Epidermal Nerve Fiber Loss Research Articles

Bortezomib-induced neuropathy is in part mediated by the sensitization of TRPV1 channels

TRPV1 is an ion channel that transduces noxious heat and chemical stimuli and is expressed in small fiber primary sensory neurons that represent almost half of skin nerve terminals. Tissue injury and inflammation result in the sensitization of TRPV1 and sustained activation of TRPV1 can lead to cellular toxicity though calcium influx. To identify signals that trigger TRPV1 sensitization after a 24-h exposure, we developed a phenotypic assay in mouse primary sensory neurons and performed an unbiased screen with a compound library of 480 diverse bioactive compounds. Chemotherapeutic agents, calcium ion deregulators and protein synthesis inhibitors were long-acting TRPV1 sensitizers. Amongst the strongest TRPV1 sensitizers were proteasome inhibitors, a class that includes bortezomib, a chemotherapeutic agent that causes small fiber neuropathy in 30–50% of patients. Prolonged exposure of bortezomib produced a TRPV1 sensitization that lasted several days and neurite retraction in vitro and histological and behavioral changes in male mice in vivo. TRPV1 knockout mice were protected from epidermal nerve fiber loss and a loss of sensory discrimination after bortezomib treatment. We conclude that long-term TRPV1 sensitization contributes to the development of bortezomib-induced neuropathy and the consequent loss of sensation, major deficits experienced by patients under this chemotherapeutic agent.

Mechanisms and treatments of neuropathic itch in a mouse model of lymphoma.

Our understanding of neuropathic itch is limited due to a lack of relevant animal models. Patients with cutaneous T cell lymphoma (CTCL) experience severe itching. Here, we characterize a mouse model of chronic itch with remarkable lymphoma growth, immune cell accumulation, and persistent pruritus. Intradermal CTCL inoculation produced time-dependent changes in nerve innervations in lymphoma-bearing skin. In the early phase (20 days), CTCL caused hyperinnervations in the epidermis. However, chronic itch was associated with loss of epidermal nerve fibers in the late phases (40 and 60 days). CTCL was also characterized by marked nerve innervations in mouse lymphoma. Blockade of C-fibers reduced pruritus at early and late phases, whereas blockade of A-fibers only suppressed late-phase itch. Intrathecal (i.t.) gabapentin injection reduced late-phase, but not early-phase, pruritus. IL-31 was upregulated in mouse lymphoma, whereas its receptor Il31ra was persistently upregulated in Trpv1-expressing sensory neurons in mice with CTCL. Intratumoral anti-IL-31 treatment effectively suppressed CTCL-induced scratching and alloknesis (mechanical itch). Finally, i.t. administration of a TLR4 antagonist attenuated pruritus in early and late phases and in both sexes. Collectively, we have established a mouse model of neuropathic and cancer itch with relevance to human disease. Our findings also suggest distinct mechanisms underlying acute, chronic, and neuropathic itch.

Development of EQ-6, a Novel Analogue of Ethoxyquin to Prevent Chemotherapy-Induced Peripheral Neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common and often dose-limiting side effect of many cancer drugs. Because the onset of neuronal injury is known, it is an ideal clinical target to develop neuroprotective strategies. Several years ago, we had identified ethoxyquin as a potent neuroprotective drug against CIPN through a phenotypic drug screening and demonstrated a novel mechanism of action, inhibition of chaperone domain of heat shock protein 90. To improve its drug-like properties we synthesized a novel analogue of ethoxyquin and named it EQ-6 (6-(5-amino)-ethoxy-2,2,4-trimethyl-1,2-dihydroquinoline hydrochloride). Here we show that EQ-6 prevents axon degeneration in primary dorsal root ganglion neurons in vitro, and this axon protection is associated with preserved levels of nicotinamide adenine dinucleotide, a key metabolite in programmed axon degeneration pathway. We also found that EQ-6 prevents loss of epidermal nerve fibers in a mouse model of CIPN induced by paclitaxel and that doses of EQ-6 that provide neuroprotection are associated with reduced tissue levels of SF3B2, a potential biomarker of target engagement. Furthermore, we show that EQ-6 is safe in vitro and in mice with daily administration for a month. We found that oral bioavailability is about 10%, partly due to rapid metabolism in liver, but EQ-6 appears to be concentrated in neural tissues. Given these findings, we propose EQ-6 as a first-in-class drug to prevent CIPN.

