Wireless Dynamic Light Scattering Sensors Detect Microvascular Changes Associated With Ageing and Diabetes.

This article presents clinical results of wireless portable dynamic light scattering sensors that implement laser Doppler flowmetry signal processing. It has been verified that the technology can detect microvascular changes associated with diabetes and ageing in volunteers. Studies were conducted primarily on wrist skin. Wavelet continuous spectrum calculation was used to analyse the obtained time series of blood perfusion recordings with respect to the main physiological frequency ranges of vasomotions. In patients with type 2 diabetes, the area under the continuous wavelet spectrum in the endothelial, neurogenic, myogenic, and cardio frequency ranges showed significant diagnostic value for the identification of microvascular changes. Aside from spectral analysis, autocorrelation parameters were also calculated for microcirculatory blood flow oscillations. The groups of elderly volunteers and patients with type 2 diabetes, in comparison with the control group of younger healthy volunteers, showed a statistically significant decrease of the normalised autocorrelation function in time scales up to 10 s. A set of identified parameters was used to test machine learning algorithms to classify the studied groups of young controls, elderly controls, and diabetic patients. Our conclusion describes and discusses the classification metrics that were found to be most effective.

Nrf2 regulates glucose uptake and metabolism in neurons and astrocytes.

The transcription factor Nrf2 and its repressor Keap1 mediate cell stress adaptation by inducing expression of genes regulating cellular detoxification, antioxidant defence and energy metabolism. Energy production and antioxidant defence employ NADH and NADPH respectively as essential metabolic cofactors; both are generated in distinct pathways of glucose metabolism, and both pathways are enhanced by Nrf2 activation. Here, we examined the role of Nrf2 on glucose distribution and the interrelation between NADH production in energy metabolism and NADPH homeostasis using glio-neuronal cultures isolated from wild-type, Nrf2-knockout and Keap1-knockdown mice. Employing advanced microscopy imaging of single live cells, including multiphoton fluorescence lifetime imaging microscopy (FLIM) to discriminate between NADH and NADPH, we found that Nrf2 activation increases glucose uptake into neurons and astrocytes. Glucose consumption is prioritized in brain cells for mitochondrial NADH and energy production, with a smaller contribution to NADPH synthesis in the pentose phosphate pathway for redox reactions. As Nrf2 is suppressed during neuronal development, this strategy leaves neurons reliant on astrocytic Nrf2 to maintain redox balance and energy homeostasis.

Brain region specificity in reactive oxygen species production and maintenance of redox balance.

The brain produces various reactive oxygen species in enzymatic and non-enzymatic reactions as a by-product of metabolism and/or for redox signaling. Effective antioxidant system in the brain cells maintains redox balance. However, neurons and glia from some brain regions are more vulnerable to oxidative stress in ischemia/reperfusion, epilepsy, and neurodegenerative disorders than the rest of the brain. Using fluorescent indicators in live cell imaging and confocal microscopy, we have measured the rate of cytosolic and mitochondrial reactive oxygen species production, lipid peroxidation, and glutathione levels in cortex, hippocampus, midbrain, brain stem and cerebellum in acute slices of rat brain. We have found that the basal rate of ROS production is at its highest in brain stem and cerebellum, and that it is mainly generated by glial cells. Activation of neurons and glia by glutamate and ATP led to maximal rates of ROS production in the midbrain compared to the rest of the brain. Mitochondrial ROS had only minor implication to the total ROS production with maximal values in the cortex and minimal in the midbrain. The basal rate of lipid peroxidation was higher in the midbrain and hippocampus, while the GSH level was similar in most brain regions with the lowest level in the midbrain. Thus, the rate of ROS production, lipid peroxidation and the level of GSH vary across brain regions.

Body Position Affects Capillary Blood Flow Regulation Measured with Wearable Blood Flow Sensors.

In this study we demonstrate what kind of relative alterations can be expected in average perfusion and blood flow oscillations during postural changes being measured in the skin of limbs and on the brow of the forehead by wearable laser Doppler flowmetry (LDF) sensors. The aims of the study were to evaluate the dynamics of cutaneous blood perfusion and the regulatory mechanisms of blood microcirculation in the areas of interest, and evaluate the possible significance of those effects for the diagnostics based on blood perfusion monitoring. The study involved 10 conditionally healthy volunteers (44 ± 12 years). Wearable laser Doppler flowmetry monitors were fixed at six points on the body: two devices were fixed on the forehead, on the brow; two were on the distal thirds of the right and left forearms; and two were on the distal thirds of the right and left lower legs. The protocol was used to record three body positions on the tilt table for orthostatic test for each volunteer in the following sequence: (a) supine body position; (b) upright body position (+75°); (c) tilted with the feet elevated above the head and the inclination of body axis of 15° (-15°, Trendelenburg position). Skin blood perfusion was recorded for 10 min in each body position, followed by the amplitude-frequency analysis of the registered signals using wavelet decomposition. The measurements were supplemented with the blood pressure and heart rate for every body position analysed. The results identified a statistically significant transformation in microcirculation parameters of the average level of skin blood perfusion and oscillations of amplitudes of neurogenic, myogenic and cardiac sensors caused by the postural changes. In paper, we present the analysis of microcirculation in the skin of the forehead, which for the first time was carried out in various positions of the body. The area is supplied by the internal carotid artery system and can be of particular interest for evaluation of the sufficiency of blood supply for the brain.

