Mitochondrial oxidative phosphorylation and energetic status are reflected by morphology of mitochondrial network in INS-1E and HEP-G2 cells viewed by 4Pi microscopy

作者：Plecita Hlavata Lydie; Lessard Mark; Santorova Jitka; Bewersdorf Joerg; Jezek Petr^*

来源：Biochimica et Biophysica Acta-Bioenergetics, 2008, 1777(7-8): 834-846.

DOI：10.1016/j.bbabio.2008.04.002

• 3D morphology of mitochondrial network
• 4Pi microscopy
• uncoupling
• oxidative phosphorylation
• insulinoma INS-1E cell
• hepatocellular carcinoma HEP-G2 cell

Mitochondria in numerous cell types, especially in cultured cells, form a reticular network undergoing constant fusion and fission. The three dimensional (3D) morphology of these networks however has not been studied in detail to our knowledge. We have investigated insulinoma INS-1E and hepatocellular carcinoma HEP-G2 cells transfected with mitochondria-addressed GFP. Using 4Pi microscopy, 3D morphology changes responding to decreased oxidative phosphorylation and/or energetic status could be observed in these cells at an unprecedented 100 nm level of detail. In INSAE cells cultivated at 11 mM glucose, the mitoreticulum appears predominantly as one interconnected mitochondrion with a nearly constant 262 26 nrn tubule diameter. If cultured at 5 mM glucose, INS-1 E cells show 311 36 nm tubules coexisting with numerous flat cisternae. Similar interconnected 284 38 nm and 417 110 nm tubules were found in HEP-G2 cells cultivated at 5 mM and hyperglycaernic 25 mM glucose, respectively. With rotenone inhibiting respiration to -10%, disintegration into several reticula and numerous -300 run spheres or short tubules was observed. Deenergization by uncoupling additionally led to formation ofrings and bulky cisternae ofl.4 0.4 pm diameter. Rotenone and uncoupler acted synergically in INS-IE cells and increased fusion (ongoing with fission) forming bowl-like shapes. In HEP-G2 cells fission partially ceased with FCCP plus rotenone. Thus we have revealed previously undescribed details for shapes upon mitochondrial disintegration and clearly demonstrate that high resolution 3D microscopy is required for visualization of mitochondrial network. We recommend 4Pi microscopy as a new standard.

• 出版日期2008-8