Huiying Yan

TLR4 inhibitor resatorvid provides neuroprotection in experimental traumatic brain injury: implication in the treatment of human brain injury.

Toll-like receptor 4 (TLR4) is considered to play an important role in neuronal death in animal models and could be an important therapeutic target following traumatic brain injury (TBI). Resatorvid is a small molecule, commonly accepted to inhibit TLR4-mediated pathway. The purpose of this study was to investigate the neuroprotective effect of resatorvid after TBI. Our data revealed that inhibition of TLR4 by resatorvid attenuated the development of TBI in mouse model. And we found that resatorvid administration dramatically reduced neuronal apoptosis. To investigate the cellular mechanism, we evaluated the expression of transforming growth factor-β-activated kinase 1 (TAK1), which plays a crucial role in TLR4 signal transduction pathway and is activated by phosphorylation in response to TBI. In addition, enzyme-linked immunosorbent assay was used to determine the expression of tumor necrosis factor-α (TNF-α) and interlukin-1β (IL-1β) at 24h after injury. Our results showed that resatorvid treatment significantly reduced the protein levels of TAK1, p-TAK1, TNF-α, and IL-1β compared with vehicle treatment. Importantly, the delayed therapy (4h post injury) once daily consecutively for 5days ameliorated brain damage and improved neurological recovery, suggesting that this drug has a wide therapeutic time window. Clinically, we observed that TLR4 and TAK1 expression was significantly increased in human contusion specimens after TBI. These data provide an experimental rationale for the evaluation of TLR4 as a clinical target and therapeutic implication of resatorvid in human traumatic brain injury.

Blockage of mitochondrial calcium uniporter prevents iron accumulation in a model of experimental subarachnoid hemorrhage

Previous studies have shown that iron accumulation is involved in the pathogenesis of brain injury following subarachnoid hemorrhage (SAH) and chelation of iron reduced mortality and oxidative DNA damage. We previously reported that blockage of mitochondrial calcium uniporter (MCU) provided benefit in the early brain injury after experimental SAH. This study was undertaken to identify whether blockage of MCU could ameliorate iron accumulation-associated brain injury following SAH. Therefore, we used two reagents ruthenium red (RR) and spermine (Sper) to inhibit MCU. Sprague-Dawley (SD) rats were randomly divided into four groups including sham, SAH, SAH+RR, and SAH+Sper. Biochemical analysis and histological assays were performed. The results confirmed the iron accumulation in temporal lobe after SAH. Interestingly, blockage of MCU dramatically reduced the iron accumulation in this area. The mechanism was revealed that inhibition of MCU reversed the down-regulation of iron regulatory protein (IRP) 1/2 and increase of ferritin. Iron-sulfur cluster dependent-aconitase activity was partially conserved when MCU was blocked. In consistence with this and previous report, ROS levels were notably reduced and ATP supply was rescued; levels of cleaved caspase-3 dropped; and integrity of neurons in temporal lobe was protected. Taken together, our results indicated that blockage of MCU could alleviate iron accumulation and the associated injury following SAH. These findings suggest that the alteration of calcium and iron homeostasis be coupled and MCU be considered to be a therapeutic target for patients suffering from SAH.

TGFβ-activated Kinase 1 (TAK1) Inhibition by 5Z-7-Oxozeaenol Attenuates Early Brain Injury after Experimental Subarachnoid Hemorrhage

Background: Role of TGFβ-activated kinase 1 (TAK1) in the pathogenesis of early brain injury after subarachnoid hemorrhage (SAH) has not been reported. Results: TAK1 inhibition attenuates early brain injury and improves neurological deficits after SAH. Conclusion: TAK1 inhibition exhibits neuro-protective effects possibly through anti-apoptotic function. Significance: These results provide a novel target for SAH treatment. Accumulating evidence suggests that activation of mitogen-activated protein kinases (MAPKs) and nuclear factor NF-κB exacerbates early brain injury (EBI) following subarachnoid hemorrhage (SAH) by provoking proapoptotic and proinflammatory cellular signaling. Here we evaluate the role of TGFβ-activated kinase 1 (TAK1), a critical regulator of the NF-κB and MAPK pathways, in early brain injury following SAH. Although the expression level of TAK1 did not present significant alternation in the basal temporal lobe after SAH, the expression of phosphorylated TAK1 (Thr-187, p-TAK1) showed a substantial increase 24 h post-SAH. Intracerebroventricular injection of a selective TAK1 inhibitor (10 min post-SAH), 5Z-7-oxozeaenol (OZ), significantly reduced the levels of TAK1 and p-TAK1 at 24 h post-SAH. Involvement of MAPKs and NF-κB signaling pathways was revealed that OZ inhibited SAH-induced phosphorylation of p38 and JNK, the nuclear translocation of NF-κB p65, and degradation of IκBα. Furthermore, OZ administration diminished the SAH-induced apoptosis and EBI. As a result, neurological deficits caused by SAH were reversed. Our findings suggest that TAK1 inhibition confers marked neuroprotection against EBI following SAH. Therefore, TAK1 might be a promising new molecular target for the treatment of SAH.

