Human-induced pluripotent stem cell-derived neural stem cell exosomes improve blood-brain barrier function after intracerebral hemorrhage by activating astrocytes via PI3K/AKT/MCP-1 axis

Cerebral edema caused by blood-brain barrier injury after intracerebral hemorrhage is an important factor leading to poor prognosis.Human-induced pluripotent stem cell-derived neural stem cell exosomes(hiPSC-NSC-Exos)have shown potential for brain injury repair in central nervous system diseases.In this study,we explored the impact of hiPSC-NSC-Exos on blood-brain barrier preservation and the underlying mechanism.Our results indicated that intranasal delivery of hiPSC-NSC-Exos mitigated neurological deficits,enhanced blood-brain barrier integrity,and reduced leukocyte infiltration in a mouse model of intracerebral hemorrhage.Additionally,hiPSC-NSC-Exos decreased immune cell infiltration,activated astrocytes,and decreased the secretion of inflammatory cytokines like monocyte chemoattractant protein-1,macrophage inflammatory protein-1α,and tumor necrosis factor-αpost-intracerebral hemorrhage,thereby improving the inflammatory microenvironment.RNA sequencing indicated that hiPSC-NSC-Exo activated the PI3K/AKT signaling pathway in astrocytes and decreased monocyte chemoattractant protein-1 secretion,thereby improving blood-brain barrier integrity.Treatment with the PI3K/AKT inhibitor LY294002 or the monocyte chemoattractant protein-1 neutralizing agent C1142 abolished these effects.In summary,our findings suggest that hiPSC-NSC-Exos maintains blood-brain barrier integrity,in part by downregulating monocyte chemoattractant protein-1 secretion through activation of the PI3K/AKT signaling pathway in astrocytes.

Cholesterol metabolism: physiological versus pathological aspects in intracerebral hemorrhage

Cholesterol is an important component of plasma membranes and participates in many basic life functions,such as the maintenance of cell membrane stability,the synthesis of steroid hormones,and myelination.Cholesterol plays a key role in the establishment and maintenance of the central nervous system.The brain contains 20%of the whole body’s cholesterol,80%of which is located within myelin.A huge number of processes(e.g.,the sterol regulatory element-binding protein pathway and liver X receptor pathway)participate in the regulation of cholesterol metabolism in the brain via mechanisms that include cholesterol biosynthesis,intracellular transport,and efflux.Certain brain injuries or diseases involving crosstalk among the processes above can affect normal cholesterol metabolism to induce detrimental consequences.Therefore,we hypothesized that cholesterol-related molecules and pathways can serve as therapeutic targets for central nervous system diseases.Intracerebral hemorrhage is the most severe hemorrhagic stroke subtype,with high mortality and morbidity.Historical cholesterol levels are associated with the risk of intracerebral hemorrhage.Moreover,secondary pathological changes after intracerebral hemorrhage are associated with cholesterol metabolism dysregulation,such as neuroinflammation,demyelination,and multiple types of programmed cell death.Intracellular cholesterol accumulation in the brain has been found after intracerebral hemorrhage.In this paper,we review normal cholesterol metabolism in the central nervous system,the mechanisms known to participate in the disturbance of cholesterol metabolism after intracerebral hemorrhage,and the links between cholesterol metabolism and cell death.We also review several possible and constructive therapeutic targets identified based on cholesterol metabolism to provide cholesterol-based perspectives and a reference for those interested in the treatment of intracerebral hemorrhage.

Mitophagy in intracerebral hemorrhage:a new target for therapeutic intervention

Intracerebral hemorrhage is a life-threatening condition with a high fatality rate and severe sequelae.However,there is currently no treatment available for intracerebral hemorrhage,unlike for other stroke subtypes.Recent studies have indicated that mitochondrial dysfunction and mitophagy likely relate to the pathophysiology of intracerebral hemorrhage.Mitophagy,or selective autophagy of mitochondria,is an essential pathway to preserve mitochondrial homeostasis by clearing up damaged mitochondria.Mitophagy markedly contributes to the reduction of secondary brain injury caused by mitochondrial dysfunction after intracerebral hemorrhage.This review provides an overview of the mitochondrial dysfunction that occurs after intracerebral hemorrhage and the underlying mechanisms regarding how mitophagy regulates it,and discusses the new direction of therapeutic strategies targeting mitophagy for intracerebral hemorrhage,aiming to determine the close connection between mitophagy and intracerebral hemorrhage and identify new therapies to modulate mitophagy after intracerebral hemorrhage.In conclusion,although only a small number of drugs modulating mitophagy in intracerebral hemorrhage have been found thus far,most of which are in the preclinical stage and require further investigation,mitophagy is still a very valid and promising therapeutic target for intracerebral hemorrhage in the long run.

