Molecular Pain

Molecular Pain

Objectives. Franks, William F. The symptoms of HIV-SN are dominated by distal neuropathic pain. Patients received relief when they took medications addressing the herpes infection. When combined or compared with transgenic approaches, this vector may provide valuable information in the mechanistic study of pain. Further studies of the mechanisms of chronic pain have opened the way for development of new treatment strategies, one of which is gene therapy. To elucidate the role of spinal TNF in the pathogenesis of neuropathic pain, we used viral vector-mediated transfer of the gene coding for the p55 sTNFR antagonist of TNF to the dorsal root ganglion (DRG) to release p55 sTNFR into the dorsal horn of spinal cord in rats with neuropathic pain created by selective SNL.

24). In previous studies we have shown that vectors constructed from nonreplicating herpes simplex virus (HSV) recombinants can be used effectively to transfer genes into DRG neurons from subcutaneous inoculation, and that release of inhibitory neurotransmitters or antiinflammatory proteins from the central afferent terminals of transduced DRG neurons in vivo can be used to modulate pain-related behaviors.11,12,17,18 In this study, we used a nonreplicating HSV vector constructed to express and release IL-10 in the formalin model of inflammatory pain. Cytokines have pleiotropic activities that can trigger several cellular responses depending on cell type, timing, and molecular environment. Curiously, transduction of the DRG following subcutaneous delivery has not been observed previously for rAAV, with rAAV serotype 2 (rAAV serotype 2 genome packaged in the serotype 2 capsid, rAAV2/2) failing to transduce DRG cells following subcutaneous injection into the hindpaw of rats [13]. This is not surprising as rAAV2/2 is inefficient at retrograde transport in comparison to rAAV2/6 [25]. Specifically, we will discuss the use of herpes simplex virus (HSV) vectors in both the treatment of chronic pain and the identification and delivery of novel therapeutic interventions. codeine) to strong opioids such as morphine.
Molecular Pain

This is in part due to the use of secreted transgenic peptides that have had the capacity to exert paracrine functions on non-transduced cells. But infection of DRG neurons by serotype 8 AAV is apparently unique; serotype 2 AAV, for example, does not infect DRG neurons when administered by the same route. A double-blind, randomized, controlled trial has demonstrated that methadone 10 to 20 mg per day has an analgesic effect in neuropathic pain (Watson et al 2003). For conditions in which the primary cause of the chronic pain is either not readily reversible or not identifiable the search for treatment has focused primarily on components of the ascending pain pathway. At a minimum, serotype 8 AAV should provide an interesting tool for animal studies investigating DRG mechanisms in chronic pain, but important issues are also raised that need to be explored further: (1) what is the rostro-caudal extent of transgene expression in the DRG following intrathecal injection? Thus, in the present study, we investigated the feasibility of HSV vector mediated GAD67 gene therapy for the treatment of DO following SCI. However, recent literature consistently reports that activation of astrocytes and microglia contributes significantly to CNP following SCI (Gwak et al., 2008, Gwak and Hulsebosch, 2009; Hains and Waxman, 2006).

With the goal of increasing the number of transduced nociceptive neurons that would be required for exploring the mechanisms of pain, we examined sciatic nerve and intrathecal routes administration of rAAV that have recently reported success in the transduction of rat DRG. Clatworthy et al. For example, the anti-allodynic effect of systemic loperamide was blocked by systemic pretreatment with naloxone hydrochloride or the peripherally acting MOR-preferring antagonist, methylnaltrexone. Among them, the ionotropic NMDA receptor is most involved in the events correlated with nociception [9], and with the maintenance of central sensitization and hyperexcitability of dorsal horn neurons. Interestingly, the efficiency and profile of transduction was similar for the three mouse strains examined, suggesting that rAAV2/6 transduction of DRG neurons can be applied to the mouse regardless of the model being used. Several lines of evidence indicate that tumor necrosis factor-alpha (TNFα) plays an key role in the development of chronic pain. With the exception of ICP47, these IE gene products trigger the expression of early (E) genes that code for proteins involved in DNA replication and late (L) genes coding for structural HSV proteins.18 As the viral thymidine kinase (tk) is required for replication in neurons, recombinants defective in tk can be propagated in culture and can replicate in skin, but not in the DRG.

