Study: NCT05048862

Intervention type: DIAGNOSTIC_TEST
Intervention: Nerve conduction studies and autonomic function tests
Description: Nerve conduction study will be performed with a Nicolet Viking IV Electromyographer (Madison, WI) in all patients following standardized methods recommended by the Consensus Development Conference on Standardized Measures in Diabetic Neuropathy. Studied nerves include sural, peroneal, tibial, median and ulnar (motor and sensory) nerves.Autonomic functions will be assessed by the SSR and RRIV with established protocol by using Nicolet Viking IV Electromyographer (Madison, WI).

Intervention type: DIAGNOSTIC_TEST
Intervention: Quantitative sensory test
Description: Quantative sensory test will be performed with a Thermal Sensory Analyzer and Vibratory Sensory Analyzer (Medoc Advanced Medical System, Minneapolis, MN). The procedure is the same as previously described 25. Briefly, the machine delivers to the patient a stimulus of constant intensity which is pre-set by the algorithm. By adjusting the intensity of stimulus (increase or decrease the intensity by a fixed ratio) according to the response of the subject (i.e. whether the subject perceives the stimulus or not), sensory thresholds of the warm, cold and vibratory modalities will be measured.

Intervention type: DIAGNOSTIC_TEST
Intervention: Quantitative sensory test
Description: Quantative sensory test will be performed with a Thermal Sensory Analyzer and Vibratory Sensory Analyzer (Medoc Advanced Medical System, Minneapolis, MN). The procedure is the same as previously described 25. Briefly, the machine delivers to the patient a stimulus of constant intensity which is pre-set by the algorithm. By adjusting the intensity of stimulus (increase or decrease the intensity by a fixed ratio) according to the response of the subject (i.e. whether the subject perceives the stimulus or not), sensory thresholds of the warm, cold and vibratory modalities will be measured.

Intervention type: DIAGNOSTIC_TEST
Intervention: Skin biopsy
Description: A skin specimen of 3 mm in diameter will be taken with a biopsy punch from the lateral side of the distal leg under 2% lidocaine local anaesthesia 26. No suturing is required, and the wounds are covered with a piece of gauze. Wound healing takes 7\~10 days, similar to a typical abrasion wound. Informed consent will be obtained from each patient before the skin biopsy. The intraepidermal nerve fiber density and sweat gland nerve innervation will be examined.

Intervention type: DIAGNOSTIC_TEST
Intervention: Muscle and nerve biopsy
Description: Two muscle specimens with 5 x 5 x 5 mm were collected in an open muscle biopsy or needle biopsy at studied muscles under 2% lidocaine local anaesthesia. The wound was about 2-3 cm long, and suture was required. Would healing usually takes 10-14 days. First specimen was undergoing snap freezing fixation in a longitudinal axis perpendicular to the cork with the liquid nitrogen and isopentane. The second specimen was divided into two equiponderous tissues, and one was freezed in liquid nitrogen for DNA and protein analysis. The another one was treated with RNAlater solution in 4°C overnight for RNA analysis. Sural nerve biopsies or superficial peroneal nerve were obtained from a standard site posterior to the lateral malleolus under local anesthesia. The nerves were then fixed in 5% glutaraldehyde in 0.1 M phosphate buffer (PB) at 4 °C overnight. All samples were stored in the -80°C refrigerator for further analysis.