TLR4-MyD88 signaling is involved in the spinal neurons during the full length of recovery from transection of the motor branch of the femoral nerve in mice.

This study was designed to see the expression of toll-like receptor 4 (TLR4) and downstream molecules including myeloid differentiation factor 88 (MyD88) and interleukin 1-ß (IL-1ß) in the spinal cord as peripheral nerve injury recovered in mice. We established a model of femoral nerve injury (FNI) in C57BL/6 mice by transection of the motor branch of the femoral nerve, followed by retrograde labeling to show the according motor neurons in the anterior horn of the spinal cord pars lumbar. We observed the motor function recovery of the injured hind limbs using behavioral tests. The expression of TLR4, MyD88, and IL-1ß was examined by immunofluorescent staining and western blot. According to the behavior test, the FNI animals fully recovered within 6-8 weeks. TLR4, MyD88, and IL-1ß were expressed in the ventral horn of the spinal cord both at 72 h till 6 weeks after the femoral nerve transection surgery, and these proteins were mostly co-localized with neurons. IL-1ß also tended to rise in the same surgery groups, but more intimate with microglia surrounding nearby retrograde labeled neurons. And western blot results were consistent with histological findings. The results indicate that peripheral nerve injury may induce innate immune reactions of the central neurons and critical signaling like TLR4/MyD88 in the spinal cord may reflect the recovery of the injury. These findings suggest that peripheral nerve injury triggered the TLR4/MyD88 signal in the soma of spinal neurons may be involved in function and nerve restoration through neuron-glia crosstalk.

Cost-Effectiveness Analysis of Camrelizumab Immunotherapy versus Docetaxel or Irinotecan Chemotherapy as Second-Line Therapy for Advanced or Metastatic Esophageal Squamous Cell Carcinoma.

PURPOSE: The aim of this study was to assess the cost-effectiveness of camrelizumab immunotherapy versus docetaxel or irinotecan chemotherapy as second-line therapy for advanced esophageal squamous cell carcinoma (ESCC), which was evaluated in the ESCORT trial. MATERIALS AND METHODS: A partitioned survival model was developed to reflect the costs and effectiveness of the ESCORT trial. The clinical efficacy data, safety data, and health-related costs and utilities were derived from published data from clinical trials or health administration departments in China. Adverse event-related costs, drug administration, and other expenses were derived from a single center of Fujian Medical University Cancer Hospital in 2021. All survival analyses were performed with SPSS software. Overall survival was estimated with the Kaplan-Meier method, and progression-free survival was estimated with the life table method. Sensitivity analyses were conducted to assess the uncertainty of the model. Incremental cost, quality-adjusted life years (QALYs), and the incremental cost-effectiveness ratio (ICER) were calculated. RESULTS: Camrelizumab therapy had 0.232 QALYs at an incremental cost of USD$9959.44 compared with the chemotherapy group with 0.158 QALYs at an incremental cost of USD$8601.67. The ICER was USD$18393.12/QALY. Probabilistic sensitivity analyses showed that when the willingness-to-pay threshold reached USD$31200/QALY, which is nearly three times the Chinese gross domestic product per capita, camrelizumab had an 80% possibility of being cost-effective versus docetaxel or irinotecan chemotherapy. CONCLUSION: Camrelizumab is a cost-effective option compared with docetaxel or irinotecan chemotherapy in patients with advanced ESCC as second-line therapy in China.

Aromatic compounds from Endocomia macrocoma.

Twenty-seven compounds, including a new diarypropane and two new lignans were isolated from the twigs and leaves of Endocomia macrocoma. Their structures were elucidated by spectroscopic analysis. Cytotoxicity evaluation of the new compounds against five human tumor lines showed no inhibitory effects.[Formula: see text].

Moderate prenatal alcohol exposure suppresses the TLR4-mediated innate immune response in the hippocampus of young rats.

