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Posterior capsule opacification (PCO) is the most common complication after cataract surgery and is likely to cause the second loss of vision. Pharmacological PCO prophylaxis has been proved to be effective, yet no clinical option is available due to the lack of a suitable mode of administration. In this work, we propose a unique concept of NIR dual-triggered drug release from black phosphorus (BP)-based implantable intraocular lens (IOL) for controlled drug release and chemo-photothermal combination therapy of PCO. Here, IOL is used as a “reservoir” of doxorubicin-loaded black phosphorus (BP-DOX), and BP is used as NIR activation agent for controlled drug release and photothermal therapy. This BP-DOX integrated IOL, namely BP-DOX@IOL, shows the characteristics of good transmittance, good mechanical property, NIR dual-triggered drug release behaviors, and excellent photothermal efficacy. In vivo studies reveal that there is no PCO occurrence in rabbits' model by using BP-DOX@IOL combined NIR irradiation, which exhibits distinct superiority on inhibiting PCO than the control group (100% PCO occurrence) 28 days post-surgery. This novel IOL drug delivery system would be a promising strategy for the future clinical application for PCO prophylaxis and treatment.

BACKGROUND: The main symptoms of chemotherapy-induced peripheral neuropathy (CIPN) include pain and numbness. Neuronal G protein–coupled receptor kinase 2 (GRK2) plays an important role in various pain models. Cisplatin treatment can induce the activation of proinflammatory microglia in spinal cord. The purpose of this study was to investigate the role of spinal neuronal GRK2 in cisplatin-induced CIPN and in the prevention of CIPN by electroacupuncture (EA). METHODS: The pain and sensory deficit behaviors of mice were examined by von Frey test and adhesive removal test. The expression of neuronal GRK2 in the spinal cord is regulated by intraspinal injection of adeno-associated virus (AAV) containing neuron-specific promoters. The protein levels of GRK2, triggering receptor expressed on myeloid cells 2 (TREM2), and DNAX-activating protein of 12 kDa (DAP12) in spinal dorsal horn were detected by Western blot, the density of intraepidermal nerve fibers (IENFs) was detected by immunofluorescence, and microglia activation were evaluated by real-time polymerase chain reaction (PCR). RESULTS: In this study, cisplatin treatment led to the decrease of GRK2 expression in the dorsal horn of spinal cord. Overexpression of neuronal GRK2 in spinal cord by intraspinal injection of an AAV vector expressing GRK2 with human synapsin (hSyn) promotor significantly inhibited the loss of IENFs and alleviated the mechanical pain and sensory deficits induced by cisplatin. Real-time PCR analysis showed that the overexpression of neuronal GRK2 significantly inhibited the messenger RNA (mRNA) upregulation of proinflammatory cytokine interleukin (IL)-1β, IL-6, inducible nitric oxide synthase (iNOS), and M1 microglia marker cluster of differentiation (CD)16 induced by cisplatin. Furthermore, the TREM2 and DAP12, which has been demonstrated to play a role in microglia activation and in the development of CIPN, were also downregulated by overexpression of neuronal GRK2 in this study. Interestingly, preventive treatment with EA completely mimics the effect of overexpression of neuronal GRK2 in the spinal cord in this mouse model of cisplatin-induced CIPN. EA increased GRK2 level in spinal dorsal horn after cisplatin treatment. Intraspinal injection of AAV vector specifically downregulated neuronal GRK2, completely reversed the regulatory effect of EA on CIPN and microglia activation. All these indicated that the neuronal GRK2 mediated microglial activation contributed to the process of CIPN. CONCLUSIONS: Neuronal GRK2 in the spinal cord contributed to the preventive effect of EA on CIPN. The neuronal GRK2 may be a potential target for CIPN intervention.

Unlike adult cardiomyocytes, neonatal cardiomyocytes can readily proliferate that contributes to a transient regenerative potential after myocardial injury in mice. We have recently reported that CD4+ regulatory T-cells promote this process; however, the role of other CD4+ T-cell subsets as well as CD8+ T-cells in postnatal heart regeneration has been less studied. Methods: by comparing the regenerating postnatal day (P) 3 and the non-regenerating P8 heart after injury, we revealed the heterogeneity of CD4+ and CD8+ T-cells in the myocardium through single cell analysis. We also specifically ablated CD4+ and CD8+ T-cells using the lytic anti-CD4 and -CD8 monoclonal antibodies, respectively, in juvenile mice at P8 after myocardial injury. Results: we observe significantly more CD4+FOXP3- conventional T-cells in the P8 heart when compared to that of the P3 heart within a week after injury. Surprisingly, such a difference is not seen in CD8+ T-cells that appear to have no function as their depletion does not reactivate heart regeneration. On the other hand, specific ablation of CD4+ T-cells contributes to mitigated cardiac fibrosis and increased cardiomyocyte proliferation after injury in juvenile mice. Single-cell transcriptomic profiling reveals a pro-fibrotic CD4+ T-cell subset in the P8 but not P3 heart. Moreover, there are likely more Th1 and Th17 cells in the P8 than P3 heart. We further demonstrate that cytokines of Th1 and Th17 cells can directly reduce the proliferation and increase the apoptosis of neonatal cardiomyocytes. Moreover, ablation of CD4+ T-cells can directly or indirectly facilitate the polarization of macrophages away from the pro-fibrotic M2-like signature in the juvenile heart. Nevertheless, ablation of CD4+ T-cells alone does not offer the same protection in the adult heart after myocardial infarction, suggesting a developmental change of immune cells including CD4+ T-cells in the regulation of age-related mammalian heart repair. Conclusions: our results demonstrate that ablation of CD4+ but not CD8+ T-cells promotes heart regeneration in juvenile mice; and CD4+ T-cells play a distinct role in the regulation of heart regeneration and repair during development.

To study the transcription factors of the spermatogenesis-related promoter mir-122-5p.SP1 and GATA4 were predicted as the possible transcription factors of the mir-122-5p promoter by bioinformatics analysis, followed by construction of the double luciferase pGL3-mir-122-5p promoter vector, pcDNA3.1 (+) -SP1 expression vector and pcDNA3.1 (+) -GATA4 expression vector, respectively. The pcDNA-SP1+pGL3-basic mixture plasmid and pcDNA-SP1+ pGL3-miR-122-5p promoter mixture plasmid, pcDNA-GATA4+pGL3-basic mixture plasmid and pcDNA-GATA4+pGL3-miR-122-5p promoter mixture plasmid were transferred into 293T cells. The enzyme activity was detected the Dual-Luciferase Reporter Assay System.The fluorescence value of the pcDNA3.1+pGL3-miR-122 promoter was 0.0362 ± 0.0004, significantly higher than that of the pcDNA3.1+pGL3-basic group (P0.05), indicating the successful construction of the mouse miR-122-5p promoter luciferase reporter plasmid. The fluorescence value was markedly higher in the pcDNA -SP1 + pGL3-miR-122-5p promoter than in the pcDNA -SP1+pGL3-basic group, suggesting that the transcription factor SP1 could promote the transcription of miR-122. There was no statistically significant difference in the fluorescence value between the pcDNA -gata4+pGL3-basic transfection and pcDNA -GATA4+pGL3-miR-122-5p promoter transfection groups, indicative of the inability of GATA4 to promote the transcription of miR-122-5p.The transcription factor SP1, rather than GATA4, can promote the transcription of miR-122-5p.

To investigate the role of occludin in tight junction (TJ) in vitro.We constructed RNA interfering lentiviral vectors and transfected them into TM4 cells. Then we detected their inhibitory effect on occuldin by RT-PCR and Western blot and analyzed the role of occuldin in TJ using an in vitro TJ cell model.The pLenti 6.3-EGFP-occludin-miR expression vector was successfully constructed. The results of RT-PCR and Western blot showed that pLenti 6.3-EGFP-occludin-miR-3 significantly inhibited the expression of occludin (P0.05), which was remarkably lower than in the blank control and the pLenti 6.3- EGFP transfection group (0.7534 ± 0.089 vs 1.000 and 1.056 ± 0.025, P0.05). The expression of occludin was markedly suppressed and the tightness of tight junctions decreased in the TM4 cells transfected with pLenti 6.3-EGFP-occludin-miR-3.The pLenti 6.3-EGFP-occludin-miR expression vector was successfully constructed, and occludin is one of the functional proteins that maintain tight junctions.

