Sirtuin 3 Mediated by Spinal cMyc-Enhancer of Zeste Homology 2 Pathway Plays an Important Role in Human Immunodeficiency Virus-Related Neuropathic Pain Model.

BACKGROUND: Clinical data demonstrate that chronic use of opioid analgesics increases neuropathic pain in people living with human immunodeficiency virus (HIV). Therefore, it is important to elucidate the molecular mechanisms of HIV-related chronic pain. In this study, we investigated the role of the transcription factor cMyc, epigenetic writer enhancer of zeste homology 2 (EZH2), and sirtuin 3 (Sirt3) pathway in HIV glycoprotein gp120 with morphine (gp120M)-induced neuropathic pain in rats. METHODS: Neuropathic pain was induced by intrathecal administration of recombinant gp120 with morphine. Mechanical withdrawal threshold was measured using von Frey filaments, and thermal latency using the hotplate test. Spinal expression of cMyc, EZH2, and Sirt3 were measured using Western blots. Antinociceptive effects of intrathecal administration of antisense oligodeoxynucleotide against cMyc, a selective inhibitor of EZH2, or recombinant Sirt3 were tested. RESULTS: In the spinal dorsal horn, gp120M upregulated expression of cMyc (ratio of gp120M versus control, 1.68 ± 0.08 vs 1.00 ± 0.14, P = .0132) and EZH2 (ratio of gp120M versus control, 1.76 ± 0.05 vs 1.00 ± 0.16, P = .006), and downregulated Sirt3 (ratio of control versus gp120M, 1.00 ± 0.13 vs 0.43 ± 0.10, P = .0069) compared to control. Treatment with intrathecal antisense oligodeoxynucleotide against cMyc, GSK126 (EZH2 selective inhibitor), or recombinant Sirt3 reduced mechanical allodynia and thermal hyperalgesia in this gp120M pain model. Knockdown of cMyc reduced spinal EZH2 expression in gp120M treated rats. Chromatin immunoprecipitation (ChIP) assay showed that enrichment of cMyc binding to the ezh2 gene promoter region was increased in the gp120M-treated rat spinal dorsal horn, and that intrathecal administration of antisense ODN against cMyc (AS-cMyc) reversed the increased enrichment of cMyc. Enrichment of trimethylation of histone 3 on lysine residue 27 (H3K27me3; an epigenetic mark associated with the downregulation of gene expression) binding to the sirt3 gene promoter region was upregulated in the gp120M-treated rat spinal dorsal horn; that intrathecal GSK126 reversed the increased enrichment of H3K27me3 in the sirt3 gene promoter. Luciferase reporter assay demonstrated that cMyc mediated ezh2 gene transcription at the ezh2 gene promoter region, and that H3K27me3 silenced sirt3 gene transcription at the gene promoter region. CONCLUSION: These results demonstrated that spinal Sirt3 decrease in gp120M-induced neuropathic pain was mediated by cMyc-EZH2/H3K27me3 activity in an epigenetic manner. This study provided new insight into the mechanisms of neuropathic pain in HIV patients with chronic opioids.

Role of Tumor Necrosis Factor Receptor 1-Reactive Oxygen Species-Caspase 11 Pathway in Neuropathic Pain Mediated by HIV gp120 With Morphine in Rats.

BACKGROUND: Recent clinical research suggests that repeated use of opioid pain medications can increase neuropathic pain in people living with human immunodeficiency virus (HIV; PLWH). Therefore, it is significant to elucidate the exact mechanisms of HIV-related chronic pain. HIV infection and chronic morphine induce proinflammatory factors, such as tumor necrosis factor (TNF)α acting through tumor necrosis factor receptor I (TNFRI). HIV coat proteins and/or chronic morphine increase mitochondrial superoxide in the spinal cord dorsal horn (SCDH). Recently, emerging cytoplasmic caspase-11 is defined as a noncanonical inflammasome and can be activated by reactive oxygen species (ROS). Here, we tested our hypothesis that HIV coat glycoprotein gp120 with chronic morphine activates a TNFRI-mtROS-caspase-11 pathway in rats, which increases neuroinflammation and neuropathic pain. METHODS: Neuropathic pain was induced by repeated administration of recombinant gp120 with morphine (gp120/M) in rats. Mechanical allodynia was assessed using von Frey filaments, and thermal latency using hotplate test. Protein expression of spinal TNFRI and cleaved caspase-11 was examined using western blots. The image of spinal mitochondrial superoxide was examined using MitoSox Red (mitochondrial superoxide indicator) image assay. Immunohistochemistry was used to examine the location of TNFRI and caspase-11 in the SCDH. Intrathecal administration of antisense oligodeoxynucleotide (AS-ODN) against TNFRI, caspase-11 siRNA, or a scavenger of mitochondrial superoxide was given for antinociceptive effects. Statistical tests were done using analysis of variance (1- or 2-way), or 2-tailed t test. RESULTS: Intrathecal gp120/M induced mechanical allodynia and thermal hyperalgesia lasting for 3 weeks ( P < .001). Gp120/M increased the expression of spinal TNFRI, mitochondrial superoxide, and cleaved caspase-11. Immunohistochemistry showed that TNFRI and caspase-11 were mainly expressed in the neurons of the SCDH. Intrathecal administration of antisense oligonucleotides against TNFRI, Mito-Tempol (a scavenger of mitochondrial superoxide), or caspase-11 siRNA reduced mechanical allodynia and thermal hyperalgesia in the gp120/M neuropathic pain model. Spinal knockdown of TNFRI reduced MitoSox profile cell number in the SCDH; intrathecal Mito-T decreased spinal caspase-11 expression in gp120/M rats. In the cultured B35 neurons treated with TNFα, pretreatment with Mito-Tempol reduced active caspase-11 in the neurons. CONCLUSIONS: These results suggest that spinal TNFRI-mtROS-caspase 11 signal pathway plays a critical role in the HIV-associated neuropathic pain state, providing a novel approach to treating chronic pain in PLWH with opioids.