Spi1 regulates the microglial/macrophage inflammatory response via the PI3K/AKT/mTOR signaling pathway after intracerebral hemorrhage

Preclinical and clinical studies have shown that microglia and macrophages participate in a multiphasic brain damage repair process following intracerebral hemorrhage.The E26 transformation-specific sequence-related transcription factor Spi1 regulates microglial/macrophage commitment and maturation.However,the effect of Spi1 on intracerebral hemorrhage remains unclear.In this study,we found that Spi1 may regulate recovery from the neuroinflammation and neurofunctional damage caused by intracerebral hemorrhage by modulating the microglial/macrophage transcriptome.We showed that high Spi1expression in microglia/macrophages after intracerebral hemorrhage is associated with the activation of many pathways that promote phagocytosis,glycolysis,and autophagy,as well as debris clearance and sustained remyelination.Notably,microglia with higher levels of Soil expression were chara cterized by activation of pathways associated with a variety of hemorrhage-related cellular processes,such as complement activation,angiogenesis,and coagulation.In conclusion,our results suggest that Spi1 plays a vital role in the microglial/macrophage inflammatory response following intracerebral hemorrhage.This new insight into the regulation of Spi1 and its target genes may advance our understanding of neuroinflammation in intracerebral hemorrhage and provide therapeutic targets for patients with intracerebral hemorrhage.

ATAT1 deficiency enhances microglia/macrophage-mediated erythrophagocytosis and hematoma absorption following intracerebral hemorrhage

MIcroglia/macrophage-mediated erythrophagocytosis plays a crucial role in hematoma clearance after intracerebral hemorrhage.Dynamic cytoskeletal changes accompany phagocytosis.However,whether and how these changes are associated with microglia/macrophage-mediated erythrophagocytosis remain unclear.In this study,we investigated the function of acetylatedα-tubulin,a stabilized microtubule form,in microglia/macrophage erythrophagocytosis after intracerebral hemorrhage both in vitro and in vivo.We first assessed the function of acetylatedα-tubulin in erythrophagocytosis using primary DiO GFP-labeled red blood cells co-cultured with the BV2 microglia or RAW264.7 macrophage cell lines.Acetylatedα-tubulin expression was significantly decreased in BV2 and RAW264.7 cells during erythrophagocytosis.Moreover,silencingα-tubulin acetyltransferase 1(ATAT1),a newly discoveredα-tubulin acetyltransferase,decreased Ac-α-tub levels and enhanced the erythrophagocytosis by BV2 and RAW264.7 cells.Consistent with these findings,in ATAT1-/-mice,we observed increased ionized calcium binding adapter molecule 1(Iba1)and Perls-positive microglia/macrophage phagocytes of red blood cells in peri-hematoma and reduced hematoma volume in mice with intracerebral hemorrhage.Additionally,knocking out ATAT1 alleviated neuronal apoptosis and pro-inflammatory cytokines and increased anti-inflammatory cytokines around the hematoma,ultimately improving neurological recovery of mice after intracerebral hemorrhage.These findings suggest that ATAT1 deficiency accelerates erythrophagocytosis by microglia/macrophages and hematoma absorption after intracerebral hemorrhage.These results provide novel insights into the mechanisms of hematoma clearance and suggest ATAT1 as a potential target for the treatment of intracerebral hemorrhage.

Correlation of free radical level and apoptosis after intracerebral hemorrhage in rats

Objective To investigate the correlation of perihematomal free radical level and neuronal apoptosis following the intracerebral hemorrhage （ICH）. Methods Animals were randomly divided into 4 groups： sham operation group, model group, 1 mg/kg edaravone group, and 3 mg/kg edaravone group. Each group was then divided into seven subgroups, in which the rats were correspondingly killed at 6 h, 12 h, 24 h, 48 h, 72 h, 7 d or 14 d （n = 1 in each subgroup of the sham group, and n = 6 in each subgroup of the other 3 groups）. By Horseley-Clarke technique, autoblood （80 μL） were administered into the left caudate putamen of SD rats in a double administration-withdrawal way. Rats in the sham group were needled in but not administered with autoblood. The ICH model was then evaluated by Bederson＇s scale. Around the hematoma, the levels of malonaldehyde （MDA） and hydroxyl radical were tested by spectrophotometer, and the process of apoptosis was tested by terminal deoxynucleofidyltransferase-mediated dUTP-biotin nick end labeling （TUNEL） method. Results （1） ICH significantly increased the levels of MDA and hydroxyl radicals. Significant differences in MDA and hydroxyl radical contents were observed among the four groups. （2） In the sham group, a small number of TUNEL-positive cells were found. In the other three groups, the TUNEL-positive cells were observed at 6 h, increased significantly at 24 h, and reached peak level at 3 d, then fell profoundly at 7 d, but remained detectable at 14 d. （3） The positive correlation existed between apoptosis and free radical level （r = 0.2003）, and existed between apoptosis and MDA content （r = 0.6563） in the brain. Conclusion Post-hemorrhagic apoptosis was related to the production of free radicals, indicating that the elevated free radicals following the ICH could induce neuron and glial cell apoptosis.

