This study protocol aims to investigate how localised cooling influences the skin's microvascular, inflammatory, structural, and perceptual tolerance to sustained mechanical loading at the sacrum, evaluating factors such as morphology, physiology, and perceptual responses. The protocol will be tested on individuals of different age, sex, skin tone and clinical status, using a repeated-measure design with three participants cohorts: i) young healthy (n = 35); ii) older healthy (n = 35); iii) spinal cord injured (SCI, n = 35). Participants will complete three testing sessions during which their sacrum will be mechanically loaded (60 mmHg; 45 min) and unloaded (20 min) with a custom-built thermal probe, causing pressure-induced ischemia and post-occlusive reactive hyperaemia. Testing sessions will differ by the probe's temperature, which will be set to either 38°C (no cooling), 24°C (mild cooling), or 16°C (strong cooling). We will measure skin blood flow (via Laser Doppler Flowmetry; 40 Hz); pro- and anti-inflammatory biomarkers in skin sebum (Sebutape); structural skin properties (Optical Coherence Tomography); and ratings of thermal sensation, comfort, and acceptance (Likert Scales); throughout the loading and unloading phases. Changes in post-occlusive reactive hyperaemia will be considered as the primary outcome and data will be analysed for the independent and interactive effects of stimuli's temperature and of participant group on within- and between-subject mean differences (and 95% Confidence Intervals) in peak hyperaemia, by means of a 2-way mixed model ANOVA (or Friedman). Regression models will also be developed to assess the relationship between absolute cooling temperatures and peak hyperaemia. Secondary outcomes will be within- and between-subject mean changes in biomarkers' expression, skin structural and perceptual responses. This analysis will help identifying physiological and perceptual thresholds for the protective effects of cooling from mechanically induced damage underlying the development of pressure ulcers in individuals varying in age and clinical status.
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