Species of Symphurus (Pleuronectiformes: Cynoglossidae) are relatively small-sized tonguefishes occurring worldwide in tropical, subtropical, and warm-temperate seas. In the Indo-West Pacific Ocean, species of Symphurus inhabiting waters shallower than 200 m are rarely reported; only five have been described, S. microrhynchus (Weber, 1913), S. holothuriae Chabanaud, 1948, S. monostigmus Munroe, 2006, S. leucochilus Lee et al. 2014, and S. longirostris Lee et al. 2016. Examination of museum and recently collected specimens yielded over 100+ Symphurus captured in relatively shallow waters off Japan, Papua New Guinea, the Philippines, Taiwan, and Vietnam. All of these specimens, except S. monostigmus (with 14 caudal-fin rays), were originally tentatively identified as S. microrhynchus because of shared similarities (small size, low meristic values, 12 caudal-fin rays, shared pigmentation traits). Detailed comparisons revealed that, although similar, specimens from allopatric locations have small differences in meristic, morphometric and pigmentation features. In previous literature, these small differences were thought to represent intraspecific variation among populations of a widespread species, S. microrhynchus. However, further study, including molecular data, has revealed that such minor differences among specimens from allopatric locations actually represent interspecific, and not population-level, variations. Where available, molecular differences among these allopatric populations, in contrast to the morphological features, were significantly different (9.0 to 26.3%), providing additional strong support for the hypothesis that more than one species is represented among fishes examined. Combined data from morphological and molecular characters, and species delimitation analysis, reveal that five, undescribed, cryptic species should be recognized: S. brachycephalus n. sp. from Vietnam, S. hongae n. sp. from Taiwan, S. leptosomus n. sp. from the Philippines, S. polylepis n. sp. from Papua New Guinea, and S. robustus n. sp. from Japan. Also, based on new information, the previous decision to place S. holothuriae Chabanaud in the synonymy of S. microrhynchus was determined to be premature. This species should be recognized as valid until additional specimens are captured and the taxonomic status of this nominal species re-evaluated. At least 10 species of Indo-West Pacific shallow-water Symphurus are now known. Eight are members of the Symphurus microrhynchus species complex with hypothesized closer relationship to each other than to the other two species of shallow-water tonguefishes. Included in this study are redescriptions of S. microrhynchus and S. holothuriae based on their holotypes, including an expanded number of morphological characters not previously used to diagnose these species; redescriptions are also provided for comparative purposes of three other shallow-water species; five new cryptic species are described; and lastly, detailed comparisons and an identification key to all 10 species of shallow-water Symphurus occurring in the Indo-West Pacific Ocean are provided. Two additional populations are also identified that likely represent other undescribed taxa belonging to the S. microrhynchus species complex. Adequate specimens are not available at this time to formally describe these nominal species. This study contributes further understanding about species diversity within Symphurus inhabiting shallow waters of the Indo-West Pacific Ocean. Several other nominal species of small-sized cynoglossid and soleid flatfishes are currently considered to have widespread distributions in the Indo-West Pacific. Many of these species also have junior synonyms available based on nominal species described from allopatric sites within their geographic ranges. How many of these presumed populations of widespread species will be resurrected from synonymy once additional specimens and their genetic information becomes available remains an interesting question for future study.

Morphine and other opiates are highly effective for treating moderate to severe pain. However, morphine-induced hyperalgesia and analgesic tolerance prevent durable efficacy in patients. Here, we investigated the underlying molecular mechanisms of this phenomenon. We found that repeated subcutaneous injections of morphine in mice increased the abundance of the cytokine interleukin-33 (IL-33) primarily in oligodendrocytes and astrocytes and that of its receptor ST2 mainly in astrocytes. Pharmacological inhibition or knockdown of IL-33 or ST2 in the spinal cord attenuated morphine-induced hyperalgesia and analgesic tolerance in mice, as did global knockout of either

The chronic neuropathic pain-associated psychiatric disorders have seriously disturbed the quality of patients' life, such as depression and anxiety. Neuroinflammation in the hippocampus plays an important role in the neuropathic pain-associated depressive and anxiety disorders, but the underlying mechanism has not been thoroughly elucidated to date. The Nod-like receptor protein (NLRP)-1 inflammasome, which controls the production of pro-inflammatory cytokines, was broadly involved in the neuroinflammation-related diseases. In the present study, we show that the NLRP1 inflammasome is significantly activated in the hippocampus of rats subjected to the chronic constriction injury (CCI)-induced neuropathic pain. Inhibiting the product of NLRP1 inflammasome not only attenuated the depression-like behaviors but also suppressed the production of mature IL-1β in the hippocampus of CCI rats. The double-stranded RNA-dependent protein kinase (PKR, also known as EIF2AK2) has been recently shown to be a pivotal regulator for the activation of inflammasome. In the rats subjected to CCI neuropathic pain, the PKR was simultaneously activated in hippocampus. Functional inhibition of PKR suppressed the NLRP1 inflammasome activation and effectively attenuated the CCI-induced depression-like behaviors. These results indicate that the hippocampal PKR/NLRP1 inflammasome pathway play an important role in the development of the depressive behaviors after chronic neuropathic pain. Thus, interrupting this pathway might provide a novel therapeutic strategy for neuropathic pain-associated depressive disorders.

Matrix attachment regions (MARs) can enhance transgene expression levels and maintain stability. However, the consensus sequence from MARs and its functional analysis remains to be examined. Here, we assessed a possible consensus sequence from MARs and assessed its activity in stably transfected Chinese hamster ovary (CHO) cells. First, we analyzed the effects of 10 MARs on transfected CHO cells and then analyzed the consensus motifs from these MARs using a bioinformatics method. The consensus sequence was synthesized and cloned upstream or downstream of the eukaryotic vector. The constructs were transfected into CHO cells and the expression levels and stability of enhanced green fluorescent protein were detected by flow cytometry. The results indicated that eight of the ten MARs increased transgene expression in transfected CHO cells. Three consensus motifs were found after bioinformatics analyses. The consensus sequence tandemly enhanced transgene expression when it was inserted into the eukaryotic expression vector; the effect of the addition upstream was stronger than that downstream. Thus, we found a MAR consensus sequence that may regulate the MAR-mediated increase in transgene expression.

Background: Chemotherapy-induced peripheral neuropathy (CIPN) is a progressive, enduring, and often irreversible adverse effect of many antineoplastic agents, among which sensory abnormities are common and the most suffering issues. The pathogenesis of CIPN has not been completely understood, and strategies for CIPN prevention and treatment are still open problems for medicine. Objectives: The objective of this paper is to review the mechanism-based therapies against sensory abnormities in CIPN. Methods: This is a literature review to describe the uncovered mechanisms underlying CIPN and to provide a summary of mechanism-based therapies for CIPN based on the evidence from both animal and clinical studies. Results: An abundance of compounds has been developed to prevent or treat CIPN by blocking ion channels, targeting inflammatory cytokines and combating oxidative stress. Agents such as glutathione, mangafodipir and duloxetine are expected to be effective for CIPN intervention, while Ca/Mg infusion and venlafaxine, tricyclic antidepressants, and gabapentin display limited efficacy for preventing and alleviating CIPN. And the utilization of erythropoietin, menthol and amifostine needs to be cautious regarding to their side effects. Conclusions: Multiple drugs have been used and studied for decades, their effect against CIPN are still controversial according to different antineoplastic agents due to the diverse manifestations among different antineoplastic agents and complex drug-drug interactions. In addition, novel therapies or drugs that have proven to be effective in animals require further investigation, and it will take time to confirm their efficacy and safety.

Trigeminal neuralgia is a debilitating condition, and the pain easily spreads to other parts of the face. Here, we established a mouse model of partial transection of the infraorbital nerve (pT-ION) and found that the Connexin 36 (Cx36) inhibitor mefloquine caused greater alleviation of pT-ION-induced cold allodynia compared to the reduction of mechanical allodynia. Mefloquine reversed the pT-ION-induced upregulation of Cx36, glutamate receptor ionotropic kainate 2 (GluK2), transient receptor potential ankyrin 1 (TRPA1), and phosphorylated extracellular signal regulated kinase (p-ERK) in the trigeminal ganglion. Cold allodynia but not mechanical allodynia induced by pT-ION or by virus-mediated overexpression of Cx36 in the trigeminal ganglion was reversed by the GluK2 antagonist NS102, and knocking down Cx36 expression in Nav1.8-expressing nociceptors by injecting virus into the orofacial skin area of Nav1.8-Cre mice attenuated cold allodynia but not mechanical allodynia. In conclusion, we show that Cx36 contributes greatly to the development of orofacial pain hypersensitivity through GluK2, TRPA1, and p-ERK signaling. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12264-020-00594-4) contains supplementary material, which is available to authorized users.

