Abstract
Background and aimsAn intra-islet incretin system has been recently suggested to operate through modulation of the expression and activity of proconvertase 1/3 and 2 (PC1/3, PC2) in pancreatic alpha-cell accounting for local release of GLP-1. Little is known, whether this alpha-cell activity can be affected by the metabolic alterations occurring in type 2 diabetes, such as hyperglycemia, hyperlipidemia or hyperglucagonemia.Materials and methodsAlphaTC1/6 cells from a mice pancreatic cell line were incubated in the presence of two glucose (G) concentration (5.5 and 16.7 mM) for 16 h with or without free fatty acid, IL6 or glucagon. GLP-1 secretion was measured by ELISA and expression of PC1/3 and PC2 by RT-PCR and western blot; cell viability was determined by MTT method, Reactive Oxygen Species generation (ROS) by H2DCFDA fluorescence and apoptosis by Annexin staining and Propidium Iodine (PI) fluorescence.ResultsUpon 16.7G incubation, GLP-1 secretion (total and active) was significantly increased in parallel with a significant increment in PC1/3 expression, a slight increase in cell viability and ROS generation and by a decrement in PC2 expression with no change in cell apoptosis. When cells were incubated at 5.5mM glucose with FFA, also an increment in GLP-1 secretion and PC1/3 expression was observed together an increment in ROS generation, a decrement in cell viability, and a modest increment in apoptosis. When incubated with 16.7mM glucose with FFA, the increment in GLP-1 secretion was reduced to basal, accompanied by an increment in apoptosis and ROS generation. This was also observed with IL-6, but in this case, no modification in ROS generation or apoptosis was observed when compared to 16.7mM glucose. The presence of glucagon did not modify any of the parameters studied.ConclusionThese data suggest that under hyperglycemic, hyperlipidemia or inflammatory conditions, alpha cells can increase expression PC1/3 and activate GLP-1 secretion, which may contribute protecting both alpha and beta-cells from glucose and lipotoxicity, while this effect seems to be lost in the presence of both pathophysiological conditions.
Highlights
Glucagon like-peptide–1 (GLP-1), one of the main incretin hormones, is secreted by the entero-endocrine L cell of the intestine [1] after nutrient ingestion [2]
GLP-1 secretion was measured by ELISA and expression of proconvertase 1/3 (PC1/3) and PC2 by RTPCR and western blot; cell viability was determined by MTT method, Reactive Oxygen Species generation (ROS) by H2DCFDA fluorescence and apoptosis by Annexin staining and Propidium Iodine (PI) fluorescence
When cells were incubated at 5.5mM glucose with FFA, an increment in GLP-1 secretion and PC1/3 expression was observed together an increment in ROS generation, a decrement in cell viability, and a modest increment in apoptosis
Summary
Glucagon like-peptide–1 (GLP-1), one of the main incretin hormones, is secreted by the entero-endocrine L cell of the intestine [1] after nutrient ingestion [2]. Marchetti et al [4] were able to confirm the co-presence of GLP-1 and PC1/3 in the alpha cells of human pancreatic islets isolated from non-diabetic and type 2 diabetic cadaveric donors They showed that PC1/3 expression and GLP-1 secretion were more pronounced in the diabetic islets [4]. Similar effects have been observed in pregnant and neonatal mouse [6] and db/db [7] and ob/ob mice [8] progressing toward overt diabetes In these conditions a shift from glucagon- to GLP-1-positive cells was found along with an increment in cells with double positivity for GLP-1 and PC1/3. Whether this alpha-cell activity can be affected by the metabolic alterations occurring in type 2 diabetes, such as hyperglycemia, hyperlipidemia or hyperglucagonemia
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