Transient receptor potential vanilloid 1 inhibition reduces brain damage by suppressing neuronal apoptosis after intracerebral hemorrhage.

Intracerebral hemorrhage (ICH) induces a complex sequence of apoptotic cascades and inflammatory responses, leading to neurological impairment. Transient receptor potential vanilloid 1 (TRPV1), a nonselective cation channel with high calcium permeability, has been implicated in neuronal apoptosis and inflammatory responses. This study used a mouse ICH model and neuronal cultures to examine whether TRPV1 activation exacerbates brain damage and neurological deficits by promoting neuronal apoptosis and neuroinflammation. ICH was induced by injecting collagenase in both wild-type (WT) C57BL/6 mice and TRPV1-/- mice. Capsaicin (CAP; a TRPV1 agonist) or capsazepine (a TRPV1 antagonist) was administered by intracerebroventricular injection 30 min before ICH induction in WT mice. The effects of genetic deletion or pharmacological inhibition of TRPV1 using CAP or capsazepine on motor deficits, histological damage, apoptotic responses, blood-brain barrier (BBB) permeability, and neuroinflammatory reactions were explored. The antiapoptotic mechanisms and calcium influx induced by TRPV1 inactivation were investigated in cultured hemin-stimulated neurons. TRPV1 expression was upregulated in the hemorrhagic brain, and TRPV1 was expressed in neurons, microglia, and astrocytes after ICH. Genetic deletion of TRPV1 significantly attenuated motor deficits and brain atrophy for up to 28 days. Deletion of TRPV1 also reduced brain damage, neurodegeneration, microglial activation, cytokine expression, and cell apoptosis at 1 day post-ICH. Similarly, the administration of CAP ameliorated brain damage, neurodegeneration, brain edema, BBB permeability, and cytokine expression at 1 day post-ICH. In primary neuronal cultures, pharmacological inactivation of TRPV1 by CAP attenuated neuronal vulnerability to hemin-induced injury, suppressed apoptosis, and preserved mitochondrial integrity in vitro. Mechanistically, CAP reduced hemin-stimulated calcium influx and prevented the phosphorylation of CaMKII in cultured neurons, which was associated with reduced activation of P38 and c-Jun NH2 -terminal kinase mitogen-activated protein kinase signaling. Our results suggest that TRPV1 inhibition may be a potential therapy for ICH by suppressing mitochondria-related neuronal apoptosis.

Surf4 collaborates with derlin-2 and derlin-1 to mediate cyclooxygenase-2 translocation to the cytosol for degradation.

Derlin family members participate in the retrotranslocation of endoplasmic reticulum (ER) lumen proteins to the cytosol for ER-associated degradation (ERAD); however, the proteins facilitating this retrotranslocation remain to be explored. Using CRISPR library screening, we have found that derlin-2 and surfeit locus protein 4 (Surf4) are candidates to facilitate degradation of cyclooxygenase-2 (COX-2, also known as PTGS2). Our results show that derlin-2 acts upstream of derlin-1 and that Surf4 acts downstream of derlin-2 and derlin-1 to facilitate COX-2 degradation. Knockdown of derlin-2 or Surf4 impedes the ubiquitylation of COX-2 and the interaction of COX-2 with caveolin-1 (Cav-1) and p97 (also known as VCP) in the cytosol. Additionally, COX-2 degradation is N-glycosylation dependent. Although derlin-2 facilitates degradation of N-glycosylated COX-2, the interaction between derlin-2 and COX-2 is independent of COX-2 N-glycosylation. Derlin-1, Surf4 and p97 preferentially interact with non-glycosylated COX-2, whereas Cav-1 preferentially interacts with N-glycosylated COX-2, regardless of the N-glycosylation pattern. Collectively, our results reveal that Surf4 collaborates with derlin-2 and derlin-1 to mediate COX-2 translocation from the ER lumen to the cytosol. The derlin-2-derlin-1-Surf4-Cav-1 machinery might represent a unique pathway to accelerate COX-2 degradation in ERAD.

