Impact of dietary zinc on stimulated zinc secretion MRI in the healthy and malignant mouse prostate

Zinc is an essential trace element with catalytic and structural roles in enzymes and transcription factors, and it participates in dynamic cellular signaling regulated by ZnT (export/storage) and Zip (import/release) transporter families. ZnT8 in pancreatic β-cells packages Zn2+ with insulin into vesicles; upon glucose-stimulated insulin secretion, Zn2+ is co-released and can be detected with optical probes or zinc-responsive MRI agents. Prior MRI studies established that a rapid rise in plasma glucose elicits glucose-stimulated zinc secretion (GSZS) detectable in vivo, not only from pancreatic islets but also from the mouse prostate. The human prostate normally accumulates high total Zn2+, whereas malignant transformation leads to reduced zinc levels. Prior work showed hypointense foci (reflecting reduced Zn2+ secretion) in TRAMP mouse prostates and in dogs with BPH, and SR-XRF indicated glucose stimulates Zn movement from glandular lumen to stromal/interstitial spaces where it can interact with GdL1. The current study asks how varying dietary zinc affects (1) total prostate zinc content (by SR-XRF) and (2) glucose-stimulated zinc efflux (by MRI) in healthy versus malignant (TRAMP) mouse prostate, given that human dietary zinc intake varies and could influence GSZS-based imaging performance.

Background literature indicates: (a) normal prostate tissue contains high zinc concentrations that decline markedly in prostate cancer; (b) zinc transporters (ZnT, Zip) regulate intracellular and extracellular zinc availability and are altered in disease; (c) GSZS MRI using Zn-responsive agents has visualized β-cell function and unexpectedly revealed Zn secretion from prostate in mice; (d) prior imaging and SR-XRF studies demonstrated glucose-triggered Zn trafficking from glandular to stromal compartments in prostate and reduced secretion in malignant tissue; (e) dietary zinc intake varies across populations and may influence systemic zinc bioavailability. Reports in TRAMP models suggest optimal—but not deficient or excessive—zinc intake may be protective, while deficiency or excess can be deleterious, aligning with the hypothesis that malignant prostate loses zinc storage/secretion capacity.

Study design: Male C57BL/6 wild-type (WT) and TRAMP mice (16–24 weeks) were fed zinc-deficient (0.05 ppm), sufficient/optimal (30 ppm), or supplemented (150 ppm) diets ad libitum for 21 days. Animals were fasted overnight prior to imaging. MRI/GSZS protocol: Under isoflurane anesthesia, two baseline 3D T1-weighted scans (ge3d; FA/TE/TR 20°/1.756/3.476 ms; NEX 4; matrix 128×128×128; FOV 30×30 mm) were acquired on a 9.4 T Varian/Agilent scanner with a custom volume coil. Mice received 0.07 mmol/kg GdL1 intravenously (zinc-responsive contrast agent; binds Zn2+ with KD ≈118 nM and forms a ternary complex with albumin, increasing r1 from 5.1 to 11.4 mM−1 s−1 at 0.5 T) and either 2.2 mmol/kg glucose intraperitoneally (to stimulate GSZS) or saline as control. Sequential 3D T1-weighted scans were collected over 10 minutes post-injection. Regions of interest covering the prostate (excluding urethra) were used to compute contrast-to-noise ratio (CNR): CNR = (SIprostate − SImuscle)/SDnoise. ΔCNR = CNR(t) − CNRbaseline; area under the CNR–time curve (AUC) was computed. An index of zinc efflux was derived by comparing glucose-stimulated enhancement to saline controls at 7 min to mitigate vascularity effects. Physiology and blood measurements: Body weight was recorded after 21 days. Tail-vein blood glucose was measured before and 10 min after GdL1 plus glucose or saline. Following imaging, mice were euthanized; blood was collected (serum prepared) and prostates were excised, frozen, and sectioned. SR-XRF elemental mapping: Adjacent cryosections (10–20 µm for H&E; 50 µm for XRF) were prepared. μSR-XRF was performed at Diamond Light Source I18 beamline. Coarse scans used 50×50 µm beam (0.75 s/pixel) at 8.2 and 11 keV; fine scans used 5×5 µm (1 s/pixel). Spectra were fitted and quantified using PyMCA (v5.6.3) with thin-film standards; custom MATLAB and IDL processed maps and ROI statistics (ventral, lateral, dorsal lobes). Elements quantified included Zn, Gd, Cu, Fe, Ca. ICP-MS: Serum samples were digested in concentrated nitric acid and analyzed on an Agilent 8800-QQQ ICP-MS for total zinc (and Gd) concentrations. Statistics: GraphPad Prism 10. One-way ANOVA with Tukey post-hoc for multiple comparisons; two-tailed Student’s t-test for pairwise comparisons. Significance: *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. Ethics: Animal procedures approved by UT Southwestern IACUC.