Prenatal alcohol exposure (PAE) could lead to developmental disorders of the central nervous system (CNS) and mental retardation. Toll-like receptor (TLR) 4 plays an important role in PAE-induced neurodevelopmental defects. However, how PAE affects TLR4 response in the brain remains controversial. Using a moderate PAE model by feeding pregnant rats with liquid ethanol diet, we investigated the TLR4-mediated response to intraventricular injection of lipopolysaccharide (LPS) in the hippocampus of PEA rats at postnatal day (PND) 30. The results showed that PAE significantly up-regulated the expression of Toll-Interleukin-1 Receptor (TIR)-domain-containing adaptor protein inducing interferon (IFN)-ß (TRIF), TNF-α, and IL-1ß in the rat hippocampus in the absence of LPS, indicated by western blot assay. LPS treatment dramatically up-regulated the expressions of TLR4 and its downstream molecules in the hippocampus of paired-food and control groups. But no such significant changes of those molecules were found in the hippocampus of PAE animals. Moreover, the LPS stimulation even down-regulated the levels of TLR4 and TRIF in the PAE group. These data suggest that the relatively moderate level of PAE may lead to a mild neuroinflammation and a suppression of TLR4-mediated response to LPS in the hippocampus of young rats. As innate immunity plays crucial roles in CNS development, moderate PAE-induced suppression of TLR4-mediated response may serve as a new candidate mechanism of CNS developmental defects.

Two new lignans from Horsfieldia kingii.

Two new dibenzylbutyrolactol lignans and three known dibenzylbutyrolactone lignans were isolated from the twigs and leaves of Horsfieldia kingii. Their structures were elucidated by spectroscopic analysis. Cytotoxicity evaluation of these compounds against five human tumour lines showed no inhibitory effects.

Inhibition of MyD88 Signaling Skews Microglia/Macrophage Polarization and Attenuates Neuronal Apoptosis in the Hippocampus After Status Epilepticus in Mice.

Inflammation is implicated in epileptogenesis. Activated microglia and macrophages (MG/MΦ) are found in the brains of patients with epilepsy-related diseases and animal models of epilepsy. It is not yet known how the MG/MΦ activation phenotype affects pathological changes in the brain after a single seizure. In this study, we had 2 main purposes: first, to characterize post-status epilepticus (SE) inflammation by tracking MG/MΦ polarization, and, second, to explore the role of an innate immune receptor adaptor protein, namely, myeloid differentiation primary response gene 88 (MyD88), in the induction of SE in a mouse model. A lithium-pilocarpine model of seizure conditions was generated in C57BL/6 mice. The intensity and distribution of MG/MΦ polarization were tracked by fluorescent immunohistochemistry and Western blotting for the polarization markers inducible nitrogen oxygenized synthase, arginase-1, CD163, and mannose receptor. We observed steadily increasing M1 MG/MΦ along with MyD88 signal upregulation after SE in the hippocampi of mice, whereas the M2 marker arginase-1 was localized mainly in astrocytes rather than in MG/MΦ. Inhibition or gene knockout of MyD88 reduced M1 MG/MΦ and gliosis although increasing M2 MG/MΦ in the hippocampi of SE mice. MyD88 inhibition also augmented glutamate transporter 1 expression and reduced N-methyl-D-aspartate receptor NR1 subunit expression in the hippocampus to protect pyramidal neurons from apoptosis. These data suggest that MG/MΦ polarization after SE impacts the pathological outcome of the hippocampus via MyD88 signaling and point to MyD88 as a potential neuroprotective target for epilepsy therapy.

RIPK3/MLKL-Mediated Neuronal Necroptosis Modulates the M1/M2 Polarization of Microglia/Macrophages in the Ischemic Cortex.