Low-intensity pulsed ultrasound ameliorates glia-mediated inflammation and neuronal damage in experimental intracerebral hemorrhage conditions.

BACKGROUND: Intracerebral hemorrhage (ICH) is a condition associated with high morbidity and mortality, and glia-mediated inflammation is a major contributor to neurological deficits. However, there is currently no proven effective treatment for clinical ICH. Recently, low-intensity pulsed ultrasound (LIPUS), a non-invasive method, has shown potential for neuroprotection in neurodegenerative diseases. This study aimed to investigate the neuroprotective effects and potential mechanisms of LIPUS on glia-mediated inflammation in ICH. METHODS: This study used 289 mice to investigate the effects of LIPUS on ICH. ICH was induced by injecting bacterial collagenase (type VII-S; 0.0375 U) into the striatum of the mice. LIPUS was applied noninvasively for 3 days, including a 2-h-delayed intervention to mimic clinical usage. The study evaluated neurological function, histology, brain water content, hemoglobin content, MRI, and protein expression of neurotrophic factors, inflammatory molecules, and apoptosis. In vitro studies investigated glia-mediated inflammation by adding thrombin (10 U/mL) or conditioned media to primary and cell line cultures. The PI3K inhibitor LY294002 was used to confirm the effects of PI3K/Akt signaling after LIPUS treatment. RESULTS: LIPUS treatment improved neurological deficits and reduced tissue loss, edema, and neurodegeneration after ICH. The protective effects of LIPUS resulted from decreased glia-mediated inflammation by inhibiting PI3K/Akt-NF-κB signaling, which reduced cytokine expression and attenuated microglial activation-induced neuronal damage in vitro. CONCLUSIONS: LIPUS treatment improved neurological outcomes and reduced glia-mediated inflammation by inhibiting PI3K/Akt-NF-κB signaling after ICH. LIPUS may provide a non-invasive potential management strategy for ICH.

Antenatal low-intensity pulsed ultrasound reduces neurobehavioral deficits and brain injury following dexamethasone-induced intrauterine growth restriction.

Intrauterine growth restriction (IUGR) is a leading cause of perinatal mortality and morbidity, and IUGR survivors are at increased risk of neurodevelopmental deficits. No effective interventions are currently available to improve the structure and function of the IUGR brain before birth. This study investigated the protective effects of low-intensity pulsed ultrasound (LIPUS) on postnatal neurodevelopmental outcomes and brain injury using a rat model of IUGR induced by maternal exposure to dexamethasone (DEX). Pregnant rats were treated with DEX (200 µg/kg, s.c.) and LIPUS daily from gestational day (GD) 14 to 19. Behavioral assessments were performed on the IUGR offspring to examine neurological function. Neuropathology, levels of neurotrophic factors, and CaMKII-Akt-related molecules were assessed in the IUGR brain, and expression of glucose and amino acid transporters and neurotrophic factors were examined in the placenta. Maternal LIPUS treatment increased fetal weight, fetal liver weight, and placental weight following IUGR. LIPUS treatment also increased neuronal number and myelin protein expression in the IUGR brain, and attenuated neurodevelopmental deficits at postnatal day (PND) 18. However, the number of oligodendrocytes or microglia was not affected. These changes were associated with the upregulation of brain-derived neurotrophic factor (BDNF) and placental growth factor (PlGF) protein expression, and enhancement of neuronal CaMKII and Akt activation in the IUGR brain at PND 1. Additionally, LIPUS treatment promoted glucose transporter (GLUT) 1 production and BDNF expression in the placenta, but had no effects on GLUT3 or amino acid transporter expression. Our findings suggest that antenatal LIPUS treatment may reduce IUGR-induced brain injury via enhancing cerebral BDNF/CaMKII/Akt signaling. These data provide new evidence that LIPUS stimulation could be considered for antenatal neuroprotective therapy in IUGR.