Akt Specific Activator SC79 Protects against Early Brain Injury following Subarachnoid Hemorrhage.

A growing body of evidence demonstrates that Akt may serve as a therapeutic target for treatment of early brain injury following subarachnoid hemorrhage (SAH). The purpose of the current study was to evaluate the neuroprotective effect of Akt specific activator SC79 in an experimental rat model of SAH. SAH was induced by injecting 300 μL of blood into the prechiasmatic cistern. Intracerebroventricular (ICV) injection of SC79 (30 min post-SAH) induced the p-Akt (Ser473) expression in a dose-dependent manner. A single ICV dose treatment of SC79 (100 μg/rat) significantly increased the expression of Bcl-2 and p-GSK-3β (Ser9), decreased the protein levels of Bax, cytoplasm cytochrome c, and cleaved caspase-3, indicating the antiapoptotic effect of SC79. As a result, the number of apoptotic cells was reduced 24 h post SAH. Moreover, SC79 treatment alleviated SAH-induced oxidative stress, restored mitochondrial morphology, and improved neurological deficits. Strikingly, treatment of SC79 provided a beneficial outcome against neurologic deficit with a therapeutic window of at least 4 h post SAH by ICV injection and 30 min post SAH by intraperitoneal injection. Collectively, SC79 exerts its neuroprotective effect likely through the dual activities of antioxidation and antiapoptosis. These data provide a basic platform to consider SC79 as a novel therapeutic agent for treatment of SAH.

Inhibition of myeloid differentiation factor 88(MyD88) by ST2825 provides neuroprotection after experimental traumatic brain injury in mice.

Myeloid differentiation factor 88(MyD88) is an endogenous adaptor protein that plays an important role in coordinating intracellular inflammatory responses induced by agonists of the Toll-like receptor and interleukin-1 receptor families. MyD88 has been reported to be essential for neuronal death in animal models and may represent a therapeutic target for pharmacologic inhibition following traumatic brain injury (TBI). The purpose of the current study was to investigate the neuroprotective effect of MyD88 specific inhibitor ST2825 in an experimental mouse model of TBI. Intracerebroventricular (ICV) injection of high concentration (20μg/μL) ST2825 (15min post TBI) attenuated the development of TBI in mice, markedly improved neurological function and reduced brain edema. Decreased neural apoptosis and increased neuronal survival were also observed. Biochemically, the high concentration of ST2825 significantly reduced the levels of MyD88, further decreased TAK1, p-TAK1, nuclear p65 and increased IκB-α. Additionally, ST2825 significantly reduced the levels of Iba-1 and inflammatory factors TNF-α and IL-1β. These data provide an experimental rationale for evaluation of MyD88 as a drug target and highlight the potential therapeutic implications of ST2825 in TBI.

Increased Expression of NLRP3 Inflammasome in Wall of Ruptured and Unruptured Human Cerebral Aneurysms: Preliminary Results.

BACKGROUND A growing body of evidence suggests that inflammation actively participates in cerebral aneurysm initiation, progression, and rupture. The primary objective of this study was to assess the expression of NLR family, Pyrin-domain containing 3 (NLRP3) inflammasome in human cerebral aneurysms. METHODS Aneurysmal domes (19 ruptured and 17 unruptured) from patients undergoing surgical treatment for ruptured or unruptured cerebral aneurysms were analyzed. A control sample comprising 4 middle cerebral arteries was obtained from autopsy subjects. The expression of NLRP3, apoptotic speck-containing protein with a card (ASC), caspase-1, and interleukin (IL)-1β were assessed by immunohistochemistry. Immunofluorescence double staining was used to determine NLRP3, ASC, and caspase-1 cellular distribution. RESULTS Expression of NLRP3, ASC, and caspase-1 were more abundant in ruptured aneurysm tissue than that in unruptured aneurysms, based on a semi-quantitative grading (P < .05). IL-1β was also overexpressed in the ruptured cerebral aneurysms and associated with increased expression of NLRP3, ASC, and caspase-1 (P < .05). Furthermore, NLRP3, ASC, and caspase-1 immunoreactivity were colocalized with immunoreactivity of CD3 in T lymphocytes and CD68 in macrophages. CONCLUSIONS NLRP3 inflammasome was expressed in the wall of human cerebral aneurysms and was more abundant in ruptured aneurysms than in unruptured. This study raises the possibility that NLRP3 inflammasome may be involved in the pathogenesis of human intracranial aneurysms, and this requires further study.