Two-photon conversion of a bacterial phytochrome.

In nature, sensory photoreceptors underlie diverse spatiotemporally precise and generally reversible biological responses to light. Photoreceptors also serve as genetically encoded agents in optogenetics to control by light organismal state and behavior. Phytochromes represent a superfamily of photoreceptors that transition between states absorbing red light (Pr) and far-red light (Pfr), thus expanding the spectral range of optogenetics to the near-infrared range. Although light of these colors exhibits superior penetration of soft tissue, the transmission through bone and skull is poor. To overcome this fundamental challenge, we explore the activation of a bacterial phytochrome by a femtosecond laser emitting in the 1 µm wavelength range. Quantum chemical calculations predict that bacterial phytochromes possess substantial two-photon absorption cross sections. In line with this notion, we demonstrate that the photoreversible Pr ↔ Pfr conversion is driven by two-photon absorption at wavelengths between 1170 and 1450 nm. The Pfr yield was highest for wavelengths between 1170 and 1280 nm and rapidly plummeted beyond 1300 nm. By combining two-photon activation with bacterial phytochromes, we lay the foundation for enhanced spatial resolution in optogenetics and unprecedented penetration through bone, skull, and soft tissue.

Adrenaline induces calcium signal in astrocytes and vasoconstriction via activation of monoamine oxidase.

Adrenaline or epinephrine is a hormone playing an important role in physiology. It is produced de-novo in the brain in very small amounts compared to other catecholamines, including noradrenaline. Although the effects of adrenaline on neurons have been extensively studied, much less is known about the action of this hormone on astrocytes. Here, we studied the effects of adrenaline on astrocytes in primary co-culture of neurons and astrocytes. Application of adrenaline induced calcium signal in both neurons and astrocytes, but only in neurons this effect was dependent on α- and ß-receptor antagonists. The effects of adrenaline on astrocytes were less dependent on adrenoreceptors: the antagonist carvedilol had only moderate effect on the calcium signal and the agonist of adrenoreceptors methoxamine induced a signal only in small proportion of the cells. We found that adrenaline in astrocytes activates phospholipase C and subsequent release of calcium from the endoplasmic reticulum. Calcium signal in astrocytes is initiated by the metabolism of adrenaline by the monoamine oxidase (MAO), which activates reactive oxygen species production and induces lipid peroxidation. Inhibitor of MAO selegiline inhibited both adrenaline-induced calcium signal in astrocytes and the vasoconstriction that indicates an important role for monoamine oxidase in adrenaline-induced signalling and function.

Multimodal Optical Diagnostics of the Microhaemodynamics in Upper and Lower Limbs.

The introduction of optical non-invasive diagnostic methods into clinical practice can substantially advance in the detection of early microcirculatory disorders in patients with different diseases. This paper is devoted to the development and application of the optical non-invasive diagnostic approach for the detection and evaluation of the severity of microcirculatory and metabolic disorders in rheumatic diseases and diabetes mellitus. The proposed methods include the joint use of laser Doppler flowmetry, absorption spectroscopy and fluorescence spectroscopy in combination with functional tests. This technique showed the high diagnostic importance for the detection of disturbances in peripheral microhaemodynamics. These methods have been successfully tested as additional diagnostic techniques in the field of rheumatology and endocrinology. The sensitivity and specificity of the proposed diagnostic procedures have been evaluated.

Optical probe pressure effects on cutaneous blood flow.

The variation of blood flow characteristics caused by the probe pressure during noninvasive studies is of particular interest within the context of fundamental and applied research. It has been shown previously that the weak local pressure induces vasodilation, whereas the increased pressure is able to stop the blood flow in the compressed area, as well as to significantly change optical signals.The blood flow oscillations measured by laser Doppler flowmetry (LDF) characterize the functional state of the microvascular system and can be used for noninvasive diagnostics of its abnormality. This study was intended to identify the patterns of the relationship between the oscillating components of blood flow registered by the LDF method under different levels of pressure applied to an optical fiber probe.For this purpose, we have developed an original optical probe capable of regulating the applied pressure. The developed protocol included six sequential records of the blood perfusion at a pressure within the 0 to 200 mmHg range with unloading at the last stage.Using wavelet analyses, we traced the variation of energy of oscillations for these records in five frequency bands associated with different vascular tone regulation mechanisms. Six young volunteers of the same age (three males and three females) were included in this preliminary study and the protocol was repeated five times in each volunteer. Accordingly, 30 LDF records were available for the analyses. As expected, the LDF signal increases at weak pressure (30 mmHg) and decreases at increased pressure. The statistically stable amplification of endothelial associated blood flow oscillations under the 90 mmHg pressure allowed us to put forward a hypothesis that the endothelial activity increases. The possible causes of this phenomenon are discussed.