Between 6 h and 3 days following injection of DNA, but not vehicle, cell counts in cerebrospinal fluid (CSF) at the site of injection are significantly elevated. Although surgical resection alleviates cancer pain, more effective analgesics are still needed; pre-surgical patients often experience extended periods of severe pain, some cancers cannot be excised, some patients are too sick to have surgery, and many patients develop an untreatable recurrence of the cancer or a second primary cancer. All the tissue culture experiments were repeated 3 times.

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Molecular Pain

Molecular Pain

45 Sprague-Dawley rats underwent chronic constriction. The patients had no visible lesion of the central nervous system on magnetic resonance imaging or abnormality in electrophysiological studies. We first investigated basal nociception, capsaicin-induced hyperalgesia, and opioid analgesia after topical infection with a recombinant herpes simplex virus type 1 (HSV-1) containing cDNA sequences for mu-opioid receptor (SGMOR), human preproenkephalin (KPE), a mix of SGMOR and KPE, or E. To determine whether an injury-specific promoter can produce neuron-specific MOR expression and enhanced antinociception, we compared animals infected with a galanin promoter virus (galMOR) or a human cytomegalovirus promoter virus (cmvMOR). When combined or compared with transgenic approaches, this vector may provide valuable information in the mechanistic study of pain. However, one needs to be aware that this “novel” approach is still in its infancy and that many of its details need to be further elucidated. Chancellor, and Naoki Yoshimura.

This series of behavioral, neurochemical and immunohistochemical studies will provide important new information about the mechanisms by which enhanced glia activation releasing proinflammatory cytokines induces spinal sensitization, and may point towards novel therapeutic approaches to treat that pain in morphine tolerance. The use of viral vectors offers a potential therapeutic option for treatment of central nervous system disorders. Gene transfer approaches may be a useful method to achieve continuous local expression of short lived analgesic peptides in the spinal cord and are being considered for clinical application. This leads to great suffering for patients and results in a heavy burden for society. Subcutaneous inoculation with T0TNFsR (30 l containing 2 108 plaque-forming units/ml) in the plantar surface of the hind foot ipsilateral to the SNL 1 week before SNL delayed the development of both mechanical allodynia and thermal hyperalgesia (Figure 3a and b) for a period of 4 weeks. The remaining sequences, e.g., shRNA-A (Figure 1a), did not significantly affect Nav1.3 expression. In a preliminary study we confirmed that transfection of dissociated primary DRG neurons in vitro with vector QHIL10 at multiplicity of infection (MOI) of 1 resulted in robust expression and release of IL-10 ().

2004). The possibility of long-term expression of therapeutic peptides in specific cells is especially attractive as a treatment option for conditions such as SCI neuropathic pain. Neuropathic pain is initiated or caused by multiple etiological factors; therefore sustained simultaneous targeting of several signalling pathways is likely necessary to obtain effective analgesia. These vectors are derived from HSV type 1 in which virus replication has been compromised by the deletion of essential viral gene functions required to initiate and sustain virus replication. In addition, continued treatment with opioids results in the development of tolerance leading to a requirement for escalating doses of medication to achieve the same result. Considering these advantages, reproducibility, and safety, we chose the lentivirus-mediated gene therapy system as an initial proof-of-concept in this study. (3) What is the effect on brain infection if the vector is injected at thoracic or higher levels of the spinal cord?

Molecular Pain
These include carbamazepine, valproic acid, lamotrigine, levetiracetam, and gabapentin. For example, selective expression of the inhibitory neurotransmitter in the amygdala, or the rostral a granular insular cortex can be used to selectively modify nociceptive neurotransmission. Meanwhile, earlier this year, the FDA approved an investigational new drug application for a phase 1 human trial of a nonreplicating herpes simplex virus vector expressing enkephalin, injected into the skin, for treatment of pain. Endomorphins inhibit nociceptive transmission in the spinal cord through μ-opioid receptors which are localized in neuronal circuits involved in processing of nociceptive information [27]. Upregulated glutamate receptors and ion channels, increased release of proinflammatory cytokines and reactive oxygen species (ROS), activation of glial cells and subsequent activation of intracellular downstream/upstream cascades (Crown et al., 2006; Gwak et al., 2008; 2009a; Hains et al., 2003, Leem et al., 2010; Tan et al., 2008; Zinck et al., 2007) are predominant events that lead to enhanced pain transmission following SCI. Antinociceptive effects of the intrathecal injection of EM-1 or EM-2 were observed in models of peripheral nerve injury [18, 31]. Cytokines are low molecular weight glycoproteins that are secreted mainly, but not exclusively, by immunological cells such as T-cells, macrophages and neutrophils.