Effectiveness of a nurse-led coaching of self-care agency intervention for elderly patients with total laryngectomy: study protocol for a randomised controlled trial.

INTRODUCTION: Due to functional defects and structural destruction after total laryngectomy, patients experienced the poor quality of life, especially for elderly. The barriers to accessing self-care in elderly patients were considered to result from complex and multifaceted interactions of biologic and social factors. Therefore, specific efforts to improve elderly patients' quality of life are needed. The purpose of our study is to verify nurse-led coaching of elderly patient self-care approaches, which can reduce logistic burden of patients and obtain the successful functional rehabilitation ultimately. METHODS AND ANALYSIS: Elderly patients (n=60) scheduled for total laryngectomy will be randomly divided into the intervention group and the control group. Patients in the control group received routinely nursing during hospitalisation and thereby at home after discharge received conventional family care without regular supervision of nurses. Patients in the intervention group will receive a series of self-care intervention based on the transtheoretical model during hospitalisation. During home after discharge, nurses will additionally evaluate and supervise the self-care effect of patients. The two groups of patients' self-care agency, self-efficacy, quality of life and nutritional status will be recorded separately at different time points. Primary outcome is the improvement of patients' self-care agency, and secondary outcome is the improvements of patients' self-efficacy, quality of life, nutritional states and 3-month unplanned readmission rate. ETHICS AND DISSEMINATION: The Ethics Committee of Hubei Cancer Hospital has approved this protocol (KYLLBA2020006). The findings of the trial will be disseminated through peer-reviewed journals, national or international conferences. TRIAL REGISTRATION NUMBER: ChiCTR2100043731.

Water quality prediction in sea cucumber farming based on a GRU neural network optimized by an improved whale optimization algorithm.

Sea cucumber farming is an important part of China's aquaculture industry, and sea cucumbers have higher requirements for aquaculture water quality. This article proposes a sea cucumber aquaculture water quality prediction model that uses an improved whale optimization algorithm to optimize the gated recurrent unit neural network(IWOA-GRU), which provides a reference for the water quality control in the sea cucumber growth environment. This model first applies variational mode decomposition (VMD) and the wavelet threshold joint denoising method to remove mixed noise in water quality time series. Then, by optimizing the convergence factor, the convergence speed and global optimization ability of the whale optimization algorithm are strengthened. Finally, the improved whale optimization algorithm is used to construct a GRU prediction model based on optimal network weights and thresholds to predict sea cucumber farming water quality. The model was trained and tested using three water quality indices (dissolved oxygen, temperature and salinity) of sea cucumber culture waters in Shandong Peninsula, China, and compared with prediction models such as support vector regression (SVR), random forest (RF), convolutional neural network (CNN), recurrent neural network (RNN), and long short-term memory neural network (LSTM). Experimental results show that the prediction accuracy and generalization performance of this model are better than those of the other compared models.

A prediction model of aquaculture water quality based on multiscale decomposition.

In the field of intensive aquaculture, the deterioration of water quality is one of the main factors restricting the normal growth of aquatic products. Predicting water quality in real time constitutes the theoretical basis for the evaluation, planning and intelligent regulation of the aquaculture environment. Based on the design principles of decomposition, recombination and integration, this paper constructs a multiscale aquaculture water quality prediction model. First, the complete ensemble empirical mode decomposition with adaptive noise (CEEMDAN) method is used to decompose the different water quality variables at different time scales step by step to generate a series of intrinsic mode function (IMF) components with the same characteristic scale. Then, the sample entropy of each IMF component is calculated, the components with similar sample entropies are combined, and the original data are recombined into several subsequences through the above operations. In this paper, a prediction model based on a long short-term memory (LSTM) neural network is constructed to predict each recombination subsequence, and the Adam optimization algorithm is used to continuously update the weight of neural network to train and optimize the prediction performance. Finally, the predicted value of each subsequence is superimposed to predict the original water quality data. The dissolved oxygen and pH data of an aquaculture base were collected for prediction experiments, the results of which show that the proposed model has a high prediction accuracy and strong generalization performance.