Multimodal optical measurement for study of lower limb tissue viability in patients with diabetes mellitus.

According to the International Diabetes Federation, the challenge of early stage diagnosis and treatment effectiveness monitoring in diabetes is currently one of the highest priorities in modern healthcare. The potential of combined measurements of skin fluorescence and blood perfusion by the laser Doppler flowmetry method in diagnostics of low limb diabetes complications was evaluated. Using Monte Carlo probabilistic modeling, the diagnostic volume and depth of the diagnosis were evaluated. The experimental study involved 76 patients with type 2 diabetes mellitus. These patients were divided into two groups depending on the degree of complications. The control group consisted of 48 healthy volunteers. The local thermal stimulation was selected as a stimulus on the blood microcirculation system. The experimental studies have shown that diabetic patients have elevated values of normalized fluorescence amplitudes, as well as a lower perfusion response to local heating. In the group of people with diabetes with trophic ulcers, these parameters also significantly differ from the control and diabetes only groups. Thus, the intensity of skin fluorescence and level of tissue blood perfusion can act as markers for various degrees of complications from the beginning of diabetes to the formation of trophic ulcers.

Combined use of laser Doppler flowmetry and skin thermometry for functional diagnostics of intradermal finger vessels.

We introduce a noninvasive diagnostic approach for functional monitoring of blood microflows in capillaries and thermoregulatory vessels within the skin. The measuring system is based on the combined use of laser Doppler flowmetry and skin contact thermometry. The obtained results suggest that monitoring of blood microcirculation during the occlusion, performed in conjunction with the skin temperature measurements in the thermally stabilized medium, has a great potential for quantitative assessment of angiospatic dysfunctions of the peripheral blood vessels. The indices of blood flow reserve and temperature response were measured and used as the primarily parameters of the functional diagnostics of the peripheral vessels of skin. Utilizing these parameters, a simple phenomenological model has been suggested to identify patients with angiospastic violations in the vascular system.

Development of a space-borne spectrometer to monitor atmospheric ozone.

A new compact satellite spectrometer dedicated to monitoring terrestrial atmospheric ozone (ozonometer) is in preparation for the Russian Geophysics Program. Four instruments at four satellites (Ionosphere) are intended to monitor the total ozone content by measuring spectra of scattered solar radiation in nadir. The spectrometer is based on the Rowland scheme with a concave holographic diffraction grating. It covers the near UV and visible range of the spectrum, 300-500 nm, with a spectral resolution of ∼0.3 nm. At present, a qualification model has been manufactured and tested. We introduce the description of the instrument and the results of laboratory and ground-based atmospheric calibrations. The ozone amount retrieved from atmospheric measurements using the differential optical absorption spectroscopy (DOAS) method is in good agreement with that measured by the collocated Brewer spectrophotometer and ozone monitoring instrument on board the Aura satellite.

Individual variability analysis of fluorescence parameters measured in skin with different levels of nutritive blood flow.

Fluorescence spectroscopy has recently become more common in clinical medicine. However, there are still many unresolved issues related to the methodology and implementation of instruments with this technology. In this study, we aimed to assess individual variability of fluorescence parameters of endogenous markers (NADH, FAD, etc.) measured by fluorescent spectroscopy (FS) in situ and to analyse the factors that lead to a significant scatter of results. Most studied fluorophores have an acceptable scatter of values (mostly up to 30%) for diagnostic purposes. Here we provide evidence that the level of blood volume in tissue impacts FS data with a significant inverse correlation. The distribution function of the fluorescence intensity and the fluorescent contrast coefficient values are a function of the normal distribution for most of the studied fluorophores and the redox ratio. The effects of various physiological (different content of skin melanin) and technical (characteristics of optical filters) factors on the measurement results were additionally studied. The data on the variability of the measurement results in FS should be considered when interpreting the diagnostic parameters, as well as when developing new algorithms for data processing and FS devices.

Substantiation of medical and technical requirements for noninvasive spectrophotometric diagnostic devices.

A scientific approach to the formulation of medical and technical requirements (MTRs) for noninvasive spectrophotometric diagnostic devices using optical technologies such as laser Doppler flowmetry and absorption spectroscopy is proposed. The theoretical modeling framework, metrological certification, and testing of these devices are still in the early stages of development. The theoretical estimation of the received signal levels for wavelengths between 514 and 940 nm is highly dependent on the blood volume level in the subject tissue. The proposed approach allows, in particular, the calculation of technical and metrological performance constraints of the instruments, such as the ranges of the sensitivity and power-related signal-to-noise ratios for different spectral channels and different biomedical (biochemical and physiological) parameters. Substantiation of specialized MTRs for the noninvasive spectrophotometric diagnostic devices can enable them to develop to the level of standardized measurement techniques.