Positron emission tomography and single photon emission computed tomography imaging of tertiary lymphoid structures during the development of lupus nephritis

Abstract

Lymphoid neogenesis occurs in tissues targeted by chronic inflammatory processes, such as infection and autoimmunity. In systemic lupus erythematosus (SLE), such structures develop within the kidneys of lupus-prone mice ((NZBXNZW)F1) and are observed in kidney biopsies taken from SLE patients with lupus nephritis (LN). The purpose of this prospective longitudinal animal study was to detect early kidney changes and tertiary lymphoid structures (TLS) using in vivo imaging. Positron emission tomography (PET) by tail vein injection of 18-F-fluoro-2-deoxy-D-glucose (¹⁸F-FDG)(PET/FDG) combined with computed tomography (CT) for anatomical localization and single photon emission computed tomography (SPECT) by intraperitoneal injection of ^99mTC labeled Albumin Nanocoll (^99mTC-Nanocoll) were performed on different disease stages of NZB/W mice (n = 40) and on aged matched control mice (BALB/c) (n = 20). By using one-way ANOVA analyses, we compared two different compartmental models for the quantitative measure of ¹⁸F-FDG uptake within the kidneys. Using a new five-compartment model, we observed that glomerular filtration of ^18FFDG in lupus-prone mice decreased significantly by disease progression measured by anti-dsDNA Ab production and before onset of proteinuria. We could not visualize TLS within the kidneys, but we were able to visualize pancreatic TLS using ^99mTC Nanocoll SPECT. Based on our findings, we conclude that the five-compartment model can be used to measure changes of FDG uptake within the kidney. However, new optimal PET/SPECT tracer administration sites together with more specific tracers in combination with magnetic resonance imaging (MRI) may make it possible to detect formation of TLS and LN before clinical manifestations.

Keywords: 18-F-fluoro-2-deoxy-D-glucose; 99mTC-Nanocoll; lupus nephritis; positron emission tomography; single photon emission computed tomography; systemic lupus erythematosus; tertiary lymphoid structures.

Figure 1.

Renal hilum SUV increased in anti-dsDNA Ab+ mice. (a) Illustrates the three-compartment model. C _B , C₁, and C₂ represent tracer in the blood (input function), activity concentration in the parenchyma, and urine, respectively. (b) Cortex and renal hilum were delineated based on CT. There was no difference in K₁ (c) and f₂ (e) between the groups, but k₂ (d) increased during SLE progression. Area-under-curve of renal hilum (f) increased during SLE progression, while AUCs of cortex (g) and whole kidney (h) were steady. (c)–(h) One-way ANOVA with post-hoc analysis (Dunn’s multiple comparisons test). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 2.

Filtration of ¹⁸F-FDG by glomeruli was lower in lupus-prone mice producing anti-dsDNA Ab. (a) Illustrates the five-compartment model, where C _a , C _f , C _t , C _u , and C _m p represent tracer in blood, extravascular, tubule, urine, and metabolized ¹⁸F-FDG, respectively. The kfa (b), k _af (c), and kmf (d) were increased in mice with anti-dsDNA Ab production, but k _ma (f) decreased. K _fm (e), k _tm (g), and k _ut (h) were steady during SLE progression. (b)–(h) One-way ANOVA with post-hoc analysis (Dunn’s multiple comparisons test). *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 3.

Histological pictures and ex vivo organ biodistribution. (a) Representative Alcian blue/periodic acid–Schiff (AB/PAS) stained kidney sections from old BALB/c, Ab- NZB/W, Ab+ NZB/W, and Ab++ NZB/W mice. (b) ¹⁸F-FDG accumulation measured by SUV (g/ml) in tissue after 24 h. ¹⁸F-FDG accumulated higher in heart (c) and renal lymph node (d). ¹⁸F-FDG was higher in right (e) and left (f) kidneys, axillary/brachial lymph node, spleen, thymus, brain, and salivary gland (b) of anti-dsDNA Ab+ mice compared to Ab neg mice and was lower for GI track (b). Unpaired t test. *<0.05, **<0.01. a, artery; us, urinary space; v, vein. Scale bar = 100 µm.

Figure 4.

SPECT imaging of pancreatic TLS. (a) SPECT and (b) CT transversal, coronal, and sagittal frames of an example (dsDNA Ab+) showing multiple ROIs generated in Pmod 3.7. A threshold of 75% of the maximum activity was defined for the delineation in all mice. (c) Total number of VOIs accumulating ^99mTC-Nanocoll over time in BALB/c, dsDNA Ab- NZB/W, and dsDNA Ab+ NZB/W groups. (d) Relation between total number of VOIs drawn and the number of VOIs that accumulated ^99mTC-Nanocoll with time. No differences among the groups could be observed. (e) Atlas representation of kidneys and pancreas in the mouse. (f) Representing a fusion SPECT/CT image of the NZB/W dsDNA Ab+ mouse with a hot spot in the pancreatic area, which could only be observed in NZB/W dsDNA Ab+ mice (n = 6) not in the (g) BALB/c or NZB/W dsDNA Ab neg mice (data not shown). (h) Autoradiography of the ^99mTC Nanocoll in dsDNA Ab+ mice, which shows a clear tracer accumulation in the pancreas in comparison to blood. (c)–(d) One-way ANOVA with post-hoc analysis (Dunn’s multiple comparisons test). p, pancreas; k, kidney; b, blood.

Figure 5.

HE staining and immunostaining of TLS in pancreatic tissue in lupus-prone (a)–(c) and BALB/c mice (d). TLS was positively stained for T cells (CD3), B cells (B220), and macrophages (F480). Some TLS was detected near pancreatic islets (black arrows, c). (d) TLS was not detected in pancreatic sections from BALB/c mice. a, artery; d, ducts.