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Photodynamic action of porphyrin on Ca2+ influx in endoplasmic reticulum: a comparison with mitochondria
F Ricchelli1, P Barbato, M Milani
1Consiglio Nazionale delle Ricerche, Centro Metalloproteine, Dipartimento di Biologia, Università di Padova, Italy. rchielli@civ.bio.unipd.it
Abstract:
We have studied the distribution properties of haematoporphyrin (HP) and protoporphyrin (PP) in mitochondria and endoplasmic reticulum after isolation from rat liver. The photosensitizing efficiency of porphyrin on the Ca2+ influx function of microsomes has been compared with that obtained on Ca2+ uptake in mitochondria. HP and PP are accumulated in microsomes to a greater extent than in mitochondria, both porphyrins binding to membrane protein sites. The Ca2+ influx functions of mitochondria and microsomes, before and after irradiation in the presence of HP or PP, were studied by following the changes in the free Ca2+ concentration in the medium as revealed by the variations in fluorescence intensity of the Ca2+ indicator Calcium Green-1. For the same amount of incorporated porphyrin, the Ca2+ influx function of microsomes is degraded by irradiation more rapidly than that of mitochondria. The protective effect of dithiothreitol suggests that thiol groups in the Ca2+-transporting enzyme are the preferential targets of the photodynamic effect. These results suggest that intracellular Ca2+ movements are altered primarily by the endoplasmic reticulum rather than by mitochondrial damage, in good agreement with other observations made in porphyrin-loaded irradiated cells.
Insights
Haematoporphyrin and protoporphyrin accumulate more in endoplasmic reticulum than mitochondria. Irradiation damages microsomes more, suggesting ER, not mitochondria, primarily alters intracellular calcium (Ca2+) movements.
Area of Science:
- Biochemistry
- Cell Biology
- Photodynamic Therapy
Background:
- Porphyrins like haematoporphyrin (HP) and protoporphyrin (PP) are photosensitizers.
- Mitochondria and endoplasmic reticulum (ER) are key cellular organelles involved in calcium (Ca2+) homeostasis.
- Understanding porphyrin distribution and photodynamic effects on these organelles is crucial for photodynamic therapy (PDT) applications.
Purpose of the Study:
- To investigate the distribution of HP and PP in rat liver mitochondria and ER.
- To compare the photosensitizing effects of HP and PP on Ca2+ transport in both organelles.
- To identify the primary cellular targets of photodynamic damage affecting Ca2+ regulation.
Main Methods:
- Isolation of mitochondria and microsomes (ER) from rat liver.
- Quantification of porphyrin accumulation in isolated organelles.
- Measurement of Ca2+ influx/uptake using the fluorescent indicator Calcium Green-1.
- Assessment of photodynamic damage after in vitro irradiation in the presence of porphyrins and dithiothreitol.
Main Results:
- HP and PP accumulated to a greater extent in microsomes than mitochondria, binding to membrane proteins.
- Irradiation of porphyrin-loaded microsomes led to more rapid degradation of Ca2+ influx function compared to mitochondria.
- Dithiothreitol offered protection, indicating thiol groups in Ca2+-transporting enzymes are targets.
Conclusions:
- Endoplasmic reticulum is primarily affected by photodynamic damage, leading to altered intracellular Ca2+ movements.
- Mitochondrial Ca2+ uptake is less sensitive to photodynamic effects compared to ER Ca2+ influx.
- These findings support the ER as the main site of photodynamic disruption of Ca2+ homeostasis in porphyrin-sensitized cells.