Viral vector-mediated gene therapy for opioid use disorders.

Chronic exposure to opioids typically results in adverse consequences. Opioid use disorder (OUD) is a disease of the CNS with behavioral, psychological, neurobiological, and medical manifestations. OUD induces a variety of changes of neurotransmitters/neuropeptides in the nervous system. Existing pharmacotherapy, such as opioid maintenance therapy (OMT) is the mainstay for the treatment of OUD, however, current opioid replacement therapy is far from effective for the majority of patients. Pharmacological therapy for OUD has been challenging for many reasons including debilitating side-effects. Therefore, developing an effective, non-pharmacological approach would be a critical advancement in improving and expanding treatment for OUD. Viral vector mediated gene therapy provides a potential new approach for treating opioid abused patients. Gene therapy can supply targeting gene products directly linked to the mechanisms of OUD to restore neurotransmitter and/or neuropeptides imbalance, and avoid the off-target effects of systemic administration of drugs. The most commonly used viral vectors in rodent studies of treatment of opioid-used disorder are based on recombinant adenovirus (AV), adeno-associated virus (AAV), lentiviral (LV) vectors, and herpes simplex virus (HSV) vectors. In this review, we will focus on the recent progress of viral vector mediated gene therapy in OUD, especially morphine tolerance and withdrawal.

Mitochondrial biogenesis factor PGC-1α suppresses spinal morphine tolerance by reducing mitochondrial superoxide.

Opioid use disorders (OUDs) have reached an epidemic level in the United States. The opioid epidemic involves illicit opioid use, prescription opioids for analgesia, counterfeit opioids, new psychoactive substances, and diverted opioids. Opioids remain the last option for the treatment of intractable clinical pain, but chronic use of opioids are limited in part due to antinociceptive/analgesic tolerance. Peroxisome proliferator-activated receptor (PPAR)-gamma coactivator-1alpha (PGC-1α), a mitochondrial biogenesis factor can reduce toxic reactive oxygen species (ROS) that play a role in morphine tolerance (MT). Decreased PGC-1α expression has been shown to contribute to various metabolic disorders or neurodegeneration diseases through increasing ROS. We examined the relationship of PGC-1α and ROS in MT. To induce MT, adult Sprague-Dawley rats received intrathecal morphine for 7 days. Mechanical threshold was measured using the von Frey test and thermal latency was examined using the heat plate test. Expression of PGC-1α in the spinal cord dorsal horn (SCDH) was examined using RT-PCR and western blots. Mitochondrial superoxide was detected using MitoSox Red, a mitochondrial superoxide indicator. The antinociceptive effect of recombinant PGC-1α (rPGC-1α) or Mito-Tempol (a mitochondria-targeted superoxide scavenger) was determined using the von Frey test and hot plate test. Furthermore, we examined the effect of rPGC-1α on mitochondrial superoxide using cultured neurons. Our findings include that: (i) spinal MT decreased the expression of spinal PGC-1α in the SCDH neurons; (ii) rPGC-1α increased mechanical threshold and thermal latency in MT animals; (iii) Mito-Tempol reduced MT behavioral response; (iv) rPGC-1α reduced MT-induced mitochondria-targeted superoxide; and (v) cultured neuronal cells treated with TNFα increased mitochondria-targeted superoxide that can be inhibited by rPGC-1α. The present findings suggest that spinal PGC-1α reduce MT through decreasing mitochondria-targeted superoxide in the SCDH.

Viral Vector-Mediated Gene Transfer of Glutamic Acid Decarboxylase for Chronic Pain Treatment: A Literature Review.

Chronic pain is long-lasting nociceptive state, impairing the patient's quality of life. Existing analgesics are generally not effective in the treatment of chronic pain, some of which such as opioids have the risk of tolerance/dependence and overdose death with higher daily opioid doses for increasing analgesic effect. Opioid use disorders have already reached an epidemic level in the United States; therefore, nonopioid analgesic approach and/or use of nonpharmacologic interventions will be employed with increasing frequency. Viral vector-mediated gene therapy is promising in clinical trials in the nervous system diseases. Glutamic acid decarboxylase (GAD) enzyme, a key enzyme in biosynthesis of γ-aminobutyric acid (GABA), plays an important role in analgesic mechanism. In the literature review, we used PubMed and bioRxiv to search the studies, and the eligible criteria include (1) article written in English, (2) use of viral vectors expressing GAD67 or GAD65, and (3) preclinical pain models. We identified 13 eligible original research articles, in which the pain models include nerve injury, HIV-related pain, painful diabetic neuropathy, and formalin test. GAD expressed by the viral vectors from all the reports produced antinociceptive effects. Restoring GABA systems is a promising therapeutic strategy for chronic pain, which provides evidence for the clinical trial of gene therapy for pain in the near future.

