Clinical, biochemical, cellular and molecular characterization of mitochondrial DNA depletion syndrome due to novel mutations in the MPV17 gene

Mitochondrial DNA (mtDNA) depletion syndromes (MDS) are severe autosomal recessive disorders associated with decreased mtDNA copy number in clinically affected tissues. The hepatocerebral form (mtDNA depletion in liver and brain) has been associated with mutations in the POLG, PEO1 (Twinkle), DGUOK and MPV17 genes, the latter encoding a mitochondrial inner membrane protein of unknown function. The aims of this study were to clarify further the clinical, biochemical, cellular and molecular genetic features associated with MDS due to MPV17 gene mutations. We identified 12 pathogenic mutations in the MPV17 gene, of which 11 are novel, in 17 patients from 12 families. All patients manifested liver disease. Poor feeding, hypoglycaemia, raised serum lactate, hypotonia and faltering growth were common presenting features. mtDNA depletion in liver was demonstrated in all seven cases where liver tissue was available. Mosaic mtDNA depletion was found in primary fibroblasts by PicoGreen staining. These results confirm that MPV17 mutations are an important cause of hepatocerebral mtDNA depletion syndrome, and provide the first demonstration of mosaic mtDNA depletion in human MPV17 mutant fibroblast cultures. We found that a severe clinical phenotype was associated with profound tissue-specific mtDNA depletion in liver, and, in some cases, mosaic mtDNA depletion in fibroblasts

Dynamic contrast optical coherence tomography images transit time and quantifies microvascular plasma volume and flow in the retina and choriocapillaris.

Despite the prevalence of optical imaging techniques to measure hemodynamics in large retinal vessels, quantitative measurements of retinal capillary and choroidal hemodynamics have traditionally been challenging. Here, a new imaging technique called dynamic contrast optical coherence tomography (DyC-OCT) is applied in the rat eye to study microvascular blood flow in individual retinal and choroidal layers in vivo. DyC-OCT is based on imaging the transit of an intravascular tracer dynamically as it passes through the field-of-view. Hemodynamic parameters can be determined through quantitative analysis of tracer kinetics. In addition to enabling depth-resolved transit time, volume, and flow measurements, the injected tracer also enhances OCT angiograms and enables clear visualization of the choriocapillaris, particularly when combined with a post-processing method for vessel enhancement. DyC-OCT complements conventional OCT angiography through quantification of tracer dynamics, similar to fluorescence angiography, but with the important added benefit of laminar resolution

Investigation of artifacts in retinal and choroidal OCT angiography with a contrast agent.

Optical coherence tomography angiography (OCTA) has recently emerged for imaging vasculature in clinical ophthalmology. Yet, OCTA images contain artifacts that remain challenging to interpret. To help explain these artifacts, we perform contrast-enhanced OCTA with a custom-designed wide-field ophthalmoscope in rats in vivo. We choose an intravascular contrast agent (Intralipid) with particles that are more isotropically scattering and more symmetrically shaped than red blood cells (RBCs). Then, by examining how OCTA artifacts change after contrast agent injection, we attribute OCTA artifacts to RBC-specific properties. In this work, we investigate retinal and choroidal OCTA in rats with or without melanosomes, both before and after contrast agent injection, at a wavelength at which scattering dominates the image contrast (1300 nm). First, baseline images suggest that high backscattering of choroidal melanosomes accounts for the relatively dark appearance of choroidal vessel lumens in OCTA. Second, Intralipid injection tends to eliminate the hourglass pattern artifact in OCTA images of vessel lumens and highlights vertical capillaries that were previously faint in OCTA, showing that RBC orientation is important in determining OCTA signal. Third, Intralipid injection increases lumen signal without significantly affecting the tails, suggesting that projection artifacts, or tails, are due to RBC multiple scattering. Fourth, Intralipid injection increases the side-to-top signal ratio less in choroidal vessel lumens of pigmented rats, suggesting that melanosome multiple scattering makes the hourglass artifact less prominent. This study provides the first direct experimental in vivo evidence to explain light scattering-related artifacts in OCTA

Dynamic Contrast Optical Coherence Tomography reveals laminar microvascular hemodynamics in the mouse neocortex in vivo

Studies of flow-metabolism coupling often presume that microvessel architecture is a surrogate for blood flow. To test this assumption, we introduce an in vivo Dynamic Contrast Optical Coherence Tomography (DyC-OCT) method to quantify layer-resolved microvascular blood flow and volume across the full depth of the mouse neocortex, where the angioarchitecture has been previously described. First, we cross-validate average DyC-OCT cortical flow against conventional Doppler OCT flow. Next, with laminar DyC-OCT, we discover that layer 4 consistently exhibits the highest microvascular blood flow, approximately two-fold higher than the outer cortical layers. While flow differences between layers are well-explained by microvascular volume and density, flow differences between subjects are better explained by transit time. Finally, from layer-resolved tracer enhancement, we also infer that microvascular hematocrit increases in deep cortical layers, consistent with predictions of plasma skimming. Altogether, our results show that while the cortical blood supply derives mainly from the pial surface, laminar hemodynamics ensure that the energetic needs of individual cortical layers are met. The laminar trends reported here provide data that links predictions based on the cortical angioarchitecture to cerebrovascular physiology in vivo

Visibility of microvessels in Optical Coherence Tomography angiography depends on angular orientation

Optical Coherence Tomography angiography (OCTA) is a widespread tool for depth-resolved imaging of chorioretinal vasculature with single microvessel resolution. To improve the clinical interpretation of OCTA, the conditions affecting visualization of microvessels must be defined. Here we inject a scattering plasma tracer (Intralipid) during OCTA imaging of the anesthetized rat eye. In the retina, we find that interlaminar (vertical) vessels that connect laminae have one-fourth to one-third the OCTA red blood cell to tracer (RBC-to-tracer) signal ratio of intralaminar (horizontal) vessels. This finding suggests that the OCTA signal from microvessels depends on angular orientation, making vertically-oriented vessels more difficult to visualize using intrinsic contrast alone. Clinicians should be aware of this potential artifact when interpreting OCTA

Incoherent excess noise spectrally encodes broadband light sources.

Across optics and photonics, excess intensity noise is often considered a liability. Here, we show that excess noise in broadband supercontinuum and superluminescent diode light sources encodes each spectral channel with unique intensity fluctuations, which actually serve a useful purpose. Specifically, we report that excess noise correlations can both characterize the spectral resolution of spectrometers and enable cross-calibration of their wavelengths across a broad bandwidth. Relative to previous methods that use broadband interferometry and narrow linewidth lasers to characterize and calibrate spectrometers, our approach is simple, comprehensive, and rapid enough to be deployed during spectrometer alignment. First, we employ this approach to aid alignment and reduce the depth-dependent degradation of the sensitivity and axial resolution in a spectrometer-based optical coherence tomography (OCT) system, revealing a new outer retinal band. Second, we achieve a pixel-to-pixel correspondence between two otherwise disparate spectrometers, enabling a robust comparison of their respective measurements. Thus, excess intensity noise has useful applications in optics and photonics

Attenuation coefficient as a quantitative parameter for analyzing cataracts with optical coherence tomography

Crystalline lenses of mice were imaged in vivo with a custom-made swept-source optical coherence tomography system. The use of the attenuation coefficient as a quantitative parameter for investigating the lens opacities magnitude is proposed, demonstrating a significant difference between the values retrieved from cataractous and normal mouse lenses