Assessing fibrotic damage to renal structure and function with T2-weighted and ASL MRI
Christopher Charles Conlin1,2, Yangyang Zhao2, Yufeng Huang3, and Jeff Lei Zhang1,4

1Utah Center for Advanced Imaging Research, University of Utah, Salt Lake City, UT, United States, 2Bioengineering, University of Utah, Salt Lake City, UT, United States, 3Nephrology, University of Utah School of Medicine, Salt Lake City, UT, United States, 4Radiology, University of Utah School of Medicine, Salt Lake City, UT, United States

Synopsis

This study examined the suitability of T2 and ASL-measured renal perfusion as biomarkers for fibrotic kidney disease. Renal perfusion was measured in healthy and fibrotic rats using a multi-TI ASL protocol and compared to renal T2 as well as urinary and histological fibrosis markers. Significantly reduced renal perfusion was observed in fibrotic rats, in parallel with increased renal T2, proteinuria, and mesangial matrix in the glomerular tuft. The sensitivity of T2 and perfusion to fibrotic kidney damage suggests that ASL and T2-weighted MRI may provide improved assessment of renal fibrosis and prove useful for the early detection of renal disease.

Motivation

The majority of chronic kidney diseases are characterized by renal fibrosis1. While biopsy is the gold standard for fibrosis assessment, it is unsuitable for early detection of fibrotic renal disease, particularly in humans. T2-weighted MRI is often used to detect tissue damage2, but it is not a direct assessment of tissue function. In this study, we examined renal perfusion in healthy and fibrotic rats using arterial spin labeling (ASL) at multiple inversion times (TIs) to directly estimate fibrotic damage to kidney function and compared it to renal T2 as well as urinary and histological fibrosis markers.

Methods

Following IACUC approval, fibrosis was induced in the renal cortex of 11 male Sprague Dawley rats (Charles River Laboratories) through injection of 1.75 mg/kg monoclonal antibody OX-7. Two other rats did not receive the injection and were used as healthy controls. Renal T2-weighted and ASL MRI, a 24-hour urine collection protocol to measure proteinuria, and renal biopsy to assess the degree of cortical fibrosis was performed 5 days after OX-7 injection for 6 of the fibrotic rats and 12 days after OX-7 injection for the other 5 fibrotic rats.

Rats were sedated with isoflurane (Fluriso; Vet One) during the MR examination. MR imaging was performed using a wrist coil on a 3T clinical scanner (TimTrio; Siemens). Sets of eight T2-weighted images were acquired from axial slices through each kidney using a turbo spin-echo sequence: TR 400ms, TE 5.8, 29, 52, 75, 98, 122, 145, 168ms, flip angle 180°, matrix 256x256, resolution 0.78x0.78mm. Each series of T2-weighted images was fit pixel-wise to the T2 decay function to generate a T2 map. Cortical T2 was determined from the T2 map by averaging values within an ROI defined over the renal cortex. ASL images were acquired from the same slices as the T2 images using a flow-sensitive alternating inversion recovery (FAIR) protocol: TR 4.7ms, TE 2.35ms, TIs 300, 500, 800, 1000, 1300, 1500ms, flip angle 180°, matrix 256x256, resolution 0.78x0.78mm, bSSFP readout, and 6s inter-image time. For the tag and control images at each TI, the signal intensity was averaged within the same cortical ROIs as for T2 analysis. Subtraction of the control from the tag signal-vs-TI curve generated the ASL difference signal. Using a tracer-kinetic model of the multi-TI ASL difference signal, similar to that proposed by Buxton et al3, renal plasma flow (RPF) was measured in the cortex by calculating the slope of the difference signal normalized by T1 relaxation.

