The prevalence of chronic widespread pain (CWP) in people with HIV is high, yet the underlying mechanisms are elusive. Leukocytes synthesize the endogenous opioid, β-endorphin, within their endoplasmic reticulum (ER). When released into plasma, β-endorphin dampens nociception by binding to opioid receptors on sensory neurons. We hypothesized that the heme-dependent redox signaling induces ER stress, which attenuates leukocyte β-endorphins levels/release, thereby increasing pain sensitivity in people with HIV. Results demonstrated that HIV positive individuals with CWP had increased plasma methemoglobin, erythrocytes membrane oxidation, hemolysis, and low plasma heme scavenging enzyme, hemopexin, compared to people with HIV without CWP and HIV-negative individuals with or without pain. In addition, the leukocytes from people with HIV with CWP had attenuated levels of the heme metabolizing enzyme, heme oxygenase-1, which metabolizes free heme to carbon-monoxide and biliverdin. These individuals also had elevated ER stress, and low β-endorphin in leukocytes. In vitro, heme exposure or heme oxygenase-1 deletion, decreased β-endorphins in murine monocytes/macrophages. Treating cells with a carbon-monoxide donor or an ER stress inhibitor, increased β-endorphins. To mimic hemolytic effects in a preclinical model, C57BL/6 mice were injected with phenylhydrazine hydrochloride (PHZ). PHZ increased cell-free heme and ER stress, decreased leukocyte β-endorphin levels and hindpaw mechanical sensitivity thresholds. Treatment of PHZ-injected mice with hemopexin blocked these effects, suggesting that heme-induced ER stress and a subsequent decrease in leukocyte β-endorphin is responsible for hypersensitivity in people with HIV.