Ryanodine Receptor to Mitochondrial Reactive Oxygen Species Pathway Plays an Important Role in Chronic Human Immunodeficiency Virus gp120MN-Induced Neuropathic Pain in Rats.

BACKGROUND: Chronic pain is one of the most common complaints in patients with human immunodeficiency virus (HIV)-associated sensory neuropathy. Ryanodine receptor (RyR) and mitochondrial oxidative stress are involved in neuropathic pain induced by nerve injury. Here, we investigated the role of RyR and mitochondrial superoxide in neuropathic pain induced by repeated intrathecal HIV glycoprotein 120 (gp120) injection. METHODS: Recombinant HIV glycoprotein gp120MN was intrathecally administered to induce neuropathic pain. Mechanical threshold was tested using von Frey filaments. Peripheral nerve fiber was assessed by the quantification of the intraepidermal nerve fiber density in the skin of the hindpaw. The expression of spinal RyR was examined using Western blots. Colocalization of RyR with neuronal nuclei (NeuN; neuron marker), glial fibrillary acidic protein (GFAP; astrocyte marker), or ionizing calcium-binding adaptor molecule 1 (Iba1; microglia marker) in the spinal cord was examined using immunohistochemistry. MitoSox-positive profiles (a mitochondrial-targeted fluorescent superoxide indicator) were examined. The antiallodynic effects of intrathecal administration of RyR antagonist, dantrolene (a clinical drug for malignant hyperthermia management), or selective mitochondrial superoxide scavenger, Mito-Tempol, were evaluated in the model. RESULTS: We found that repeated but not single intrathecal injection of recombinant protein gp120 induced persistent mechanical allodynia. Intraepidermal nerve fibers in repeated gp120 group was lower than that in sham at 2 weeks, and the difference in means (95% confidence interval) was 8.495 (4.79-12.20), P = .0014. Repeated gp120 increased expression of RyR, and the difference in means (95% confidence interval) was 1.50 (0.504-2.495), P = .007. Repeated gp120 also increased mitochondrial superoxide cell number in the spinal cord, and the difference in means (95% confidence interval) was 6.99 (5.99-8.00), P < .0001. Inhibition of spinal RyR or selective mitochondrial superoxide scavenger dose dependently reduced mechanical allodynia induced by repeated gp120 injection. RyR and mitochondrial superoxide were colocalized in the neuron, but not glia. Intrathecal injection of RyR inhibitor lowered mitochondrial superoxide in the spinal cord dorsal horn in the gp120 neuropathic pain model. CONCLUSIONS: These data suggest that repeated intrathecal HIV gp120 injection induced an acute to chronic pain translation in rats, and that neuronal RyR and mitochondrial superoxide in the spinal cord dorsal horn played an important role in the HIV neuropathic pain model. The current results provide evidence for a novel approach to understanding the molecular mechanisms of HIV chronic pain and treating chronic pain in patients with HIV.

Probing DNA Hybridization Equilibrium by Cationic Conjugated Polymer for Highly Selective Detection and Imaging of Single-Nucleotide Mutation.

Hybridization-based probes emerge as a promising tool for nucleic acid target detection and imaging. However, the single-nucleotide selectivity is still challenging because the specificity of hybridization reaction is typically low at room temperature. We disclose an effective and simple method for highly selective detection and in situ imaging of single-nucleotide mutation (SNM) by taking the advantages of the specific hybridization of short duplex and the signal amplifying effect of cationic conjugated polymer (CCP). Excellent discrimination of the nucleic acid strands only differing by single nucleotide was achieved enabling the sensitive detection of SNM at the abundance as low as 0.1%. Single-molecule fluorescence resonance energy transfer (smFRET) study reveals that the presence of CCP enhances the perfect matched duplex and the mismatched duplex to a different extent, which can be an explanation for the high single-nucleotide selectivity. Due to the simple design of the probe and the stable brightness of CCP, highly selective mRNA in situ imaging was achieved in fixed cells. Melanoma cell line A375 with BRAF V600E point mutation exhibits higher FRET efficiency than liver cancer cell line HegG2 that was not reported having the mutation at this point.