Cell death and subsequent inflammation are 2 key pathological changes occurring in cerebral ischemia. Active microglia/macrophages play a double-edged role depending on the balance of their M1/M2 phenotypes. Necrosis is the predominant type of cell death following ischemia. However, how necrotic cells modulate the M1/M2 polarization of microglia/macrophages remains poorly investigated. Here, we reported that ischemia induces a rapid RIPK3/MLKL-mediated neuron-dominated necroptosis, a type of programmed necrosis. Ablating RIPK3 or MLKL could switch the activation of microglia/macrophages from M1 to the M2 type in the ischemic cortex. Conditioned medium of oxygen-glucose deprivation (OGD)-treated wild-type (WT) neurons induced M1 polarization, while that of RIPK3-/- neurons favored M2 polarization. OGD treatment induces proinflammatory IL-18 and TNFα in WT but not in RIPK3-/- neurons, which in turn upregulate anti-inflammatory IL-4 and IL-10. Furthermore, the expression of Myd88-a common downstream adaptor of toll-like receptors-is significantly upregulated in the microglia/macrophages of ischemic WT but not of RIPK3-/- or MLKL-/- cortices. Antagonizing the function of Myd88 could phenocopy the effects of RIPK3/MLKL-knockout on the polarization of microglia/macrophages and was neuroprotective. Our data revealed a novel role of necroptotic neurons in modulating the M1/M2 balance of microglia/macrophages in the ischemic cortex, possibly through Myd88 signaling.

A new dimeric diarylpropane from Horsfieldia tetratepala.

Five compounds, including a new dimeric diarylpropane, were isolated from the petroleum ether extract of the twigs and leaves of Horsfieldia tetratepala. The structures of these compounds were elucidated by spectroscopic analysis. Moreover, the antiproliferative activities of these compounds were tested on cancer cell lines, but none is active.

Reactive astrocytes undergo M1 microglia/macrohpages-induced necroptosis in spinal cord injury.

BACKGROUND: A unique feature of the pathological change after spinal cord injury (SCI) is the progressive enlargement of lesion area, which usually results in cavity formation and is accompanied by reactive astrogliosis and chronic inflammation. Reactive astrocytes line the spinal cavity, walling off the lesion core from the normal spinal tissue, and are thought to play multiple important roles in SCI. The contribution of cell death, particularly the apoptosis of neurons and oligodendrocytes during the process of cavitation has been extensively studied. However, how reactive astrocytes are eliminated following SCI remains largely unclear. RESULTS: By immunohistochemistry, in vivo propidium iodide (PI)-labeling and electron microscopic examination, here we reported that in mice, reactive astrocytes died by receptor-interacting protein 3 and mixed lineage kinase domain-like protein (RIP3/MLKL) mediated necroptosis, rather than apoptosis or autophagy. Inhibiting receptor-interacting protein 1 (RIP1) or depleting RIP3 not only significantly attenuated astrocyte death but also rescued the neurotrophic function of astrocytes. The astrocytic expression of necroptotic markers followed the polarization of M1 microglia/macrophages after SCI. Depleting M1 microglia/macrophages or transplantation of M1 macrophages could significantly reduce or increase the necroptosis of astrocytes. Further, the inflammatory responsive genes Toll-like receptor 4 (TLR4) and myeloid differentiation primary response gene 88 (MyD88) are induced in necroptotic astrocytes. In vitro antagonizing MyD88 in astrocytes could significantly alleviate the M1 microglia/macrophages-induced cell death. Finally, our data showed that in human, necroptotic markers and TLR4/MyD88 were co-expressed in astrocytes of injured, but not normal spinal cord. CONCLUSION: Taken together, these results reveal that after SCI, reactive astrocytes undergo M1 microglia/macrophages-induced necroptosis, partially through TLR/MyD88 signaling, and suggest that inhibiting astrocytic necroptosis may be beneficial for preventing secondary SCI.

MicroRNA-141 regulates the expression level of ICAM-1 on endothelium to decrease myocardial ischemia-reperfusion injury.

