacological activator 5-aminoimidazole-4 carboxamide ribonucleoside (1 mM AICAR). Cell viability was measured by MTT, and apoptosis was analyzed by the expression of Annexin V by Muse cell analyser. ROS was measured by DCFHD probe by FACS. Oxygen consumption was measured using high-resolution respirometry (Oroboros). Hexokinase (HK) tertiary structure was evaluated on a spectrofluorometer (Jasco).

BCPAP cells constitutively express high pAMPK levels, which correlates with the high glucose consumption rate (2 fold) and lactate production (2 fold), but lower oxygen consumption (30%) in these cells when compared to the nontumoral NTHYori cell line. AICAR exposure further stimulated these metabolic parameters, but decreased BCPAP cell viability (4 fold), with increased (1.5 fold) production of reactive oxygen species (ROS). HK is an enzyme of the glycolysis pathway that, upon binding to mitochondria, protects against ROS. We observed an increase in HK activity (1.4 fold) produced by AICAR, but not in its association with HK-mitochondrial activity. Purified HK experiments confirmed that ROS (100 μM Z2O2) alters the tertiary structure of HK decreasing its mitochondrial binding. The presence of NAC prevented cell death induced by AICAR treatment.

Overall, these results suggest that, despite the upregulation of glucose metabolism by AMPK, chronic activation of the enzyme with AICAR increases ROS levels promoting a negative regulation of HK-mitochondrial binding. The further increase in ROS production induced by AICAR might play a role in the induction of tumor cell apoptosis.


O2k-Network Lab: BR Rio de Janeiro Galina A


Labels: MiParea: Respiration  Pathology: Cancer  Stress:Cell death, Oxidative stress;RONS 




HRR: Oxygraph-2k 


Affiliations

UFRJ, Rio de Janeiro, Brazil