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On-line monitoring ...
On-line monitoring of apoptosis in insulin-secreting cells
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- Kohler, M (författare)
- Karolinska Institutet
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- Zaitsev, SV (författare)
- Karolinska Institutet
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- Zaitseva, II (författare)
- Karolinska Institutet
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- Leibiger, B (författare)
- Karolinska Institutet
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- Leibiger, IB (författare)
- Karolinska Institutet
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Turunen, M (författare)
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Kapelioukh, IL (författare)
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- Bakkman, L (författare)
- Karolinska Institutet
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Appelskog, IB (författare)
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de Monvel, JB (författare)
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- Imreh, G (författare)
- Karolinska Institutet
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- Berggren, PO (författare)
- Karolinska Institutet
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(creator_code:org_t)
- American Diabetes Association, 2003
- 2003
- Engelska.
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 52:12, s. 2943-2950
- Relaterad länk:
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http://kipublication...
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https://doi.org/10.2...
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Abstract
Ämnesord
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- Apoptosis was monitored in intact insulin-producing cells both with microfluorometry and with two-photon laser scanning microscopy (TPLSM), using a fluorescent protein based on fluorescence resonance energy transfer (FRET). TPLSM offers three-dimensional spatial information that can be obtained relatively deep in tissues. This provides a potential for future in vivo studies of apoptosis. The cells expressed a fluorescent protein (C-DEVD-Y) consisting of two fluorophores, enhanced cyan fluorescent protein (ECFP) and enhanced yellow fluorescent protein (EYFP), linked by the amino acid sequence DEVD selectively cleaved by caspase-3–like proteases. FRET between ECFP and EYFP in C-DEVD-Y could therefore be monitored on-line as a sensor of caspase-3 activation. The relevance of using caspase-3 activation to indicate β-cell apoptosis was demonstrated by inhibiting caspase-3–like proteases with Z-DEVD-fmk and thereby showing that caspase-3 activation was needed for high-glucose–and cytokine-induced apoptosis in the β-cell and for staurosporine-induced apoptosis in RINm5F cells. In intact RINm5F cells expressing C-DEVD-Y and in MIN6 cells expressing the variant C-DEVD-Y2, FRET was lost at 155 ± 23 min (n = 9) and 257 ± 59 min (n = 4; mean ± SE) after activation of apoptosis with staurosporine (6 μmol/l), showing that this method worked in insulin-producing cells.
Publikations- och innehållstyp
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- art (ämneskategori)
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Diabetes
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Till lärosätets databas
- Av författaren/redakt...
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Kohler, M
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Zaitsev, SV
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Zaitseva, II
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Leibiger, B
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Leibiger, IB
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Turunen, M
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visa fler...
-
Kapelioukh, IL
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Bakkman, L
-
Appelskog, IB
-
de Monvel, JB
-
Imreh, G
-
Berggren, PO
-
visa färre...
- Artiklar i publikationen
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Diabetes
- Av lärosätet
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Karolinska Institutet