Topical Delivery of Muscarinic Receptor Antagonists Prevents and Reverses Peripheral Neuropathy in Female Diabetic Mice.

Muscarinic antagonists promote sensory neurite outgrowth in vitro and prevent and/or reverse multiple indices of peripheral neuropathy in rodent models of diabetes, chemotherapy-induced peripheral neuropathy, and HIV protein-induced neuropathy when delivered systemically. We measured plasma concentrations of the M1 receptor-selective muscarinic antagonist pirenzepine when delivered by subcutaneous injection, oral gavage, or topical application to the skin and investigated efficacy of topically delivered pirenzepine against indices of peripheral neuropathy in diabetic mice. Topical application of 2% pirenzepine to the paw resulted in plasma concentrations 6 hours postdelivery that approximated those previously shown to promote neurite outgrowth in vitro. Topical delivery of pirenzepine to the paw of mice with streptozotocin-induced diabetes dose-dependently (0.1%-10.0%) prevented tactile allodynia, thermal hypoalgesia, and loss of epidermal nerve fibers in the treated paw and attenuated large fiber motor nerve conduction slowing in the ipsilateral limb. Efficacy against some indices of neuropathy was also noted in the contralateral limb, indicating systemic effects following local treatment. Topical pirenzepine also reversed established paw heat hypoalgesia, whereas withdrawal of treatment resulted in a gradual decline in efficacy over 2-4 weeks. Efficacy of topical pirenzepine was muted when treatment was reduced from 5 to 3 or 1 day/wk. Similar local effects were noted with the nonselective muscarinic receptor antagonist atropine when applied either to the paw or to the eye. Topical delivery of muscarinic antagonists may serve as a practical therapeutic approach to treating diabetic and other peripheral neuropathies. SIGNIFICANCE STATEMENT: Muscarinic antagonist pirenzepine alleviates diabetic peripheral neuropathy when applied topically in mice.

Therapeutic Effect of Y-27632 on Tumorigenesis and Cisplatin-Induced Peripheral Sensory Loss through RhoA-NF-κB.

Chemotherapy-induced peripheral neuropathy (CIPN) is a major side effect of cancer therapy that frequently requires a reduction or cessation of treatments and negatively impacts the patient's quality of life. There is currently no effective means to prevent or treat CIPN. In this study, we developed and applied CIPN in an immunocompetent, syngeneic murine Lewis Lung Carcinoma (LLCab) model that enabled the elucidation of both tumor and host responses to cisplatin and treatments of Y-27632, a selective inhibitor of Rho kinase/p160ROCK. Y-27632 not only preserved cisplatin's efficacy toward tumor suppression but also the combination treatment inhibited tumor cell proliferation and increased cellular apoptosis. By alleviating the cisplatin-induced loss of epidermal nerve fibers (ENFs), Y-27632 protected tumor-bearing mice from cisplatin-induced reduction of touch sensation. Furthermore, quantitative proteomic analysis revealed the striking cisplatin-induced dysregulation in cellular stress (inflammation, mitochondrial deficiency, DNA repair, etc.)-associated proteins. Y-27632 was able to reverse the changes of these proteins that are associated with Rho GTPase and NF-κB signaling network, and also decreased cisplatin-induced NF-κB hyperactivation in both footpad tissues and tumor. Therefore, Y-27632 is an effective adjuvant in tumor suppression and peripheral neuroprotection. These studies highlight the potential of targeting the RhoA-NF-κB axis as a combination therapy to treat CIPN. IMPLICATIONS: This study, for the first time, demonstrated the dual antineoplastic and neuroprotective effects of Rho kinase/p160ROCK inhibition in a syngeneic immunocompetent tumor-bearing mouse model, opening the door for further clinical adjuvant development of RhoA-NF-κB axis to improve chemotherapeutic outcomes.

