Beta cell chromogranin B is partially segregated in distinct granules and can be released separately from insulin in response to stimulation

T. Giordano, C. Brigatti, P. Podini, E. Bonifacio, J. Meldolesi, M. L. Malosio

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Aims/hypothesis: We investigated, in three beta cell lines (INS-1E, RIN-5AH, betaTC3) and in human and rodent primary beta cells, the storage and release of chromogranin B, a secretory protein expressed in beta cells and postulated to play an autocrine role. We asked whether chromogranin B is stored together with and discharged in constant ratio to insulin upon various stimuli. Methods: The intracellular distribution of insulin and chromogranin B was revealed by immunofluorescence followed by three-dimensional image reconstruction and by immunoelectron microscopy; their stimulated discharge was measured by ELISA and immunoblot analysis of homogenates and incubation media. Results: Insulin and chromogranin B, co-localised in the Golgi complex/trans-Golgi network, appeared largely segregated from each other in the secretory granule compartment. In INS-1E cells, the percentage of granules positive only for insulin or chromogranin B and of those positive for both was 66, 7 and 27%, respectively. In resting cells, both insulin and chromogranin B were concentrated in the granule cores; upon stimulation, chromogranin B (but not insulin) was largely redistributed to the core periphery and the surrounding halo. Strong stimulation with a secretagogue mixture induced parallel release of insulin and chromogranin B, whereas with 3-isobutyl-1-methylxantine and forskolin ± high glucose release of chromogranin B predominated. Weak, Ca2+-dependent stimulation with ionomycin or carbachol induced exclusive release of chromogranin B, suggesting a higher Ca2+ sensitivity of the specific granules. Conclusions/interpretation: The unexpected complexity of the beta cell granule population in terms of heterogeneity, molecular plasticity and the differential discharge, could play an important role in physiological control of insulin release and possibly also in beta cell pathology.

Original languageEnglish
Pages (from-to)997-1007
Number of pages11
JournalDiabetologia
Volume51
Issue number6
DOIs
Publication statusPublished - Jun 2008

Fingerprint

Chromogranin B
Insulin
trans-Golgi Network
Ionomycin
Computer-Assisted Image Processing
Immunoelectron Microscopy
Three-Dimensional Imaging
Secretory Vesicles
Carbachol
Golgi Apparatus
Colforsin
Fluorescent Antibody Technique
Rodentia

Keywords

  • Beta cell lines
  • Beta cells
  • Carbachol
  • Chromogranin B
  • INS-1E
  • Insulin
  • Ionomycin
  • Islets, dissociation of storage and secretion
  • Regulated secretion

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Internal Medicine

Cite this

Beta cell chromogranin B is partially segregated in distinct granules and can be released separately from insulin in response to stimulation. / Giordano, T.; Brigatti, C.; Podini, P.; Bonifacio, E.; Meldolesi, J.; Malosio, M. L.

In: Diabetologia, Vol. 51, No. 6, 06.2008, p. 997-1007.

Research output: Contribution to journalArticle

Giordano, T. ; Brigatti, C. ; Podini, P. ; Bonifacio, E. ; Meldolesi, J. ; Malosio, M. L. / Beta cell chromogranin B is partially segregated in distinct granules and can be released separately from insulin in response to stimulation. In: Diabetologia. 2008 ; Vol. 51, No. 6. pp. 997-1007.
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T1 - Beta cell chromogranin B is partially segregated in distinct granules and can be released separately from insulin in response to stimulation

AU - Giordano, T.

AU - Brigatti, C.

AU - Podini, P.

AU - Bonifacio, E.

AU - Meldolesi, J.

AU - Malosio, M. L.

PY - 2008/6

Y1 - 2008/6

N2 - Aims/hypothesis: We investigated, in three beta cell lines (INS-1E, RIN-5AH, betaTC3) and in human and rodent primary beta cells, the storage and release of chromogranin B, a secretory protein expressed in beta cells and postulated to play an autocrine role. We asked whether chromogranin B is stored together with and discharged in constant ratio to insulin upon various stimuli. Methods: The intracellular distribution of insulin and chromogranin B was revealed by immunofluorescence followed by three-dimensional image reconstruction and by immunoelectron microscopy; their stimulated discharge was measured by ELISA and immunoblot analysis of homogenates and incubation media. Results: Insulin and chromogranin B, co-localised in the Golgi complex/trans-Golgi network, appeared largely segregated from each other in the secretory granule compartment. In INS-1E cells, the percentage of granules positive only for insulin or chromogranin B and of those positive for both was 66, 7 and 27%, respectively. In resting cells, both insulin and chromogranin B were concentrated in the granule cores; upon stimulation, chromogranin B (but not insulin) was largely redistributed to the core periphery and the surrounding halo. Strong stimulation with a secretagogue mixture induced parallel release of insulin and chromogranin B, whereas with 3-isobutyl-1-methylxantine and forskolin ± high glucose release of chromogranin B predominated. Weak, Ca2+-dependent stimulation with ionomycin or carbachol induced exclusive release of chromogranin B, suggesting a higher Ca2+ sensitivity of the specific granules. Conclusions/interpretation: The unexpected complexity of the beta cell granule population in terms of heterogeneity, molecular plasticity and the differential discharge, could play an important role in physiological control of insulin release and possibly also in beta cell pathology.

AB - Aims/hypothesis: We investigated, in three beta cell lines (INS-1E, RIN-5AH, betaTC3) and in human and rodent primary beta cells, the storage and release of chromogranin B, a secretory protein expressed in beta cells and postulated to play an autocrine role. We asked whether chromogranin B is stored together with and discharged in constant ratio to insulin upon various stimuli. Methods: The intracellular distribution of insulin and chromogranin B was revealed by immunofluorescence followed by three-dimensional image reconstruction and by immunoelectron microscopy; their stimulated discharge was measured by ELISA and immunoblot analysis of homogenates and incubation media. Results: Insulin and chromogranin B, co-localised in the Golgi complex/trans-Golgi network, appeared largely segregated from each other in the secretory granule compartment. In INS-1E cells, the percentage of granules positive only for insulin or chromogranin B and of those positive for both was 66, 7 and 27%, respectively. In resting cells, both insulin and chromogranin B were concentrated in the granule cores; upon stimulation, chromogranin B (but not insulin) was largely redistributed to the core periphery and the surrounding halo. Strong stimulation with a secretagogue mixture induced parallel release of insulin and chromogranin B, whereas with 3-isobutyl-1-methylxantine and forskolin ± high glucose release of chromogranin B predominated. Weak, Ca2+-dependent stimulation with ionomycin or carbachol induced exclusive release of chromogranin B, suggesting a higher Ca2+ sensitivity of the specific granules. Conclusions/interpretation: The unexpected complexity of the beta cell granule population in terms of heterogeneity, molecular plasticity and the differential discharge, could play an important role in physiological control of insulin release and possibly also in beta cell pathology.

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KW - Carbachol

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KW - Insulin

KW - Ionomycin

KW - Islets, dissociation of storage and secretion

KW - Regulated secretion

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