The HSV vector targets primary afferent neurons that innervate the inoculation site, thus bypassing the need for surgical delivery of virus. Is the neuropathic pain a complete disease and not only the result of an other syndrome or injury? SHG was identified in a screen for compounds that enhanced adrenal transplant-mediated analgesia in rat pain models [62]. The activity of sTNFα can be terminated in part by binding to cleaved soluble forms of the TNFR (p55 or p75 sTNFR). shows the design of HSV vectors applied to gene therapy studies in animal models and humans and vectors with improved capacity and stability. Moreover, if cell influx is blocked through IL-10 protein pre- and co-treatment, therapeutic benefit is observed only for several days following a second pDNA-IL-10F129S injection as opposed to the normal duration of greater than 3 months (). Studies in other cancers suggest that adding NSAIDs to the treatment regimen improves pain control.

In the current study, deleterious motor side effects induced by the lentiviral injection encoding SHG were not observed. Similarly, in previous studies using bolus intrathecal SHG injection, locomotor side effects were not observed in contrast to other NMDA antagonists [17]. Using the clip compression model, rats develop a partial paralysis due to SCI with modest progressive recovery, as observed with the BBB test. The BBB test scores of animals injected with EMs and SHG lentiviruses were comparable to those without injections and with the animals injected with the control GFP lentivirus. Since IL-4 can bind to type I and II IL-4-R and IL-13 binds to type II IL-4R, both receptors seem to contribute to the antinociceptive effects observed in the studies mentioned above. The analgesic effect was observed as soon as 1 week post injection for some modalities, and sustained for at least up to 7 weeks, when the behavioral study was terminated. Substances released in the vicinity of spared fibers by Wallerian degeneration (e.g., nerve growth factor) or calcium influx into neurons may underlie substantial transcriptome remodeling in neuropathic pain development [10].

To adequately assess and diagnose a patient with neuropathic pain, the HCP will need to perform a focused physical exam. Previous findings in our lab have indicated that the blockade of spinal NMDA receptors alone, via bolus intrathecal injection of either SHG or another NMDA antagonist ketamine, is insufficient in reducing neuropathic pain symptoms in the SCI clip compression model [57]. In wild type infection, the virus is transmitted by direct contact, replicating initially in epithelial cells of skin or mucous membranes. Both SHG and the EMs have reported effectiveness in numerous other pain models, although recent findings in our lab has shown that these effects are variable depending on the selected models and outcome measures [18]. It is likely that the labile nature of small neuropeptides limits their time course and efficacy; thus a gene therapy approach may be ideal in overcoming this limitation. Although the intrathecal injection of endomorphins alone have not been evaluated in the SCI clip compression model, a possible explanation for their limited effectiveness alone using the viral vector approach in the current study is the potential development of tolerance to continued exposure to these mu-opioids mediated by sustained delivery from transduced spinal cells. Thus, it is currently believed that the proinflammatory cytokines that drive chronic pain behavior may be derived from the cellular elements of the nervous system itself, and that these molecules can act directly on receptors expressed by neurons and other cells of the nervous system.

In contrast to the singly delivered peptides, when co-delivered using the combined SHG and EM viral vectors, potent analgesic effects, once initiated, were maintained without decrement through the study duration. The contributions of both the EMs and SHG in mediating the sustained analgesic effects of the viral vectors is supported by the reversal using intrathecal injection of opioid (naloxone) or NMDA antagonist (anti-SHG). The time of delivery of therapeutic compounds in SCI model is an important issue because delayed onset of the pain treatment after SCI is clinically more relevant than early interventions. In this study we showed that the delayed treatment using viral vector-mediated delivery of analgesic peptides could still be effective in alleviating SCI neuropathic pain. While systemically applied ibuprofen reduced both mechanical allodynia (MA) and hypersensitivity [105], another group failed to see any relief using the anti-inflammatory NSAID diclofenac [103]. This suggests that delayed injection of lentiviral vectors encoding the appropriate analgesic peptides can still alleviate chronic pain in SCI models.

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