Chronic kidney disease (CKD) is characterized by a progressive loss of renal function over time. Histopathological analysis of the condition of glomeruli and the proximal convolutional tubules over time can provide valuable insights into the progression of CKD. Optical coherence tomography (OCT) is a technology that can analyze the microscopic structures of a kidney in a nondestructive manner. Recently, we have shown that OCT can provide real-time imaging of kidney microstructures in vivo without administering exogenous contrast agents. A murine model of CKD induced by intravenous Adriamycin (ADR) injection is evaluated by OCT. OCT images of the rat kidneys have been captured every week up to eight weeks. Tubular diameter and hypertrophic tubule population of the kidneys at multiple time points after ADR injection have been evaluated through a fully automated computer-vision system. Results revealed that mean tubular diameter and hypertrophic tubule population increase with time in post-ADR injection period. The results suggest that OCT images of the kidney contain abundant information about kidney histopathology. Fully automated computer-aided diagnosis based on OCT has the potential for clinical evaluation of CKD conditions.
Chronic Kidney Disease (CKD) is characterized by a progressive loss in renal function over time. Pathology can provide valuable insights into the progression of CKD by analyzing the status of glomeruli and the uriniferous tubules over time. Optical coherence tomography (OCT) is a new procedure that can analyze the microscopic structure of the kidney in a non-invasive manner. This is especially important because there are significant artifacts associated with excision biopsies and immersion fixation procedures. Recently, we have shown that OCT can provide real time images of kidney microstructure and Doppler OCT (DOCT) can image glomerular renal blood flow in vivo without administrating exogenous contrast agents. In this study, we used OCT to evaluate CKD in a model induced by intravenous Adriamycin injection into Munich-Wistar rats. We evaluated tubular density and tubular diameter from OCT images at several post- Adriamycin induction time points and compared them with conventional light microscopic histological imaging. Proteinurea and serum creatinine were used as physiological markers of the extent of CKD. Preliminary OCT results revealed changes in tubular density due to tubular necrosis and interstitial fibrosis within the first 4 weeks following Adriamycin injection. From week 4 to 8 after Adriamycin induction, changes in tubular density and diameter occurred due to both tubular loss and tubular dilation. The results suggest OCT can provide additional information about kidney histopathology in CKD. DOCT revealed reduced blood flow in some glomeruli probably as a consequence of focal glomerularsclerosis.
KEYWORDS: Luminescence, Two photon imaging, Hydrogen, Absorption, Oxygen, Sapphire lasers, Confocal microscopy, Medical research, Molecules, In vivo imaging
We present the application of two-photon fluorescence (TPF) imaging to monitor intracellular hydrogen peroxide (H 2 O 2 ) production in brain cells. For selective imaging of H 2 O 2 over other reactive oxygen species, we employed small-molecule fluorescent probes that utilize a chemoselective boronate deprotection mechanism. Peroxyfluor-6 acetoxymethyl ester detects global cellular H 2 O 2 and mitochondria peroxy yellow 1 detects mitochondrial H 2 O 2 . Two-photon absorption cross sections for these H 2 O 2 probes are measured with a mode-locked Ti:sapphire laser in the wavelength range of 720 to 1040 nm. TPF imaging is demonstrated in the HT22 cell line to monitor both cytoplasmic H 2 O 2 and localized H 2 O 2 production in mitochondria. Endogenous cytoplasmic H 2 O 2 production is detected with TPF imaging in rat astrocytes modified with d-amino acid oxidase. The TPF H 2 O 2 imaging demonstrated that these chemoselective probes are powerful tools for the detection of intracellular H 2 O 2 .
We have developed a multi-kilowatt peak power 1-μm optical pulse source for two-photon microscopy. Utilizing an
external-cavity hybrid mode-locked semiconductor laser, we were able to generate picosecond optical pulses at a
500-MHz repetition rate. With a semiconductor optical amplifier driven by synchronized electronic gating pulses, the optical
pulse repetition rate was sub-harmonically extracted at 1-100 MHz. At a 10-MHz repetition rate, optical pulses were then
amplified to a peak power of greater than 2 kW with a two-stage
Yb-doped fiber amplifier. Using this light source, we
successfully obtained clear two-photon images of mouse brain neurons expressing green fluorescent proteins.
We developed a novel scheme for two-photon fluorescence bioimaging. We generated supercontinuum (SC) light at wavelengths of 600 to 1200 nm with 774-nm light pulses from a compact turn-key semiconductor laser picosecond light pulse source that we developed. The supercontinuum light was sliced at around 1030- and 920-nm wavelengths and was amplified to kW-peak-power level using laboratory-made low-nonlinear-effects optical fiber amplifiers. We successfully demonstrated two-photon fluorescence bioimaging of mouse brain neurons containing green fluorescent protein (GFP).
We present the microfabrication of high refractive index-modulated structures written by line scan inside bulk of fused silica with a femtosecond laser at wavelength of 810 nm. The femtosecond laser beam, with duration between 130 fs and 500 fs at a repetition rate of 1 kHz, was focused through a microscope objective with numerical aperture (NA) of 0.10 or 0.25 into the sample. To fabricate high refractive index-modulated structures in fused silica, we investigated the dependence of refractive index change on laser pulse energy, pulse duration, scan speed, and scanning repetitions. The results showed a "triangle region", with pulse duration of 130 fs to 230 fs and pulse energy of 0.35 μJ to 1.5 μJ, for the fabrication of refractive index modulation structures. The refractive index modulation was increased to 3×10-3 after several scanning passes. Diffractive optical components such as grid, square, circle gratings and Fresnel zone plates have been fabricated by direct writing technique. The structures could be used as diffractive beam splitters, beam shaper and micro-lens.
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