Spinal bromodomain-containing protein 4 contributes to neuropathic pain induced by HIV glycoprotein 120 with morphine in rats.

The symptoms of HIV-sensory neuropathy are dominated by neuropathic pain. Recent data show that repeated use of opiates enhances the chronic pain states in HIV patients. Limited attention has so far been devoted to exploring the exact pathogenesis of HIV painful disorder and opiate abuse in vivo, for which there is no effective treatment. Bromodomain-containing protein 4 (Brd4) is a member of the bromodomain and extraterminal domain protein (BET) family and functions as a chromatin 'reader' that binds acetylated lysines in histones in brain neurons to mediate the transcriptional regulation underlying learning and memory. Here, we established a neuropathic pain model of interaction of intrathecal HIV envelope glycoprotein 120 (gp120) and chronic morphine in rats. The combination of gp120 and morphine (gp120/M, for 5 days) induced persistent mechanical allodynia compared with either gp120 or morphine alone. Mechanical allodynia reached the lowest values at day 10 from gp120/M application, beginning to recover from day 21. In the model, gp120/M induced overexpression of Brd4 mRNA and protein at day 10 using RT-qPCR and western blots, respectively. Immunohistochemical studies showed that Brd4 at day 10 was expressed in the neurons of spinal cord dorsal horn. BET inhibitor I-BET762 dose-dependently increased the mechanical threshold in the gp120/M pain state. The present study provides preclinical evidence for treating HIV neuropathic pain with opioids using the BET inhibitor.

Phosphorylated CCAAT/Enhancer Binding Protein ß Contributes to Rat HIV-Related Neuropathic Pain: In Vitro and In Vivo Studies.

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/EBPß, one of the C/EBPs, is involved in the progression of HIV/AIDS, but the exact role of C/EBPß 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/EBPß (pC/EBPß) and its upstream pathway in the spinal cord dorsal horn (SCDH). HIV gp120 induced overexpression of pC/EBPß in the ipsilateral SCDH compared with contralateral SCDH. Inhibition of C/EBPß using siRNA against C/EBPß 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/EBPß 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/EBPß. Intrathecal Mito-tempol (a mitochondria-targeted O2·-scavenger) reduced mechanical allodynia and decreased pCREB and pC/EBPß. Knockdown of CREB with antisense oligodeoxynucleotide against CREB reduced mechanical allodynia and lowered pC/EBPß. These results suggested that the pathway of TNFα/TNFRI-mtO2·--pCREB triggers pC/EBPß in the HIV gp120-induced neuropathic pain state. Furthermore, we confirmed the pathway using both cultured neurons treated with recombinant TNFα in vitro 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.SIGNIFICANCE STATEMENT Painful HIV-associated sensory neuropathy is a neurological complication of HIV infection. Phosphorylated C/EBPß (pC/EBPß) influences AIDS progression, but it is still not clear about the exact role of pC/EBPß and the detailed upstream factors of pC/EBPß in HIV-related pain. In a neuropathic pain model of perineural HIV gp120 application onto the sciatic nerve, we found that pC/EBPß was triggered by TNFα/TNFRI-mtO2·--pCREB signaling pathway. The pathway was confirmed by using cultured neurons treated with recombinant TNFα in vitro, and by repeated intrathecal injection of recombinant TNFα in naive rats. The present results revealed the functional significance of TNFα/TNFRI-mtO2·--pCREB-pC/EBPß signaling in HIV neuropathic pain, and should help in the development of more specific treatments for neuropathic pain.

Gene therapy with HSV encoding p55TNFR gene for HIV neuropathic pain: an evidence-based mini-review.

While effective antiretroviral treatment makes human immunodeficiency virus (HIV)-related death decreased dramatically, neuropathic pain becomes one of the most common complications in patients with HIV/acquired immunodeficiency syndrome (AIDS). The exact mechanisms of HIV-related neuropathic pain are not well understood yet, and no effective therapy is for HIV-pain. Evidence has shown that proinflammatory factors (e.g., tumor necrosis factor alpha (TNFα)) released from glia, are critical to contributing to chronic pain. Preclinical studies have demonstrated that non-replicating herpes simplex virus (HSV)-based vector expressing human enkephalin reduces inflammatory pain, neuropathic pain, or cancer pain in animal models. In this review, we describe recent advances in the use of HSV-based gene transfer for the treatment of HIV pain, with a special focus on the use of HSV-mediated soluble TNF receptor I (neutralizing TNFα in function) in HIV neuropathic pain model.

Spinal CPEB-mtROS-CBP signaling pathway contributes to perineural HIV gp120 with ddC-related neuropathic pain in rats.

