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Effects of free fatty acid on polymerization of islet amyloid polypeptide (IAPP) in vitro and on amyloid fibril formation in cultivated isolated islets of transgenic mice overexpressing human IAPP
Linköping University, Department of Molecular and Clinical Medicine, Molecular and Immunological Pathology. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Biomedicine and Surgery, Cell biology. Linköping University, Faculty of Health Sciences.
2002 (English)In: Molecular medicine (Cambridge, Mass. Print), ISSN 1076-1551, Vol. 8, no 12, 863-868 p.Article in journal (Refereed) Published
Abstract [en]

BACKGROUND: Islet amyloid polypeptide (IAPP) is deposited as amyloid in the islets of Langerhans in type 2 diabetes. The mechanism behind the formation of the cytotoxic fibrils is unknown. Islet amyloid develops in a mouse IAPP null mouse strain that expresses human IAPP (+hIAPP/-mIAPP) after 9 months on a high-fat diet. Herein we investigate the effect that individual free fatty acids (FFAs) exert on formation of amyloid-like fibrils from synthetic IAPP and the effects of FFAs on IAPP polymerization in +hIAPP/-mIAPP islets cultivated in vitro.

MATERIALS AND METHODS: In the study myristic acid, palmitic acid, stearic acid, oleic acid, and linoleic acid were used together with albumin. Thioflavin T (Th T) assay was used for quantification of amyloid-like fibrils. Islets were isolated from the +hIAPP/-mIAPP transgenic strain and cultured in the presence of the FFAs for 2 days. Immuno-electron microscopy was used for evaluation.

RESULTS: The Th T assay showed that all studied FFAs potentiated fibril formation but that myristic acid revealed the highest capacity. In some cells from cultured islets, intragranular aggregates were present. These aggregates had a filamentous appearance and labeled with antibodies against IAPP. In some cells cultured in the presence of linoleic acid, large amounts of intracellular amyloid were present. Earlier, this has not been observed after such a short incubation period.

CONCLUSIONS: Our studies suggest that FFAs can potentiate amyloid formation in vitro, probably without being integrated in the fibril. Cultivation of +hIAPP/-mIAPP transgenic mouse islets with FFAs results in altered morphology of the secretory granules with appearance of IAPP- immunoreactive fibrillar material. We suggest that such fibrillar material may seed extracellular amyloid formation after exocytosis.

Place, publisher, year, edition, pages
2002. Vol. 8, no 12, 863-868 p.
National Category
Medical and Health Sciences
URN: urn:nbn:se:liu:diva-26538Local ID: 11099OAI: diva2:247087
On the day of the defence day the status of this article was a manuscriptAvailable from: 2009-10-08 Created: 2009-10-08 Last updated: 2012-08-17Bibliographically approved
In thesis
1. Islet amyloid polypeptide (IAPP): Mechanisms of Amyloidogenesis in the Pancreatic Islets and Potential Roles in Diabetes Mellitus
Open this publication in new window or tab >>Islet amyloid polypeptide (IAPP): Mechanisms of Amyloidogenesis in the Pancreatic Islets and Potential Roles in Diabetes Mellitus
2001 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Islet amyloid is the most common characteristic feature of the islets in type 2 diabetes, being found in up to 90% of diabetic patients at post-mortem. lt has as its unique component the islet beta-cell peptide islet amyloid polypeptide (IAPP), which is eo-secreted with insulin. Because all human subjects produce and secrete the amyloidogenic form of IAPP, yet not all develop islet amyloid, some other factors must be involved in islet amyloid formation. The aim of the research presented in this thesis was to study factors of importance for the IAPP amyloidogenesis in type 2 diabetes. We developed a mouse monoclonal antibody to raVmouse IAPP (MAb4A5). MAb4A5 shows reactivity with IAPP in different species without detecting its close relative CGRP. In the pancreatic islets from patients with type 2 diabetes and diabetic cat, MAb4A5 labels immunohistochemically cellular IAPP but not IAPP in islet amyloid deposits. In contrast to MAb4A5 polyclonal rabbitiAPP antisera label beta cells close to amyloid only weakly, but label strongly IAPP in its amyloid form. The varying findings of IAPP immunoreactivity in pancreatic islets indicate that IAPP undergoes structural changes (impaired cleavage of proiAPP, conformational change, or post-transitional modifications) during the amyloidogenesis. A potentially important finding was the increased IAPP immunoreactivity in beta cells in islets of impaired glucose tolerant cats, irrespective of presence of amyloid in these islets. The finding may indicate that the formation of first islet amyloid occurs before Type 2 diabetes is manifest. Given the immunohistochemical results with MAb 4A5, we investigated whether the altered immunoreactivity of IAPP in association with the amyloidogenesis is due to a modification of IAPP (e.g. non-enzymatic glycation). We used synthetic IAPP fibrils glycated in vitro to study if non-enzymatic glycation may result in loss of an antigenic epitope. The results showed that a possible explanation of the lack of immunoreactivity of islet amyloid with MAb 4A5 actually is an nonenzymatic glycation. Association of an IAPP gene mutation with Type 2 diabetes has been found in the Japanese and Chinese population. We studied the possible enhanced fibril formation capacity of the mutant IAPP in vitro. Full-length and truncated IAPPS20G can form more amyloid-like fibrils and do this faster compared to wild type IAPP in vitro. We concluded that mutant (S20G) IAPP is a more amyloidogenic IAPP molecule and may be associated with an increased islet amyloid formation in vivo.

Based on the reports on the occurrence of islet amyloid in transgenic mice fed high fat diet, we investigated effects of free fatty acids on IAPP amyloid formation in isolated islets from transgenic mice expressing the gene for human IAPP but deficient of endogenous murine IAPP, and effects of FFAs on polymerization of IAPP in vitro. We found free fatty acids accelerate and increase polymerization of IAPP in vitro and promote amyloid like aggregation occurring in cultivated transgenic mouse isolated islets. All these results indicate that the pathogenesis of the islet amyloid may be a complex process involving many different mechanisms.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet, 2001. 70 p.
Linköping University Medical Dissertations, ISSN 0345-0082 ; 655
National Category
Medical and Health Sciences
urn:nbn:se:liu:diva-28607 (URN)13762 (Local ID)91-7219-756-0 (ISBN)13762 (Archive number)13762 (OAI)
Public defence
2001-01-12, Berzeliussalen, Universitetssjukhuset, Linköping, 13:30 (Swedish)
Available from: 2009-10-09 Created: 2009-10-09 Last updated: 2012-08-17Bibliographically approved

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