Approach to non-invasive assessment of vascular circulation

introduction

The treatment of vascular diseases entails enormous costs for health systems. Current approaches largely focus on symptom management. Over the past decade, substantial evidence has linked vascular disease to a dysfunctional response to hypoxia. A distinct reaction to hypoxia can be identified from the macro- and microvessels. Based on this effect, we have developed a novel non-invasive diagnostic technique called Flow Mediated Skin Fluorescence (FMSF) to be used for the assessment of vascular circulation and metabolic regulation.1 The FMSF technique has been used successfully for the noninvasive diagnosis of many vascular diseases, including cardiovascular disease, PAD, diabetes, and hypertension.2–4 The FMSF technique has also generated considerable interest among sports physiologists.5

A conclusion from our studies using the FMSF technique is that diagnosis of vascular circulation should focus not only on dysfunctional blood flow in major arteries, but also on microcirculatory responses to hypoxia. In this communication, we present two key parameters derived from FMSF measurements: reactive response to hyperemia (RHR) and sensitivity to hypoxia. [log(HS)]. These parameters can be used for effective characterization of vascular flow based on the response to transient ischemia.

Methods

The measurements were carried out using the AngioExpert, a device manufactured by Angionica Ltd. The AngioExpert device uses the Flow Mediated Skin Fluorescence (FMSF) technique, which measures changes in nicotinamide adenine dinucleotide (NADH) fluorescence intensity of the skin on the forearm in response to block and release blood flow . The skin is the largest organ of the human body and is characterized by a specific metabolism. The epidermal layer of the skin is not directly vascularized, and oxygen and nutrients are transported from the dermis by diffusion. Therefore, epidermal cell metabolism can be considered as a unique and sensitive marker of early dysfunction of vascular circulation and metabolic regulation.

The AngioExpert assesses the patient’s condition by analyzing changes in the NADH fluorescence signal emitted by epidermal cells in response to forced occlusion ischemia and subsequent hyperemia due to occlusion removal. By interpreting the parameters and shape of the obtained NADH fluorescence curve, disturbances in vascular circulation can be identified which may be indicative of chronic diseases. The AngioExpert measures NADH fluorescence excited by ultraviolet (UV) radiation with a wavelength of 340 nm (UVB). As the maximum UVB light penetration of 340nm is about 0.3-0.5mm, the results are determined by NADH fluorescence in the epidermis. The emitted wavelength of NADH fluorescence is 460 nm (blue light). The test is performed in a comfortable seated position, after a minimum adaptation period of 5 minutes, in a quiet room with controlled air temperature (24 ± 1°C). The resting NADH fluorescence value emitted by the epidermal layer of the forearm is recorded during the first 3 minutes (180 s). The brachial artery is then occluded by inflating the device cuff to 60 mmHg above systolic pressure. The ischemic response is recorded over a period of 3 minutes (180 s). During this time, ischemic changes in the NADH fluorescence signal are recorded. At the end of the occlusion, the cuff pressure is released abruptly, restoring flow in the brachial artery and inducing a hyperemic response, for a minimum duration of 4 minutes (240 s).

RHR is a newly introduced parameter (defined in Figure 1), which characterizes endothelial function related mainly to the production of nitric oxide (NO) in the vasculature due to reactive hyperemia. RHR is a single parameter, based on the combined response of the ischemic and hyperemic portions of the measured FMSF trace.

Figure 1 Definition of the RHR parameter.

The HS parameter has been used previously in FMSF trace analysis and represents a direct measure of the intensity of microcirculatory oscillations related to myogenic oscillations with frequencies between 0.052 and 0.15 Hz, recorded during reperfusion.6.7 Myogenic microcirculatory oscillations are a very sensitive measure of the microcirculatory response to hypoxia, which can be monitored with great precision using the FMSF technique. Since the values ​​of the HS parameter can vary over a fairly wide range, it is more practical to use a normally distributed log(HS).

The study was conducted at the Medical University of Lodz, Poland, and the University of Physical Education in Poznan, Poland. It complied with the principles set out in the Declaration of Helsinki and the study protocol was approved by the University Bioethics Committee. All subjects gave written informed consent prior to participation.