Human immunodeficiency virus (HIV) patients treated with nucleoside reverse transcriptase inhibitors (NRTIs), have been known to develop neuropathic pain. While there has been a major shift away from some neurotoxic NRTIs in current antiretroviral therapy, a large number of HIV patients alive today have previously received them, and many have developed painful peripheral neuropathy. The exact mechanisms by which HIV with NRTIs contribute to the development of neuropathic pain are not known. Previous studies suggest that cytoplasmic polyadenylation element-binding protein (CPEB), reactive oxygen species (ROS), and cAMP-response element-binding protein (CREB)-binding protein (CBP), are involved in the neuroimmunological diseases including inflammatory/neuropathic pain. In this study, we investigated the role of CPEB, mitochondrial ROS (mtROS), or CBP in neuropathic pain induced by HIV envelope protein gp120 combined with antiretroviral drug. The application of recombinant gp120 into the sciatic nerve plus systemic ddC (one of NRTIs) induced mechanical allodynia. Knockdown of CPEB or CBP using intrathecal antisense oligodeoxynucleotide (AS-ODN) reduced mechanical allodynia. Intrathecal mitochondrial superoxide scavenger mito-tempol (Mito-T) increased mechanical withdrawal threshold. Knockdown of CPEB using intrathecal AS-ODN, reduced the up-regulated mitochondrial superoxide in the spinal dorsal horn in rats with gp120 combined with ddC. Intrathecal Mito-T lowered the increased expression of CBP in the spinal dorsal horn. Immunostaining studies showed that neuronal CPEB positive cells were co-localized with MitoSox positive profiles, and that MitoSox positive profiles were co-localized with neuronal CBP. Our studies suggest that neuronal CPEB-mtROS-CBP pathway in the spinal dorsal horn, plays an important role in the gp120/ddC-induced neuropathic pain in rats.

Inhibition of Mitochondrial Fission Protein Reduced Mechanical Allodynia and Suppressed Spinal Mitochondrial Superoxide Induced by Perineural Human Immunodeficiency Virus gp120 in Rats.

BACKGROUND: Mitochondria play an important role in many cellular and physiologic functions. Mitochondria are dynamic organelles, and their fusion and fission regulate cellular signaling, development, and mitochondrial homeostasis. The most common complaint of human immunodeficiency virus (HIV)-sensory neuropathy is pain on the soles in patients with HIV, but the exact molecular mechanisms of HIV neuropathic pain are not clear. In the present study, we investigated the role of mitochondrial dynamin-related protein 1 (Drp1, a GTPase that mediates mitochondrial fission) in the perineural HIV coat glycoprotein gp120-induced neuropathic pain state. METHODS: Neuropathic pain was induced by the application of recombinant HIV-1 envelope protein gp120 into the sciatic nerve. Mechanical threshold was tested using von Frey filaments. The mechanical threshold response was assessed over time using the area under curves. Intrathecal administration of antisense oligodeoxynucleotide (ODN) against Drp1, mitochondrial division inhibitor-1 (mdivi-1), or phenyl-N-tert-butylnitrone (a reactive oxygen species scavenger) was given. The expression of spinal Drp1 was examined using western blots. The expression of mitochondrial superoxide in the spinal dorsal horn was examined using MitoSox imaging. RESULTS: Intrathecal administration of either antisense ODN against Drp1 or mdivi-1 decreased mechanical allodynia (a sensation of pain evoked by nonpainful stimuli) in the gp120 model. Intrathecal ODN or mdivi-1 did not change basic mechanical threshold in sham surgery rats. Intrathecal Drp1 antisense ODN decreased the spinal expression of increased Drp1 protein induced by peripheral gp120 application. Intrathecal phenyl-N-tert-butylnitrone reduced mechanical allodynia. Furthermore, both intrathecal Drp1 antisense ODN and mdivi-1 reversed the upregulation of mitochondrial superoxide in the spinal dorsal horn in the gp120 neuropathic pain state. CONCLUSIONS: These data suggest that mitochondrial division plays a substantial role in the HIV gp120-related neuropathic pain state through mitochondrial reactive oxygen species and provides evidence for a novel approach to treating chronic pain in patients with HIV.

[Analysis of diagnosis and treatment towards CHB patients complicated with fatty liver with complete virological response while biochemical suboptimal response].