Trigeminal neuralgia (TN) is debilitating and is usually accompanied by mood disorders. The lateral habenula (LHb) is considered to be involved in the modulation of pain and mood disorders, and the present study aimed to determine if and how the LHb participates in the development of pain and anxiety in TN. To address this issue, a mouse model of partial transection of the infraorbital nerve (pT-ION) was established. pT-ION induced stable and long-lasting primary and secondary orofacial allodynia and anxiety-like behaviors that correlated with the increased excitability of LHb neurons. Adeno-associated virus (AAV)-mediated expression of hM4D(Gi) in glutamatergic neurons of the unilateral LHb followed by clozapine-N-oxide application relieved pT-ION-induced anxiety-like behaviors but not allodynia. Immunofluorescence validated the successful infection of AAV in the LHb, and microarray analysis showed changes in gene expression in the LHb of mice showing allodynia and anxiety-like behaviors after pT-ION. Among these differentially expressed genes was Tacr3, the downregulation of which was validated by RT-qPCR. Rescuing the downregulation of Tacr3 by AAV-mediated Tacr3 overexpression in the unilateral LHb significantly reversed pT-ION-induced anxiety-like behaviors but not allodynia. Whole-cell patch clamp recording showed that Tacr3 overexpression suppressed nerve injury-induced hyperexcitation of LHb neurons, and western blotting showed that the pT-ION-induced upregulation of p-CaMKII was reversed by AAV-mediated Tacr3 overexpression or chemicogenetic inhibition of glutamatergic neurons in the LHb. Moreover, not only anxiety-like behaviors, but also allodynia after pT-ION were significantly alleviated by chemicogenetic inhibition of bilateral LHb neurons or by bilateral Tacr3 overexpression in the LHb. In conclusion, Tacr3 in the LHb plays a protective role in treating trigeminal nerve injury-induced allodynia and anxiety-like behaviors by suppressing the hyperexcitability of LHb neurons. These findings provide a rationale for suppressing unilateral or bilateral LHb activity by targeting Tacr3 in treating the anxiety and pain associated with TN.

Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New Background Cancer pain is a pervasive clinical symptom impairing life quality. Vascular endothelial growth factor A has been well studied in tumor angiogenesis and is recognized as a therapeutic target for anti-cancer treatment. This study tested the hypothesis that vascular endothelial growth factor A and vascular endothelial growth factor receptor 2 contribute to bone cancer pain regulation associated with spinal central sensitization. Methods This study was performed on female rats using a metastatic breast cancer bone pain model. Nociceptive behaviors were evaluated by mechanical allodynia, thermal hyperalgesia, spontaneous pain, and CatWalk gait analysis. Expression levels were measured by real-time quantitative polymerase chain reaction, western blot, and immunofluorescence analysis. Excitatory synaptic transmission was detected by whole-cell patch-clamp recordings. The primary outcome was the effect of pharmacologic intervention of spinal vascular endothelial growth factor A/vascular endothelial growth factor receptor 2–signaling on bone cancer pain behaviors. Results The mRNA and protein expression of vascular endothelial growth factor A and vascular endothelial growth factor receptor 2 were upregulated in tumor-bearing rats. Spinal blocking vascular endothelial growth factor A or vascular endothelial growth factor receptor 2 significantly attenuated tumor-induced mechanical allodynia (mean ± SD: vascular endothelial growth factor A, 7.6 ± 2.6 g vs. 5.3 ± 3.3 g; vascular endothelial growth factor receptor 2, 7.8 ± 3.0 g vs. 5.2 ± 3.4 g; n = 6; P < 0.0001) and thermal hyperalgesia (mean ± SD: vascular endothelial growth factor A, 9.0 ± 2.4 s vs. 7.4 ± 2.7 s; vascular endothelial growth factor receptor 2, 9.3 ± 2.5 s vs. 7.5 ± 3.1 s; n = 6; P < 0.0001), as well as spontaneous pain and abnormal gaits. Exogenous vascular endothelial growth factor A enhanced excitatory synaptic transmission in a vascular endothelial growth factor receptor 2–dependent manner, and spinal injection of exogenous vascular endothelial growth factor A was sufficient to cause pain hypersensitivity via vascular endothelial growth factor receptor 2–mediated activation of protein kinase C and Src family kinase in naïve rats. Moreover, spinal blocking vascular endothelial growth factor A/vascular endothelial growth factor receptor 2 pathways suppressed protein kinase C-mediated N-methyl-d-aspartate receptor activation and Src family kinase-mediated proinflammatory cytokine production. Conclusions Vascular endothelial growth factor A/vascular endothelial growth factor receptor 2 contributes to central sensitization and bone cancer pain via activation of neuronal protein kinase C and microglial Src family kinase pathways in the spinal cord.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common adverse side effect of many antineoplastic agents. Patients treated with chemotherapy often report pain and paresthesias in a “glove-and-stocking” distribution. Diverse mechanisms contribute to the development and maintenance of CIPN. However, the role of spinal microglia in CIPN is not completely understood. In this study, cisplatin-treated mice displayed persistent mechanical allodynia, sensory deficits and decreased density of intraepidermal nerve fibers (IENFs). In the spinal cord, activation of microglia, but not astrocyte, was persistently observed until week five after the first cisplatin injection. Additionally, mRNA levels of inflammation related molecules including IL-1β, IL-6, tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS) and CD16, were increased after cisplatin treatment. Intraperitoneal (i.p.) or intrathecal (i.t.) injection with minocycline both alleviated cisplatin-induced mechanical allodynia and sensory deficits, and prevented IENFs loss. Furthermore, cisplatin enhanced triggering receptor expressed on myeloid cells 2 (TREM2) /DNAX-activating protein of 12 kDa (DAP12) signaling in the spinal cord microglia. The blockage of TREM2 by i.t. injecting anti-TREM2 neutralizing antibody significantly attenuated cisplatin-induced mechanical allodynia, sensory deficits and IENFs loss. Meanwhile, anti-TREM2 neutralizing antibody prominently suppressed the spinal IL-6, TNF-α, iNOS and CD16 mRNA level, but it dramatically up-regulated the anti-inflammatory cytokines IL-4 and IL-10. The data demonstrated that cisplatin triggered persistent activation of spinal cord microglia through strengthening TREM2/DAP12 signaling, which further resulted in CIPN. Functional blockage of TREM2 or inhibition of microglia both benefited for cisplatin-induced peripheral neuropathy. Microglial TREM2/DAP12 may serve as a potential target for CIPN intervention.

Orofacial pain is characterized by its easy spread to adjacent areas, thus presenting with primary hyperalgesia (hypersensitivity at the site of injury) and secondary hyperalgesia (extraterritorial hypersensitivity outside the injured zone). However, the mechanisms behind the secondary hyperalgesia are poorly understood. In the present study, we used a mouse model of partial transection of the infraorbital nerve (pT-ION) to study whether calcium channel subunit α2δ1 (Cavα2δ1) and its downstream signaling contributes to the development of secondary hyperalgesia in the orofacial area. pT-ION caused primary (V2 skin) and secondary (V3 skin) hyperalgesia, which was reversed by the Cavα2δ1 antagonist gabapentin and by the expression of Cavα2δ1-targeting interfering RNA in trigeminal ganglion (TG)-V3 neurons. pT-ION induced increased expression of PKC and TRPA1, which was reversed by Cavα2δ1-targeting interfering RNA, and PKC inhibition reversed the upregulation of TRPA1 and gap junction (GJ) proteins induced by pT-ION. Cavα2δ1 overexpression in TG-V2 neurons induced the upregulation of PKC, TRPA1, and the GJ proteins in the TG and trigeminal subnucleus caudalis and induced hypersensitivity in the V3 skin area, which was reversed by TRPA1, GJ, or PKC blockade. Thus, we conclude that Cavα2δ1 contributes to the development of secondary hyperalgesia through its downstream PKC-TRPA1/GJ signaling pathways. PERSPECTIVE: This study demonstrates that the activation of Cavα2δ1 and the downstream PKC-TRPA1/GJ signaling pathway contributes greatly to trigeminal nerve injury-induced secondary mechanical and cold hyperalgesia. This suggests that inhibitors of Cavα2δ1, TRPA1, or GJs might be effective treatments for nerve injury-induced spreading of orofacial pain.

Human amniotic membrane mesenchymal stem cells (hAMSCs) are considered ideal candidate stem cells for cell-based therapy. In this study, we assessed whether hAMSCs transplantation promotes neurological functional recovery in rats after traumatic spinal cord injury (SCI). In addition, the potential mechanisms underlying the possible benefits of this therapy were investigated. Female Sprague–Dawley rats were subjected to SCI using a weight drop device and then hAMSCs, or phosphate-buffered saline (PBS) were immediately injected into the contused dorsal spinal cord at 2 mm rostral and 2 mm caudal to the injury site. Our results indicated that transplanted hAMSCs migrated in the host spinal cord without differentiating into neuronal or glial cells. Compared with the control group, hAMSCs transplantation significantly decreased the numbers of ED1+ macrophages/microglia and caspase-3+ cells. In addition, hAMSCs transplantation significantly increased the levels of brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) in the injured spinal cord, and promoted both angiogenesis and axonal regeneration. These effects were associated with significantly improved neurobehavioral recovery in the hAMSCs transplantation group. These results show that transplantation of hAMSCs provides neuroprotective effects in rats after SCI, and could be candidate stem cells for the treatment of SCI.

