Related Experiment Videos
Differentiation between glucose-induced desensitization of insulin secretion and beta-cell exhaustion in the HIT-T15
E D Kilpatrick1, R P Robertson
1Pacific Northwest Research Foundation and the Division of Metabolism, Endocrinology, and Nutrition, University of Washington, Seattle 98122, USA.
Abstract:
Refractoriness of the pancreatic beta-cell to glucose stimulation plays a role in the secretory defect of NIDDM, but the mechanisms underlying this refractoriness are not clear. The purpose of this study was to determine whether the HIT-T15 pancreatic beta-cell line can be used as an investigative model for refractoriness of glucose-induced insulin secretion and, if so, whether the mechanism for this refractoriness involves alteration in stimulus-secretion coupling (desensitization) or results from exhaustion of insulin stores. In perifusion experiments, acute insulin responses (AIRs) in HIT-T15 cells progressively diminished during consecutive 5-min glucose (11.1 mmol/l) pulses (G) given every 20 min (G1=9.2+/-1.3, G2=4.1+/-1.0, G3=2.7+/-0.7, G4=2.5+/-1.1 microU/ml). To determine whether this refractoriness for glucose extended to the potentiating effects of glucose on nonglucose secretagogues, cells were challenged with arginine after desensitization with glucose. In HIT-T15 cells, the response to the arginine pulse (16.7+/-5.2 microU/ml) after three glucose pulses was significantly less (P < 0.01) than the response to a control arginine pulse (29.6+/-12.1 microU/ml) preceded by an infusion of buffer in the absence of glucose pulses. Variable rest periods after desensitization allowed recovery of the AIR in HIT-T15 cells; responses 30, 60, 90, and 120 min after the desensitization procedure increased in a stepwise fashion (3.8+/-2.7, 4.5+/-2.7, 7.8+/-5.2, and 9.7+/-5.3 microU/ml, respectively). To differentiate desensitization from exhaustion of insulin stores, studies were performed in the presence of epinephrine, a potent inhibitor of insulin secretion. In HIT-T15 cells, after three pulses of glucose during the epinephrine infusion, epinephrine was discontinued and the insulin response to a fourth pulse was assessed. The G4 AIR (1.9+/-0.6 microU/ml) was markedly less than a control G4 AIR (5.4+/-1.2 microU/ml) that followed an epinephrine infusion alone with no concurrent glucose pulses. Beta-cell refractoriness was also induced in the HIT-T15 cell using 45-min steady-state infusions of glucose. Cells were exposed to a 45-min infusion of either 3.7 or 11.1 mmol/l glucose, rested for 20 min in the absence of glucose, and then challenged with a 5-min, 11.1 mmol/l glucose pulse. In both cases, the AIR to the 5-min pulse (10.2+/-5.1 and 2.9+/-1.4 microU/ml after the 3.7 and 11.1 mmol/l infusion, respectively) was lower than the AIR to a control pulse (27.4+/-5.9 microU/ml) not preceded by glucose infusion. These studies demonstrated that the HIT-T15 cell line is an appropriate model for studying mechanisms of beta-cell refractoriness to glucose signaling. The short-term intensive glucose stimulation paradigms used in our studies induced an abnormality in insulin secretion that is consistent with desensitization but not beta-cell exhaustion.
Insights
The HIT-T15 pancreatic beta-cell line effectively models glucose-induced insulin secretion refractoriness. This refractoriness stems from desensitization, not insulin store exhaustion, offering insights into NIDDM mechanisms.
Area of Science:
- Endocrinology
- Cell Biology
- Metabolic Research
Background:
- Pancreatic beta-cell refractoriness to glucose contributes to NIDDM secretory defects.
- Mechanisms underlying beta-cell glucose refractoriness remain unclear.
Purpose of the Study:
- To validate the HIT-T15 cell line as a model for studying glucose-induced insulin secretion refractoriness.
- To elucidate whether refractoriness results from desensitization or insulin store exhaustion.
Main Methods:
- Perifusion experiments with sequential glucose pulses to assess acute insulin responses (AIRs).
- Challenge with arginine post-glucose desensitization to evaluate potentiating effects.
- Studies with epinephrine to differentiate desensitization from exhaustion.
- Induction of refractoriness using steady-state glucose infusions.
Main Results:
- HIT-T15 cells showed progressively diminished AIRs with repeated glucose stimulation, indicating refractoriness.
- Glucose desensitization reduced the response to arginine.
- Insulin secretion recovered after rest periods, suggesting reversibility.
- Epinephrine studies and steady-state infusions supported desensitization over exhaustion.
Conclusions:
- The HIT-T15 cell line serves as a suitable model for investigating beta-cell refractoriness to glucose.
- Short-term intensive glucose stimulation induces desensitization, not exhaustion, of insulin secretion.