Quantitative multi‐spectral oxygen saturation measurements independent of tissue optical properties

Imaging of tissue oxygenation is important in several applications associated with patient care. Optical sensing is commonly applied for assessing oxygen saturation but is often restricted to local measurements or else it requires spectral and spatial information at the expense of time. Many methods proposed so far require assumptions on the properties of measured tissue. In this study we investigated a computational method that uses only multispectral information and quantitatively computes tissue oxygen saturation independently of tissue optical properties. The method is based on linear transformations of measurements in three isosbestic points. We investigated the ideal isosbestic point combination out of six isosbestic points available for measurement in the visible and near‐infrared region that enable accurate oxygen saturation computation. We demonstrate this method on controlled tissue mimicking phantoms having different optical properties and validated the measurements using a gas analyzer. A mean error of 2.9 ± 2.8% O₂Sat was achieved. Finally, we performed pilot studies in tissues in‐vivo by measuring dynamic changes in fingers subjected to vascular occlusion, the vasculature of mouse ears and exposed mouse organs.

Selected steps of spectral transformations applied to oxygenation spectra. The original reflectance spectrum M(λ) is transformed in step 1 to overlap with reference spectra (grey) in three isosbestic points, resulting in M″(λ). In step 2, the gradient of M″(λ) is computed resulting in M″_grad(λ), which can be used for quantitative oxygenation computation.     

应变率成像技术评价吗啡预处理对兔心肌缺血再灌注损伤时左室功能的影响

目的:应用应变率成像技术(SRI)动态观察和评价吗啡预处理对兔心肌缺血再灌注损伤时左室长轴功能的影响。方法:建立新西兰大白兔心肌缺血再灌注模型,随机分为假手术组(SH组)、吗啡预处理组(MF组)和生理盐水预处理组(NS组)。于手术前1天及手术术后28天内不同时间点测量超声心动图常规数据及左室各壁收缩期峰值速度与应变率参数。结果:术后1天,MF组与NS组左室整体与局部心肌收缩功能较术前和SH组明显减低(P0.05),并随缺血再灌注时间的延长呈减低趋势(P0.05),且NS组减低更明显(P0.05)。结论:SRI技术能够客观、准确地评价缺血再灌注损伤心肌的局部收缩功能的变化,而吗啡预处理对兔心肌缺血再灌注损伤存在明显的弱化作用。     

Quantitative imaging of fibrotic and morphological changes in liver of non-alcoholic steatohepatitis (NASH) model mice by second harmonic generation (SHG) and auto-fluorescence (AF) imaging using two-photon excitation microscopy (TPEM)

Non-alcoholic steatohepatitis (NASH) is a common liver disorder caused by fatty liver. Because NASH is associated with fibrotic and morphological changes in liver tissue, a direct imaging technique is required for accurate staging of liver tissue. For this purpose, in this study we took advantage of two label-free optical imaging techniques, second harmonic generation (SHG) and auto-fluorescence (AF), using two-photon excitation microscopy (TPEM). Three-dimensional ex vivo imaging of tissues from NASH model mice, followed by image processing, revealed that SHG and AF are sufficient to quantitatively characterize the hepatic capsule at an early stage and parenchymal morphologies associated with liver disease progression, respectively.     

Pyrene-labeled lipids: versatile probes of membrane dynamics in vitro and in living cells

Pyrene-labeled analogs of fatty acids have been studied as probes of lipid metabolism in vitro and in cultured cells. Procedures for the synthesis of complex pyrenyl lipids and the analytical methods for their separation and quantification are described. Pyrenyl-lipids have been used to quantify the relationship between lipid structure and the rates of spontaneous lipid transfer. Modifications of these methods have also been used to monitor protein-mediated lipid transfer, lipolysis and lipid translocation across bilayer membranes. According to several criteria, pyrene dodecanoic acid has been identified as a good analog of some naturally occurring fatty acids. Digital imaging microscopy has been used to monitor the rate of accumulation of pyrenyl lipids in living cells.     

Clinical applications of assessments of myocardial substrate utilization with positron emission tomography

Summary Positron emission tomography (PET) permits sequential, noninvasive assessment of myocardial perfusion and metabolism. Based on the pattern of substrate use, clinical studies have utilized PET to define the location and extent of myocardial infarction, to identify areas of jeopardized but viable myocardium, and to assess the metabolic response of the myocardium to pharmacological therapy as well as to interventions such as coronary thrombolysis and coronary artery bypass surgery. The ability to noninvasively assess specific metabolic pathways should facilitate our understanding of normal myocardial metabolism, its perturbations with cardiac disease, and thereby improve the diagnosis and treatment of the biochemical processes underlying cardiac dysfunction.     

Measurement of internal pH in the coccolithophoreEmiliania huxleyi using 2′,7′-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein acetoxymethylester and digital imaging microscopy

Internal pH (pH_i) was determined inEmiliania huxleyi (Lohmann) using the probe 2,7-bis-(2-carboxyethyl)-5(and-6)carboxyfluoresceinacetoxymethylester (BCEF-AM) and digital imaging microscopy. The probe BCECF-AM was taken up and hydrolysed to the free acid by the cells. A linear relationship was established between pH_i and the 490/450 fluorescence ratio of BCECF-AM over the pH range 6.0 to 8.0 using the ionophore nigericin. Two distinct pH domains were identified within the cell, the cytoplasmic domain (approx. pH 7.0) and the chloroplast domain (approx. pH 8.0). The average pH_i was 7.29 (±0.11) for cells in the presence of 2 mM HCO ₃ ^– . In the absence of HCO ₃ ^– the pH_i was decreased by 0.8 pH unit. The importance of these changes in pH_i is considered in relation to inorganic-carbon uptake.Abbreviations AM acetoxymethylester - BCECF 2,7-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein - Hepes 4-(2-hydroxyethyl)-1-piperazine ethanesulfonic acid - pH_i intracellular pH     

