Neuroprotective Effect of Sonic Hedgehog Mediated PI3K/AKT Pathway in Amyotrophic Lateral Sclerosis Model Mice.

The Sonic Hedgehog (SHH) signaling pathway is related to the progression of various tumors and nervous system diseases. Still, its specific role in neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS), remains studied. This research investigates the role of SHH and PI3K/AKT signaling pathway proteins on ALS development in a SOD1-G93A transgenic mouse model. After injection of SHH and PI3K/AKT signaling pathway inhibitors or agonists in hSOD1-G93A (9 weeks of age) transgenic mice, we studied skeletal muscle pathology using immunohistochemical staining and Western blot methods. In addition, recorded data on rotation time, weight, and survival were analyzed for these mice. Our study showed that the expression of SHH, Gli-1 and p-AKT in ALS mice decreased with the progression of the disease. The expression of p-AKT changed together with Gli-1 while injecting PI3K/AKT signaling pathway inhibitor or agonist; SHH and Gli-1 protein expression remained unchanged; p-AKT protein expression significantly decreased while injecting PI3K/AKT signaling pathway inhibitor. These results indicate that SHH has a regulatory effect on PI3K/AKT signaling pathway. In behavioral experiments, we found that the survival time of hSOD1-G93A mice was prolonged by injection of SHH agonist while shortened by injection of SHH inhibitor. In conclusion, we confirmed that the SHH pathway played a neuroprotective role in ALS by mediating PI3K/AKT signaling pathway.

The Role of Sonic Hedgehog Pathway in the Development of the Central Nervous System and Aging-Related Neurodegenerative Diseases.

The Sonic hedgehog (SHH) pathway affects neurogenesis and neural patterning during the development of the central nervous system. Dysregulation of the SHH pathway in the brain contributes to aging-related neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. At present, the SHH signaling pathway can be divided into the canonical signaling pathway and non-canonical signaling pathway, which directly or indirectly mediates other related pathways involved in the development of neurodegenerative diseases. Hence, an in-depth knowledge of the SHH signaling pathway may open an avenue of possibilities for the treatment of neurodegenerative diseases. Here, we summarize the role and mechanism of the SHH signaling pathway in the development of the central nervous system and aging-related neurodegenerative diseases. In this review, we will also highlight the potential of the SHH pathway as a therapeutic target for treating neurodegenerative diseases.

Upregulation of heme oxygenase-1 protected against brain damage induced by transient cerebral ischemia-reperfusion injury in rats.

The aim of the present study was to identify the effect of heme oxygenase (HO)-1 gene on cerebral ischemia-reperfusion injury. Sprague-Dawley rats were divided randomly into four groups: Sham group, vehicle group, empty adenovirus vector (Ad) group and recombinant HO-1 adenovirus (Ad-HO-1) transfection group. Rats in the vehicle, Ad and Ad-HO-1 groups were respectively injected with saline, Ad or Ad-HO-1 for 3 days prior to cerebral ischemia-reperfusion injury. Subsequently, the middle cerebral artery occlusion method was used to establish the model of cerebral ischemia-reperfusion injury. Following the assessment of neurological function, rats were sacrificed, and the infarction volume and apoptotic index in rat brains were measured. Furthermore, the protein expression levels of HO-1 in brain tissues were detected using western blot analysis. Results indicated that the neurological score of the Ad-HO-1 group was significantly increased compared with the Ad or vehicle groups, respectively (P<0.001). The volume of cerebral infarction and the index score of neuronal apoptosis in the vehicle and Ad groups was significantly increased compared with the Ad-HO-1 group (P<0.01). The death of neuronal cells following cerebral ischemia-reperfusion injury reduced remarkably induced by over-expression of HO-1. These findings suggest a neuroprotective role of HO-1 against brain injury induced by transient cerebral ischemia-reperfusion injury.

NGF protects against oxygen and glucose deprivation-induced oxidative stress and apoptosis by up-regulation of HO-1 through MEK/ERK pathway.

Both nerve growth factor (NGF) and heme oxygenases-1 (HO-1) promotes neuron survival from cerebral ischemic lesions. NGF protects neurons from oxygen-glucose deprivation (OGD), and HO-1 expression can be induced by some growth factors like NGF. This work attempted to identify the contribution of HO-1 on the neuroprotection role of NGF in OGD model, which is an injury simulation of ischemic neuron in vitro. The viability of cortical neurons cells treated with OGD restored significantly by pretreatment with NGF in a dose dependent manner. Moreover, NGF provided obvious protective effects against OGD-induced neurons apoptosis. It identified that NGF could prevent apoptosis and ROS (reactive oxygen species) accumulation in the primary cortical neurons exposed to OGD. NGF could up-regulate the expression level of HO-1, and then afford neuroprotection against OGD insult. In addition, we found that MEK/ERK pathway participated NGF-induced over-expression of HO-1, and was involved in the transcriptional activity or neuroprotection effect of NGF.

Effect Comparison of Both Iron Chelators on Outcomes, Iron Deposit, and Iron Transporters After Intracerebral Hemorrhage in Rats.