A growing number of studies have suggested microRNAs (miRNAs) are involved in the modulation of myocardial ischemia-reperfusion (MI/R) injury; however, the role of endogenous miRNAs targeting endothelial cells (ECs) and its interaction with ICAM-1 in the setting of MI/R remain poorly understood. Our microarray results showed that miR-146a, miR-146b-5p, miR-155*, miR-155, miR-497, and miR-451 were significantly upregulated, whereas, miR-141 and miR-564 were significantly downregulated in the ECs challenged with TNF-α for 6 h. Real-time PCR analyses additionally validated that the expression levels of miR-146a, miR-155*, and miR-141 were consistent with the microarray results. Then, ICAM-1 was identified as a novel target of miR-141 by Target Scan software and the reporter gene system. Further functional experiments showed that elevated levels of miR-141 inhibited ICAM-1 expression and diminished leukocytes adhesion to ECs in vitro. In an in vivo murine model of MI/R injury, pretreatment with miR-141 mimics through the tail vein downregulated the expression level of ICAM-1 in heart and attenuated MI/R injury as evidenced by decreased infarct size and decline of serum cardial troponin I (cTnI) and lactate dehydrogenase (LDH) concentration. The cardioprotective effects of miR-141 mimics may be attributed to the decreased infiltration of CD11b(+) cells and F4/80(+) macrophages into ischemic myocardium tissue. In conclusion, our results demonstrate that miR-141, as a novel repressor of ICAM-1, is involved in the attenuation of MI/R injury via antithetical regulation of ICAM-1 and inflammatory cells infiltration. Thus miR-141 may constitute a new therapeutic target in the setting of ischemic heart disease.

Age-dependent loss of parvalbumin-expressing hippocampal interneurons in mice deficient in CHL1, a mental retardation and schizophrenia susceptibility gene.

In humans, deletions/mutations in the CHL1/CALL gene are associated with mental retardation and schizophrenia. Juvenile CHL1-deficient (CHL1(-/-) ) mice have been shown to display abnormally high numbers of parvalbumin-expressing (PV(+) ) hippocampal interneurons and, as adults, display behavioral traits observed in neuropsychiatric disorders. Here, we addressed the question whether inhibitory interneurons and synaptic plasticity in the CHL1(-/-) mouse are affected during brain maturation and in adulthood. We found that hippocampal, but not neocortical, PV(+) interneurons were reduced with age in CHL1(-/-) mice, from a surplus of +27% at 1 month to a deficit of -20% in adulthood compared with wild-type littermates. This loss occurred during brain maturation, correlating with microgliosis and enhanced interleukin-6 expression. In parallel with the loss of PV(+) interneurons, the inhibitory input to adult CA1 pyramidal cells was reduced and a deficit in short- and long-term potentiation developed at CA3-CA1 excitatory synapses between 2 and 9 months of age in CHL1(-/-) mice. This deficit could be abrogated by a GABAA receptor agonist. We propose that region-specific aberrant GABAergic synaptic connectivity resulting from the mutation and a subsequently enhanced synaptic elimination during brain maturation lead to microgliosis, increase in pro-inflammatory cytokine levels, loss of interneurons, and impaired synaptic plasticity. Close homolog of L1-deficient (CHL1(-/-) ) mice have abnormally high numbers of parvalbumin (PV)-expressing hippocampal interneurons in juvenile animals, but in adult animals a loss of these cells is observed. This loss correlates with an increased density of microglia (M), enhanced interleukin-6 (IL6) production and a deficit in short- and long-term potentiation at CA3-CA1 excitatory synapses. Furthermore, adult CHL1(-/-) mice display behavioral traits similar to those observed in neuropsychiatric disorders of humans.

Toll-like receptor 4 signaling in neurons of trigeminal ganglion contributes to nociception induced by acute pulpitis in rats.

Pain caused by acute pulpitis (AP) is a common symptom in clinical settings. However, its underlying mechanisms have largely remained unknown. Using AP model, we demonstrated that dental injury caused severe pulp inflammation with up-regulated serum IL-1ß. Assessment from head-withdrawal reflex thresholds (HWTs) and open-field test demonstrated nociceptive response at 1 day post injury. A consistent up-regulation of Toll-like receptor 4 (TLR4) in the trigeminal ganglion (TG) ipsilateral to the injured pulp was found; and downstream signaling components of TLR4, including MyD88, TRIF and NF-κB, and cytokines such as TNF-α and IL-1ß, were also increased. Retrograde labeling indicated that most TLR4 positve neuron in the TG innnervated the pulp and TLR4 immunoreactivity was mainly in the medium and small neurons. Double labeling showed that the TLR4 expressing neurons in the ipsilateral TG were TRPV1 and CGRP positive, but IB4 negative. Furthermore, blocking TLR4 by eritoran (TLR4 antagonist) in TGs of the AP model significantly down-regulated MyD88, TRIF, NF-κB, TNF-α and IL-1ß production and behavior of nociceptive response. Our findings suggest that TLR4 signaling in TG cells, particularly the peptidergic TRPV1 neurons, plays a key role in AP-induced nociception, and indicate that TLR4 signaling could be a potential therapeutic target for orofacial pain.