Increased Soluble Epoxide Hydrolase in Human Gestational Tissues from Pregnancies Complicated by Acute Chorioamnionitis.

Chorioamnionitis (CAM) is primarily a polymicrobial bacterial infection involving chorionic and amniotic membranes that is associated with increased risk of preterm delivery. Epoxyeicosatrienoic acids (EETs) are eicosanoids generated from arachidonic acid by cytochrome P450 enzymes and further metabolized mainly by soluble epoxide hydrolase (sEH) to produce dihydroxyeicosatrienoic acids (DHETs). As a consequence of this metabolism of EETs, sEH reportedly exacerbates several disease states; however, its role in CAM remains unclear. The objectives of this study were to (1) determine the localization of sEH and compare the changes it undergoes in the gestational tissues (placentas and fetal membranes) of women with normal-term pregnancies and those with pregnancies complicated by acute CAM; (2) study the effects of lipopolysaccharide (LPS) on the expression of sEH in the human gestational tissues; and (3) investigate the effect of 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA), a specific sEH inhibitor, on LPS-induced changes in 14,15-DHET and cytokines such as interleukin- (IL-) 1ß and IL-6 in human gestational tissues in vitro and in pregnant mice. We found that women with pregnancies complicated by acute CAM had higher levels of sEH mRNA and protein in fetal membranes and villous tissues compared to those in women with normal-term pregnancies without CAM. Furthermore, fetal membrane and villous explants treated with LPS had higher tissue levels of sEH mRNA and protein and 14,15-DHET than those present in the vehicle controls, while the administration of AUDA in the media attenuated the LPS-induced production of 14,15-DHET in tissue homogenates and IL-1ß and IL-6 in the media of explant cultures. Administration of AUDA also reduced the LPS-induced changes of 14,15-DHET, IL-1ß, and IL-6 in the placentas of pregnant mice. Together, these results suggest that sEH participates in the inflammatory changes in human gestational tissues in pregnancies complicated by acute CAM.

Activation of TrkB/Akt signaling by a TrkB receptor agonist improves long-term histological and functional outcomes in experimental intracerebral hemorrhage.

BACKGROUND: Intracerebral hemorrhage (ICH) induces a complex sequence of apoptotic cascades that contribute to secondary neuronal damage. Tropomyosin-related kinase receptor B (TrkB) signaling plays a crucial role in promoting neuronal survival following brain damage. METHODS: The present study investigated the protective effects and underlying mechanisms of TrkB activation by the specific TrkB agonist, 7,8-dihydroxyflavone (7,8-DHF), in a model of collagenase-induced ICH and in neuronal cultures. Mice subjected to collagenase-induced ICH were intraperitoneally injected with either 7,8-DHF or vehicle 10 min after ICH and, subsequently, daily for 3 days. Behavioral studies, brain edema measurement, and histological analysis were conducted. Levels of TrkB signaling-related molecules and apoptosis-related proteins were analyzed by western blots. RESULTS: Treatment with 20 mg/kg 7,8-DHF significantly improved functional recovery and reduced brain damage up to 28 days post-ICH. Reduction in neuronal death, apoptosis, and brain edema were also observed in response to 7,8-DHF treatment at 3 days post-ICH. These changes were accompanied by a significant increase in the phosphorylation of TrkB and Akt (Ser473/Thr308) at 1 and 3 days, but had no effect on Erk 44/42 phosphorylation. 7,8-DHF also enhanced the phosphorylation of Ask-1 Ser967 and FOXO-1, downstream targets of Akt at 1 and 3 days. Moreover, 7,8-DHF increased brain-derived neurotrophic factor levels at 1 day. In primary cultured neurons stimulated with hemin, 7,8-DHF promoted survival and reduced apoptosis. Furthermore, delaying the administration of 7,8-DHF to 3 h post-ICH reduced brain tissue damage and neuronal death. CONCLUSIONS: Our findings demonstrate that the activation of TrkB signaling by 7,8-DHF protects against ICH via the Akt, but not the Erk, pathway. These data provide new insights into the role of TrkB signaling deficit in the pathophysiology of ICH and highlight TrkB/Akt as possible therapeutic targets in this disease.

