Objective Skin cholesterol is an important biomarker for early atherosclerosis screening. Atherosclerosis is the leading cause of disability and death from cardiovascular disease. Effective control of pathogenic factors in the early pathological stage may delay or prevent the development of asymptomatic atherosclerosis into cardiovascular diseases. Thus, skin cholesterol detection becomes relevant in the prevention of cardiovascular diseases. Traditional skin cholesterol detection methods, such as skin biopsy or tape stripping, are invasive and usually time consuming. Alternatively, the recent three-drop method is being widely studied. In this method, three specific concentrations of reagents that bind to skin cholesterol are used on the skin surface of a subject, and atherosclerosis can be diagnosed by analyzing the reagent color changes. However, the three-drop method is sensitive to the application habits of the operator. Moreover, the detection reagents contain enzymes, polymers, and small molecule compounds, hindering quality control and increasing the sensitivity to environmental factors such as temperature and pH levels. We report a non-invasive skin cholesterol detection technique based on fluorescent spectrometry. By measuring the fluorescence spectrum of fluorescent-labeled skin, the cholesterol content can be calculated from the fluorescence spectra. This method corrects the influence of temperature on the test results and provides stability under various environmental conditions. Moreover, the skin cholesterol content can be obtained within 4 minutes. The proposed method provides a rapid non-invasive and stable method for skin cholesterol detection and corresponding applications including early atherosclerosis screening.
Methods The proposed non-invasive skin cholesterol detection system is composed of a light source, fiber probe, spectrometer, photodiode, infrared temperature sensor, and computer. The fluorescence fluctuation of the detection reagent caused by temperature variation is corrected by establishing the relation between temperature and the fluorescence intensity of the detection reagent. To confirm the accuracy of the proposed skin cholesterol detection system, we extract skin cholesterol with absolute ethanol after the non-invasive measurement. The cholesterol content in the extraction liquid is determined by gas chromatography, and the correlation between the two results are analyzed. Finally, the clinical applicability of the proposed system is confirmed by measuring skin cholesterol content from both healthy subjects and subjects with high risk of presenting atherosclerosis.
Results and Discussions The schematic of the proposed non-invasive skin cholesterol detection system based on fluorescent spectrometry is shown in Fig. 1. The system accurately detects skin cholesterol content after correcting for temperature. The average fluorescence intensity of the detection reagent in the 462--520 nm wavelength band decreases with increasing temperature, resulting in a significant negative correlation between fluorescence intensity and temperature (r=-0.995, p<0.0001). This relation can be used to establish a calibration curve to correct for temperature (Fig. 5). We recruited 80 subjects to verify the accuracy of the proposed system. The skin cholesterol content measured using the proposed temperature-corrected system is highly correlated (correlation coefficient of 0.905) with that measured using gas chromatography (Fig. 6). These results verify the accuracy of the proposed system to measure skin cholesterol. To verify whether the proposed system can distinguish healthy subjects from subjects with high risk of presenting atherosclerosis, we used the system in 43 and 46 subjects from the respective groups. There is a significant difference in skin cholesterol content between the healthy and high risk samples (p=0.0004) (Fig. 7). The proposed non-invasive skin cholesterol detection system can screen subjects with high risk of presenting atherosclerosis. Nevertheless, clinical trials are required for verification given the small sample size used in this study.
Conclusions We propose a rapid non-invasive detection system for skin cholesterol based on fluorescent spectrometry. The system quickly provides the skin cholesterol content on-site from the fluorescence spectrum of detection reagents that specifically bind to skin cholesterol. The proposed system performs temperature correction to prevent deviations of the measurement results and improve accuracy and stability. The system and its detection accuracy are verified through comparisons with skin cholesterol results obtained from gas chromatography. The proposed system may be used to screen people with high risk of presenting atherosclerosis by detecting skin cholesterol content in healthy subjects and subjects at high risk. Overall, the proposed system can detect skin cholesterol accurately, non-invasively, and quickly. We expect that the widespread adoption of this technology will contribute to the prevention and control of cardiovascular diseases.
