Intravital imaging of adriamycin-induced renal pathology using two-photon microscopy and optical coherence tomography

Abstract

Adriamycin (doxorubicin), a common cancer chemotherapeutic drug, can be used to induce a model of chronic progressive glomerular disease in rodents. In our studies, we evaluated renal changes in a rat model after Adriamycin injection using two-photon microscopy (TPM), optical coherence tomography (OCT) and Doppler OCT (DOCT). Taking advantage of deep penetration and fast scanning speed for three-dimensional (3D) label-free imaging, OCT/DOCT system was able to reveal glomerular and tubular pathology noninvasively and in real time. By imaging renal pathology following the infusion of°uorophore-labeled dextrans of di®erent molecular weights, TPM can provide direct views of glomerular and tubular°ow dynamics with the onset and progression of renal disease. Speci¯cally, glomerular permeability and¯ltration, proximal and distal tubular°ow dynamics can be revealed. 6-8 weeks after injection of Adriamycin, TPM and OCT/DOCT imaging revealed glomerular sclerosis, compromised°ow across the glomerular wall, tubular atrophy, tubular dilation, and variable intra-tubular°ow dynamics. Our results indicate that TPM and OCT/DOCT provide real-time imaging of renal pathology in vivo that has not been previously available using conventional microscopic procedures. §

Morphological and functional characteristics of aging kidneys based on two‐photon microscopy in vivo

“…The renal tubules occur both atrophic and dilated in our study, it agrees well with previous OCT observations in aging kidneys and OCT/TPM observations in Adriamycin induced chronic kidney disease models . The diameters of normal renal tubules are consistent with previous study .…”

“…From Figure C, the urine protein value reaches an average of 1000 mL/dL in the 17 to 18 months group, while it is only about 300 mL/dL for the 3 to 4 months group. Figure D shows that serum creatinine is within the normal range (less than or equal to 1.3 mg/dL) for the 3 to 4 months group . The values increase slightly for the 11 to 12 months group, but they are still within the normal range.…”

“…In the similar manner, tubular dilation was defined as an increase of greater than 50% in tubular diameter compared to nonatrophic tubules. From the previous data, the normal sizes of the proximal and distal tubules are around 30 to 40 μm . Cell nuclei of renal tubules were labeled by Hoechst 33342 (2 mg/mL).…”

“…TPM images were acquired using a nonlinear optical imaging system based on a custom‐built multiphoton microscope and a Ti:Sapphire laser (Mira 900‐F, Coherent, Santa Clara, California) operating at 790 nm wavelength and 120 fs pulse width . A silicon oil immersion objective (Olympus ×60, NA = 1.3) was employed for focusing the excitation beam into tissue samples and was also used to collect the backscattered two‐photon excited fluorescence signals.…”

“…Fluorescent dyes were dissolved in 1.0 mL sterile isotonic saline and intravenously injected into rats. High‐MW 500‐kDa dextran‐fluorescein (5 mg/mL) was used for the visualization of blood flow, as well as glomerular filtration and permeability . Since high‐MW dextran is not filtered by normal glomeruli and remains in the vasculature, it labels the peritubular and glomerular capillaries.…”