Background Emerging evidence indicates that nerve damage–initiated neuroinflammation and immune responses, which are evidenced by the up-regulation of proinflammatory cytokines, contribute to the development of neuropathic pain. This study investigated the role of spinal interleukin (IL)-33 and its receptor ST2 in spared nerve injury (SNI)-induced neuropathic pain. Methods The von Frey test and acetone test were performed to evaluate neuropathic pain behaviors (n = 8 to 12), and Western blot (n = 4 to 6), immunohistochemistry, real-time polymerase chain reaction (n = 5), and Bio-Plex (n = 5) assays were performed to understand the molecular mechanisms. Results Intrathecal administration of ST2-neutralizing antibody or ST2 gene knockout (ST2−/−) significantly attenuated the SNI-induced mechanical and cold allodynia. On the 7th day after SNI, the expression of spinal IL-33 and ST2 was increased by 255.8 ± 27.3% and 266.4 ± 83.5% (mean ± SD), respectively. Mechanistic studies showed that the increased expression of the spinal N-methyl-d-aspartate (NMDA) receptor subunit 1 after SNI was reduced by ST2 antibody administration or ST2−/−. The induction of nociceptive behaviors in naive mice due to recombinant IL-33 was reversed by the noncompetitive NMDA antagonist MK-801. ST2 antibody administration or ST2−/− markedly inhibited the increased activation of the astroglial janus kinase 2 (JAK2)–signal transducer and activator of transcription 3 (STAT3) cascade and the neuronal calcium–calmodulin-dependent kinase II (CaMKII)–cyclic adenosine monophosphate response element–binding protein (CREB) cascade after SNI. Moreover, intrathecal pretreatment with the CaMKII inhibitor KN-93 or the JAK2–STAT3 cascade inhibitor AG490 attenuated recombinant IL-33-induced nociceptive behaviors and NMDA subunit 1 up-regulation in naive mice. Conclusion Spinal IL-33/ST2 signaling contributes to neuropathic pain by activating the astroglial JAK2–STAT3 cascade and the neuronal CaMKII–CREB cascade.

Left-eyed flounders of the genus Chascanopsetta Alcock 1894 (Bothidae) occur in the Indian, Pacific, and Atlantic oceans at depths ranging from 120 to 1500 meters. They possess some unique features in bothid fishes including a strongly compressed and elongated body and a tremendously large mouth. Currently, nine species of Chascanopsetta are recognized, and three of them (C. micrognatha Amaoka & Yamamoto 1984, C. lugubris Alcock 1894 and C. prognatha Norman 1939) are distributed in the West Pacific. We collected 25 specimens of Chascanopsetta during 11 biodiversity expeditions carried out mainly in the West Pacific. Among them, eight specimens taken off Papua New Guinea present morphological features that differ from those of the three nominal species known in the West Pacific. In this study, we examined these eight specimens of unknown affinity and compared their morphology to that of specimens of other congeneric species. Results of these comparisons showed that these specimens represent an undescribed species of Chascanopsetta, named herein, C. novaeguineae sp. nov.. The new species resembles C. elski Foroshchuk 1991, which is known only from the Saya de Malha Bank in the western Indian Ocean, in having a high number of gill rakers (> 13). However, the combination of the following characters further distinguishes C. novaeguineae sp. nov. from C. elski: longer jaws, narrower interorbital width, and number of pseudobranches (21–25 vs. 26–27). The DNA sequences from the mitochondrial cytochrome oxidase subunit I (COI) gene from C. novaeguineae sp. nov. and other species were obtained and compared to confirm its taxonomic status and to infer its tentative phylogenetic position within the Chascanopsetta.

Interleukin-33 (IL-33) and its receptor ST2 contribute to spinal glial activation and chronic pain. A recent study showed that peripheral IL-33 plays a pivotal role in the pathogenesis of chronic itch induced by poison ivy. However, how IL-33/ST2 signaling in the spinal cord potentially mediates chronic itch remains elusive. Here, we determined that St2-/- substantially reduced scratching behaviors in 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD) as well as acetone and diethylether followed by water-induced dry skin in mice. Intrathecal administration of the neutralizing anti-ST2 or anti-IL-33 antibody remarkably decreased the scratching response in DNFB-induced ACD mice. Expression of spinal IL-33 and ST2 significantly increased in ACD mice, as evidenced by increased mRNA and protein levels. Immunofluorescence and in situ hybridization demonstrated that increased expression of spinal IL-33 was predominant in oligodendrocytes and astrocytes, whereas ST2 was mainly expressed in astrocytes. Further studies showed that in ACD mice, the activation of astrocytes and increased phosphorylation of signal transducer and activator of transcription 3 (STAT3) were markedly attenuated by St2-/- . Intrathecal injection of Janus Kinase 2 Inhibitor AG490 significantly alleviated scratching behaviors in ACD mice. rIL-33 pretreatment exacerbated gastrin-releasing peptide (GRP)-evoked scratching behaviors. This increased gastrin-releasing peptide receptor (GRPR) expression was abolished by St2-/- . Tnf-α upregulation was suppressed by St2-/- . Our results indicate that the spinal IL-33/ST2 signaling pathway contributes to chronic itch via astrocytic JAK2-STAT3 cascade activation, promoting TNF-α release to regulate the GRP/GRPR signaling-related itch response. Thus, these findings provide a potential therapeutic option for treating chronic pruritus.

Acupoint catgut embedding (ACE) is a widely used traditional Chinese medicine method to manage various diseases, including chronic inflammatory pain. We sought to assess the possible analgesic effects of ACE in comparison with electroacupuncture (EA) and to study the analgesic mechanisms of ACE in a rat model of inflammatory pain induced by injection of complete Freund's adjuvant (CFA) into the hind paw of rats. The von Frey, radiant heat, and gait analysis tests were performed to evaluate the analgesic effects of ACE and EA, and Western blot and immunohistochemistry assays were carried out to determine the molecular mechanisms of ACE. ACE treatments were administered every 4 days or every week with different acupoints (ipsilateral, contralateral, or bilateral ST36 and GB30 acupoints). The most effective ACE strategy for attenuating the nocifensive response induced by CFA injection was performing ACE once a week at ipsilateral ST36 in combination with GB30. EA treatment every other day at ipsilateral ST36 and GB30 showed comparable analgesic effects. ACE inhibited the increased activation of the GluN1 subunit of the N-methyl-d-aspartate receptor and the subsequent Ca2+-dependent signals (CaMKII, ERK, and CREB) that take place in response to CFA. The effects of ACE were similar to intrathecal injection of vilazodone (a serotonin 1A receptor [5-HT1AR] agonist) and were blocked by WAY-100635 (a 5-HT1AR antagonist). In summary, we show that ACE attenuates CFA-induced inflammatory pain in rats by activating spinal 5-HT1AR and by inhibiting the phosphorylation of GluN1, thus, inhibiting the activation of Ca2+-dependent signaling cascades. PERSPECTIVE: This article presents the novel evidence concerning the spinal 5-HT1AR activation-related molecular signaling of ACE analgesia in a rat model of CFA-induced inflammatory pain. This work may help clinicians to verify the effectiveness of ACE analgesia and to better understand the underlying mechanism.

Accumulating evidence suggests that obesity is associated with chronic pain. However, whether obesity is associated with acute inflammatory pain is unknown. Using a well-established obese mouse model induced by a high-fat diet, we found that: (1) the acute thermal pain sensory threshold did not change in obese mice; (2) the model obese mice had fewer nociceptive responses in formalin-induced inflammatory pain tests; restoring the obese mice to a chow diet for three weeks partly recovered their pain sensation; (3) leptin injection induced significant phosphorylation of STAT3 in control mice but not in obese mice, indicating the dysmodulation of topical leptin–leptin receptor signaling in these mice; and (4) leptin–leptin receptor signaling-deficient mice (ob/ob and db/db) or leptin–leptin receptor pathway blockade with a leptin receptor antagonist and the JAK2 inhibitor AG 490 in wild-type mice reduced their nociceptive responses in formalin tests. These results indicate that leptin plays a role in nociception induced by acute inflammation and that interference in the leptin–leptin receptor pathway could be a peripheral target against acute inflammatory pain.

