OBJECTIVE-Human islet amyloid polypeptide (hIAPP) aggregation plays a major role in the development of islet amyloidosis in type 2 diabetes. It is known that extracellular hIAPP oligomers are toxic to pancreatic ␤-cells and associated with apoptosis. We therefore investigated the molecular mechanism by which extracellular hIAPP mediates pancreatic ␤-cell apoptosis. RESEARCH DESIGN AND METHODS-MIN6cells and primary cultures of human pancreatic islets were treated with freshly dissolved hIAPP peptide. Morphology of the cultures was evaluated by electron microscopy. Gene expression was analyzed by microarray, RT-PCR, and immunoblot. Calcium levels were measured in fura-2-loaded cells. Apoptosis was quantified by cytometry.RESULTS-Increased expression of several heat shock proteins and activation of the spliced form of XBP-1, a transcription factor for overexpression of chaperones during endoplasmic reticulum (ER) stress, were detected together with morphological evidence of ER dysfunction. Intracellular calcium overload was detected in association with this process. Moreover, reduction in the proteasome activity, which was detected over time, contributed to the intracellular accumulation of ubiquitinated proteins, leading to a functional suppression of the ubiquitinproteasome pathway. In addition, impairment of the proteasome function contributed to apoptosis, while, despite the presence of hIAPP, cell viability improved when a proteasome activator was overexpressed. The key cytotoxic events induced by extracellular hIAPP were also observed in treated human islets. T he presence of amyloid deposits within the pancreatic islets is a pathophysiological hallmark of type 2 diabetes (1,2). Islet amyloid polypeptide (IAPP), also known as amylin, is the main component of islet amyloid (3,4). It is produced by ␤-cells and cosecreted with insulin in response to nutrient stimuli (5-7). A target region between positions 20 and 29 is thought to be responsible for amyloid fibril formation by the human peptide (8). Several point mutations located in the human IAPP (hIAPP) promoter and missense point mutations on the encoding gene region have been identified by our group and others with different prevalence according to ethnic origin (9,10). However, because the sequence of hIAPP is identical in diabetic and nondiabetic individuals, the process of amyloid formation remains poorly understood. CONCLUSIONS-OurIslet amyloid colocalizes with areas of cell degeneration, and the process of amyloidosis has been associated with progressive loss of pancreatic ␤-cell mass by apoptosis and, thus, much of the pathology of type 2 diabetes (11)(12)(13)(14). A number of studies showed that the toxicity of hIAPP and other amyloidogenic peptides lies in the oligomeric intermediates rather than in the mature fibrils (15-17).The endoplasmic reticulum (ER) integrates protein synthesis and folding and calcium storage and signaling. Several cellular stress conditions can cause ER dysfunction and protein misfolding (18). To overcome ER stress, cells ini...

Alterations in [Ca(2+)](i) play a key role in hIAPP-induced beta cell cytotoxicity. By electron microscopy, we detected extracellular hIAPP aggregates adjacent to irregular invaginated regions of the plasma membrane. We propose that TRPV4 channels may sense physical changes in the plasma membrane induced by hIAPP aggregation, enabling Ca(2+) entry, membrane depolarisation and activation of L-type Ca(2+) channels. Decreasing the activity of TRPV4 prevented hIAPP-induced [Ca(2+)](i) changes, reduced hIAPP-triggered ER stress and improved cell viability.

Previous gene function analyses have indicated that HOXA9, DEK, CBL and CSF1R are aberrantly expressed in acute myeloid leukemia (AML). We analyzed the expression of these genes in a series of 41 adult patients with AML using quantitative real-time RT-PCR, and tested the association of the expression with the following hematologic and clinical parameters: age, FAB, immunophenotype and karyotype aberrations. A high proportion of the patients showed over- or underexpression of the analyzed genes. DEK was overexpressed in 98% of the cases, whereas CBL, CSF1R and HOXA9 were either overexpressed in 20%, 17% and 78% or underexpressed in 20%, 42% and 15% of the cases, respectively. Patients whose karyotype contained t(8;21)(q22;q22), showed lower relative expression of HOXA9 at a statistically significant level (p < 0.05). Bone marrow samples without expression of CD34 antigen were associated with either overexpression of DEK or HOXA9. Furthermore, an association was found between the AML-M2 subtype and lower expression of CBL, CSF1R or HOXA9, and between the AML-M5 subtype and CBL or CSF1R overexpression.