SIV-Induced Immune Activation and Metabolic Alterations in the Dorsal Root Ganglia During Acute Infection.

Human immunodeficiency virus-associated sensory neuropathy (HIV-SN) remains a frequent neurologic complication of HIV infection. Little is known about alterations in the peripheral nervous system during the early stages of HIV, a time when neuroprotective interventions may be most beneficial. We performed Nanostring gene expression analysis on lumbar dorsal root ganglia (DRG) from 6 simian immunodeficiency virus (SIV)-infected pigtailed macaques killed at 7 days post-inoculation and 8 uninfected controls. We found significant upregulation of many genes involved in immune signaling and activation in the DRG. Among genes related to glutamate metabolism, there was significant upregulation of glutamine synthetase (GS), while glutaminase (GLS) was downregulated. Several genes involved in the oxidative stress response also showed significant differential regulation in the DRG of 7d SIV-infected animals, with superoxide dismutase-2 (SOD2) showing the greatest median fold change compared to controls. Novel findings in the DRG were compared to corresponding brain data and further investigated at the protein level by Western blotting and immunohistochemistry. Together with our previous finding of significant epidermal nerve fiber loss at 14 days post-SIV infection, results of this study demonstrate that immune activation and altered cellular metabolism at in the DRG precede and likely contribute to early sensory nerve injury in HIV-SN.

Structural, functional, and symptom relations in painful distal symmetric polyneuropathies: a systematic review.

The diagnosis of distal symmetric polyneuropathies (DSPs) relies on the presenting symptomatology and neurological sensory examination, supported by objectively quantified structural and functional changes in sensory nerves. Although these separate components have important diagnostic utility, the associations between the structural vs the symptomatic and functional findings in painful DSP are still unclear. It is assumed that delineation of the correlations, or lack of such, between structure, clinical presentation, and function will contribute to a better understanding and treatment of DSP. This systematic review assessed small fiber morphology in patients with different types of painful DSP, and compared it with symptoms, signs, and nerve fiber function. Overall, 111 papers met the inclusion criteria for the systematic review. The results indicate that epidermal nerve fiber loss, in isolation, is not a useful indicator of painful symptoms or their severity in DSP. Intraepidermal nerve fiber density correlated reasonably well with neuropathy scores on tools assessing signs and symptoms (such as the Michigan Neuropathy Screening Instrument and the Total Neuropathy Score), but less so with symptom measures only. Among various psychophysical sensory measures, warmth detection and heat pain thresholds correlated best with intraepidermal nerve fiber density, particularly when assessed at the same anatomical site. The observed sources of heterogeneity, and the lack of associations between structural and functional measures in several studies are discussed. A framework is proposed for uniform assessment of nerve fiber parameters for investigating clinically relevant mechanisms of neuropathic pain in DSP.

NRP-1 Receptor Expression Mismatch in Skin of Subjects with Experimental and Diabetic Small Fiber Neuropathy.