Lamotrigine (LTG), a broad-spectrum anti-epileptic drug widely used in treatment for seizures, shows potential efficacy in Alzheimer's disease (AD) therapy. Chronic LTG treatment rescues the suppressed long-term potentiation, loss of spines and cognitive deficits in AβPP/PS1 mice, known to overexpress a chimeric mouse/human mutant amyloid-β protein precursor (AβPP) and a mutant human presenilin 1 (PS1). These changes are accompanied by reduction of amyloid-β (Aβ) plaques density and of levels of β-C-terminal fragment of AβPP (β-CTF), a fragment of AβPP cleaved by β-secretase. These results suggest LTG treatment reduces Aβ production, possibly through modulation of cleavage of AβPP by β-secretase. However, the underlying mechanisms still remain unclear. In this study, decreased protein levels, but not mRNA levels of β-site AβPP-cleaving enzyme 1 (BACE1), were observed in cultured HEK293 cells and the brains of AβPP/PS1 transgenic mice upon LTG treatment. Moreover, LTG treatment suppressed mammalian target of rapamycin (mTOR) signaling, while enhancing activation of cAMP response element binding protein (CREB), two signaling pathways essential for autophagy induction. LTG treatment increased the numbers of LC3-GFP + puncta and LC3-II levels in HEK293 cells, indicating an induction of autophagy. The downregulation of BACE1 by LTG treatment was prevented by the autophagy inhibitor 3-Methyladenine. Therefore, this study shows that LTG treatment reduces the protein levels of BACE1 through activation of autophagy, possibly via inhibition of mTOR signaling and activation of CREB.

Transplantation of human amniotic mesenchymal stem cells (hAM-MSCs) seems to be a promising strategy for the treatment of neurodegenerative disorders, including Alzheimer's disease (AD). However, the clinical therapeutic effects of hAM-MSCs and their mechanisms of action in AD remain to be determined. Here, we used amyloid precursor protein (APP) and presenilin1 (PS1) double-transgenic mice to evaluate the effects of hAM-MSC transplantation on AD-related neuropathology and cognitive dysfunction. We found that hAM-MSC transplantation into the hippocampus dramatically reduced amyloid-β peptide (Aβ) deposition and rescued spatial learning and memory deficits in APP/PS1 mice. Interestingly, these effects were associated with increasing in Aβ-degrading factors, elevations in activated microglia, and the modulation of neuroinflammation. Furthermore, enhanced hippocampal neurogenesis in the subgranular zone (SGZ) of the dentate gyrus (DG) and enhanced synaptic plasticity following hAM-MSC treatment could be another important factor in reversing the cognitive decline in APP/PS1 mice. Instead, the mechanism underlying the improved cognition apparently involves a robust increase in hippocampal synaptic density and neurogenesis that is mediated by brain-derived neurotrophic factor (BDNF). In conclusion, our data suggest that hAM-MSCs may be a new and effective therapy for the treatment of AD.

Cancer pain, particularly bone cancer pain, affects the quality of life of cancer patients, and current treatments are limited. Interleukin (IL)-33, a new member of the IL-1 super family, has been reported to be involved in the modulation of inflammatory pain. However, studies focused on its role in the modulation of cancer pain have been rare. The present study was designed to investigate whether spinal IL-33/ST2 signaling was involved in bone cancer-induced pain in mice. Bone cancer was induced via intra-femoral inoculation of 4T1 mammary carcinoma cells. The mice inoculated with carcinoma cells showed mechanical allodynia, heat hyperalgesia and a reduction in limb use, whereas phosphate-buffered saline or heat-killed cells-injected mice showed no significant difference compared to non-treated mice. The pain hypersensitive behaviors worsened over time and with bone destruction. Both the mRNA and the protein levels of IL-33 and relative cytokines (IL-1β, IL-6, TNF-a) were significantly increased in the spinal cord after the inoculation of carcinoma cells. Intrathecal administration of ST2 antibody to block IL-33/ST2 signaling alleviated pain behaviors in a dose-dependent manner in bone cancer pain mice compared with vehicle-injected mice. Moreover, the ST2(-/-) mice showed a significant amelioration of limb use and heat hyperalgesia compared to wild-type mice. Meanwhile, concentrations of spinal IL-1β, IL-6 and TNF-a in the cancer-bearing ST2(-/-) mice had no significant changes. These data further suggested that IL-33/ST2 signaling played a vital role in cancer pain. Our results provided evidence that IL-33 and its receptor ST2 may be a potential therapeutic target for the treatment of pain in bone cancer patients.

Curcumin has been shown to possess strong anti-inflammatory activity in many diseases. It has been demonstrated that the janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) cascade and the NAcht leucine-rich-repeat protein 1 (NALP1) inflammasome are important for the synthesis of Pro-Interleukin (IL)-1β and the processing of the inactive protein to its mature form, which plays an active role in the pathogenesis of neuropathic pain. The present study showed that repeated intraperitoneal injection of curcumin ameliorated SNI-induced mechanical and cold allodynia in a dose-dependent manner and inhibited the elevation of spinal mature IL-1β protein levels. Additionally, repeated curcumin treatment significantly inhibited the aggregation of the NALP1 inflammasome and the activation of the JAK2-STAT3 cascade in spinal astrocytes. Furthermore, the genetic down-regulation of NALP1 inflammasome activation by NALP1 siRNA and the pharmacological inhibition of the JAK2-STAT3 cascade by AG490 markedly inhibited IL-1β maturation and Pro-IL-1β synthesis, respectively and reduced SNI-induced pain hypersensitivity. Our results suggest that curcumin attenuated neuropathic pain and down-regulated the production of spinal mature IL-1β by inhibiting the aggregation of NALP1 inflammasome and the activation of the JAK2-STAT3 cascade in astrocytes.

Species identification based on the DNA sequence of a fragment of the cytochrome c oxidase subunit I gene in the mitochondrial genome, DNA barcoding, is widely applied to assist in sustainable exploitation of fish resources and the protection of fish biodiversity. The aim of this study was to establish a reliable barcoding reference database of the native ray-finned fishes in Taiwan. A total of 2993 individuals, belonging to 1245 species within 637 genera, 184 families and 29 orders of ray-finned fishes and representing approximately 40% of the recorded ray-finned fishes in Taiwan, were PCR amplified at the barcode region and bidirectionally sequenced. The mean length of the 2993 barcodes is 549 bp. Mean congeneric K2P distance (15.24%) is approximately 10-fold higher than the mean conspecific one (1.51%), but approximately 1.4-fold less than the mean genetic distance between families (20.80%). The Barcode Index Number (BIN) discordance report shows that 2993 specimens represent 1275 BINs and, among them, 86 BINs are singletons, 570 BINs are taxonomically concordant, and the other 619 BINs are taxonomically discordant. Barcode gap analysis also revealed that more than 90% of the collected fishes in this study can be discriminated by DNA barcoding. Overall, the barcoding reference database established by this study reveals the need for taxonomic revisions and voucher specimen rechecks, in addition to assisting in the management of Taiwan's fish resources and diversity.

Cancer pain, especially cancer-induced bone pain, affects the quality of life of cancer patients, and current treatments for this pain are limited. The present study demonstrates that spinal extracellular signal-regulated kinase (ERK) activation in glial cells plays a crucial role in cancer-induced bone pain. From day 4 to day 21 after the intra-tibia inoculation with Walker 256 mammary gland carcinoma cells, significant mechanical allodynia was observed as indicated by the decrease of mechanical withdrawal thresholds in the von Frey hair test. Intra-tibia inoculation with carcinoma cells induced a vast and persistent (>21 D) activation of ERK in the bilateral L2-L3 and L4-L5 spinal dorsal horn. The increased pERK1/2-immunoreactivity was observed in both Iba-1-expressing microglia and GFAP-expressing astrocytes but not in NeuN-expressing neurons. A single intrathecal injection of the selective MEK (ERK kinase) inhibitors PD98059 (10 μg) on day 12 and U0126 (1.25 and 3 μg) on day 14, attenuated the bilateral mechanical allodynia in the von Frey hair test. Altogether, our results suggest that ERK activation in spinal microglia and astrocytes is correlated with the onset of allodynia and is important for allodynia maintenance in the cancer pain model. This study indicated that inhibition of the ERK pathway may provide a new therapy for cancer-induced bone pain.