Systemic injection of low-dose lipopolysaccharide fails to break down the blood-brain barrier or activate the TLR4-MyD88 pathway in neonatal rat brain.

We aimed to investigate whether peripheral low-dose lipopolysaccharide (LPS) induces the breakdown of the blood-brain barrier (BBB) and/or the activation of toll-like receptor 4 (TLR4) in the neonatal rat brain. Neonatal rats received intraperitoneal injections of low-dose LPS (0.3 mg/kg∙bw), and the BBB compromise was detected by Evans Blue extravasation and electron microscopy. Meanwhile, TLR4, adaptin myeloid differentiation factor 88 (MyD88), nuclear transcription factor kappa-B (NF-κB) p50 and tumor necrosis factor alpha (TNFα) in the neonatal rat brain were determined by quantitative real-time polymerase chain reaction (PCR) and Western Blot. Immunohistochemistry was used to determine the distribution and activation of microglia in the brain after LPS administration. It was demonstrated that Evans Blue extravasation was not observed in the brain parenchyma, and that tight junctions of cerebral endothelial cells remained intact after systemic injections of LPS in neonatal rats. Although intracerebroventricular injections of LPS activated microglia and up-regulated the expression of TLR4, MyD88, NF-κB p50 and TNFα in the neonatal rat brain, systemic LPS did not induce these responses. These findings indicate that while the neonatal rat brain responds to the direct intra-cerebral administration of LPS through robust TLR4 activation, systemic low-dose LPS does not induce the innate immune reaction or compromise the BBB in neonatal rats.

Beneficial effects of thymosin ß4 on spinal cord injury in the rat.

Thymosin ß4 (Tß4) has many physiological functions that are highly relevant to spinal cord injury (SCI), including neuronal survival, anti-inflammation, wound repair promotion, and angiogenesis. The present study investigated the therapeutic value of Tß4 in SCI, with a focus on its neuroprotective, anti-inflammatory, and vasculoprotective properties. Tß4 or a saline control was administered by intraperitoneal injection 30 min, 3 days, or 5 days after SCI with mild compression in rat. Locomotor recovery was tested with the Basso-Beattie-Bresnahan scale and a footprint analysis. All behavioral assessments were markedly improved with Tß4 treatment. Histological examination at 7 days post injury showed that the numbers of surviving neurons and oligodendrocytes were significantly increased in Tß4-treated animals compared to saline-treated controls. Levels of myelin basic protein, a marker of mature oligodendrocytes, in Tß4-treated rats were 57.8% greater than those in saline-treated controls. The expression of ED1, a marker of activated microglia/macrophages, was reduced by 36.9% in the Tß4-treated group compared to that of the saline-treated group. Tß4 treatment after SCI was also associated with a significant decrease in pro-inflammatory cytokine gene expression and a significant increase in the mRNA levels of IL-10 compared to the control. Moreover, the size of lesion cavity delineated by astrocyte scar in the injured spinal cord was markedly reduced in Tß4-treated animals compared to saline-treated controls. Given the known safety of Tß4 in clinical trials and its beneficial effects on SCI recovery, the results of this study suggested that Tß4 is a good candidate for SCI treatment in humans.

Different TLR4 expression and microglia/macrophage activation induced by hemorrhage in the rat spinal cord after compressive injury.