The in vivo cutaneous nerve regeneration model using capsaicin is applied extensively to study the regenerative mechanisms and therapeutic efficacy of disease modifying molecules for small fiber neuropathy (SFN). Since mismatches between functional and morphological nerve fiber recovery are described for this model, we aimed at determining the capability of the capsaicin model to truly mimic the morphological manifestations of SFN in diabetes. As nerve and blood vessel growth and regenerative capacities are defective in diabetes, we focused on studying the key regulator of these processes, the neuropilin-1 (NRP-1)/semaphorin pathway. This led us to the evaluation of NRP-1 receptor expression in epidermis and dermis of subjects presenting experimentally induced small fiber neuropathy, diabetic polyneuropathy and of diabetic subjects without clinical signs of small fiber neuropathy. The NRP-1 receptor was co-stained with CD31 vessel-marker using immunofluorescence and analyzed with Definiens® technology. This study indicates that capsaicin application results in significant loss of epidermal NRP-1 receptor expression, whereas diabetic subjects presenting small fiber neuropathy show full epidermal NRP-1 expression in contrast to the basal expression pattern seen in healthy controls. Capsaicin induced a decrease in dermal non-vascular NRP-1 receptor expression which did not appear in diabetic polyneuropathy. We can conclude that the capsaicin model does not mimic diabetic neuropathy related changes for cutaneous NRP-1 receptor expression. In addition, our data suggest that NRP-1 might play an important role in epidermal nerve fiber loss and/or defective regeneration and that NRP-1 receptor could change the epidermal environment to a nerve fiber repellant bed possibly through Sem3A in diabetes.

Elevated pain threshold in patients with asymptomatic diabetic neuropathy: an intraepidermal electrical stimulation study.

The loss of epidermal nerve fibers is regarded as an early pathological change in human diabetes. We investigated epidermal Aδ nerve fiber function by examining pain threshold by means of intraepidermal electrical stimulation (IES) in early diabetic neuropathy. We recruited 20 asymptomatic diabetic patients. Eighteen age-matched, healthy subjects served as controls. We placed the IES electrode onto the skin of the foot dorsum and delivered weak electrical stimulation. We defined pain threshold as the minimum electrical intensity at which a subject felt a pricking sensation. The mean pain thresholds in the patient group were significantly higher (0.053 ± 0.036 mA; P < 0.01) than in the control group (0.027 ± 0.006 mA). We confirmed that the pain threshold was elevated in early diabetic neuropathy. We conclude that the IES electrode is a useful tool to evaluate early diabetic polyneuropathy. Muscle Nerve 54: 146-149, 2016.

Severe Epidermal Nerve Fiber Loss in Diabetic Neuropathy Is Not Reversed by Long-Term Normoglycemia After Simultaneous Pancreas and Kidney Transplantation.

Whether nerve fiber loss, a prominent feature of advanced diabetic neuropathy, can be reversed by reestablishment of normal glucose control remains questionable. We present 8-year follow-up data on epidermal nerve fiber (ENF) density and neurological function in patients with type 1 diabetes after simultaneous pancreas and kidney transplantation (SPK) with long-term normoglycemia. Distal thigh skin biopsies with ENF counts, vibration perception thresholds (VPTs), autonomic function testing (AFT) and electrophysiological examinations were performed at time of SPK and 2.5 and 8 years after SPK in 12 patients with type 1 diabetes. In comparison to controls, baseline ENF density, VPT and AFT results of patients indicated severe neuropathy. At follow-up, all SPK recipients were insulin independent with excellent glycemic control and kidney graft function; however, the severe ENF depletion present at baseline had not improved, with total ENF absence in 11 patients at 8-year follow-up. Similarly, no amelioration occurred in the VPT and AFT results. Numerical improvement was seen in some electrophysiological parameters; however, statistical significance was achieved only in median motor nerve conduction velocity. ENF loss and functional deficits in advanced diabetic peripheral neuropathy are rarely reversible, even by long-term normoglycemia, which underscores the importance of neuropathy prevention by early optimal glycemic control.

Neuroinflammation and virus replication in the spinal cord of simian immunodeficiency virus-infected macaques.