In prior studies, Eph/ephrin system was demonstrated to be involved in inflammatory and neuropathic pain modulation. The present study was to investigate whether the spinal Eph/ephrin signaling was involved in modulation of spinal inflammatory cytokines in bone cancer pain (BCP) of rats. BCP was induced by intra-tibial inoculation of Walker 256 mammary gland carcinoma cells. The expressions of EphB1/ephrinB1 in spinal cord (SC) and dorsal root ganglia (DRG) were determined. At 16 days post inoculation, the pain relieving effect and the mRNA levels of inflammatory cytokines were detected after intrathecal administration of EphB1-Fc (blocker of EphB1 receptor, 10μg). The results showed that the EphB1/ephrinB1 expression was significantly increased in SC, but ephrinB1 was decreased in DRG after Walker 256 inoculation. The mechanical allodynia induced by bone cancer was significantly alleviated by intrathecal administration of EphB1-Fc. Furthermore, the RT-PCR analysis showed that the mRNA levels of IL-1β, IL-6 and TNF-α were significantly increased at 16 days post Walker 256 inoculation and were significantly suppressed by intrathecal administration of EphB1-Fc in SC. We concluded that Eph/ephrin might be involved in the maintenance of mechanical allodynia, via modulating the expression of spinal inflammatory cytokines, in the present rat model of BCP. This study suggested that Eph/ephrin signaling would be a potential target for the treatment of BCP.

Objective To establish a proper experimental model of bone cancer pain in rat for acupuncture research, and observe the pain-relieving effect of electroacupuncture (EA) and/or Celebrex on bone cancer pain in rats. Methods The rat model of bone cancer pain was established by percutaneous direct puncture technique and inoculating the rat mammary gland carcinoma cells Walker 256 into tibial medullary cavity directly, and evaluated by detecting the bone tumor growth and mechanical allodynia. The effects of daily EA treatment and/or Celebrex treatment on the rat mechanical allodynia after intratibial Walker 256 inoculation was observed in this study. Results Significant mechanical allodynia in ipsilateral hind paw and tumor growth in proximal end of tibial bone of rats in the untreated group were observed after intratibial Walker 256 inoculation. The mechanical allodynia thresholds in rats that received EA or 5 mg/(kg.d) Celebrex treatment showed no significant difference as compared with that of rats in the untreated group. However, the mechanical allodynia thresholds of rats in 10 mg/(kg.d) Celebrex group showed significant increase after 22- and 26-day treatment as compared with that in the methyl cellulose (MC) group. There was significant difference between rats with EA combined with 5 mg/(kg.d) Celebrex treatment and rats in the untreated group after 10-, 18- and 23-day treatment. Conclusion EA and 5 mg/(kg.d) Celebrex have synergistic effect on pain relieving and their combined use may enhance the analgesic effect on bone cancer pain.

Adult hippocampal neurogenesis has been demonstrated in several species and is regulated by both environmental and pharmacological stimuli. Repeated exposure to stress is known to induce the reduction of neurogenesis in the dentate gyrus (DG) of hippocampus. The present study aimed at determining whether the clinically effective antidepressant clomipramine may influence hippocampal proliferation and neurogenesis in adult rats subjected to the chronic unpredictable stress (CUS) procedure, a model of depression with predictive validity. Repeated administration of clomipramine (5 mg kg(-1), intraperitoneal) for 3 weeks, starting 2 weeks after the beginning of the stress procedure, significantly reversed the reduction of behavior measured by open-field test and forced swimming test. Moreover, rats subjected to stress exhibited a 49.9% reduction of cell proliferation at the end of a 5-week stress period, an effect which was suppressed by clomipramine treatment. These results demonstrated that exposure to CUS, which results in a state of behavioral depression, decreases hippocampal cell proliferation and that these effects can be counteracted by chronic clomipramine treatment.

The present study was designed to investigate whether electroacupuncture (EA) was beneficial to extenuate the behavioral deficit in a rat model of depression induced by chronic unpredictable stress (CUS) and to observe the effect of EA on progenitor cell proliferation in the dentate gyrus (DG) of hippocampus. EA was performed on acupoints "Bai-Hui" (Du 20) and unilateral "An-Mian" (EX 17) once daily for 3 consecutive weeks, 2 weeks post CUS procedure. Open field test and forced swimming test were employed to evaluate the behavioral activity during a stress period or EA treatment. The results revealed that exposure to CUS resulted in a decrease of behavioral activity, whilst a daily session of EA treatment significantly reversed the behavioral deficit of these depression model rats. Moreover, as shown by 5-bromo-2-deoxyuridine (BrdU) labeling immunohistochemistry, hippocampal progenitor cell proliferation was decreased in the DG of depression model rats. Intriguingly, EA treatment effectively blocked this decrease. The study demonstrated a potential antidepressant-like effect of EA treatment on CUS induced depression model rats, which might be mediated by up-regulating the hippocampal progenitor cell proliferation.

Spinal microglia play a key role for creating exaggerated pain following tissues inflammation or injury. Electroacupuncture (EA) can effectively control the exaggerated pain both in humans with inflammatory disease and animals with experimental inflammatory pain. However, little is known about the relationship between spinal glial activation and EA analgesia. Using immunohistochemistry, RT-PCR analysis, and behavioral testing, the present study demonstrated that (1) Unilateral intra-articular injection of CFA produced a robust microglial activation and the up-regulation of the tumor necrosis factor (TNF)-alpha, interleukin (IL-1beta), and IL-6 mRNA levels in the spinal cord; (2) Repeated intrathecal (i.t.) injection of minocycline (100 microg), a microglial inhibitor, or EA stimulation of ipsilateral "Huantiao"(GB30) and "Yanglingquan" (GB34) acupoints significantly suppressed CFA-induced nociceptive behavioral hypersensitivity and spinal microglial activation; (3) Combination of EA with minocycline significantly enhanced the inhibitory effects of EA on allodynia and hyperalgesia. For the first time, these data provide direct evidence for the involvement of spinal microglial functional state in anti-nociception of EA. Thus, anti-neuroinflammatory effect of EA might be considered as one of the mechanisms of its anti-arthritic pain effects, and thereby a multidisciplinary integrated approach to treating symptoms related to arthritis might be raised.

This study was designed to explore the different expression of L-PGDS (lipocalin-type prostaglandin D synthase) in rat epididymidis and to gain further insight into the potential function of L-PGDS in male reproduction. The expression of L-PGDS in rat epididymidis was assessed using real-time quantitative PCR and immunoblotting. The distribution of L-PGDS in rat epididymidis was explored by immunohistochemical methods. The result of immunohistochemistry displayed that L-PGDS was mainly distributed in epididymidis and localized within the cytoplasm and the cilia of the epithelial cells. Real-time quantitative PCR and immunoblotting showed that L-PGDS was strikingly expressed in the caput epididymidis, while a moderate to weak expression was observed in the corpus and cauda epididymidis, the level of mRNA was 0.52 ± 0.02 in the caput, 0.48 ± 0.03 in the corpus and 0.32 ± 0.01 in the cauda epididymidis, the level of protein expression in caput, corpus and the cauda groups was 1, 0.89 ± 0.03 and 0.62 ± 0.01, which suggested that L-PGDS may play certain kind of role during the process of the spermatozoa maturation.

Aphoristia (= Symphurus) orientalis Bleeker 1879, collected from an unspecified depth and location in Japanese waters, is the first described species of symphurine tonguefish from Indo-Pacific waters. The original description with accompanying illustration is based on the unique holotype specimen and provides limited diagnostic characters for this taxon. Subsequent to its description, the holotype of A. orientalis has been lost. Limited diagnostic information and loss of the holotype have caused considerable confusion to subsequent systematic studies regarding the identity of this and similar tonguefish species occurring in the Indo-West Pacific region. Several, often-cited, taxonomic accounts purportedly redescribing S. orientalis are erroneous because they include more than one species in these redescriptions. These erroneous redescriptions not only confused the species concept of S. orientalis (Bleeker), but also confounded the systematics of similar Indo-West Pacific tonguefishes. Symphurus novemfasciatus Shen and Lin, described on two specimens collected in southern Taiwan, shares many morphological and pigmentation features similar to those of S. orientalis. Morphological data from a large series of tonguefishes collected in Taiwanese and Japanese waters, as well as molecular data from a smaller number of specimens from these locations, including the type locality of S. novemfasciatus, confirm the presence of only one species, S. orientalis (Bleeker), among these specimens. Symphurus novemfasciatus Shen and Lin is therefore regarded as ajunior subjective synonym of S. orientalis. Symphurus orientalis is redefined based on a large series of specimens identified by a consistent set of morphological criteria, and a neotype is designated to stabilize nomenclature and systematics of this species. Symphurus orientalis differs from congeners by its combination of: a predominant 1-2-2-2-2 pattern of interdigitation of proximal dorsal-fin pterygiophores and neural spines, 12 caudal-fin rays, 9 abdominal and 52-55 total vertebrae, four hypurals, 96-101 dorsal-fin rays, 82-89 anal-fin rays, 87-99 longitudinal scale rows, 37-42 transverse scales, 5-11 (usually) distinct, complete or incomplete, blackish-brown crossbands on the ocular side, uniformly white blind side, and conspicuous bluish-black peritoneum. Documenting morphological variation for S. orientalis represents the most important step towards clarification of the identity of this and other symphurine tonguefish species from this region. Reliable identification of specimens of S. orientalis also provides the foundation for evaluating the status of several other, poorly-known, nominal species of Indo-West Pacific tonguefishes that have features similar to those of S. orientalis. Improved identifications will lead to better knowledge on the geographic distribution of S. orientalis and these other species, as well as to improve estimates of biodiversity and the biogeography of Indo-West Pacific symphurine tonguefishes.

