2.4 Animal study
To test whether chronic arsenic exposure upregulates β-endorphin, 18 C57BL/6J male mice (6–8 week) were purchased from the Department of Laboratory Animal Medicine of Central South University and raised under a controlled environment with constant temperature (22±2 °C) and a 12-hour light-dark cycle. The mice were randomly assigned to vehicle control, 5 mg/L arsenite-treated group, or 15 mg/L arsenite-treated group, and each group contained 6 mice. The arsenite-treated mice were administered arsenite for 6 months by the drinking water. Mice in vehicle control group were fed with sterilized water correspondingly. Arsenic trioxide was purchased from Sigma-Aldrich (St. Louis, MO, USA). After 6 months treatment, the mice were killed and blood sampled by eyeball extirpating for the detection of β-endorphin.
2.5 Randomized, double-blind, placebo-controlled trial
To test whether the inhibition of β-endorphin (μ-opioid) receptor alleviates itch in patients under chronic arsenic exposure, we conducted a randomized, double-blind, placebo-controlled trial (ClinicalTrial.gov ID: NCT03751111). We randomly assigned patients to naloxone (0.4 mg/qd, sublingual) and placebo (0.4 mg/qd, sublingual) using a computer-generated sequence of random numbers during January 2019 to March 2019. The trial consisted of a one-week treatment period and a one-week withdrawl period. The primary outcome (severity of itch) was evaluated using the NRS through a face-to-face interview on baseline (Day 0) and the last day of the treatment period (Day 7), and a telephone interview on the last day of the withdrawl period (Day 14). The participants were also inquired about the possible adverse events of naloxone such as headache, sleep difficulty, sickness, and dizziness in each interview.
The inclusion criteria can be found in the protocol (supplementary file S1). In brief, the participants must be above 18 years, be able to understand the study protocol and sign informed consent voluntarily, and report moderate-to-severe itch (NRS≥3). Participants were excluded if they used anti-histamines, anti-inflammatory, anti-pruritic, analgesic, antidepressant, or anti-epileptic agents within 2 weeks prior to the study, or had a history of pruritic skin diseases such as eczema and psoriasis.