Transcranial Ultrasound Stimulation Improves Long-Term Functional Outcomes and Protects Against Brain Damage in Traumatic Brain Injury.

The purpose of this study was to assess the long-term treatment efficacy of low-intensity pulsed ultrasound (LIPUS) on functional outcomes, brain edema, and the possible involvement of reactions in mice following traumatic brain injury (TBI). Mice subjected to controlled cortical impact injury received LIPUS treatment daily for a period of 4 weeks. The effects of LIPUS on edema were detected by MR imaging in the mouse brain at 148 days following TBI. Long-term functional outcomes of LIPUS stimulation were evaluated by behavioral analyses. One-way or two-way analysis of variance and Student's t test were used for statistical analyses, with a significant level of .05. Up to post-injury day 148, treatment with LIPUS significantly improved functional outcomes (all p < 0.05). LIPUS also significantly attenuated brain edema and neuronal death at day 148 after TBI (all p < 0.05). Furthermore, LIPUS reduced MMP9 activity, neutrophil infiltration, and microglial activation at day 1 or day 4 following TBI (all p < 0.05). Meanwhile, LIPUS increased the Bcl-2/Bax ratio and enhanced the phosphorylation of Bad and FOXO-1 at day 1 or day 4 following TBI (all p < 0.05). Almost 5 months of follow-up showed that the treatment efficacy of post-injury LIPUS stimulation on reduced brain edema and improved functional outcomes persisted over time after TBI. The neuroprotective effects of LIPUS are associated with a reduction of early inflammatory events and inhibition of apoptotic progression.

Deletion or inhibition of soluble epoxide hydrolase protects against brain damage and reduces microglia-mediated neuroinflammation in traumatic brain injury.

Traumatic brain injury (TBI) induces a series of inflammatory processes that contribute to neuronal damage. The present study investigated the involvement of soluble epoxide hydrolase (sEH) in neuroinflammation and brain damage in mouse TBI and in microglial cultures. The effects of genetic deletion of sEH and treatment with an sEH inhibitor, 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA), on brain damage and inflammatory responses were evaluated in mice subjected to controlled cortical impact. The anti-inflammatory mechanism of sEH inhibition/deletion was investigated in vitro. TBI-induced an increase in sEH protein level in the injured cortex from 1 h to 4 days and sEH was expressed in microglia. Genetic deletion of sEH significantly attenuated functional deficits and brain damage up to 28 days post-TBI. Deletion of sEH also reduced neuronal death, apoptosis, brain edema, and BBB permeability at 1 and 4 day(s). These changes were associated with markedly reduced microglial/macrophage activation, neutrophil infiltration, matrix metalloproteinase-9 activity, inflammatory mediator expression at 1 and 4 day(s), and epoxyeicosatrienoic acid (EET) degradation at 1 and 4 day(s). Administration of AUDA attenuated brain edema, apoptosis, inflammatory mediator upregulation and EET degradation at 4 days. In primary microglial cultures, AUDA attenuated both LPS- or IFN-γ-stimulated nitric oxide (NO) production and reduced LPS- or IFN-γ-induced p38 MAPK and NF-κB signaling. Deletion of sEH also reduced IFN-γ-induced NO production. Moreover, AUDA attenuated N2A neuronal death induced by BV2 microglial-conditioned media. Our results suggest that inhibition of sEH may be a potential therapy for TBI by modulating the cytotoxic functions of microglia.

Low-intensity pulsed ultrasound improves behavioral and histological outcomes after experimental traumatic brain injury.