OBJECTIVE: To analyze the reason of biochemical suboptimal response in CHB patients with complete virological response after more than 2 years standard treatment with Nucleos(t)-ide analogs (NUCs).To evaluate the efficacy and safety profiles of bicyclol tablets plus on the basis of the original treatment and lifestyle intervention. in CHB patients complicated with fatty liver. METHODS: In 40 patients with chronic hepatitis B meeting the inclusion criteria,the liver biopsy was conducted.And patients complicated with fatty liver were treated with bicyclol tablets (25 mg, t.i.d) additional consecutive 48 weeks. The changes of serum biochemistry indexes and liver fibrosis index were observed before and after treatment. RESULTS: Among 40 patients, 27 were complicated with fatty liver(69.23%), fatty degree in liver cell and liver inflammatory were closely related to the advanced fibrosis (x² =4.746, P=0.029; x² =5.072, P=0.024). The expression of HBsAg in serum and liver tissue showed no correlation with the advanced fibrosis (x² = 0.273, P=0.601; x² = 0.020, P =0.887) After bicyclol tablets treatment, serum biochemistry of patients complicated with fatty liver significantly decreased (F=58.045, P =0.000), plasma GST-PX significantly increased (t=15.109, P =0.000), plasma MDA significantly decreased (t=-10.786, P=0.000); LSM significantly decreased (t=2.255, P=0.036; t =5.376, P =0.002). CONCLUSION: For the antiviral purpose of guide treatment, CHB patients treated with Nucleos(t)-ide analogs (NUCs) with biochemical suboptimal response, other risk factors should be considered as early as possible. Bicyclol plus lifestyle intervention was effective for chronic hepatitis B combined fatty liver patients with poor biochemical responses.

Current gene therapy using viral vectors for chronic pain.

The complexity of chronic pain and the challenges of pharmacotherapy highlight the importance of development of new approaches to pain management. Gene therapy approaches may be complementary to pharmacotherapy for several advantages. Gene therapy strategies may target specific chronic pain mechanisms in a tissue-specific manner. The present collection of articles features distinct gene therapy approaches targeting specific mechanisms identified as important in the specific pain conditions. Dr. Fairbanks group describes commonly used gene therapeutics (herpes simplex viral vector (HSV) and adeno-associated viral vector (AAV)), and addresses biodistribution and potential neurotoxicity in pre-clinical models of vector delivery. Dr. Tao group addresses that downregulation of a voltage-gated potassium channel (Kv1.2) contributes to the maintenance of neuropathic pain. Alleviation of chronic pain through restoring Kv1.2 expression in sensory neurons is presented in this review. Drs Goins and Kinchington group describes a strategy to use the replication defective HSV vector to deliver two different gene products (enkephalin and TNF soluble receptor) for the treatment of post-herpetic neuralgia. Dr. Hao group addresses the observation that the pro-inflammatory cytokines are an important shared mechanism underlying both neuropathic pain and the development of opioid analgesic tolerance and withdrawal. The use of gene therapy strategies to enhance expression of the anti-pro-inflammatory cytokines is summarized. Development of multiple gene therapy strategies may have the benefit of targeting specific pathologies associated with distinct chronic pain conditions (by Guest Editors, Drs. C. Fairbanks and S. Hao).

Gene Transfer of Glutamic Acid Decarboxylase 67 by Herpes Simplex Virus Vectors Suppresses Neuropathic Pain Induced by Human Immunodeficiency Virus gp120 Combined with ddC in Rats.

BACKGROUND: Human immunodeficiency virus (HIV)-related painful sensory neuropathies primarily consist of the HIV infection-related distal sensory polyneuropathy and antiretroviral toxic neuropathies. Pharmacotherapy provides only partial relief of pain in patients with HIV/acquired immune deficiency syndrome because little is known about the exact neuropathological mechanisms for HIV-associated neuropathic pain (NP). Hypofunction of γ-aminobutyric acid (GABA) GABAergic inhibitory mechanisms has been reported after peripheral nerve injury. In this study, we tested the hypothesis that HIV gp120 combined with antiretroviral therapy reduces spinal GABAergic inhibitory tone and that restoration of GABAergic inhibitory tone will reduce HIV-related NP in a rat model. METHODS: The application of recombinant HIV-1 envelope protein gp120 into the sciatic nerve plus systemic ddC (one antiretroviral drug) induced mechanical allodynia. The hind paws of rats were inoculated with replication-defective herpes simplex virus (HSV) vectors genetically encoding gad1 gene to express glutamic acid decarboxylase 67 (GAD67), an enzyme that catalyzes the decarboxylation of glutamate to GABA. Mechanical threshold was tested using von Frey filaments before and after treatments with the vectors. The expression of GAD67 in both the lumbar spinal cord and the L4-5 dorsal root ganglia was examined using western blots. The expression of mitochondrial superoxide in the spinal dorsal horn was examined using MitoSox imaging. The immunoreactivity of spinal GABA, pCREB, and pC/EBPß was tested using immunohistochemistry. RESULTS: In the gp120 with ddC-induced neuropathic pain model, GAD67 expression mediated by the HSV vector caused an elevation of mechanical threshold that was apparent on day 3 after vector inoculation. The antiallodynic effect of the single HSV vector inoculation expressing GAD67 lasted >28 days. The area under the time-effect curves in the HSV vector expressing GAD67 was increased compared with that in the control vectors (P = 0.0005). Intrathecal GABA-A/B agonists elevated mechanical threshold in the pain model. The HSV vectors expressing GAD67 reversed the lowered GABA immunoreactivity in the spinal dorsal horn in the neuropathic rats. HSV vectors expressing GAD67 in the neuropathic rats reversed the increased signals of mitochondrial superoxide in the spinal dorsal horn. The vectors expressing GAD67 reversed the upregulated immunoreactivity expression of pCREB and pC/EBPß in the spinal dorsal horn in rats exhibiting NP. CONCLUSIONS: Based on our results, we suggest that GAD67 mediated by HSV vectors acting through the suppression of mitochondrial reactive oxygen species and transcriptional factors in the spinal cord decreases pain in the HIV-related neuropathic pain model, providing preclinical evidence for gene therapy applications in patients with HIV-related pain states.

