CD4+ αß T cell infiltration into the leptomeninges of lumbar dorsal roots contributes to the transition from acute to chronic mechanical allodynia after adult rat tibial nerve injuries.

BACKGROUND: Antigen-specific and MHCII-restricted CD4+ αß T cells have been shown or suggested to play an important role in the transition from acute to chronic mechanical allodynia after peripheral nerve injuries. However, it is still largely unknown where these T cells infiltrate along the somatosensory pathways transmitting mechanical allodynia to initiate the development of chronic mechanical allodynia after nerve injuries. Therefore, the purpose of this study was to ascertain the definite neuroimmune interface for these T cells to initiate the development of chronic mechanical allodynia after peripheral nerve injuries. METHODS: First, we utilized both chromogenic and fluorescent immunohistochemistry (IHC) to map αß T cells along the somatosensory pathways for the transmission of mechanical allodynia after modified spared nerve injuries (mSNIs), i.e., tibial nerve injuries, in adult male Sprague-Dawley rats. We further characterized the molecular identity of these αß T cells selectively infiltrating into the leptomeninges of L4 dorsal roots (DRs). Second, we identified the specific origins in lumbar lymph nodes (LLNs) for CD4+ αß T cells selectively present in the leptomeninges of L4 DRs by two experiments: (1) chromogenic IHC in these lymph nodes for CD4+ αß T cell responses after mSNIs and (2) fluorescent IHC for temporal dynamics of CD4+ αß T cell infiltration into the L4 DR leptomeninges after mSNIs in prior lymphadenectomized or sham-operated animals to LLNs. Finally, following mSNIs, we evaluated the effects of region-specific targeting of these T cells through prior lymphadenectomy to LLNs and chronic intrathecal application of the suppressive anti-αßTCR antibodies on the development of mechanical allodynia by von Frey hair test and spinal glial or neuronal activation by fluorescent IHC. RESULTS: Our results showed that during the sub-acute phase after mSNIs, αß T cells selectively infiltrate into the leptomeninges of the lumbar DRs along the somatosensory pathways responsible for transmitting mechanical allodynia. Almost all these αß T cells are CD4 positive. Moreover, the temporal dynamics of CD4+ αß T cell infiltration into the lumbar DR leptomeninges are specifically determined by LLNs after mSNIs. Prior lymphadenectomy to LLNs specifically reduces the development of mSNI-induced chronic mechanical allodynia. More importantly, intrathecal application of the suppressive anti-αßTCR antibodies reduces the development of mSNI-induced chronic mechanical allodynia. In addition, prior lymphadenectomy to LLNs attenuates mSNI-induced spinal activation of glial cells and PKCγ+ excitatory interneurons. CONCLUSIONS: The noteworthy results here provide the first evidence that CD4+ αß T cells selectively infiltrate into the DR leptomeninges of the somatosensory pathways transmitting mechanical allodynia and contribute to the transition from acute to chronic mechanical allodynia after peripheral nerve injuries.

[Early Systemic Administration of IL-10 Inhibits Neuropathic Pain in Adult Rats with Tibial Nerve Injury].

OBJECTIVES: To determine the effect of early systemic administration of IL-10 on peripheral neuropathic pain induced by tibial nerve permanent transection [modified spared nerve injury (mSNI)]in adult rats. METHODS: Male adult Sprague-Dawley (SD) rats (ten-week old, 250-300 g) with mSNI were randomly divided into mSNI, sham-operated, IL-10 intervention (intraperitoneal injection), PBS intervention (intraperitoneal injection) groups, each containing six rats. Intraperitoneally injections (IL-10 or PBS) were given immediately after surgeries for a single regime with a dosage of 500 uL (0.1 mg/mL). Plantar test, von Frey hairs test, pinprick test and acetone test were performed before and after tibial nerve injuries (0 d, 4/5 d, 7/8 d, 14/15 d) to evaluate region-specific pain responses of the rats on the plantar sural and saphenous skin territories of ipsilateral and contralateral hindpaws. The hindpaw position (on 8 d) of six additional rats with standard SNI was compared with those with mSNI. RESULTS: The rats with standard SNI showed an eversion posture of hindpaws, more prominent than those with mSNI. Region-specific pathological pain evoked by mechanical and thermal stimuli on the sural and saphenous skin territories of the plantar surfaces of rat hindpaws was demonstrated on the ipsilateral rather than contralateral hindpaws. This effect was shown in the rats with mSNI but not in those with sham operations. Compared with PBS, early intraperitoneal injection of IL-10 significantly and persistently attenuated either allodynia or hyperalgesia in the rats with mSNI. CONCLUSIONS: Tibial nerve permanent transection models of adult rats can be used as a simple but useful rodent model of peripheral neuropathic pain. Early systemic administration of IL-10 impairs the pathogenesis of neuropathic pain induced by tibial nerve injuries.

