Effect of Environmental Conditions on Intracellular pH of Hybridoma Cells Cultured in Bioreactors

To test the hypothesis that hypoxia may induce cellular acidification during chemotransduction in the carotid body, we compared the effects of hypoxia and of extracellular acidosis on intracellular pH (pHi) of glomus cells cultured from rat and cat carotid bodies. The cells were prepared and cultured for 2–7 days. The plated cells were loaded with a pH-sensitive fluorescent probe, SNARF-1-acetoxymethyl ester, and were placed in a closed chamber and superfused. The effects of lowering PO2 and pH in the superfusion medium containing CO2-HCO3- buffer on the glomus cell pHi were measured at 37 degrees C. The pHi was measured in a single or a few isolated cells with single excitation at 540 nm and dual emission at 590 and 640 nm, after the exposure to different PO2 levels from 132 to 43, 14, and 1–2 Torr for 10–12 min in the closed chamber. The resting pHi values were 7.263 +/- 0.008 for rat and 7.175 +/- 0.004 for cat carotid body glomus cells. For a decrease of PO2 from 132 Torr to 14 Torr, the change in pHi values, on average, for cat and rat glomus cells was 0.034 lower, and with PO2 decrease to 1–2 Torr for the cat glomus cells, the change in pHi values was 0.051 lower. On the other hand, when the perfusate pH values were decreased from 7.4 to 6.9 during normoxia, the reduction of change in pHi values were 0.327 for the rat and 0.397 for the cat. Thus glomus cell pHi change due to low PO2 exposure was not significant and was not commensurate with the large increases in the chemosensory activity.

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Low Radiation Environment Switches the Overgrowth-Induced Cell Apoptosis Toward Autophagy

Frontiers in Public Health ◽

10.3389/fpubh.2020.594789 ◽

2021 ◽

Vol 8 ◽

Author(s):

Mariafausta Fischietti ◽

Emiliano Fratini ◽

Daniela Verzella ◽

Davide Vecchiotti ◽

Daria Capece ◽

...

Keyword(s):

National Laboratory ◽

Growth Stress ◽

Radiation Environment ◽

Hybridoma Cells ◽

Environmental Radiation ◽

Biological Responses ◽

Italian National Health ◽

Set Up ◽

Cells Cultured

Low radiation doses can affect and modulate cell responses to various stress stimuli, resulting in perturbations leading to resistance or sensitivity to damage. To explore possible mechanisms taking place at an environmental radiation exposure, we set-up twin biological models, one growing in a low radiation environment (LRE) laboratory at the Gran Sasso National Laboratory, and one growing in a reference radiation environment (RRE) laboratory at the Italian National Health Institute (Istituto Superiore di Sanità, ISS). Studies were performed on pKZ1 A11 mouse hybridoma cells, which are derived from the pKZ1 transgenic mouse model used to study the effects of low dose radiation, and focused on the analysis of cellular/molecular end-points, such as proliferation and expression of key proteins involved in stress response, apoptosis, and autophagy. Cells cultured up to 4 weeks in LRE showed no significant differences in proliferation rate compared to cells cultured in RRE. However, caspase-3 activation and PARP1 cleavage were observed in cells entering to an overgrowth state in RRE, indicating a triggering of apoptosis due to growth-stress conditions. Notably, in LRE conditions, cells responded to growth stress by switching toward autophagy. Interestingly, autophagic signaling induced by overgrowth in LRE correlated with activation of p53. Finally, the gamma component of environmental radiation did not significantly influence these biological responses since cells grown in LRE either in incubators with or without an iron shield did not modify their responses. Overall, in vitro data presented here suggest the hypothesis that environmental radiation contributes to the development and maintenance of balance and defense response in organisms.

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