Epidermal patterns of the hands and feet of the pygmy chimpanzee (Pan paniscus)

The characteristics of the epidermal ridge system were studied in a series of eighteen lesser or pygmy chimpanzees (Pan paniscus). The general ridge alignments are very similar to those of the chimpanzee (Pan troglodytes); Biegert ('61). On the average the pattern intensity (P.I.) of the palm configurations is considerably higher in the pygmy chimpanzee than in the chimpanzee, thus representing the highest total palm pattern intensity of all species of the Hominoidea. The sole configurations show parallel main results to those of the palm; however, the decreased sole pattern frequency of the pygmy chimpanzee is of a smaller predominance only as compared to the values of the other species of this superfamily. The preliminary data on the finger tip patterns, translated into P.I. values, are much higher than in chimpanzees and within the range of the mean values of gorillas (Brehme, '73), while those of the toes of pygmy chimpanzees seem to possess the lowest P.I. values of the African apes.     

Determination of cytoplasmic calcium concentration inDryopteris spores

Germination of Dryopteris spores is mediated by the physiologically active, far-red-absorbing form of phytochrome, Pfr, and external Ca²⁺ is necessary for the transduction of the light signal. Because knowledge about the cytoplasmic calcium ion concentration, [Ca²⁺]_i, is of great importance for understanding the role of calcium during signal transduction, this value was measured using fura-2 in fern spores undergoing the normal developmental progression into germination. Fura-2 was loaded into the spores by electroporation, which does not disrupt the normal process of germination. The intensity of the fluorescence emission of the loaded fura-2 was analysed by a microspectrophotometric assay of single spores, and successful loading could be obtained by the application of ten electrical pulses (field strength 7.5 kV · cm^–1, half-life (time constant) 230 s). Fura-2 was alternately excited by light of wavelengths 355 and 385 nm through an inverted fluorescence microscope, and the emitted fura-2 fluorescence was collected by a silicon-intensified video camera. The cytoplasmic calcium ion concentration was calculated from the ratio of the camera output obtained for both wavelengths and displayed by a pseudo-color technique. Spores responded to changes of the extracellular Ca²⁺ concentration, and this observation is considered as evidence that fura-2 is loaded into the cytoplasm. The substitution of a low external [Ca²⁺] (1 mM ethyleneglycol-bis(2-aminoethyl-ether) {ie166-01},N-tetraacetic acid (EGTA)) by 1 mM CaCl₂ caused a fast increase of [Ca²⁺]_i from approx. 50 nM to above 500 nM. In contrast, the subsequent substitution of CaCl₂ by EGTA decreased [Ca²⁺]_i again below 100 nM within 0.5 h. Furthermore, the application of ionomycin could initiate a change in [Ca2+]_i according to the Ca²⁺ gradient established between the extracellular medium and cytoplasm. In spores sown on a Ca²⁺-free medium, [Ca²⁺]_i, analysed in a buffer containing EGTA, was found to be around 50 nM during the first days of cultivation, independent of the irradiation protocol. However, if spores were grown in darkness on a Ca²⁺-containing medium and analysed in EGTA, [Ca²⁺]_i was significantly higher ( 500 nM). In red-light-irradiated spores, [Ca²⁺]_i was found to decrease with increasing time after irradiation, and was determined to be less than 100 nM when analysis was done 44 h after germination was initiated by the light treatment.Dedicated to Professor H. Mohr on the occasion of his 60th birthday     

One- and Two-Dimensional Electron Paramagnetic Resonance Imaging in Skin

EPR imaging with modulated field gradients provides the possibility for obtaining an EPR spectrum in a selected volume We demonstrate the feasibility of X-band (9.5GHz) electron paramagnetic resonance (EPR) imaging in skin biopsies of hairless mice. One- (ID) and two-dimensional (2D) EPR images of the persistent free radical di-tertiary-butyl-nitroxide are measured. At a microwave frequency of 9.5 GHz (X-band), 2D images are obtained in skin biopsies with an actual point distinction resolution of 25 μm. In a biological model system. 2D images are measured at L-band frequency (2.0 GHz) with a pixel resolution of 61 μm. and a theoretical spatial resolution of 12.5 μm. In combination with the spin labeling and spin trapping technique. EPR imaging is the most direct approach to analyzing spatial distribution of physico-chemical properties in skin, such as membrane fluidity and polarity. as well as detection of free radicals.     

Intraepidermal nerves in human digital skin

Summary Intraepidermal nerve fibers of human glabrous digital skin were investigated using a new silver impregnation method. Nerves were observed to enter the epidermis without regional preference, and to extend into the stratum granulosum. They are non-varicose (smooth) or varicose and range from less than 0.2 m to approximately 2 m in diameter, with varicosities up to 3 m in diameter. Some axons branch profusely within the epithelium, giving off fine branches of differing diameters, while others appear to remain unbranched. At least some intraepidermal axons are fine branches of larger axons taking a horizontal course below the epithelial layer. Others are, at least topically, closely associated with Meissner's corpuscles. At 57 nerves per mm² surface area, the density of intraepidermal nerves found in this investigation is much greater than that reported in recent publications, and agrees closely with values given in several older studies.Dedicated to Prof. Dr. A. Hopf on the occasion of his 65th birthday