Results and discussion

The results summarized in Figure 2 were collected for three study groups: A – endurance athletes; B – healthy middle-aged individuals; C – type 2 diabetic patients. The RHR parameter distinguishes these three groups with high statistical significance. Such observations indicate that the use of the RHR parameter has adequate sensitivity to be used for the characterization of vascular circulation. A similar conclusion can be made based on the analysis of the log(HS) parameter, representing the reaction of the microcirculation to transient hypoxia. The RHR and log(HS) parameters describe quite distinctive properties of the vasculature, and the two parameters should be used together for effective diagnosis. Some correlation exists between log(HS) and RHR parameters for healthy individuals (groups A + B), as shown in Figure 2D. However, these parameters must be interpreted separately when analyzing patients with various diseases and disorders of vascular origin. For example, very low values ​​(less than 1) for the log(HS) parameter can effectively predict a limited chance of healing in patients with diabetic foot ulcers, regardless of the measured value of the RHR parameter.4 The diagnostic use of the RHR parameter may be more important in cardiovascular diseases, where macrocirculatory dysfunction prevails.

Figure 2 Evaluation of RHR and log(HS) parameters in groups A, B and C: A – highly trained endurance athletes (distance runners – 22, triathletes – 14, rowers – 14), n = 50 (33 m, 17 f), mean age 22.0 (16–35 years); B – healthy middle-aged individuals, n = 32 (19 m, 13 w), mean age 38.2 (30–50 years); C – type 2 diabetic patients, n = 70 (40 m, 32 f), mean age 63.1 (45–80 years). The statistical analysis of the differences between the parameters in the compared groups was based on the one-way ANOVA test with Scheffe’s post hoc test. (A) Statistical differentiation of groups A, B and C based on RHR parameter. (B) Distribution of the RHR parameter. (VS) Statistical differentiation of groups A, B and C based on the log(HS) parameter. (D) Correlation between log(HS) and RHR parameters for groups A + B (Pearson correlation).

Based on our experience using the FMSF diagnostic technique, we conclude that characterization of vascular circulation based on RHR and log(HS) parameters can be effective in a large segment of the population, healthy individuals. physically active health to individuals with serious health problems. related to vascular dysfunction. We believe that this simple two-parameter approach based on distinguishable macro and microcirculatory responses to hypoxia will be recognized in the near future as a powerful diagnostic tool for the characterization of vascular circulation.

Funding

This work was supported by the European Union from the resources of the European Regional Development Fund under the Smart Growth Operational Programme, Grant No. POIR. 01.01.01-00-0540/15-00.

Disclosure

JG and AM are the inventors of patents protecting the use of FMSF technology (EP2713860B1) issued to Angionica Ltd. Joanna Katarzynska, Andrzej Marcinek and Jerzy Gebicki affiliated with Angionica Ltd. The authors report no other conflicts of interest in this work.

The references

1. Katarzynska J, Lipinski Z, Cholewinski T, et al. Noninvasive evaluation of microcirculation and metabolic regulation by flow-mediated skin fluorescence (FMSF): technical aspects and methodology. Rev Sci Instrument. 2019;90:104104. doi:10.1063/1.5092218

2. Katarzynska J, Borkowska A, Czajkowski P, et al. The Flow Mediated Skin Fluorescence technique reveals a remarkable effect of age on microcirculation and metabolic regulation in type 1 diabetes. Res Microbasin. 2019;124:19–24. doi:10.1016/j.mvr.2019.02.005

3. Katarzynska J, Borkowska A, Los A, et al. Flow-mediated skin fluorescence technique (FMSF) for the study of vascular complications of type 2 diabetes. J Diabetes Sci Technol. 2020;14:693–694. doi:10.1177/1932296819895544

4. Los-Stegienta A, Katarzynska J, Borkowska A, et al. Differentiation of diabetic foot ulcers based on stimulation of myogenic oscillations by transient ischemia. Vasc health risk management. 2021;17:145–152. doi:10.2147/VHRM.S307366

5. Bugaj O, Zielinski J, Kusy K, et al. The effect of exercise on skin content of reduced form NAD and its response to transient ischemia and reperfusion in highly trained athletes. Before Physiol. 2019;10:600. doi:10.3389/fphys.2019.00600

6. Gebicki J, Katarzynska J, Cholewinski T, et al. Flowmotion monitored by Flow Mediated Skin Fluorescence (FMSF): a tool for characterizing the microcirculatory state. Before Physiol. 2020;11:702. doi:10.3389/fphys.2020.00702

7. Gebicki J, Marcinek A, Zielinski J. Assessment of microcirculatory status based on stimulation of myogenic oscillations by transient ischemia: from health to disease. Vasc health risk management. 2021;17:33–36. doi:10.2147/VHRM.S292087

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