The purpose of this study was to investigate the neuroprotective effects of low-intensity pulsed ultrasound (LIPUS) on behavioral and histological outcomes in a mouse model of traumatic brain injury (TBI). Mice subjected to controlled cortical impact injury were treated with LIPUS in the injured region daily for a period of 4 weeks. The effects of LIPUS on edema were observed by MR imaging in the mouse brain at 1 and 4 days following TBI. Brain water content, blood-brain barrier permeability, histology analysis, and behavioral studies were performed to assess the effects of LIPUS. Two-way analysis of variance and Student t test were used for statistical analyses, with a significant level of 0.05. Treatment with LIPUS significantly attenuated brain edema, blood-brain barrier permeability, and neuronal degeneration beginning at day 1. Compared with the TBI group, LIPUS also significantly improved functional recovery and reduced contusion volumes up to post-injury day 28. Post-injury LIPUS treatment reduced brain edema and improved behavioral and histological outcomes following TBI. The neuroprotective effects of LIPUS may be a promising new technique for treating TBI.

Genetic deletion or pharmacological inhibition of soluble epoxide hydrolase reduces brain damage and attenuates neuroinflammation after intracerebral hemorrhage.

BACKGROUND: Inflammatory responses significantly contribute to neuronal damage and poor functional outcomes following intracerebral hemorrhage (ICH). Soluble epoxide hydrolase (sEH) is known to induce neuroinflammatory responses via degradation of anti-inflammatory epoxyeicosatrienoic acids (EET), and sEH is upregulated in response to brain injury. The present study investigated the involvement of sEH in ICH-induced neuroinflammation, brain damage, and functional deficits using a mouse ICH model and microglial cultures. METHODS: ICH was induced by injecting collagenase in both wild-type (WT) C57BL/6 mice and sEH knockout (KO) mice. WT mice were injected intracerebroventricularly with 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA), a selective sEH inhibitor, 30 min before ICH. Expression of sEH in the hemorrhagic hemisphere was examined by immunofluorescence and Western blot analysis. The effects of genetic deletion or pharmacological inhibition of sEH by AUDA on neuroinflammatory responses, EET degradation, blood-brain barrier (BBB) permeability, histological damage, and functional deficits were evaluated. The anti-inflammatory mechanism of sEH inactivation was investigated in thrombin- or hemin-stimulated cultured microglia. RESULTS: ICH induced an increase in sEH protein levels in the hemorrhagic hemisphere from 3 h to 4 days. sEH was expressed in microglia/macrophages, astrocytes, neurons, and endothelial cells in the perihematomal region. Genetic deletion of sEH significantly attenuated microglia/macrophage activation and expression of inflammatory mediators and reduced EET degradation at 1 and 4 days post-ICH. Deletion of sEH also reduced BBB permeability, matrix metalloproteinase (MMP)-9 activity, neutrophil infiltration, and neuronal damage at 1 and 4 days. Likewise, administration of AUDA attenuated proinflammatory microglia/macrophage activation and EET degradation at 1 day post-ICH. These findings were associated with a reduction in functional deficits and brain damage for up to 28 days. AUDA also ameliorated neuronal death, BBB disruption, MMP-9 activity, and neutrophil infiltration at 1 day. However, neither gene deletion nor pharmacological inhibition of sEH altered the hemorrhage volume following ICH. In primary microglial cultures, genetic deletion or pharmacological inhibition of sEH by AUDA reduced thrombin- and hemin-induced microglial activation. Furthermore, AUDA reduced thrombin- and hemin-induced P38 MAPK and NF-κB activation in BV2 microglia cultures. Ultimately, AUDA attenuated N2A neuronal death that was induced by BV2 microglial conditioned media. CONCLUSIONS: Our results suggest that inhibition of sEH may provide a potential therapy for ICH by suppressing microglia/macrophage-mediated neuroinflammation.

Transcranial ultrasound stimulation promotes brain-derived neurotrophic factor and reduces apoptosis in a mouse model of traumatic brain injury.

