3D virtual histology of murine kidneys –high resolution visualization of pathological alterations by micro computed tomography

The increasing number of patients with end stage chronic kidney disease not only calls for novel therapeutics but also for pioneering research using convincing preclinical disease models and innovative analytical techniques. The aim of this study was to introduce a virtual histology approach using micro computed tomography (µCT) for the entire murine kidney in order to close the gap between single slice planar histology and a 3D high resolution dataset. An ex vivo staining protocol based on phosphotungstic acid diffusion was adapted to enhance renal soft tissue x-ray attenuation. Subsequent CT scans allowed (i) the detection of the renal cortex, medulla and pelvis in greater detail, (ii) the analysis of morphological alterations, (iii) the quantification of the volume as well as the radio-opacity of these portions and (iv) the quantification of renal fibrotic remodeling based on altered radio-opacity using the unilateral ureteral obstruction model. Thus, virtual histology based on PTA contrast enhanced CT will in future help to refine the outcome of preclinical research on kidney associated murine disease models.

[1]  B. Thornhill,et al.  Ureteral obstruction as a model of renal interstitial fibrosis and obstructive nephropathy. , 2009, Kidney international.

[2]  R. Mark Henkelman,et al.  Structural Stabilization of Tissue for Embryo Phenotyping Using Micro-CT with Iodine Staining , 2013, PloS one.

[3]  S. Kauppinen,et al.  Determining collagen distribution in articular cartilage using contrast-enhanced micro-computed tomography , 2015, Osteoarthritis and cartilage.

[4]  Z. Endre,et al.  Early detection of acute kidney injury: Emerging new biomarkers (Review Article) , 2008, Nephrology.

[5]  D. Bulmer,et al.  Observations on Histological Methods Involving the use of Phosphotungstic and Phosphomolybdic Acids, with Particular Reference to Staining with Phosphotungstic Acid/Haematoxylin , 1962 .

[6]  F. Pfeiffer,et al.  Qualitative and Quantitative Imaging Evaluation of Renal Cell Carcinoma Subtypes with Grating-based X-ray Phase-contrast CT , 2017, Scientific Reports.

[7]  Giuliana Tromba,et al.  μCT of ex-vivo stained mouse hearts and embryos enables a precise match between 3D virtual histology, classical histology and immunochemistry , 2017, PloS one.

[8]  R. Cardiff,et al.  Manual hematoxylin and eosin staining of mouse tissue sections. , 2014, Cold Spring Harbor protocols.

[9]  S. Obenauer,et al.  Flat-panel detector-based volume computed tomography: a novel 3D imaging technique to monitor osteolytic bone lesions in a mouse tumor metastasis model. , 2007, Neoplasia.

[10]  T. Tamada,et al.  Measurement of renal cortical thickness using noncontrast‐enhanced steady‐state free precession MRI with spatially selective inversion recovery pulse: Association with renal function , 2015, Journal of magnetic resonance imaging : JMRI.

[11]  Brian D. Metscher,et al.  X-ray microtomographic imaging of intact vertebrate embryos. , 2011, Cold Spring Harbor protocols.

[12]  M. Perazella,et al.  Drug-induced glomerular disease: attention required! , 2015, Clinical journal of the American Society of Nephrology : CJASN.

[13]  R. Kalluri,et al.  Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression. , 2017, Kidney international.

[14]  R. Kalluri,et al.  Epithelial to Mesenchymal Transition induces cell cycle arrest and parenchymal damage in renal fibrosis , 2015, Nature Medicine.

[15]  K. Johnson An Update. , 1984, Journal of food protection.

[16]  E L Ritman,et al.  Three-dimensional microcomputed tomography of renal vasculature in rats. , 1998, Hypertension.

[17]  D. Poppas,et al.  Interstitial Fibrosis of Unilateral Ureteral Obstruction is Exacerbated in Kidneys of Mice Lacking the Gene for Inducible Nitric Oxide Synthase , 2000, Laboratory Investigation.

[18]  V. LeBleu,et al.  Thrombospondin-1 deficiency causes a shift from fibroproliferative to inflammatory kidney disease and delays onset of renal failure. , 2014, The American journal of pathology.

[19]  L. Leatherbury,et al.  Microcomputed Tomography Provides High Accuracy Congenital Heart Disease Diagnosis in Neonatal and Fetal Mice , 2013, Circulation. Cardiovascular imaging.

[20]  B. Metscher MicroCT for developmental biology: A versatile tool for high‐contrast 3D imaging at histological resolutions , 2009, Developmental dynamics : an official publication of the American Association of Anatomists.

[21]  L. Pardo,et al.  SK3 Channel Overexpression in Mice Causes Hippocampal Shrinkage Associated with Cognitive Impairments , 2016, Molecular Neurobiology.

[22]  B. Shrestha,et al.  Experimental rat models of chronic allograft nephropathy: a review , 2014, International journal of nephrology and renovascular disease.

[23]  R. Lindeman,et al.  The aging kidney. , 1986, Comprehensive therapy.

[24]  R. Kalluri,et al.  Induction of Tet3-dependent Epigenetic Remodeling by Low-dose Hydralazine Attenuates Progression of Chronic Kidney Disease , 2014, EBioMedicine.

[25]  J. Keggin,et al.  Structure of the Molecule of i2-Phosphotungstic , 1933, Nature.

[26]  Ahmer V. Farooq,et al.  Three-Dimensional Renal Parenchymal Volume as a Surrogate for Renal Function Estimation in Obstructed Kidneys Undergoing Surgical Repair. , 2015, Journal of endourology.

[27]  F. Strutz,et al.  Renal fibrosis: an update , 2001, Current opinion in nephrology and hypertension.

[28]  J. Bonventre,et al.  The Aging Kidney: Increased Susceptibility to Nephrotoxicity , 2014, International journal of molecular sciences.

[29]  J. Baker,et al.  Principles of biological microtechnique; a study of fixation and dyeing , 2011 .

[30]  P. Masson Some histological methods: Trichrome staining and their preliminary technique , 1929 .

[31]  J. Epstein,et al.  Rapid 3D Phenotyping of Cardiovascular Development in Mouse Embryos by Micro-CT With Iodine Staining , 2010, Circulation. Cardiovascular imaging.

[32]  Marta Zientkowska,et al.  Morphologic changes of mammary carcinomas in mice over time as monitored by flat-panel detector volume computed tomography. , 2008, Neoplasia.

[33]  J. Goldner A modification of the masson trichrome technique for routine laboratory purposes. , 1938, The American journal of pathology.

[34]  Gerd B Müller,et al.  MicroCT for molecular imaging: Quantitative visualization of complete three‐dimensional distributions of gene products in embryonic limbs , 2011, Developmental dynamics : an official publication of the American Association of Anatomists.

[35]  J. Kellum,et al.  Acute kidney injury: what's the prognosis? , 2011, Nature Reviews Nephrology.

[36]  P. Boor,et al.  Quantitative Micro-Computed Tomography Imaging of Vascular Dysfunction in Progressive Kidney Diseases. , 2016, Journal of the American Society of Nephrology : JASN.

[37]  M. L. Watson Staining of Tissue Sections for Electron Microscopy with Heavy Metals , 1958, The Journal of biophysical and biochemical cytology.

[38]  N. GuptaGopal,et al.  Three-Dimensional Renal Parenchymal Volume as a Surrogate for Renal Function Estimation in Obstructed Kidneys Undergoing Surgical Repair , 2014 .

[39]  Robert C. Wolpert,et al.  A Review of the , 1985 .