BACKGROUND: Hemorrhage is a direct consequence of traumatic injury to the central nervous system and may cause innate immune reactions including cerebral Toll-like receptor (TLR) 4 upregulation which usually leads to poor outcome in the traumatic brain injury. In spinal cord injury (SCI), however, how hemorrhage induces innate immune reaction in spinal parenchyma remains unknown. The present study aimed to see whether blood component and/or other factor(s) induce TLR4 and microglia/macrophages involved innate immune reactions in the rat spinal cord after traumatic injury. METHODS: Using the compressive SCI model of the rat, hemorrhage in the spinal cord was identified by hematoxylin-eosin staining. Microglia/macrophage activation, TLR4 expression, and cell apoptosis were investigated by immunohistochemistry. Nuclear factor (NF)-κB p50 level of the two segments of the cord was detected by western blotting assay. With carbon powder injection, blood origination of the hematoma was explored. The blood-spinal cord barrier (BSCB) states of the lesion site and the hematoma were compared with immunohistochemistry and tannic acid-ferric chloride staining. RESULTS: Histological observation found blood accumulated in the center of compression lesion site (epicenter) and in the hematoma approximately 1.5 cm away from the epicenter. TLR4 expression, microglia//macrophage activation, and subsequent apoptosis in the area of far-away hematoma were late and weak in comparison to that in epicenter. In addition, TLR4 positive microglia/macrophages appeared to be phagocytotic in the far-away hematoma more obviously than that in the epicenter. Injected carbon powder indicated that accumulated blood of the far-away hematoma originated from the bleeding of the lesion epicenter, and the BSCB around the hematoma was not compromised in the early phase. Accordingly, at 3 days post injury, NF-κB p50 was upregulated based on the similar levels of blood component hemoglobin, and cell apoptosis was obvious in the epicenter but not in the far-away hematoma. CONCLUSION: These data suggest that besides blood component, BSCB compromise and the extent of tissue injury contribute more to TLR4 and microglia/macrophage responses to the spinal cord hemorrhage. Therefore, the innate immune environment is a necessary consideration for the SCI therapy targeting TLR4 and microglia/macrophages.

Electromagnetic pulse exposure induces overexpression of beta amyloid protein in rats.

BACKGROUND AND AIMS: With the developing and widely used electromagnetic field (EMF) technology, more and more studies are focusing on the relationship between EMF and Alzheimer's disease (AD). Electromagnetic pulse (EMP) is one type of widely used EMF. This study aimed to clarify whether EMP exposure could induce cognitive and memory impairment, thus finding a possible relationship between EMP and AD. METHODS: Forty healthy male Sprague Dawley rats were randomly divided into four groups. Animals, respectively, received 100, 1000, and 10,000 pulses EMP (field strength 50 kV/m, repetition rate 100 Hz) exposure and sham exposure when 2 months old. Monthly Morris water maze (MWM) was used to test the changes of cognitive and memory ability. Superoxide dismutase (SOD) activity and glutathione (GSH) content were used as oxidative stress indexes. Expressions of some types of Alzheimer's disease-related proteins were also detected. RESULTS: After exposure, EMP exposure caused clear cognitive and memory impairment compared with sham exposure group (p <0.05). Determination of oxidation indexes showed decreased SOD activity and GSH content in exposure groups compared with sham group. Immunohistochemical (IHC) staining showed increased beta amyloid protein (Aß) in EMP exposure groups compared with sham group. Western blot experiments showed increased expressions of Aß oligomer and beta amyloid protein precursor (APP) in EMP exposure groups. Increased expression of microtubule-associated protein 1 light chain 3-II (LC3-II) was also found. CONCLUSIONS: The present results showed that EMP exposure can cause long-term impairment in impaired cognition and memory of rats, resulting in AD-like symptoms. This may be induced by enhancing oxidative stress and is related to autophagy dysfunction.

Early Blockade of TLRs MyD88-Dependent Pathway May Reduce Secondary Spinal Cord Injury in the Rats.

To determine the role of toll-like receptors (TLRs) myeloid differentiation factor 88 (MyD88) dependent pathway in the spinal cord secondary injury, compression injury was made at T8 segment of the spinal cord in adult male Sprague-Dawley rats. Shown by RT-PCR, TLR4 mRNA in the spinal cord was quickly elevated after compression injury. Intramedullary injection of MyD88 inhibitory peptide (MIP) resulted in significant improvement in locomotor function recovery at various time points after surgery. Meanwhile, injury area, p38 phosphorylation, and proinflammation cytokines in the injured spinal cord were significantly reduced in MIP-treated animals, compared with control peptide (CP) group. These data suggest that TLRs MyD88-dependent pathway may play an important role in the development of secondary spinal cord injury, and inhibition of this pathway at early time after primary injury could effectively protect cells from inflammation and apoptosis and therefore improve the functional recovery.