• Body weight: Zinc-deficient diets reduced weight in both WT and TRAMP mice. WT: 0.05 ppm Zn, 28.0 ± 2.20 g vs 30 ppm Zn, 30.4 ± 1.44 g (p = 0.0061). TRAMP: 0.05 ppm Zn, 26.9 ± 2.85 g vs 30 ppm Zn, 30.1 ± 3.08 g (p = 0.0199) and vs 150 ppm Zn, 29.1 ± 3.4 g (p = 0.0208).
• Blood glucose: In saline groups, BG 10 min post GdL1 decreased significantly only in zinc-supplemented animals (−27.1 ± 14.4%, p = 0.013), suggesting GdL1 may influence glucose homeostasis when zinc is abundant. No significant differences 10 min after exogenous glucose in either genotype across diets.
• Serum zinc (ICP-MS): Total serum Zn was relatively insensitive to dietary level overall. However, under deficiency, WT had higher serum Zn than TRAMP (23.33 ± 3.08 µM vs 20.07 ± 3.01 µM, p = 0.0148). Under high zinc, TRAMP exhibited higher serum Zn than WT (24.51 ± 5.13 µM vs 20.44 ± 2.11 µM, p = 0.0081). At 30 ppm, WT and TRAMP were similar (21.09 ± 3.98 µM vs 22.38 ± 2.06 µM, p = 0.4554). These suggest WT increases serum Zn bioavailability under deficiency, while TRAMP does not; conversely, WT reduces serum Zn under excess whereas TRAMP normalizes less effectively.
• SR-XRF tissue zinc: Across diets, total prostate zinc content varied only modestly among 0.05, 30, 150 ppm groups. Zinc tended to be higher in the lateral lobe of WT; Ca was more concentrated in ventral lobes. Zinc content was reduced in all lobes under zinc-deficient diet in both WT and TRAMP. Gd distribution did not differ among lobes.
• GSZS MRI: WT prostates showed glucose-enhanced signal in all diet groups, with additional enhancement beyond GdL1-alone (saline). TRAMP prostates had higher saline enhancement (greater vascularity), reducing apparent glucose–saline differences. After normalization (index of zinc efflux: glucose minus saline at 7 min), WT exhibited clear increases in GSZS proportional to dietary zinc (0.05 < 30 < 150 ppm), while TRAMP showed overall reduced GSZS with no parallel diet-dependent increase, indicating malignant tissue’s diminished capacity to store/release dietary zinc.
• Lobe-specific variability: WT lateral lobes showed wider zinc variation (with glucose) than TRAMP, consistent with lateral lobe involvement in zinc storage/secretion. Overall, dietary zinc supplementation increased stimulated zinc efflux detectable by MRI in healthy prostate but not in TRAMP, while total tissue zinc content by SR-XRF changed little across diets.

Findings support that malignant prostate tissue (TRAMP) loses the capacity to regulate zinc homeostasis, particularly to store zinc and to secrete it upon glucose stimulation. Despite limited changes in total tissue zinc with altered dietary intake, healthy prostate increased glucose-stimulated zinc efflux as dietary zinc rose, whereas TRAMP did not. Systemically, WT mice adapt to deficiency by elevating serum zinc availability and to excess by reducing it, while TRAMP mice show impaired regulation in both contexts. The observed decrease in blood glucose after GdL1 plus saline under high zinc suggests the zinc probe may modulate glycemic control, aligning with non-human primate data where a Gd-based zinc probe potentiated insulin and C-peptide secretion; mechanistically, extracellular Zn2+ and insulin could modulate glucagon via α-cell zinc uptake, and GdL1 binding to Zn2+ may influence transporter activity. For imaging, vascularity confounds in TRAMP elevated saline enhancement; normalization via a zinc efflux index mitigated this. Clinically, slower clearance of low-MW agents in humans suggests a practical GSZS protocol using a single zinc-responsive agent with two T1-weighted scans (pre- and post-glucose) to directly index zinc secretion. Importantly, given that healthy prostate increases stimulated zinc efflux under higher zinc supply while malignant tissue cannot, short-term zinc supplementation prior to GSZS MRI might enhance contrast between malignant and non-malignant regions, improving prostate cancer detection.

Dietary zinc level has minimal impact on total prostate tissue zinc content but strongly modulates glucose-stimulated zinc efflux in healthy prostate, not in malignant TRAMP prostate. Zinc deficiency causes systemic and organ-level dysregulation (weight loss, altered serum zinc bioavailability), while zinc supplementation enhances GSZS in WT but not TRAMP. These results indicate that short-term zinc supplementation before GSZS MRI could increase T1-weighted contrast between malignant and non-malignant prostate, potentially improving cancer detection. Future work should validate GSZS with zinc supplementation in larger animal models and humans, incorporate vascularity-matched control agents where needed, and further elucidate zinc–glucose endocrine interactions and transporter dynamics in prostate.

• Vascularity confound: TRAMP prostates exhibited higher baseline enhancement with saline due to greater vascularity, reducing apparent glucose-induced changes; although an index of zinc efflux was used, separating vascular effects from zinc secretion remains challenging in rodent models.
• Control agent: A non–zinc-responsive Gd agent with similar relaxivity was not used in parallel to control for vascular differences, limiting definitive attribution of signal changes solely to zinc secretion.
• Pharmacokinetics: Rapid clearance of low-MW agents in mice constrains timing and may differ from human kinetics, affecting translational comparability.
• Dietary extrapolation: While zinc supplementation enhanced GSZS contrast in WT, prior reports suggest both deficiency and excess zinc can have deleterious effects; results do not imply supplementation is protective/therapeutic for cancer.
• Sample sizes in some SR-XRF subgroup analyses were small (e.g., N=2 in certain TRAMP diet/glucose conditions), limiting statistical power for lobe-specific comparisons.