Phosphorylated CCAAT/Enhancer Binding Protein β Contributes to Rat HIV-Related Neuropathic Pain:<i>In Vitro</i>and<i>In Vivo</i>Studies

Abstract

Chronic pain is increasingly recognized as an important comorbidity of HIV-infected patients, however, the exact molecular mechanisms of HIV-related pain are still elusive. CCAAT/enhancer binding proteins (C/EBPs) are expressed in various tissues, including the CNS. C/EBPβ, one of the C/EBPs, is involved in the progression of HIV/AIDS, but the exact role of C/EBPβ and its upstream factors are not clear in HIV pain state. Here, we used a neuropathic pain model of perineural HIV envelope glycoprotein gp120 application onto the rat sciatic nerve to test the role of phosphorylated C/EBPβ (pC/EBPβ) and its upstream pathway in the spinal cord dorsal horn (SCDH). HIV gp120 induced overexpression of pC/EBPβ in the ipsilateral SCDH compared with contralateral SCDH. Inhibition of C/EBPβ using siRNA against C/EBPβ reduced mechanical allodynia. HIV gp120 also increased TNFα, TNFRI, mitochondrial superoxide (mtO₂^·-), and pCREB in the ipsilateral SCDH. ChIP-qPCR assay showed that pCREB enrichment on the C/EBPβ gene promoter regions in rats with gp120 was higher than that in sham rats. Intrathecal TNF soluble receptor I (functionally blocking TNFα bioactivity) or knockdown of TNFRI using antisense oligodeoxynucleotide against TNFRI reduced mechanical allodynia, and decreased mtO₂^·-, pCREB and pC/EBPβ. Intrathecal Mito-tempol (a mitochondria-targeted O₂^·-scavenger) reduced mechanical allodynia and decreased pCREB and pC/EBPβ. Knockdown of CREB with antisense oligodeoxynucleotide against CREB reduced mechanical allodynia and lowered pC/EBPβ. These results suggested that the pathway of TNFα/TNFRI-mtO₂^·--pCREB triggers pC/EBPβ in the HIV gp120-induced neuropathic pain state. Furthermore, we confirmed the pathway using both cultured neurons treated with recombinant TNFα <i>in vitro</i> and repeated intrathecal injection of recombinant TNFα in naive rats. This finding provides new insights in the understanding of the HIV neuropathic pain mechanisms and treatment.<b>SIGNIFICANCE STATEMENT</b> Painful HIV-associated sensory neuropathy is a neurological complication of HIV infection. Phosphorylated C/EBPβ (pC/EBPβ) influences AIDS progression, but it is still not clear about the exact role of pC/EBPβ and the detailed upstream factors of pC/EBPβ in HIV-related pain. In a neuropathic pain model of perineural HIV gp120 application onto the sciatic nerve, we found that pC/EBPβ was triggered by TNFα/TNFRI-mtO₂^·--pCREB signaling pathway. The pathway was confirmed by using cultured neurons treated with recombinant TNFα <i>in vitro</i>, and by repeated intrathecal injection of recombinant TNFα in naive rats. The present results revealed the functional significance of TNFα/TNFRI-mtO₂^·--pCREB-pC/EBPβ signaling in HIV neuropathic pain, and should help in the development of more specific treatments for neuropathic pain.