Studies of neurologic diseases induced by simian immunodeficiency virus (SIV) in Asian macaques have contributed greatly to the current understanding of human immunodeficiency virus pathogenesis in the brain and peripheral nervous system. Detailed investigations into SIV-induced alterations in the spinal cord, a critical sensorimotor relay point between the brain and the peripheral nervous system, have yet to be reported. In this study, lumbar spinal cords from SIV-infected pigtailed macaques were examined to quantify SIV replication and associated neuroinflammation. In untreated SIV-infected animals, there was a strong correlation between amount of SIV RNA in the spinal cord and expression of the macrophage marker CD68 and the key proinflammatory mediators tumor necrosis factor and CCL2. We also found a significant correlation between SIV-induced alterations in the spinal cord and the degree of distal epidermal nerve fiber loss among untreated animals. Spinal cord changes (including elevated glial fibrillary acidic protein immunostaining and enhanced CCL2 gene expression) also were present in SIV-infected antiretroviral drug-treated animals despite SIV suppression. A fuller understanding of the complex virus and host factor dynamics in the spinal cord during human immunodeficiency virus infection will be critical in the development of new treatments for human immunodeficiency virus-associated sensory neuropathies and studies aimed at eradicating the virus from the central nervous system.

Clinical staging of type 2 diabetes: The time has come

Clinical staging of type 2 diabetes: The time has come

Anesthesia Literature Review

Anesthesia Literature Review

Poly(ADP-ribose) polymerase inhibition as a novel therapeutic approach against intraepidermal nerve fiber loss and neuropathic pain associated with advanced diabetic neuropathy: A commentary on “PARP Inhibition or gene deficiency counteracts intraepidermal nerve fiber loss and neuropathic pain in advanced diabetic neuropathy”

Poly(ADP-ribose) polymerase inhibition as a novel therapeutic approach against intraepidermal nerve fiber loss and neuropathic pain associated with advanced diabetic neuropathy: A commentary on “PARP Inhibition or gene deficiency counteracts intraepidermal nerve fiber loss and neuropathic pain in advanced diabetic neuropathy”

Painful small-fiber neuropathy in Sjögren syndrome

Of 20 consecutive patients with Sjögren neuropathy, 16 (80%) presented with burning feet and 12 (60%) with non-length-dependent sensory symptoms. Leg and thigh skin biopsies, performed in 13 patients, including 7 with normal electrophysiology, showed either reduced epidermal nerve fiber (ENF) density or abnormal morphology. ENF loss was frequently non length dependent, suggesting that patients with this disorder commonly have a small-fiber sensory neuronopathy rather than a "dying-back" axonopathy.

INTRAVESICAL RESINIFERATOXIN FOR THE TREATMENT OF INTERSTITIAL CYSTITIS: A RANDOMIZED, DOUBLE-BLIND, PLACEBO CONTROLLED TRIAL

Interstitial cystitis is a painful bladder condition of unknown etiology and poorly understood pathophysiology. Current therapies have met with limited success. Vanilloid receptor agonists such as resiniferatoxin (RTX) desensitize C-fibers that transmit pain; it is hypothesized that such drugs will be effective in the treatment of interstitial cystitis and painful bladder syndrome by decreasing the pain that leads to urinary frequency and urgency. A randomized, double-blind, placebo controlled study was conducted in 163 patients with interstitial cystitis. Participants were randomly assigned to receive a single intravesical dose of 50 ml of either RTX 0.01 microM, 0.05 microM, 0.10 microM, or placebo. Safety and efficacy was evaluated over 12 weeks. The primary efficacy endpoint was the Global Response Assessment, a 7-point scale rating overall change in symptoms of interstitial cystitis after 4 weeks. Secondary efficacy endpoints included reduction in pain, urgency, frequency, nocturia, average void volume, and the O'Leary-Sant Symptom and Problem Indexes. RTX did not improve overall symptoms, pain, urgency, frequency, nocturia, or average void volume during 12 weeks followup. RTX resulted in a dose-dependent increase in the incidence of instillation pain, but was otherwise generally well tolerated. In the largest prospective, randomized clinical trial reported to date with intravesical vanilloid therapy, single administration of RTX at doses of 0.01 microM to 0.10 microM was not effective in patients with interstitial cystitis.

