Suppression of neuropathic pain in the circadian clock-deficient Per2m/m mice involves up-regulation of endocannabinoid system.

Neuropathic pain often results from injuries and diseases that affect the somatosensory system. Disruption of the circadian clock has been implicated in the exacerbation of the neuropathic pain state. However, in this study, we report that mice deficient in a core clock component Period2 (Per2m/m mice) fail to develop tactile pain hypersensitivity even following peripheral nerve injury. Similar to male wild-type mice, partial sciatic nerve ligation (PSL)-Per2m/m male mice showed activation of glial cells in the dorsal horn of the spinal cord and increased expression of pain-related genes. Interestingly, α1D-adrenergic receptor (α1D-AR) expression was up-regulated in the spinal cord of Per2m/m mice, leading to increased production of 2-arachidonoylglycerol (2-AG), an endocannabinoid receptor ligand. This increase in 2-AG suppressed the PSL-induced tactile pain hypersensitivity. Furthermore, intraspinal dorsal horn injection of adeno-associated viral vectors expressing α1D-AR also attenuated pain hypersensitivity in PSL-wild-type male mice by increasing 2-AG production. Our findings reveal an uncovered role of the circadian clock in neuropathic pain disorders and suggest a link between α1D-AR signaling and the endocannabinoid system.

Inhibition of Tumor-Derived C-C Motif Chemokine Ligand 2 Expression Attenuates Tactile Allodynia in NCTC 2472 Fibrosarcoma-Inoculated Mice.

Neuropathic pain associated with cancers is caused by tumor growth compressing and damaging nerves, which would also be enhanced by inflammatory factors through sensitizing nociceptor neurons. A troublesome hallmark symptom of neuropathic pain is hypersensitivity to innocuous stimuli, a condition known as "tactile allodynia", which is often refractory to NSAIDs and opioids. The involvement of chemokine CCL2 (monocyte chemoattractant protein-1) in cancer-evoked neuropathic pain is well established, but opinions remain divided as to whether CCL2 is involved in the production of tactile allodynia with tumor growth. In this study, we constructed Ccl2 knockout NCTC 2472 (Ccl2-KO NCTC) fibrosarcoma cells and conducted pain behavioral test using Ccl2-KO NCTC-implanted mice. Implantation of naïve NCTC cells around the sciatic nerves of mice produced tactile allodynia in the inoculated paw. Although the growth of Ccl2 KO NCTC-formed tumors was comparable to that of naïve NCTC-formed tumors, Ccl2-KO NCTC-bearing mice failed to show tactile pain hypersensitivity, suggesting the involvement of CCL2 in cancer-induced allodynia. Subcutaneous administration of controlled-release nanoparticles containing the CCL2 expression inhibitor NS-3-008 (1-benzyl-3-hexylguanidine) significantly attenuated tactile allodynia in naïve NCTC-bearing mice accompanied by a reduction of CCL2 content in tumor masses. Our present findings suggest that inhibition of CCL2 expression in cancer cells is a useful strategy to attenuate tactile allodynia induced by tumor growth. Development of a controlled-release system of CCL2 expression inhibitor may be a preventative option for the treatment of cancer-evoked neuropathic pain. SIGNIFICANCE STATEMENT: The blockade of chemokine/receptor signaling, particularly for C-C motif chemokine ligand 2 (CCL2) and its high-affinity receptor C-C chemokine receptor type 2 (CCR2), has been implicated to attenuate cancer-induced inflammatory and nociceptive pain. This study demonstrated that continuous inhibition of CCL2 production from cancer cells also prevents the development of tactile allodynia associated with tumor growth. Development of a controlled-release system of CCL2 expression inhibitor may be a preventative option for management of cancer-evoked tactile allodynia.

Senescence-induced alteration of circadian phagocytic activity of retinal pigment epithelium cell line ARPE-19.

Renewal of retinal photoreceptor outer segments is conducted through daily shedding of distal photoreceptor outer segment tips and subsequent their phagocytosis by the adjacent retinal pigment epithelium (RPE) monolayer. Dysregulation of the diurnal clearance of photoreceptor outer segment tips has been implicated in age-related retinal degeneration, but it remains to be clarified how the circadian phagocytic activity of RPE cells is modulated by senescence. In this study, we used the human RPE cell line ARPE-19 to investigate whether hydrogen peroxide (H2O2)-induced senescence in ARPE-19 cells alters the circadian rhythm of their phagocytic activity. After synchronization of the cellular circadian clock by dexamethasone treatment, the phagocytic activity of normal ARPE-19 cells exhibited significant 24-h oscillation, but this oscillation was modulated by senescence. The phagocytic activity of senescent ARPE-19 cells increased constantly throughout the 24-h period, which still exhibited blunted circadian oscillation, accompanied by an alteration in the rhythmic expression of circadian clock genes and clock-controlled phagocytosis-related genes. The expression levels of REV-ERBα, a molecular component of the circadian clock, were constitutively increased in senescent ARPE-19 cells. Furthermore, pharmacological activation of REV-ERBα by its agonist SR9009 enhanced the phagocytic activity of normal ARPE-19 cells and increased the expression of clock-controlled phagocytosis-related genes. Our present findings extend to understand the role of circadian clock in the alteration of phagocytic activity in RPE during aging. Constitutive enhancement of phagocytic activity of senescent RPE may contribute to age-related retinal degeneration.