Mitophagy in acute central nervous system injuries:regulatory mechanisms and therapeutic potentials

Acute central nervous system injuries,including ischemic stro ke,intracerebral hemorrhage,subarachnoid hemorrhage,traumatic brain injury,and spinal co rd injury,are a major global health challenge.Identifying optimal therapies and improving the long-term neurological functions of patients with acute central nervous system injuries are urgent priorities.Mitochondria are susceptible to damage after acute central nervous system injury,and this leads to the release of toxic levels of reactive oxygen species,which induce cell death.Mitophagy,a selective form of autophagy,is crucial in eliminating redundant or damaged mitochondria during these events.Recent evidence has highlighted the significant role of mitophagy in acute central nervous system injuries.In this review,we provide a comprehensive overview of the process,classification,and related mechanisms of mitophagy.We also highlight the recent developments in research into the role of mitophagy in various acute central nervous system injuries and drug therapies that regulate mitophagy.In the final section of this review,we emphasize the potential for treating these disorders by focusing on mitophagy and suggest future research paths in this area.

Epigenetic regulation of the inflammatory response in stroke

Stroke is classified as ischemic or hemorrhagic,and there are few effective treatments for either type.Immunologic mechanisms play a critical role in secondary brain injury following a stroke,which manifests as cytokine release,blood–brain barrier disruption,neuronal cell death,and ultimately behavioral impairment.Suppressing the inflammatory response has been shown to mitigate this cascade of events in experimental stroke models.However,in clinical trials of anti-inflammatory agents,longterm immunosuppression has not demonstrated significant clinical benefits for patients.This may be attributable to the dichotomous roles of inflammation in both tissue injury and repair,as well as the complex pathophysiologic inflammatory processes in stroke.Inhibiting acute harmful inflammatory responses or inducing a phenotypic shift from a pro-inflammatory to an anti-inflammatory state at specific time points after a stroke are alternative and promising therapeutic strategies.Identifying agents that can modulate inflammation requires a detailed understanding of the inflammatory processes of stroke.Furthermore,epigenetic reprogramming plays a crucial role in modulating post-stroke inflammation and can potentially be exploited for stroke management.In this review,we summarize current findings on the epigenetic regulation of the inflammatory response in stroke,focusing on key signaling pathways including nuclear factor-kappa B,Janus kinase/signal transducer and activator of transcription,and mitogen-activated protein kinase as well as inflammasome activation.We also discuss promising molecular targets for stroke treatment.The evidence to date indicates that therapeutic targeting of the epigenetic regulation of inflammation can shift the balance from inflammation-induced tissue injury to repair following stroke,leading to improved post-stroke outcomes.

Influence of Hypertension, Lipometabolism Disorders,Obesity and Other Lifestyles on SpontaneousIntracerebral Hemorrhage