Influence of thermode size for detecting heat pain dysfunction in a capsaicin model of epidermal nerve fiber loss

Quantitative sensory testing of heat pain sensation has become an important tool to evaluate small caliber afferent nerve function in peripheral neuropathy. In earlier studies, we found that topical application of capsaicin in humans results in the loss of epidermal nerve fibers (ENFs) with a corresponding decrease in detection of heat pain sensation. Capsaicin may therefore be a useful model for developing optimal psychophysical testing procedures for detection of neuropathy in its early stages. Here we determined the influence of thermal probe (thermode) size in detecting the diminished heat pain sensation following capsaicin application. Twelve healthy volunteers applied 0.075% capsaicin topically to the volar forearm four times daily for 7 days. Psychophysical measures of heat pain, mechanical (sharp) pain, and tactile threshold were obtained daily from untreated control skin and from capsaicin-treated skin during capsaicin application, and once weekly for 5 weeks following discontinuation of capsaicin. Heat pain sensation was assessed using a large (30×30 mm) and small (3×3 mm) thermode and different algorithms to assess pain threshold and suprathreshold heat pain. Skin biopsies were obtained and were processed for immunohistochemical localization of (ENFs) using the pan neuronal marker protein gene product 9.5. Capsaicin produced a rapid decrease in the number of ENFs, with nearly complete disappearance after 3 days of treatment. Heat pain evoked by the small, but not the large, thermode decreased dramatically after capsaicin treatment. The sensation of heat pain returned toward normal after 2–3 weeks following discontinuation of capsaicin treatment concordant with gradual reinnervation of the epidermis. Regression analysis indicated that the sensation of heat pain evoked by the small thermode correlated much better with the number of ENFs than heat pain evoked by the large thermode. The detection of sharp pain decreased moderately after capsaicin treatment. Assessment of heat pain sensation using small thermodes has potential for detecting sensory deficits in early stages of small fiber neuropathy.

Topical capsaicin in humans: parallel loss of epidermal nerve fibers and pain sensation

Capsaicin applied topically to human skin produces itching, pricking and burning sensations due to excitation of nociceptors. With repeated application, these positive sensory responses are followed by a prolonged period of hypalgesia that is usually referred to as desensitization, or nociceptor inactivation. Consequently, capsaicin has been recommended as a treatment for a variety of painful syndromes. The precise mechanisms that account for nociceptor desensitization and hypalgesia are unclear. The present study was performed to determine if morphological changes of intracutaneous nerve fibers contribute to desensitization and hypalgesia. Capsaicin (0.075%) was applied topically to the volar forearm four times daily for 3 weeks. At various time intervals tactile, cold, mechanical and heat pain sensations were assessed in the treated and in contralateral untreated areas. Skin blisters and skin biopsies were collected and immunostained for protein gene product (PGP) 9.5 to assess the morphology of cutaneous nerves and to quantify the number of epidermal nerve fibers (ENFs). Capsaicin resulted in reduced sensitivity to all cutaneous stimuli, particularly to noxious heat and mechanical stimuli. This hypalgesia was accompanied by degeneration of epidermal nerve fibers as evidenced by loss of PGP 9.5 immunoreactivity. As early as 3 days following capsaicin application, there was a 74% decrease in the number of nerve fibers in blister specimens. After 3 weeks of capsaicin treatment, the reduction was 79% in blisters and 82% in biopsies. Discontinuation of capsaicin was followed by reinnervation of the epidermis over a 6-week period with a return of all sensations, except cold, to normal levels. We conclude that degeneration of epidermal nerve fibers contributes to the analgesia accredited to capsaicin. Furthermore, our data demonstrate that ENFs contribute to the painful sensations evoked by noxious thermal and mechanical stimuli.