[Clinical value of viral detection of liquid chip method in community-acquired pneumonia].

OBJECTIVE: To evaluate the clinical value of viral detection of liquid chip method in community-acquired pneumonia (CAP). METHODS: A total of 342 swabs were collected from 171 patients of community-acquired pneumonia from October 2013 to September 2014. The methods of xTAG(RVP) and Seeplex RV15 ACE were employed to detect respiratory viruses. And traditional methods of indirect immunofluorescence and specific antigen were used for comparison. All results were validated by realtime-PCR and statistically analyzed. RESULTS: Of 171 CAP patients with an average age of 49.17 years, 35.7% (61/171) were virus positive.Influenza A (FluA), influenza B (FluB), respiratory syncytial virus (RSV), human rhinovirus (HRV) and human metapneumovirus (hMPV) accounted for 90.5% of all detected viruses. The detection rates of mouth swabs and nasopharyngeal swabs were 31.6% and 33.9% respectively. Two specimen types showed no significant differences (Kappa = 0.714, P < 0.001; McNemar χ(2) = 0, P = 1.000). The positive rates of viral detection by xTAG(RVP) and Seeplex RV15 were 32.5% and 29.5% respectively. And the consistence rate of results was up to 85.4% (292/342) (Kappa = 0.66). The positive rate of traditional methods was 14.0%.However, xTAG(RVP) had a higher sensitivity (93.3%), higher consistence rate (92.4%) and negative predictive value (96.9%) compared with Seeplex RV15 and traditional methods. Also xTAG(RVP) had a high consistent rate of realtime-PCR (Kappa = 0.83). CONCLUSIONS: Liquid chip is superior to other detection methods. And it may be used routinely for viral detection of CAP.

IL-10 mediated by herpes simplex virus vector reduces neuropathic pain induced by HIV gp120 combined with ddC in rats.

BACKGROUND: HIV-associated sensory neuropathy affects over 50% of HIV patients and is a common peripheral nerve complication of HIV infection and highly active antiretroviral therapy (HAART). Evidence shows that painful HIV sensory neuropathy is influenced by neuroinflammatory events that include the proinflammatory molecules, MAP Kinase, tumor necrosis factor-α (TNFα), stromal cell-derived factor 1-α (SDF1α), and C-X-C chemokine receptor type 4 (CXCR4). However, the exact mechanisms of painful HIV sensory neuropathy are not known, which hinders our ability to develop effective treatments. In this study, we investigated whether inhibition of proinflammatory factors reduces the HIV-associated neuropathic pain state. RESULTS: Neuropathic pain was induced by peripheral HIV coat protein gp120 combined with 2',3'-dideoxycytidine (ddC, one of the nucleoside reverse transcriptase inhibitors (NRTIs)). Mechanical threshold was tested using von Frey filament fibers. Non-replicating herpes simplex virus (HSV) vectors expressing interleukin 10 (IL10) were inoculated into the hindpaws of rats. The expression of TNFα, SDF1α, and CXCR4 in the lumbar spinal cord and L4/5 dorsal root ganglia (DRG) was examined using western blots. IL-10 expression mediated by the HSV vectors resulted in a significant elevation of mechanical threshold. The anti-allodynic effect of IL-10 expression mediated by the HSV vectors lasted more than 3 weeks. The area under the effect-time curves (AUC) in mechanical threshold in rats inoculated with the HSV vectors expressing IL-10, was increased compared with the control vectors, indicating antinociceptive effect of the IL-10 vectors. The HSV vectors expressing IL-10 also concomitantly reversed the upregulation of p-p38, TNFα, SDF1α, and CXCR4 induced by gp120 in the lumbar spinal dorsal horn and/or the DRG at 2 and/or 4 weeks. CONCLUSION: The blocking of the signaling of these proinflammatory molecules is able to reduce HIV-related neuropathic pain, which provide a novel mechanism-based approach to treating HIV-associated neuropathic pain using gene therapy.