After MR examination, rats were housed in metabolic cages for 24 hours to allow for urine collection. Urinary albumin and general protein concentrations were measured using a DC2000+ reagent kit (Bayer) and the Bradford method (Bio-Rad), respectively. Rats were then sacrificed and their kidneys removed and fixed in formalin. Sections of cortical tissue were embedded in paraffin and stained with periodic acid-Schiff (PAS) solution to visualize the glomeruli. Subsections of these samples containing 20 glomeruli were photographed under 400x magnification. Using semi-automatic color-image analysis software (Image J; NIH), fibrosis was quantified as the PAS-positive mesangial area over the total area of the glomerular tuft. RPF, T2, urinary albumin and protein excretion, and histological fibrosis measurements were compared between control rats and those 5-days and 12-days after fibrosis induction. Two-sided t-tests (α=0.05) were used to test for significant differences between groups.

Results

Compared to the controls, significantly elevated levels of albumin and general protein were measured in the urine of fibrotic rats (Figure 1). Photomicrographs of glomeruli from healthy and fibrotic rats are shown in Figures 2A-2C, demonstrating increased PAS-positive mesangial matrix in the diseased groups. The average degree of fibrosis (percentage of glomerular tuft) is quantified in Figure 2D, showing significantly greater fibrosis in rats treated with monoclonal antibody OX-7. ASL images from healthy and diseased rats are compared in Figures 3A and 3B, illustrating a significant drop in average cortical perfusion compared to controls (Figure 3C). Cortical T2 values were also significantly greater in fibrotic rats than in controls (Figure 4). No significant difference was observed between the 5-day and 12-day fibrosis groups with any metric. These results are summarized in Table 1.

Discussion

Cortical RPF and T2 changes both revealed fibrotic damage to the kidneys, in parallel with observations from urinary and histological fibrosis markers. Unlike T2, however, RPF is a direct measurement of renal function. ASL may therefore provide improved assessment of fibrotic kidney damage, and in conjunction with structural imaging techniques like T2-weighted MRI, prove useful for the early detection of renal disease.

Acknowledgements

We would like to thank Niels Oesingmann at Siemens Healthcare for contributing his pulse sequence programming expertise.

This work was made possible by funding from the NKF Young Investigator Award and RSNA Research Scholar Grant programs.

References

1. Liu Y. Renal fibrosis: new insights into the pathogenesis and therapeutics. Kidney international. 2006;69(2):213-7.

2. Mathur S, Vohra RS, Germain SA, et al. Changes in muscle T2 and tissue damage following downhill running in mdx mice. Muscle & nerve. 2011;43(6):878-886. doi:10.1002/mus.21986.

3. Buxton, R. B., L. R. Frank, et al. A general kinetic model for quantitative perfusion imaging with arterial spin labeling. Magnetic resonance in medicine 1998;40(3): 383-396.

Figures

Figure 1: Urinary fibrosis markers in healthy and fibrotic rats. Albumin and protein levels were significantly elevated in the urine of fibrotic rats compared to healthy controls. Asterisks indicate a significant (P<0.05) difference from the values observed in the control group.

Figure 2: Photomicrographs of glomeruli (arrows) in healthy (A) and fibrotic rats 5 days (B) and 12 days (C) after fibrosis induction. Increased PAS-positive matrix is seen in B and C. D shows that fibrotic rats have a significantly greater degree of fibrosis than controls (P<0.05; indicated by asterisks).

Figure 3: ASL signal intensity is higher in the cortex (indicated by arrows) of healthy rats (A) than those with fibrosis (B). C shows that cortical RPF is significantly reduced in fibrotic rats. Asterisks indicate a significant (P < 0.05) difference from the values observed in healthy rats.

Figure 4: Cortical T2 is elevated in fibrotic rats. A) T2 map of control rat. B) T2 map of fibrotic rat. Arrows point to the right kidney in each image. C) Average cortical T2 among the three rodent groups. Asterisks indicate a significant (P<0.05) difference from control values.

Table 1: Comparison of cortical perfusion, T2, and urinary and histological fibrosis markers in control and fibrotic rats. Asterisks indicate a significant (P < 0.05) difference from the values observed in the control group.



Proc. Intl. Soc. Mag. Reson. Med. 24 (2016)
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