BACKGROUND: The protein expressions of brain-derived neurotrophic factor (BDNF) can be elevated by transcranial ultrasound stimulation in the rat brain. OBJECTIVE: The purpose of this study was to investigate the effects and underlying mechanisms of BDNF enhancement by low-intensity pulsed ultrasound (LIPUS) on traumatic brain injury (TBI). METHODS: Mice subjected to controlled cortical impact injury were treated with LIPUS in the injured region daily for a period of 4 days. Western blot analysis and immunohistochemistry were performed to assess the effects of LIPUS. RESULTS: The results showed that the LIPUS treatment significantly promoted the neurotrophic factors BDNF and vascular endothelial growth factor (VEGF) at day 4 after TBI. Meanwhile, LIPUS also enhanced the phosphorylation of Tropomyosin-related kinase B (TrkB), Akt, and cAMP-response element binding protein (CREB). Furthermore, treatment with LIPUS significantly decreased the level of cleaved caspase-3. The reduction of apoptotic process was inhibited by the anti-BDNF antibody. CONCLUSIONS: In short, post-injury LIPUS treatment increased BDNF protein levels and inhibited the progression of apoptosis following TBI. The neuroprotective effects of LIPUS may be associated with enhancements of the protein levels of neurotrophic factors, at least partially via the TrkB/Akt-CREB signaling pathway.

Performing Repeated Quantitative Small-Animal PET with an Arterial Input Function Is Routinely Feasible in Rats.

Performing quantitative small-animal PET with an arterial input function has been considered technically challenging. Here, we introduce a catheterization procedure that keeps a rat physiologically stable for 1.5 mo. We demonstrated the feasibility of quantitative small-animal 18F-FDG PET in rats by performing it repeatedly to monitor the time course of variations in the cerebral metabolic rate of glucose (CMRglc). Methods: Aseptic surgery was performed on 2 rats. Each rat underwent catheterization of the right femoral artery and left femoral vein. The catheters were sealed with microinjection ports and then implanted subcutaneously. Over the next 3 wk, each rat underwent 18F-FDG quantitative small-animal PET 6 times. The CMRglc of each brain region was calculated using a 3-compartment model and an operational equation that included a k*4Results: On 6 mornings, we completed 12 18F-FDG quantitative small-animal PET studies on 2 rats. The rats grew steadily before and after the 6 quantitative small-animal PET studies. The CMRglc of the conscious brain (e.g., right parietal region, 99.6 ± 10.2 µmol/100 g/min; n = 6) was comparable to that for 14C-deoxyglucose autoradiographic methods. Conclusion: Maintaining good blood patency in catheterized rats is not difficult. Longitudinal quantitative small-animal PET imaging with an arterial input function can be performed routinely.

Treatment with TO901317, a synthetic liver X receptor agonist, reduces brain damage and attenuates neuroinflammation in experimental intracerebral hemorrhage.

BACKGROUND: Intracerebral hemorrhage (ICH) induces a series of inflammatory processes that contribute to neuronal damage and neurological deterioration. Liver X receptors (LXRs) are nuclear receptors that negatively regulate transcriptional processes involved in inflammatory responses, but their role in the pathology following ICH remains unclear. The present study investigated the neuroprotective effects and anti-inflammatory actions of TO901317, a synthetic LXR agonist, in a model of collagenase-induced ICH and in microglial cultures. METHODS: Mice subjected to collagenase-induced ICH injury were injected with either TO901317 (30 mg/kg) or vehicle 10 min after ICH and subsequently daily for 2 days. Behavioral studies, histology analysis, and assessments of hematoma volumes, brain water content, and blood-brain barrier (BBB) permeability were performed. The protein expression of LXR-α, LXR-ß, ATP binding cassette transporter-1 (ABCA-1), and inflammatory molecules was analyzed. The anti-inflammatory mechanism of TO901317 was investigated in cultured microglia that were stimulated with either lipopolysaccharide (LPS) or thrombin. RESULTS: ICH induced an increase in LXR-α protein levels in the hemorrhagic hemisphere at 6 h whereas LXR-ß expression remained unaffected. Both LXR-α and LXR-ß were expressed in neurons and microglia in the peri-ICH region and but rarely in astrocytes. TO901317 significantly attenuated functional deficits and brain damage up to 28 days post-ICH. TO901317 also reduced neuronal death, BBB disruption, and brain edema at day 4 post-ICH. These changes were associated with marked reductions in microglial activation, neutrophil infiltration, and expression levels of inflammatory mediators at 4 and 7 days. However, TO901317 had no effect on matrix metalloproteinase-9 activity. In BV2 microglial cultures, TO901317 attenuated LPS- and thrombin-stimulated nitric oxide production and reduced LPS-induced p38, JNK, MAPK, and nuclear factor-kappa B (NF-κB) signaling. Moreover, delaying administration of TO901317 to 3 h post-ICH reduced brain tissue damage and neuronal death. CONCLUSIONS: Our results suggest that enhancing LXR activation may provide a potential therapy for ICH by modulating the cytotoxic functions of microglia.

