Two-photon microscopes have been successfully translated into clinical imaging tools to obtain high-resolution optical biopsies for in vivo histology. We report on clinical multiphoton coherent anti-Stokes Raman spectroscopy (CARS) tomography based on two tunable ultrashort near-infrared laser beams for label-free in vivo multimodal skin imaging. The multiphoton biopsies were obtained with the compact tomograph "MPTflex-CARS" using a photonic crystal fiber, an optomechanical articulated arm, and a four-detector-360 deg measurement head. The multiphoton tomograph has been employed to patients in a hospital with diseased skin. The clinical study involved 16 subjects, 8 patients with atopic dermatitis, 4 patients with psoriasis vulgaris, and 4 volunteers served as control. Two-photon cellular autofluorescence lifetime, second harmonic generation (SHG) of collagen, and CARS of intratissue lipids/proteins have been detected with single-photon sensitivity, submicron spatial resolution, and picosecond temporal resolution. The most important signal was the autofluorescence from nicotinamide adenine dinucleotide [NAD(P)H]. The SHG signal from collagen was mainly used to detect the epidermal-dermal junction and to calculate the ratio elastin/collagen. The CARS/Raman signal provided add-on information. Based on this view on the disease-affected skin on a subcellular level, skin areas affected by dermatitis and by psoriasis could be clearly identified. Multimodal multiphoton tomographs may become important label-free clinical high-resolution imaging tools for in vivo skin histology to realize rapid early diagnosis as well as treatment control. © The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.

First-order proximal methods that solve linear and bilinear elliptic optimal control problems with a sparsity cost functional are discussed. In particular, fast convergence of these methods is proved. For benchmarking purposes, inexact proximal schemes are compared to an inexact semismooth Newton method. Results of numerical experiments are presented to demonstrate the computational effectiveness of proximal schemes applied to infinite-dimensional elliptic optimal control problems and to validate the theoretical estimates.

The diagnosis of corneal diseases may be improved by monitoring the metabolism of cells and the structural organization of the stroma using two-photon imaging (TPI). We used TPI to assess the differences between nonpathological (NP) human corneas and corneas diagnosed with either keratoconus, Acanthamoeba keratitis, or stromal corneal scars. Images were acquired using a custom-built five-dimensional laser-scanning microscope with a broadband sub-15 femtosecond near-infrared pulsed excitation laser and a 16-channel photomultiplier tube detector in combination with a time-correlated single photon counting module. Morphological alterations of epithelial cells were observed for all pathologies. Moreover, diseased corneas showed alterations to the cells' metabolism that were revealed using the NAD(P)H free to protein-bound ratios. The mean autofluorescence lifetime of the stroma and the organization of the collagen fibers were also significantly altered due to the pathologies. We demonstrate that TPI can be used to distinguish between NP and diseased human corneas, based not only on alterations of the cells' morphology, which can also be evaluated using current clinical devices, but on additional morphological and functional features such as the organization of the stroma and the cells' metabolism. Therefore, TPI could become an efficient tool for diagnosing corneal diseases and better understanding the biological processes of the diseases.

PURPOSE. The purpose of this study was to evaluate the feasibility of using two-photon imaging (TPI) to assess the condition of human corneas for transplantation.METHODS. Human corneas were imaged after different storage times: short-term (STS), mediumterm (MTS), and long-term (LTS) storage. A high-resolution, custom-built 5-dimensional multiphoton microscope with 12-fs pulsed laser excitation was used for image acquisition. RESULTS.Optical discrimination between different corneal layers and sublayers based on their morphologic characteristics revealed by two-photon autofluorescence (AF) is possible. Furthermore, all layers were characterized based on AF lifetimes to gain information on metabolic activities of cells. The NAD(P)H free to protein-bound ratio (a 1 /a 2 ) of epithelial cells increased significantly in both MTS and LTS corneas compared with STS corneas. In endothelial cells, NAD(P)H a 1 /a 2 was significantly increased in MTS samples. For keratocytes, the NAD(P)H a 1 /a 2 decreased significantly with storage time. This could indicate that the metabolic activity of the epithelial and endothelial cells reduces, whereas the activity of keratocytes increases with storage time. The analysis of the stroma SHG images indicated that the organization of collagen fibers decreases with storage time. The feasibility of measuring the endothelial cell density (ECD) using TPI was demonstrated. An ECD of 1461 6 190 cells/ mm 2 was obtained for MTS samples based on TPI.CONCLUSIONS. TPI can provide information not accessible by current clinical methods, such as the cells' metabolic state and structural organization of the stroma, with subcellular resolution. Thus, it may improve the screening process of corneas prior to transplantation and might help to optimize the storage conditions.

A novel method for real-time magnetic resonance imaging for the assessment of cardiac function in mice at 9.4 T is proposed. The technique combines a highly undersampled radial gradient echo acquisition with an image reconstruction utilizing both parallel imaging and compressed sensing. Simulations on an in silico phantom were performed to determine the achievable acceleration factor and to optimize regularization parameters. Several parameters characterizing the quality of the reconstructed images (such as spatial and temporal image sharpness or compartment areas) were calculated for this purpose. Subsequently, double-gated segmented cine data as well as non-gated undersampled real-time data using only six projections per timeframe (temporal resolution ~ 10 ms) were acquired in a mid-ventricular slice of four normal mouse hearts in vivo. The highly accelerated data sets were then subjected to the introduced reconstruction technique and results were validated against the fully sampled references. Functional parameters obtained from real-time and fully sampled data agreed well with a comparable accuracy for left-ventricular volumes and a slightly larger scatter for mass. This study introduces and validates a real-time cine-MRI technique, which significantly reduces scan time in preclinical cardiac functional imaging and has the potential to investigate mouse models with abnormal heart rhythm.