Objective To investigate whether hypertension, abnormal lipometabolism, obesity, cigarette smoking and alcohol drinking affect the intracerebral hemorrhagic volumes (IHV) in patients with spontaneous intracerebral hemorrhage (SIHP), and to explore the roles of these factors in spontaneous intracerebral hemorrhage (SIH). Methods Five hundred patients with acute SIH and 200 healthy adult volunteers (HAV) were enrolled in a study of independently randomized controlled design, in which the levels of systolic pressure (SP) and diastolic pressure (DP), and total cholesterol (TCH), triacylglycerols (triglycerides, TG), high density lipoprotein cholesterol (HDL-CH), low density lipoprotein cholesterol (LDL-CH) in serum as well as the level of erythrocytic membrane cholesterol (EM-CH) were measured, and the body mass index (BMI), daily cigarette smoking consumption (DCSC) and daily pure alcohol consumption (DPAC) were calculated. Results Compared with the average parameters in the HAV group, those of SP, DP, TG, LDL-CH and BMI in the SIHP group were significantly increased (P<0.0001), while those of HDL-CH and EM-CH were significantly decreased (P<0.0001). The linear regression and correlation analysis showed that with increased SP, DP, LDL-CH, BMI, DCSC, DPAC and aging as well as decreased HDL-CH and EM-CH, the IHV levels in SIHP were increased gradually (P<0.0001-0.01). The linear stepwise regression analysis suggested that there existed a close correlation among the values of SP, DP, TCH, TG, HDL-CH, LDL-CH, EM-CH, BMI, DCSC, DPAC, age and fflV of the SIH patients, and that Y = - 12.4583 + 0.1127SP -1.1977EM-CH + 0.9788LDL-CH + 0.2477BMI + 0.0382DCSC + 0.0248DP,P<0.0001-0.05. Conclusions The findings in the present study suggest that significantly increased systolic and diastolic pressure, low density lipoprotein cholesterol, body mass index and daily cigarette smoking consumption, and significantly decreased erythrocytic membrane cholesterol may be likely the main factors affecting intracerebral hemorrhagic volumes in patients with acute spontaneous intracerebral hemorrhage.

Acupuncture through Baihui (DU20) to Qubin (GB7) mitigates neurological impairment after intracerebral hemorrhage

Inflammation plays an important role in nerve defects caused by intracerebral hemorrhage. Repairing brain damage by inhibiting the macrophage-inducible C-type lectin/spleen tyrosine kinase （Mincle/Syk） signaling pathway is a potential new target for treating cerebral hemorrhage. In this study, we aimed to determine whether acupuncture through Baihui （DU20） to Qubin （GBT） is an effective treatment for intracerebral hemorrhage through the Mincle/Syk signaling pathway. An intracerebral hemorrhage rat model was established by autol- ogous blood infusion into the caudate nucleus. Acupuncture through Baihui to Qubin was performed for 30 minutes, once every 12 hours, for a total of three times. Piceatannol （34.62 mg/kg）, a Syk inhibitor, was intraperitoneally injected as a control. Modified neurological severity score was used to assess neurological function. Brain water content was measured. Immunohistochemistry and western blot assay were used to detect immunoreactivity and protein expression levels of Minde, Syk, and CARD9. Real-time polymerase chain reaction was used to determine interleukin-1[~ mRNA levels. Hematoxylin-eosin staining was performed to observe histopathological changes. Our re- suits showed that acupuncture through Baihui to Qubin remarkably improved neurological function and brain water content, and inhibited immunoreactivity and expression of Mincle, Syk, CARDg, and interkeukin-1β Moreover, this effect was similar to piceatannol. These find- ings suggest that acupuncture through Baihui to Qubin can improve neurological impairment after cerebral hemorrhage by inhibiting the Mincle/Syk signaling pathway.

Combined Transplantation of Neural Stem Cells and Olfactory Ensheathing Cells Improves the Motor Function of Rats with Intracerebral Hemorrhage

Objective To investigate the effects of combined transplantation of neural stem cells （NSC） and olfactory ensheathing cells （OEC） on the motor function of rats with intracerebral hemorrhage. Methods In three days after a rat model of caudate nucleus hemorrhage was established, NSCs and OEC, NSC, OEC （from embryos of Wistar rats） or normal saline were injected into bematomas of rats in combined transplantation group, NSC group, OEC group, and control group, respectively. Damage of neural function was scored before and in 3, 7, 14, 30 days after operation. Tissue after transplantation was observed by immunocytochemistry staining. Results The scores for the NSC, OEC and co-transplantation groups were significantly lower in 14 and 30 days after operation than in 3 days after operation （P〈0.05）. The scores for the NSC and OEC groups were significantly lower than those for the control group only in 30 days after operation （P〈0.05）, while the difference for the NSC-OEC group was significant in 14 days after operation （P〈0.05）. Immunocytochemistry staining revealed that the transplanted OEC and NSC could survive, migrate and differentiate into neurons, astrocytes, and oligodendrocytes. The number of neural precursor cells was greater in the NSC and combined transplantation groups than in the control group. The number of neurons differentiated from NSC was significantly greater in the co-transplantation group than in the NSC group. Conclusion Co-transplantation of NSC and OEC can promote the repair of injured tissue and improve the motor fimction of rats with intracerebral hemorrhage.