Behavioral characterization of neuropathic pain on the glabrous skin areas reinnervated solely by axotomy-regenerative axons after adult rat sciatic nerve crush.

In cranial and spinal nerve ganglia, both axotomized primary sensory neurons without regeneration (axotomy-nonregenerative neurons) and spared intact primary sensory neurons adjacent to axotomized neurons (axotomy-spared neurons) have been definitely shown to participate in pain transmission in peripheral neuropathic pain states. However, whether axotomized primary sensory neurons with regeneration (axotomy-regenerative neurons) would be integral components of neural circuits underlying peripheral neuropathic pain states remains controversial. In the present study, we utilized an adult rat sciatic nerve crush model to systematically analyze pain behaviors on the glabrous plantar surface of the hindpaw sural nerve skin territories. To the best of our knowledge, our results for the first time showed that heat hyperalgesia, cold allodynia, mechanical allodynia, and mechanical hyperalgesia emerged and persisted on the glabrous sural nerve skin areas after adult rat sciatic nerve crush. Interestingly, mechanical hyperalgesia was sexually dimorphic. Moreover, with our optimized immunofluorescence staining protocol of free-floating thick skin sections for wide-field epifluorescence microscopic imaging, changes in purely regenerative reinnervation on the same skin areas by axotomized primary sensory afferents were shown to be paralleled by those pathological pain behaviors. To our surprise, Protein Gene Product 9.5-immunoreactive nerve fibers with regular and large varicosities ectopically emigrated into the upper dermis of the glabrous sural nerve skin territories after adult rat sciatic nerve crush. Our results indicated that axotomy-regenerative primary sensory neurons could be critical elements in neural circuits underlying peripheral neuropathic pain states. Besides, our results implied that peripheral neuropathic pain transmitted by axotomy-regenerative primary sensory neurons alone might be a new dimension in the clinical therapy of peripheral nerve trauma beyond regeneration.

ProBDNF inhibits collective migration and chemotaxis of rat Schwann cells.

Schwann cell migration, including collective migration and chemotaxis, is essential for the formation of coordinate interactions between Schwann cells and axons during peripheral nerve development and regeneration. Moreover, limited migration of Schwann cells imposed a serious obstacle on Schwann cell-astrocytes intermingling and spinal cord repair after Schwann cell transplantation into injured spinal cords. Recent studies have shown that mature brain-derived neurotrophic factor, a member of the neurotrophin family, inhibits Schwann cell migration. The precursor form of brain-derived neurotrophic factor, proBDNF, was expressed in the developing or degenerating peripheral nerves and the injured spinal cords. Since "the yin and yang of neurotrophin action" has been established as a common sense, proBDNF would be expected to promote Schwann cell migration. However, we found, in the present study, that exogenous proBDNF also inhibited in vitro collective migration and chemotaxis of RSC 96 cells, a spontaneously immortalized rat Schwann cell line. Moreover, proBDNF suppressed adhesion and spreading of those cells. At molecular level, proBDNF inhibits F-actin polymerization and focal adhesion dynamics in cultured RSC 96 cells. Therefore, our results suggested a special case against the classical opinion of "the yin and yang of neurotrophin action" and implied that proBDNF might modulate peripheral nerve development or regeneration and spinal cord repair through perturbing native or transplanted Schwann cell migration.