Clinical usage of opioids in pain relief is dampened by analgesic tolerance after chronic exposure, which is related to opioid-associated neuroinflammation. In the current study, which is based on a chronic morphine tolerance rat model and sustained morphine treatment on primary neuron culture, it was observed that Akt phosphorylation, cleaved-Caspase-1-dependent NALP1 inflammasome activation and IL-1β maturation in spinal cord neurons were significantly enhanced by morphine. Moreover, treatment with LY294002, a specific inhibitor of PI3k/Akt signaling, significantly reduced Caspase-1 cleavage, NALP1 inflammasome activation and attenuated morphine tolerance. Tail-flick tests demonstrated that pharmacological inhibition on Caspase-1 activation or antagonizing IL-1β dramatically blocked the development of morphine tolerance. The administration of an exogenous analogue of lipoxin, Aspirin-triggered Lipoxin (ATL), caused a decline in Caspase-1 cleavage, inflammasome activation and mature IL-1β production and thus attenuated the development of morphine tolerance by inhibiting upstream Akt phosphorylation. Additionally, treatment with DAMGO, a selective μ-opioid receptor peptide, significantly induced Akt phosphorylation, Caspase-1 cleavage and anti-nociception tolerance, all of which were attenuated by ATL treatment. Taken together, the present study revealed the involvement of spinal NALP1 inflammasome activation in the development of morphine tolerance and the role of the μ-receptor/PI3k–Akt signaling/NALP1 inflammasome cascade in this process. By inhibiting this signaling cascade, ATL blocked the development of morphine tolerance.

Chronic pain frequently co-occurs with major depressive disorder but the mechanisms are poorly understood. We investigated the contribution of indoleamine 2,3-dioxygenase-1 (IDO1), a rate-limiting enzyme in the conversion of tryptophan to neurotoxic metabolites, to this comorbidity using the spared nerve injury (SNI) model of neuropathic pain in mice. SNI resulted in unilateral mechanical allodynia, reduced social interaction, and increased immobility in the forced swim test without changes in locomotor activity. These findings indicate SNI-induced pain and comorbid depression-like behavior. These behavioral responses were accompanied by increases in plasma kynurenine/tryptophan ratios and increased expression of Ido1 and Il1b mRNA in the liver. Interestingly, SNI did not induce detectable changes in spinal cord or brain Ido1 mRNA levels. SNI was associated with spinal cord inflammatory activity as evidenced by increased Il1b mRNA expression. The SNI-induced increase of liver Ido1 and Il1b mRNA was abrogated by intrathecal administration of the IL-1 inhibitor IL-1RA. Intrathecal IL-1RA also inhibited both mechanical allodynia and depression-like behavior. We also show that Ido1 is required for the development of depression-like behavior because Ido1 −/− mice do not develop increased immobility in the forced swim test or decreased social exploration in response to SNI. Mechanical allodynia was similar in WT and Ido1 −/− mice. In conclusion, our findings show for the first time that neuropathic pain is associated with an increase of Ido1 in liver, but not brain, downstream of spinal cord IL-1β signaling and that Ido1 mediates comorbid depression. Moreover, comorbidity of neuropathic pain and depression are only partially mediated by a common mechanism because mechanical hyperalgesia develops independently of Ido1 .

This study described a modified rat model of bone cancer pain. Syngeneic Walker 256 mammary gland carcinoma cells were injected into the tibia medullary cavity via intercondylar eminence. Series of tests were carried out including bone radiology, bone histology, ambulatory pain, thermal hyperalgesia, mechanical allodynia, weight bearing ability, and electrophysiological recording from primary afferent fibers. The rats inoculated with carcinoma cells showed significant ambulatory pain, mechanical allodynia, and reduction in weight bearing, as well as increased incidence of spontaneous activity in Abeta fibers in affected limb, whereas PBS (vehicle) or heat-killed cells (sham) injected rats showed no significant difference in comparison to normal rats. The pain hypersensitive behaviors were aggravated with time and destruction of bone. Interestingly, mechanical allodynia was also observed in the contralateral limb, indicating the involvement of 'mirror image' pain in bone cancer pain. In summary, the present study provided a useful and easily established rat model of bone cancer pain which will contribute to further study of the mechanisms underlying cancer pain.

Alzheimer's disease (AD) is associated with the inflammatory response in response to amyloid β-peptide (Aβ). Previous studies have suggested that paeoniflorin (PF) shows anti-inflammatory and neuroprotective effects in inflammation-related diseases. However, the impacts of PF on AD have not been investigated. In the present study, we showed that a 4-week treatment with PF could significantly inhibit Aβ burden, Aβ-induced over activation of astrocytes and microglia, downregulation of proinflammatory cytokines, and upregulation of anti-inflammatory cytokines in the brain. In addition, we demonstrated that chronic treatment with PF inhibited the activation of glycogen synthase kinase 3β (GSK-3β) and reversed neuroinflammtory-induced activation of nuclear factor-kappa B (NF-κB) signaling pathways. Moreover, PF exerted inhibitory effects on NALP3 inflammasome, caspase-1, and IL-1β. Collectively, in the present study, we demonstrated that PF exhibits neuroprotective effects in amyloid precursor protein (APP) and presenilin 1 (PS1) double-transgenic (APP/PS1) mice via inhibiting neuroinflammation mediated by the GSK-3β and NF-κB signaling pathways and nucleotide-binding domain-like receptor protein 3 inflammasome. Thus, these results suggest that PF might be useful to intervene in development or progression of neurodegeneration in AD through its anti-inflammatory and anti-amyloidogenic effects.

Hyperactivity and its compensatory mechanisms may causally contribute to synaptic and cognitive deficits in Alzheimer's disease (AD). Blocking the overexcitation of the neural network, with levetiracetam (LEV), a sodium channel blocker applied in the treatment of epilepsy, prevented synaptic and cognitive deficits in human amyloid precursor protein (APP) transgenic mice. This study has brought the potential use of antiepileptic drugs (AEDs) in AD therapy. We showed that the chronic treatment with lamotrigine (LTG), a broad-spectrum AED, suppressed abnormal spike activity, prevented the loss of spines, synaptophysin immunoreactivity, and neurons, and thus attenuated the deficits in synaptic plasticity and learning and memory in APP and presenilin 1 (PS1) mice, which express human mutant APP and PS1. In contrast with LEV, which failed to reduce the generation of amyloid β, the chronic LTG treatment reduced the cleavage of APP by β-secretase and thus the numbers and the size of amyloid plaques in the brains of APP and PS1 mice. Moreover, the levels of brain-derived neurotrophic growth factor (BDNF) and nerve growth factor (NGF) were enhanced in the brains of APP and PS1 mice by the chronic LTG treatment. Therefore, these observations demonstrate that LTG attenuates AD pathology through multiple mechanisms, including modulation of abnormal network activity, reduction of the generation of amyloid beta and upregulation of BDNF and NGF.

Chemotherapy-induced peripheral neuropathy (CIPN) characterized by loss of sensory sensitivity and pain in hands and feet is the major dose-limiting toxicity of many chemotherapeutics. At present, there are no FDA-approved treatments for CIPN. The anti-diabetic drug metformin is the most widely used prescription drug in the world and improves glycemic control in diabetes patients. There is some evidence that metformin enhances the efficacy of cancer treatment. The aim of this study was to test the hypothesis that metformin protects against chemotherapy-induced neuropathic pain and sensory deficits. Mice were treated with cisplatin together with metformin or saline. Cisplatin induced increased sensitivity to mechanical stimulation (mechanical allodynia) as measured using the von Frey test. Co-administration of metformin almost completely prevented the cisplatin-induced mechanical allodynia. Co-administration of metformin also prevented paclitaxel-induced mechanical allodynia. The capacity of the mice to detect an adhesive patch on their hind paw was used as a novel indicator of chemotherapy-induced sensory deficits. Co-administration of metformin prevented the cisplatin-induced increase in latency to detect the adhesive patch indicating that metformin prevents sensory deficits as well. Moreover, metformin prevented the reduction in density of intra-epidermal nerve fibers (IENFs) in the paw that develops as a result of cisplatin treatment. We conclude that metformin protects against pain and loss of tactile function in a mouse model of CIPN. The finding that metformin reduces loss of peripheral nerve endings indicates that mechanism underlying the beneficial effects of metformin includes a neuroprotective activity. Because metformin is widely used for treatment of type II diabetes, has a broad safety profile, and is currently being tested as an adjuvant drug in cancer treatment, clinical translation of these findings could be rapidly achieved.

