Human islet mass, morphology, and survival after cryopreservation using the Edmonton protocol.
The aim of this study was to assess recovery, cell death, and cell composition of post-thaw cultured human islets. Cryopreserved islets were provided by the Clinical Islet Transplant Program, Edmonton, Canada. Islets were processed using media prepared in accordance with Pre-Edmonton and Edmonton protocols. Cryopreserved islets were rapidly thawed and cultured for 24 h, 3 d, 5 d, and 7 d, following which they were processed for histology. Islet quantification, integrity, morphology and tissue turnover were studied via hematoxylin and eosin stained sections. Ultrastructure was studied by electron microscopy and endocrine cell composition by immunohistochemistry. Using the Pre-Edmonton protocol, islet recovery was 50.1% and islet survival was 50% at 24 h while for the Edmonton protocol, the islet recovery was 69.4% (p<0.001) and islet survival, 50% at ≈2.5 d. With an increasing culture duration although the physical integrity was retained there was an increasing loss of cohesivity both at light microscopic and at ultrastructure level regardless of the protocols used. Percentage islet survival and tissue turnover correlated negatively with culture duration in both protocols. The Edmonton protocol appears to preserve the islets better. However, culture duration adversely affects islet survival and quality, indicating the need for more optimal cryopreservation and culture techniques.
Madras Diabetes Research Foundation & Dr. Mohan's Diabetes Specialties Centre; WHO Collaborating Centre for Noncommunicable Diseases-Prevention and Control; Chennai, Tamilnadu, India.
Priya M
PM
Miranda
Madras Diabetes Research Foundation & Dr. Mohan's Diabetes Specialties Centre; WHO Collaborating Centre for Noncommunicable Diseases-Prevention and Control; Chennai, Tamilnadu, India.
Viswanathan
V
Mohan
Saveetha Medical College; Thandalam, Chennai, India.
Sekhar
S
Ganthimathy
Madras Diabetes Research Foundation & Dr. Mohan's Diabetes Specialties Centre; WHO Collaborating Centre for Noncommunicable Diseases-Prevention and Control; Chennai, Tamilnadu, India.
Ranjit M
RM
Anjana
Christian Medical College; Vellore, Tamil Nadu, India.
S
S
Gunasekaran
Tamilnadu Veterinary and Animal Science University; Chennai, India.
Venkatachalam
V
Thiagarajan
University of Alberta; Edmonton, Alberta, Canada.
Thomas A
TA
Churchill
University of Alberta; Edmonton, Alberta, Canada.
Tatsuya
T
Kin
University of Alberta; Edmonton, Alberta, Canada.
A M James
AM
Shapiro
University of California; Irvine, CA USA.
Jonathan R T
JR
Lakey
D017209:Apoptosis; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods*; D005934:Glucagon / Q000032:analysis; D006801:Humans; D007328:Insulin / Q000032:analysis; D007515:Islets of Langerhans / Q000033:anatomy & histology* / Q000737:chemistry / Q000166:cytology; D008940:Mitotic Index; D009336:Necrosis; D013004:Somatostatin / Q000032:analysis; D046509:Tissue Culture Techniques
D007328:Insulin; D013004:Somatostatin; D005934:Glucagon
clinical phase; cryopreservation; immunohistochemistry; islet isolation; islet transplantation; post-thaw culture; pre-clinical phase; ultrastructure TUNEL