The protective effects of inosine against chemical hypoxia on cultured rat oligodendrocytes.

Inosine is a purine nucleoside and is considered protective to neural cells including neurons and astrocytes against hypoxic injury. However, whether oligodendrocytes (OLs) could also be protected from hypoxia by inosine is not known. Here we investigated the effects of inosine on primarily cultured rat OLs injured by rotenone-mediated chemical hypoxia, and the mechanisms of the effects using ATP assay, MTT assay, PI-Hoechst staining, TUNEL, and immunocytochemistry. Results showed that rotenone exposure for 24 h caused cell death and impaired viability in both immature and mature OLs, while pretreatment of 10 mM inosine 30 min before rotenone administration significantly reduced cell death and improved the viability of OLs. The same concentration of inosine given 120 min after rotenone exposure also improved viability of injured mature OLs. Immunocytochemistry for nitrotyrosine and cellular ATP content examination indicated that inosine may protect OLs by providing ATP and scavenging peroxynitrite for cells. In addition, immature OLs were more susceptible to hypoxia than mature OLs; and at the similar degree of injury, inosine protected immature and mature OLs differently. Quantitative real-time PCR revealed that expression of adenosine receptors was different between these two stages of OLs. These data suggest that inosine protect OLs from hypoxic injury as an antioxidant and ATP provider, and the protective effects of inosine on OLs vary with cell differentiation, possibly due to the adenosine receptors expression profile. As OLs form myelin in the central nervous system, inosine could be used as a promising drug to treat demyelination-involved disorders.

Neutralization of BDNF attenuates the in vitro protective effects of olfactory ensheathing cell-conditioned medium on scratch-insulted retinal ganglion cells.

Transplantation of olfactory ensheathing cells (OECs) becomes one of the promising strategies in restoring lost functions of injured central nervous system. Elevated level of expressed brain-derived neurotrophic factor (BDNF) was revealed in the previous studies to be related to the protective effects of OECs on injured cortical and brain stem neurons as well as retinal ganglion cells (RGCs), but no evidence has been obtained to demonstrate whether transplanted OECs protect injured central neurons directly by their secreted BDNF. In the present study, the effects of BDNF neutralization on the neuroprotection of adult OEC-conditioned medium (OEC-CM) on scratch-insulted RGCs were examined. The results showed that OEC-CM protected cultured RGCs from scratch insult, and neutralization of BDNF by BDNF neutralizing antibody attenuated such neuroprotection of the medium. It is thus concluded that neurotrophic factors including BDNF secreted by OECs can protect injured OECs in vitro and BDNF plays a major role in such a protection of OECs.

Jingzhaotoxin-IX, a novel gating modifier of both sodium and potassium channels from Chinese tarantula Chilobrachys jingzhao.

Tarantula Chilobrachys jingzhao is one of the most venomous species distributed in China. In this study, we have isolated and characterized a novel neurotoxin named Jingzhaotoxin-IX (JZTX-IX) from the venom of the tarantula. JZTX-IX is a C-terminally amidated peptide composed of 35 amino acid residues. The toxin shows 74% sequence identity with CcoTx3 from southeastern Africa tarantula Ceratogyrus cornuatus. JZTX-IX was found to interact with multiple types of ion channels including voltage-gated sodium channels (both tetrodotoxin-resistant and tetrodotoxin-sensitive isoforms) and Kv2.1 channel. The toxin had no effect on delayed rectifier potassium channel Kv1.1, 1.2 and 1.3. JZTX-IX shifted the voltage dependence of channel activation to more positive voltages, but binding of toxin to ion channels was not reversible by extreme depolarization. In addition, JZTX-IX could bias the activities of ion channels towards closed state because the time constant for decay (channel deactivation) of tail currents became faster in the presence of toxin. Taken together with the finding that 10 microM JZTX-IX completely blocked ion channels at resting potential without pulsing, we propose that JZTX-IX is a gating modifier showing low selectivity for ion channel types and trapping voltage sensor at closed state.