Berberine protects against neuronal damage via suppression of glia-mediated inflammation in traumatic brain injury.

Traumatic brain injury (TBI) triggers a series of neuroinflammatory processes that contribute to evolution of neuronal injury. The present study investigated the neuroprotective effects and anti-inflammatory actions of berberine, an isoquinoline alkaloid, in both in vitro and in vivo TBI models. Mice subjected to controlled cortical impact injury were injected with berberine (10 mg·kg(-1)) or vehicle 10 min after injury. In addition to behavioral studies and histology analysis, blood-brain barrier (BBB) permeability and brain water content were determined. Expression of PI3K/Akt and Erk signaling and inflammatory mediators were also analyzed. The protective effect of berberine was also investigated in cultured neurons either subjected to stretch injury or exposed to conditioned media with activated microglia. Berberine significantly attenuated functional deficits and brain damage associated with TBI up to day 28 post-injury. Berberine also reduced neuronal death, apoptosis, BBB permeability, and brain edema at day 1 post-injury. These changes coincided with a marked reduction in leukocyte infiltration, microglial activation, matrix metalloproteinase-9 activity, and expression of inflammatory mediators. Berberine had no effect on Akt or Erk 1/2 phosphorylation. In mixed glial cultures, berberine reduced TLR4/MyD88/NF-κB signaling. Berberine also attenuated neuronal death induced by microglial conditioned media; however, it did not directly protect cultured neurons subjected to stretch injury. Moreover, administration of berberine at 3 h post-injury also reduced TBI-induced neuronal damage, apoptosis and inflammation in vivo. Berberine reduces TBI-induced brain damage by limiting the production of inflammatory mediators by glial cells, rather than by a direct neuroprotective effect.

Post-injury treatment with 7,8-dihydroxyflavone, a TrkB receptor agonist, protects against experimental traumatic brain injury via PI3K/Akt signaling.

Tropomyosin-related kinase B (TrkB) signaling is critical for promoting neuronal survival following brain damage. The present study investigated the effects and underlying mechanisms of TrkB activation by the TrkB agonist 7,8-dihydroxyflavone (7,8-DHF) on traumatic brain injury (TBI). Mice subjected to controlled cortical impact received intraperitoneal 7,8-DHF or vehicle injection 10 min post-injury and subsequently daily for 3 days. Behavioral studies, histology analysis and brain water content assessment were performed. Levels of TrkB signaling-related molecules and apoptosis-related proteins were analyzed. The protective effect of 7,8-DHF was also investigated in primary neurons subjected to stretch injury. Treatment with 20 mg/kg 7,8-DHF attenuated functional deficits and brain damage up to post-injury day 28. 7,8-DHF also reduced brain edema, neuronal death, and apoptosis at day 4. These changes were accompanied by a significant decrease in cleaved caspase-3 and increase in Bcl-2/Bax ratio. 7,8-DHF enhanced phosphorylation of TrkB, Akt (Ser473/Thr308), and Bad at day 4, but had no effect on Erk 1/2 phosphorylation. Moreover, 7,8-DHF increased brain-derived neurotrophic factor levels and promoted cAMP response element-binding protein (CREB) activation. This beneficial effect was attenuated by inhibition of TrkB or PI3K/Akt. 7,8-DHF also promoted survival and reduced apoptosis in cortical neurons subjected to stretch injury. Remarkably, delayed administration of 7,8-DHF at 3 h post-injury reduced brain tissue damage. Our study demonstrates that activation of TrkB signaling by 7,8-DHF protects against TBI via the PI3K/Akt but not Erk pathway, and this protective effect may be amplified via the PI3K/Akt-CREB cascades.

