Noninvasive MRI of β-cell function using a Zn2+-responsive contrast agent

Abstract

Elevation of postprandial glucose stimulates release of insulin from granules stored in pancreatic islet β-cells. We demonstrate here that divalent zinc ions coreleased with insulin from β-cells in response to high glucose are readily detected by MRI using the Zn(2+)-responsive T(1) agent, GdDOTA-diBPEN. Image contrast was significantly enhanced in the mouse pancreas after injection of a bolus of glucose followed by a low dose of the Zn(2+) sensor. Images of the pancreas were not enhanced by the agent in mice without addition of glucose to stimulate insulin release, nor were images enhanced in streptozotocin-treated mice with or without added glucose. These observations are consistent with MRI detection of Zn(2+) released from β-cells only during glucose-stimulated insulin secretion. Images of mice fed a high-fat (60%) diet over a 12-wk period and subjected to this same imaging protocol showed a larger volume of contrast-enhanced pancreatic tissue, consistent with the expansion of pancreatic β-cell mass during fat accumulation and progression to type 2 diabetes. This MRI sensor offers the exciting potential for deep-tissue monitoring of β-cell function in vivo during development of type 2 diabetes or after implantation of islets in type I diabetic patients.

Fig. 1.

Schematic illustration of the steps involved in insulin (brown spheres) and Zn²⁺ (blue triangles) release from β-cells during GSIS. Zn²⁺ released from β-cells during GSIS forms a 2:1 complex with GdDOTA-diBPEN (yellow squares) which, upon forming a complex with albumin [human serum albumin (HSA), pink], shows a ≈3.5-fold increase in r₁ at 23 MHz³ and a ≈1.5-fold increase in r₁ at 400 MHz. The two plots show the increase in water proton relaxivity, r₁, of GdDOTA-diBPEN (1 mM) after incremental addition of ZnCl₂ either in the presence (squares) or absence (diamonds) of albumin (titrations were performed in 100 mM Tris buffer, pH 7.6, 37 °C, ±600 μM albumin at 23 and 400 MHz).

Fig. 2.

Ex vivo imaging of freshly isolated islets. (A) Each well contains ≈50 islets in Krebs-Henseleit-Hepes buffer, pH 7.4, plus either high (17.5 mM) or low (2.5 mM) glucose at 37 °C. (B) T₁-weighted fast spin-echo images of six wells containing ≈50 islets each (repetition time 500 ms, echo time 10 ms, averaging 6; 1-mm-thick slice above the level of the islets as shown). The images were collected ≈20 min after exposure of islets to the components indicated next to each image. For those wells containing the Zn²⁺ sensor and albumin, the concentrations were 50 μM of GdDOTA-diBPEN and 600 μM albumin. (C) Analytical amounts of Zn²⁺ and insulin in buffer solutions collected from a different set of wells containing variable numbers of islets. Data are expressed in pmol Zn²⁺ or insulin divided by the number of islet equivalents (IEQ) to normalize the data. (D) The change in water proton relaxation rate (ΔT₁⁻¹) in buffer solutions collected from the same wells as in C containing variable numbers of islets. As in C, the relaxation rate changes before and after addition of glucose (Δ1/T₁ in s⁻¹) was divided by the number of IEQ to normalize for differences in islet number in each well. Data are presented as average ± SD per IEQ (i.e., the islet quantity normalized to a 150-μm-diameter islet). All wells contained 50 μM GdDOTA-diBPEN and 600 μM albumin.

Fig. 3.

Representative grayscale T₁-weighted MR images of a single slice through the abdomen that contains a portion of pancreatic tissue (1 mm slice without fat saturation) of 12-wk-old control animals after injection of either saline (A) or glucose (B) followed by GdDOTA-diBPEN. The colored overlays represent a 3D composite of those pixels in each of the 14 slices where the water image intensity increased by threefold or more over the average noise (N) after injection of saline plus agent or glucose plus agent.

Fig. 4.

Representative grayscale T₁-weighted MR images of the abdomen (1-mm slice without fat saturation) of 24-wk-old mice fed either a standard 10% fat diet (A) or a 60% fat diet (B) over 12 wk. The colored overlays reflect a 3D composite of those pixels in each of the 14 slices where the water image intensity increased by threefold or more over the average noise after injection of glucose plus GdDOTA-diBPEN. (C) Total functional volume (mm³) of pancreatic tissue (regions with enhanced signal intensity by threefold or more summed over 14 separate MRI slices) in the three groups of animals as detected by GdDOTA-diBPEN. The average functional volumes, SDs, and P values are shown above each bar (n = 10 for control_12-wk; n = 6 for control_24-wk; n = 6 for DIO_24-wk).

Fig. 5.

Images of Zn²⁺ release during GSIS in a control (A) vs. a STZ-treated mouse (B). The color overlay represents the tissue areas where a contrast enhancement was observed after a bolus injection of GdDOTA-diBPEN and glucose. The colored image overlays reflect the same changes as noted in Figs. 3 and 4. F, fundus stomach; K, kidneys; S, spleen. Images were collected from the same mouse before and 5 d after a single high-dose treatment of STZ.