Iron overload plays a key role in brain injury after intracerebral hemorrhage (ICH). We explored the roles of ferric iron chelator-deferiprone (DFP)-and ferrous iron chelator-clioquinol (CQ)-in ICH rats through the outcomes, iron deposits, reactive oxygen species (ROS), brain water content, and related iron transporters. One hundred eight Sprague-Dawley rats received intra-striatal infusions of 0.5 U of type IV collagenase to establish ICH models. The rats were randomly assigned to the sham, vehicle, DFP, and CQ groups. We evaluated the outcomes, iron levels, brain water content, and ROS; meanwhile, immunohistochemistry and real-time quantitative polymerase chain reaction (RT-qPCR) were utilized to determine ferritin, transferrin, transferrin receptor, divalent metal transport 1 (DMT1), and ferroportin at 48 and 72 h, 7 and 14 days after surgery. Our results showed ICH induced iron overload, brain edema, ROS formation, and neurological deficits. Both iron chelators decreased iron levels; CQ improved the neurological outcome, attenuated brain edema, and ROS production. DFP reduced iron contents but not brain water content and ROS generation. DFP failed to improve the outcome. ICH initiated endogenous iron chelators and transporters, both exogenous iron chelators enhanced expression of transferrin and transferrin receptor. CQ enhanced expression of ferroportin but not DMT1, while DFP enhanced expression of DMT1 but not ferroportin. Ferrous iron contributed to brain injury, and binding ferrous iron can modestly improve outcome after ICH in rats. The exogenous ferrous iron chelator possibly functioned via endogenous ferrous iron transporters on ICH. Therefore, ferrous iron may be a promising target for ICH in future.

Changes of ferrous iron and its transporters after intracerebral hemorrhage in rats.

OBJECTIVE: Ferrous iron is a major source inducing oxidative stress after intracerebral hemorrhage (ICH). Divalent metal transporter1 (DMT1) is the important and well-known plasma membrane transport protein which was proved to be involved in the transport of free ferrous iron in mammals. Ferroportin 1 (FPN1) is the unique exporter of ferrous iron from mammalian cells. The role of DMT1 and FPN1 in brain after ICH is still not elucidated. Therefore, we measure the expression of DMT1 and FPN1, to explore the correlations between ferrous iron and its specific transporters after ICH. METHODS: Ninety-six Sprague-Dawley rats received intra-striatal infusions of 0.5 U type IV collagenase to establish ICH model. Ferrous iron content in brain was determined using Turnbull's method. DMT1 and FPN1 expression were examined by immunohistochemical staining and Real-Time quantitative polymerase chain reaction (RT-PCR). With the use of confocal laser microscopy, we determined the colocalization of DMT1 and FPN1 at 1, 3, 7 and 14 days after ICH. RESULTS: Ferrous iron deposition was shown in the perihematomal zone as early as 1 day after ICH; it reached a peak after 7 days and was not elevated within 14 days following ICH. The expression of the DMT1 upregulated and reached to peak at day 7 after ICH. FPN1 reached a plateau at 3 days post-ICH. Expression levels of DMT1 and FPN1 were in parallel with ferrous iron deposition. There was a positive correlation between FPN1 and DMT1. DMT1 mainly localized in the cytoplasm of glias and neurons. FPN1 were mostly distributed on the membrane of endothelial cells and glias. Confocal microscope showed that DMT1 colocalized with FPN1. CONCLUSIONS: DMT1 and FPN1 are positively influenced by ferrous iron status in brain after ICH. DMT1 and FPN1 attenuate iron overload after ICH via increasing transmembrane iron export.

Time course of heme oxygenase-1 and oxidative stress after experimental intracerebral hemorrhage.

BACKGROUND: Heme oxygenase-1 (HO-1), the rate-limiting enzyme for heme catabolism and iron production, its role in intracerebral hemorrhage (ICH) is controversial. The study was to investigate correlations between brain oxidative injury and HO-1 after experimental ICH. METHOD: Sprague-Dawley rats received intra-striatal infusions of 100 µl autologous whole blood as ICH models. HO-1 were examined by immunohistochemical and reverse transcription polymerase chain reaction (RT-PCR) analysis. Brain oxidative stress was quantitated by malondialdehyde (MDA); antioxidation were measured by copper-zinc superoxide dismutase (Cu/Zn-SOD) activity using RT-PCR assay. RESULTS: The expression of the HO-1 upregulated and reached its peak at days 3 and 7 after ICH (P < 0.01). There was a significant increase of MDA and a top at 3-day post-ICH (P < 0.01); Cu/Zn-SOD was upregulated post-ICH and reached the top at day 7 (P < 0.001); HO-1 was correlated significantly with brain MDA content at days 7 and 14 following ICH (r = 0.435-0.501, P < 0.001) but there is no definite correlation between them on 1 to 3 days (P > 0.05); conversely, HO-1 was correlated significantly with Cu/Zn-SOD on 1 to 3 days after ICH (r = 0.433-0.621, P < 0.001) but there is no definite correlation between them at days 7 and 14 (P > 0.05). CONCLUSIONS: HO-1 has both antioxidant and prooxidant properties in ICH. The early upregulation of HO-1 possibly fit with the events and be protective against oxidative stress, whereas its overexpression in the late stages may result in its dysfunction and be toxic. So it should be prudent to intervene ICH with the inhibitor/activator of HO-1.