Inhibition of myeloid differentiation primary response protein 88 provides neuroprotection in early brain injury following experimental subarachnoid hemorrhage

Accumulating of evidence suggests that activation of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPKs) exacerbates early brain injury (EBI) following subarachnoid hemorrhage (SAH) by provoking pro-inflammatory and pro-apoptotic signaling. Myeloid differentiation primary response protein 88 (MyD88) is an endogenous adaptor protein in the toll-like receptors (TLRs) and interleukin (IL) -1β family signaling pathways and acts as a bottle neck in the NF-κB and MAPK pathways. Here, we used ST2825, a selective inhibitor of MyD88, to clarify whether inhibiting MyD88 could provide neuroprotection in EBI following SAH. Our results showed that the expression of MyD88 was markedly increased at 24 h post SAH. Intracerebroventricular injection of ST2825 significantly reduced the expression of MyD88 at 24 h post SAH. Involvement of MAPKs and NF-κB signaling pathways was revealed that ST2825 inhibited SAH-induced phosphorylation of TAK1, p38 and JNK, the nuclear translocation of NF-κB p65, and degradation of IκBα. Further, ST2825 administration diminished the SAH-induced inflammatory response and apoptosis. As a result, SAH-induced EBI was alleviated and neurological deficits caused by SAH were reversed. Our findings suggest that MyD88 inhibition confers marked neuroprotection against EBI following SAH. Therefore, MyD88 might be a promising new molecular target for the treatment of SAH.

Biphasic activation of nuclear factor-κB and expression of p65 and c-Rel following traumatic neuronal injury

The transcription factor nuclear factor-κB (NF-κB) has been shown to function as a key regulator of cell death or survival in neuronal cells. Previous studies indicate that the biphasic activation of NF-κB occurs following experimental neonatal hypoxia-ischemia and subarachnoid hemorrhage. However, the comprehensive understanding of NF-κB activity following traumatic brain injury (TBI) is incomplete. In the current study, an in vitro model of TBI was designed to investigate the NF-κB activity and expression of p65 and c-Rel subunits following traumatic neuronal injury. Primary cultured neurons were assigned to control and transected groups. NF-κB activity was detected by electrophoretic mobility shift assay. Western blotting and immunofluorescence were used to investigate the expression and distribution of p65 and c-Rel. Reverse transcription-quantitative polymerase chain reaction was performed to assess the downstream genes of NF-κB. Lactate dehydrogenase (LDH) quantification and trypan blue staining were used to estimate the neuronal injury. Double peaks of elevated NF-κB activity were observed at 1 and 24 h following transection. The expression levels of downstream genes exhibited similar changes. The protein levels of p65 also presented double peaks while c-Rel was elevated significantly in the late stage. The results of the trypan blue staining and LDH leakage assays indicated there was no sustained neuronal injury during the late peak of NF-κB activity. In conclusion, biphasic activation of NF-κB is induced following experimental traumatic neuronal injury. The elevation of p65 and c-Rel levels at different time periods suggests that within a single neuron, NF-κB may participate in different pathophysiological processes.

Expression of the long non-coding RNA H19 and MALAT-1 in growth hormone-secreting pituitary adenomas and its relationship to tumor behavior

Aggressive growth hormone‐secreting pituitary adenomas (GHPAs) represent an important clinical problem in patients with acromegaly. Surgical therapy, although often the mainstay of treatment for GHPAs, is less effective in aggressive GHPAs due to their invasive and destructive growth patterns, and their proclivity for infrasellar invasion. LncRNAs are important players in cancer development and emerging in various fundamental biological processes. In the present study, qRT‐PCR was performed to examine the expression of lncRNA H19 and MALAT‐1 in invasive and non‐invasive GHPAs. Our results revealed that the expression of lncRNA H19 was remarkably higher in invasive GHPAs, however, there was no significant differences between the expression of lncRNA MALAT‐1 in invasive GHPAs and non‐invasive GHPAs, suggesting that lncRNA H19 may play an important role in GHPA invasion. LncRNA H19 might be a target for the study of GHPAs invasion, and a potential index for the diagnosis or prognosis of GHPAs.