Caveolin-1 interacts with Derlin-1 and promotes ubiquitination and degradation of cyclooxygenase-2 via collaboration with p97 complex.

Caveolin-1 (Cav-1) interacts with and mediates protein trafficking and various cellular functions. Derlin-1 is a candidate for the retrotranslocation channel of endoplasmic reticulum proteins. However, little is known about how Derlin-1 mediates glycosylated protein degradation. Here, we identified Cav-1 as a key player in Derlin-1- and p97-mediated cyclooxygenase 2 (COX-2) ubiquitination and degradation. Derlin-1 augmented the interaction of Cav-1 and COX-2 and mediated the degradation of COX-2 in a COX-2 C terminus-dependent manner. Suppression of Cav-1 decreased the ubiquitination of COX-2, and mutation of Asn-594 to Ala to disrupt N-glycosylation at the C terminus of COX-2 reduced the interaction of COX-2 with Cav-1 but not Derlin-1. Moreover, suppression of p97 increased the ubiquitination of COX-2 and up-regulated COX-2 but not COX-1. Cav-1 enhanced the interaction of p97 with Ufd1 and Derlin-1 and collaborated with p97 to interact with COX-2. Cav-1 may be a cofactor in the interaction of Derlin-1 and N-glycosylated COX-2 and may facilitate Derlin-1- and p97 complex-mediated COX-2 ubiquitination, retrotranslocation, and degradation.

Salidroside improves behavioral and histological outcomes and reduces apoptosis via PI3K/Akt signaling after experimental traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) induces a complex sequence of apopototic cascades that contribute to secondary tissue damage. The aim of this study was to investigate the effects of salidroside, a phenolic glycoside with potent anti-apoptotic properties, on behavioral and histological outcomes, brain edema, and apoptosis following experimental TBI and the possible involvement of the phosphoinositide 3-kinase/protein kinase B (PI3K)/Akt signaling pathway. METHODOLOGY/PRINCIPAL FINDINGS: Mice subjected to controlled cortical impact injury received intraperitoneal salidroside (20, or 50 mg/kg) or vehicle injection 10 min after injury. Behavioral studies, histology analysis and brain water content assessment were performed. Levels of PI3K/Akt signaling-related molecules, apoptosis-related proteins, cytochrome C (CytoC), and Smac/DIABLO were also analyzed. LY294002, a PI3K inhibitor, was administered to examine the mechanism of protection. The protective effect of salidroside was also investigated in primary cultured neurons subjected to stretch injury. Treatment with 20 mg/kg salidroside significantly improved functional recovery and reduced brain tissue damage up to post-injury day 28. Salidroside also significantly reduced neuronal death, apoptosis, and brain edema at day 1. These changes were associated with significant decreases in cleaved caspase-3, CytoC, and Smac/DIABLO at days 1 and 3. Salidroside increased phosphorylation of Akt on Ser473 and the mitochondrial Bcl-2/Bax ratio at day 1, and enhanced phosphorylation of Akt on Thr308 at day 3. This beneficial effect was abolished by pre-injection of LY294002. Moreover, delayed administration of salidroside at 3 or 6 h post-injury reduced neuronal damage at day 1. Salidroside treatment also decreased neuronal vulnerability to stretch-induced injury in vitro. CONCLUSIONS/SIGNIFICANCE: Post-injury salidroside improved long-term behavioral and histological outcomes and reduced brain edema and apoptosis following TBI, at least partially via the PI3K/Akt signaling pathway.