Levels of expression of the nm23 gene inversely correlated with metastatic potential in several rodent tumor model systems and human breast carcinoma. In the present study, we examined nm23 mRNA levels in two murine ascites hepatoma models (H22-16A3-F and H22-A2-P) with different metastatic potentials. Metastatic H22-16A3-F (80% metastatic rate) and non-metastatic H22-A2-P clones were both derived from murine ascites hepatoma (H22). We found that a 0.8-kb nm23 transcript was expressed in both cell clones. The nm23 gene was expressed at a higher level in non-metastatic H22-A2-P: approximately 8.6-fold higher than in metastatic H22-16A3-F. The present data suggest that the expression of nm23 mRNA might be associated with metastasis of murine ascites hepatoma (H22), though heterogeneity of nm23 steady-state expression levels among the H22 clones remains to be investigated.

Background The neuroinflammatory responses in the spinal cord following bone cancer development have been shown to play an important role in cancer-induced bone pain (CIBP). Lipoxins (LXs), endogenous lipoxygenase-derived eicosanoids, represent a unique class of lipid mediators that possess a wide spectrum of anti-inflammatory and pro-resolving actions. In this study, we investigated the effects of intrathecal injection with lipoxin and related analogues on CIBP in rats. Methods The CIBP model was induced by intra-tibia inoculation of Walker 256 mammary gland carcinoma cells. Mechanical thresholds were determined by measuring the paw withdrawal threshold to probing with a series of calibrated von Frey filaments. Lipoxins and analogues were administered by intrathecal (i.t.) or intravenous (i.v.) injection. The protein level of LXA4 receptor (ALX) was tested by western blot. The localization of lipoxin receptor in spinal cord was assessed by fluorescent immunohistochemistry. Real-time PCR was carried out for detecting the expression of pro-inflammatory cytokines. Results Our results demonstrated that: 1) i.t. injection with the same dose (0.3 nmol) of lipoxin A4 (LXA4), lipoxin B4 (LXB4) or aspirin-triggered-15-epi-lipoxin A4 (ATL) could alleviate the mechanical allodynia in CIBP on day 7 after surgery. ATL showed a longer effect than the others and the effect lasted for 6 hours. ATL administered through i.v. injection could also attenuate the allodynia in cancer rats. 2) The results from western blot indicate that there is no difference in the expression of ALX among the naive, sham or cancer groups. 3) Immunohistochemistry showed that the lipoxin receptor (ALX)-like immunoreactive substance was distributed in the spinal cord, mainly co-localized with astrocytes, rarely co-localized with neurons, and never co-localized with microglia. 4) Real-time PCR analysis revealed that, compared with vehicle, i.t. injection with ATL could significantly attenuate the expression of the mRNA of proinflammatory cytokines (IL-1β and TNF-α) in the spinal cord in CIBP. Conclusions Taken together, the results of our study suggest that LXs and analogues exert strong analgesic effects on CIBP. These analgesic effects in CIBP are associated with suppressing the expression of spinal proinflammatory cytokines.

Although human amnion derived mesenchymal stem cells (AMSC) are a promising source of stem cells, their therapeutic potential for traumatic brain injury (TBI) has not been widely investigated. In this study, we evaluated the therapeutic potential of AMSC using a rat TBI model. AMSC were isolated from human amniotic membrane and characterized by flow cytometry. After induction, AMSC differentiated in vitro into neural stem-like cells (AM-NSC) that expressed higher levels of the neural stem cell markers, nestin, sox2 and musashi, in comparison to undifferentiated AMSC. Interestingly, the neurotrophic factors, brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), neurotrophin 3 (NT-3), glial cell derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF) were markedly upregulated after neural stem cell induction. Following transplantation in a rat TBI model, significant improvements in neurological function, brain tissue morphology, and higher levels of BDNF, NGF, NT-3, GDNF and CNTF, were observed in the AM-NSC group compared with the AMSC and Matrigel groups. However, few grafted cells survived with minimal differentiation into neural-like cells. Together, our results suggest that transplantation of AM-NSC promotes functional rehabilitation of rats with TBI, with enhanced expression of neurotrophic factors a likely mechanistic pathway.

The herbal analgesic gel Tong-Luo-San-Jie (TLSJ) and its modifications are used in traditional Chinese medicine to manage cancer pain. However, its mechanisms are still unknown.To investigate the effects and mechanisms of TLSJ gel on bone cancer pain in a rat model.A bone cancer pain rat model was established by inoculating Walker 256 rat carcinoma cells directly into the right tibial medullary cavity of Sprague-Dawley rats (150-170 g); Phosphate buffered saline (PBS) tibial inoculation was used as control. Cancer-bearing rats were treated twice a day with external TLSJ gel (0.5 g/cm(2)/day) or inert gel control for 21 day (n=10/group). Behavioral tests such as mechanical threshold and paw withdrawal latency (PWL) were carried out. Osteoclastic activities were determined and carboxyterminal pyridinoline cross-linked type I collagen telopeptides (ICTP) and bone-specific alkaline phosphatase (BAP) concentrations were detected with ELISA after treatment. Adverse effects were monitored, and biochemical and histological tests were performed in naïve rats treated with local TLSJ gel for six weeks.TLSJ treatment significantly restored bone cancer-induced decrease of PWL and mechanical threshold compared to inert gel. It also decreased the level of blood serum ICTP and BAP and inhibited osteoclast activities. No adverse effects or abnormal biochemical and histological changes were detected after TLSJ treatment.The present study shows that TLSJ significantly inhibits bone cancer-induced thermal and mechanical sensitization. It suggests that the gel may be useful in managing cancer pain and that it may act by inhibiting osteoclastic activity.

Background Orexin A (OXA, hypocretin/hcrt 1) is a newly discovered potential analgesic substance. However, whether OXA is involved in acupuncture analgesia remains unknown. The present study was designed to investigate the involvement of spinal OXA in electroacupuncture (EA) analgesia. Methods A modified rat model of post-laparotomy pain was adopted and evaluated. Von Frey filaments were used to measure mechanical allodynia of the hind paw and abdomen. EA at 2/15 Hz or 2/100 Hz was performed once on the bilateral ST36 and SP6 for 30 min perioperatively. SB-334867, a selective orexin 1 receptor (OX1R) antagonist with a higher affinity for OXA than OXB, was intrathecally injected to observe its effect on EA analgesia. Results OXA at 0.3 nmol and EA at 2/15 Hz produced respective analgesic effects on the model (P0.05). In addition, naloxone, a selective opioid receptor antagonist, failed to antagonize OXA-induced analgesia (P>0.05). Conclusions The results of the present study indicate the involvement of OXA in EA analgesia via OX1R in an opioid-independent way.

Background The most frequent pain in patients with metastatic breast and prostate cancer is bone pain, which can be severe and difficult to treat. The mechanisms underlying this pain remain unclear. Here we investigated the role of c-jun N-terminal kinase (JNK) pathway in the spinal cord in cancer-induced bone pain (CIBP). Results In this study, we used an established rat CIBP model to investigate the possible role of JNK activation in the spinal cord. After intra-tibial inoculation with Walker 256 rat mammary gland carcinoma cells, the rats displayed mechanical allodynia on day 5, which lasted to day 16. The activation of JNK in neurons and astrocytes in the spinal cord was found on day 12 and day 16 after intra-tibial inoculation with carcinoma cells. A single intrathecal injection with JNK inhibitor SP600125 by lumbar puncture attenuated mechanical allodynia on day 12, and repeated intrathecal injection of SP600126 from day 10 to day 14 had a cumulative analgesic effect on CIBP. Conclusions Taken together, our results demonstrated for the first time that JNK activation in the spinal cord is required in the maintenance of CIBP. Inhibition of the spinal JNK pathway may provide a new therapy for CIBP management.

Tenascin-R (TN-R) is a neural specific protein and an important molecule involved in inhibition of axonal regeneration after spinal cord injury (SCI). Here we report on rabbit-derived TN-R polyclonal antibody, which acts as a TN-R antagonist with high titer and high specificity, promoted neurite outgrowth and sprouting of rat cortical neurons cultured on the inhibitory TN-R substrate in vitro. When locally administered into the lesion sites of rats received spinal cord dorsal hemisection, these TN-R antibodies could significantly decrease RhoA activation and improve functional recovery from corticospinal tract (CST) transection. Thus, passive immunotherapy with specific TN-R antagonist may represent a promising repair strategy following acute SCI.