Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry

Ivan Todorov, Indu Nair, Alina Avakian-Mansoorian, Jeffrey Rawson, Keiko Omori, Taihei Ito, Luis Valiente, Itzia Iglesias-Meza, Chris Orr, Keh Dong Shiang, Kevin Ferreri, Ismail H. Al-Abdullah, Yoko Mullen, Fouad Kandeel

Research output: Contribution to journalArticle

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Abstract

Background.: Assays for assessing human islet cell quality, which provide results before transplantation, would be beneficial to improve the outcomes for islet transplantation therapy. Parameters such as percent β-cell apoptosis and cell composition are found to vary markedly between different islet preparations and may serve as markers of islet quality. We have developed fluorescence-based assays using laser scanning cytometry for assessing β-cell apoptosis and islet cell composition on serial sections of intact isolated islets. Methods.: Isolated human islets were fixed in formalin and embedded in paraffin. Serial sections were immunostained for the pancreatic hormones and acinar and ductal cell markers. DNA fragmentation was used to label apoptotic cells. Stained cells were quantified using an iCys laser scanning cytometer. Results.: Islet preparations from 102 human pancreatic islet isolations were analyzed. For the whole set of islet preparations, we found a mean islet cell composition of 54.5%±1.2% insulin-positive, 33.9%±1.2% glucagon, 12.1%±0.7% somatostatin, and 1.5%±0.2% pancreatic polypeptide-positive cells. The apoptotic β cells were 2.85%±0.4% with a range of 0.27% to 18.3%. The percentage of apoptotic β cells correlated well (P<0.0001, n=59) with results obtained in vivo by transplantation of the corresponding islets in diabetic NODscid mice. Conclusions.: The analysis of whole, nondissociated islets for cell composition and β-cell apoptosis using laser scanning cytometry gives reliable and reproducible results and could be performed both before islet transplantation and on preserved cell blocks at any time in future. Thus, they can be a powerful tool for islet quality assessment.

Original languageEnglish
Pages (from-to)836-842
Number of pages7
JournalTransplantation
Volume90
Issue number8
DOIs
Publication statusPublished - 27-10-2010

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Laser Scanning Cytometry
Apoptosis
Islets of Langerhans
Islets of Langerhans Transplantation
Pancreatic Polypeptide-Secreting Cells
Pancreatic Hormones
Acinar Cells
DNA Fragmentation
Somatostatin
Glucagon
Paraffin
Formaldehyde
Lasers
Transplantation
Fluorescence

All Science Journal Classification (ASJC) codes

  • Transplantation

Cite this

Todorov, Ivan ; Nair, Indu ; Avakian-Mansoorian, Alina ; Rawson, Jeffrey ; Omori, Keiko ; Ito, Taihei ; Valiente, Luis ; Iglesias-Meza, Itzia ; Orr, Chris ; Shiang, Keh Dong ; Ferreri, Kevin ; Al-Abdullah, Ismail H. ; Mullen, Yoko ; Kandeel, Fouad. / Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry. In: Transplantation. 2010 ; Vol. 90, No. 8. pp. 836-842.
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title = "Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry",
abstract = "Background.: Assays for assessing human islet cell quality, which provide results before transplantation, would be beneficial to improve the outcomes for islet transplantation therapy. Parameters such as percent β-cell apoptosis and cell composition are found to vary markedly between different islet preparations and may serve as markers of islet quality. We have developed fluorescence-based assays using laser scanning cytometry for assessing β-cell apoptosis and islet cell composition on serial sections of intact isolated islets. Methods.: Isolated human islets were fixed in formalin and embedded in paraffin. Serial sections were immunostained for the pancreatic hormones and acinar and ductal cell markers. DNA fragmentation was used to label apoptotic cells. Stained cells were quantified using an iCys laser scanning cytometer. Results.: Islet preparations from 102 human pancreatic islet isolations were analyzed. For the whole set of islet preparations, we found a mean islet cell composition of 54.5{\%}±1.2{\%} insulin-positive, 33.9{\%}±1.2{\%} glucagon, 12.1{\%}±0.7{\%} somatostatin, and 1.5{\%}±0.2{\%} pancreatic polypeptide-positive cells. The apoptotic β cells were 2.85{\%}±0.4{\%} with a range of 0.27{\%} to 18.3{\%}. The percentage of apoptotic β cells correlated well (P<0.0001, n=59) with results obtained in vivo by transplantation of the corresponding islets in diabetic NODscid mice. Conclusions.: The analysis of whole, nondissociated islets for cell composition and β-cell apoptosis using laser scanning cytometry gives reliable and reproducible results and could be performed both before islet transplantation and on preserved cell blocks at any time in future. Thus, they can be a powerful tool for islet quality assessment.",
author = "Ivan Todorov and Indu Nair and Alina Avakian-Mansoorian and Jeffrey Rawson and Keiko Omori and Taihei Ito and Luis Valiente and Itzia Iglesias-Meza and Chris Orr and Shiang, {Keh Dong} and Kevin Ferreri and Al-Abdullah, {Ismail H.} and Yoko Mullen and Fouad Kandeel",
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Todorov, I, Nair, I, Avakian-Mansoorian, A, Rawson, J, Omori, K, Ito, T, Valiente, L, Iglesias-Meza, I, Orr, C, Shiang, KD, Ferreri, K, Al-Abdullah, IH, Mullen, Y & Kandeel, F 2010, 'Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry', Transplantation, vol. 90, no. 8, pp. 836-842. https://doi.org/10.1097/TP.0b013e3181f1db5d

Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry. / Todorov, Ivan; Nair, Indu; Avakian-Mansoorian, Alina; Rawson, Jeffrey; Omori, Keiko; Ito, Taihei; Valiente, Luis; Iglesias-Meza, Itzia; Orr, Chris; Shiang, Keh Dong; Ferreri, Kevin; Al-Abdullah, Ismail H.; Mullen, Yoko; Kandeel, Fouad.

In: Transplantation, Vol. 90, No. 8, 27.10.2010, p. 836-842.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Quantitative assessment of β-cell apoptosis and cell composition of isolated, undisrupted human islets by laser scanning cytometry

AU - Todorov, Ivan

AU - Nair, Indu

AU - Avakian-Mansoorian, Alina

AU - Rawson, Jeffrey

AU - Omori, Keiko

AU - Ito, Taihei

AU - Valiente, Luis

AU - Iglesias-Meza, Itzia

AU - Orr, Chris

AU - Shiang, Keh Dong

AU - Ferreri, Kevin

AU - Al-Abdullah, Ismail H.

AU - Mullen, Yoko

AU - Kandeel, Fouad

PY - 2010/10/27

Y1 - 2010/10/27

N2 - Background.: Assays for assessing human islet cell quality, which provide results before transplantation, would be beneficial to improve the outcomes for islet transplantation therapy. Parameters such as percent β-cell apoptosis and cell composition are found to vary markedly between different islet preparations and may serve as markers of islet quality. We have developed fluorescence-based assays using laser scanning cytometry for assessing β-cell apoptosis and islet cell composition on serial sections of intact isolated islets. Methods.: Isolated human islets were fixed in formalin and embedded in paraffin. Serial sections were immunostained for the pancreatic hormones and acinar and ductal cell markers. DNA fragmentation was used to label apoptotic cells. Stained cells were quantified using an iCys laser scanning cytometer. Results.: Islet preparations from 102 human pancreatic islet isolations were analyzed. For the whole set of islet preparations, we found a mean islet cell composition of 54.5%±1.2% insulin-positive, 33.9%±1.2% glucagon, 12.1%±0.7% somatostatin, and 1.5%±0.2% pancreatic polypeptide-positive cells. The apoptotic β cells were 2.85%±0.4% with a range of 0.27% to 18.3%. The percentage of apoptotic β cells correlated well (P<0.0001, n=59) with results obtained in vivo by transplantation of the corresponding islets in diabetic NODscid mice. Conclusions.: The analysis of whole, nondissociated islets for cell composition and β-cell apoptosis using laser scanning cytometry gives reliable and reproducible results and could be performed both before islet transplantation and on preserved cell blocks at any time in future. Thus, they can be a powerful tool for islet quality assessment.

AB - Background.: Assays for assessing human islet cell quality, which provide results before transplantation, would be beneficial to improve the outcomes for islet transplantation therapy. Parameters such as percent β-cell apoptosis and cell composition are found to vary markedly between different islet preparations and may serve as markers of islet quality. We have developed fluorescence-based assays using laser scanning cytometry for assessing β-cell apoptosis and islet cell composition on serial sections of intact isolated islets. Methods.: Isolated human islets were fixed in formalin and embedded in paraffin. Serial sections were immunostained for the pancreatic hormones and acinar and ductal cell markers. DNA fragmentation was used to label apoptotic cells. Stained cells were quantified using an iCys laser scanning cytometer. Results.: Islet preparations from 102 human pancreatic islet isolations were analyzed. For the whole set of islet preparations, we found a mean islet cell composition of 54.5%±1.2% insulin-positive, 33.9%±1.2% glucagon, 12.1%±0.7% somatostatin, and 1.5%±0.2% pancreatic polypeptide-positive cells. The apoptotic β cells were 2.85%±0.4% with a range of 0.27% to 18.3%. The percentage of apoptotic β cells correlated well (P<0.0001, n=59) with results obtained in vivo by transplantation of the corresponding islets in diabetic NODscid mice. Conclusions.: The analysis of whole, nondissociated islets for cell composition and β-cell apoptosis using laser scanning cytometry gives reliable and reproducible results and could be performed both before islet transplantation and on preserved cell blocks at any time in future. Thus, they can be a powerful tool for islet quality assessment.

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U2 - 10.1097/TP.0b013e3181f1db5d

DO - 10.1097/TP.0b013e3181f1db5d

M3 - Article

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AN - SCOPUS:78349295959

VL - 90

SP - 836

EP - 842

JO - Transplantation

JF - Transplantation

SN - 0041-1337

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