Neurotoxic role of interleukin-17 in neural stem cell differentiation after intracerebral hemorrhage

Interleukin 17(IL-17)and its main producer,T cell receptorγδcells,have neurotoxic effects in the pathogenesis of intracerebral hemorrhage(ICH),aggravating brain injuries.To investigate the correlation between IL-17 and ICH,we dynamically screened serum IL-17 concentrations using enzyme-linked immunosorbent assay and explored the clinical values of IL-17 in ICH patients.There was a significant negative correlation between serum IL-17 level and neurological recovery status in ICH patients(r=–0.498,P<0.01).To study the neurotoxic role of IL-17,C57 BL/6 mice were used to establish an ICH model by injecting autologous blood into the caudate nucleus.Subsequently,the mice were treated with mouse neural stem cells(NSCs)and/or IL-17 neutralizing antibody for 72 hours.Flow cytometry,brain water content detection,Nissl staining,and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling results indicated that NSC transplantation significantly reduced IL-17 expression in peri-hematoma tissue,but there was no difference in T cell receptorγδcells.Compared with the ICH group,there were fewer apoptotic bodies and more Nissl bodies in the ICH+NSC group and the ICH+NSC+IL-17 group.To investigate the potential effect of IL-17 on directional differentiation of NSCs,we cultured mouse NSCs(NE-4 C)alone or co-cultured them with T cell receptorγδcells,which were isolated from mouse peripheral blood mononuclear cells,for 7 days.The results of western blot assays revealed that IL-17 secreted by T cell receptorγδcells reduced the differentiation of NSCs into astrocytes and neurons,while IL-17 neutralization relieved the inhibition of directional differentiation into astrocytes rather than neurons.In conclusion,serum IL-17 levels were elevated in the early stage of ICH and were negatively correlated with outcome in ICH patients.Animal experiments and cytological investigations therefore demonstrated that IL-17 probably has neurotoxic roles in ICH because of its inhibitory effects on the directional differentiation of NSCs.The application of IL-17 neutralizing antibody may promote the directional differentiation of NSCs into astrocytes.This study was approved by the Clinical Research Ethics Committee of Anhui Medical University of China(For human study:Approval No.20170135)in December 2016.All animal handling and experimentation were reviewed and approved by the Institutional Animal Care and Use Committee of Anhui Medical University(approval No.20180248)in December 2017.

MicroRNA-181c provides neuroprotection in an intracerebral hemorrhage model

Apoptosis is an important factor during the early stage of intracerebral hemorrhage.MiR-181 c plays a key regulatory role in apoptosis.However,whether miR-181 c is involved in apoptosis of prophase cells after intracerebral hemorrhage remains unclear.Therefore,in vitro and in vivo experiments were conducted to test this hypothesis.In vivo experiments:collagenase type VII was injected into the basal ganglia of adult Sprague-Dawley rats to establish an intracerebral hemorrhage model.MiR-181 c mimic or inhibitor was injected in situ 4 hours after intracerebral hemorrhage.Neurological functional defects(neurological severity scores)were assessed 1,7,and 14 days after model establishment.Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling and western blot assay were conducted 14 days after model establishment.In vitro experiments:PC12 cells were cultured under oxygen-glucose deprivation,and hemins were added to simulate intracerebral hemorrhage in vitro.MiR-181 c mimic or inhibitor was added to regulate miR-181 c expression.3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay,luciferase reporter system,and western blot assay were performed.Experimental results revealed differences in miR-181 c expression in brain tissues of both patients and rats with cerebral hemorrhage.In addition,in vitro experiments found that miR-181 c overexpression could upregulate the Bcl-2/Bax ratio to inhibit apoptosis,while inhibition of miR-181 c expression could reduce the Bcl-2/Bax ratio and aggravate apoptosis of cells.Regulation of apoptosis occurred through the phosphoinositide 3 kinase(PI3 K)/Akt pathway by targeting of phosphatase and tensin homolog deleted on chromosome ten(PTEN).Higher miR-181 c overexpression correlated with lower neurological severity scores,indicating better recovery of neurological function.In conclusion,miR-181 c affects the prognosis of intracerebral hemorrhage by regulating apoptosis,and these effects might be directly mediated and regulated by targeting of the PTEN\PI3 K/Akt pathway and Bcl-2/Bax ratio.Furthermore,these results indicated that miR-181 c played a neuroprotective role in intracerebral hemorrhage by regulating apoptosis of nerve cells,thus providing a potential target for the prevention and treatment of intracerebral hemorrhage.Testing of human serum was authorized by the Ethics Committee of China Medical University(No.2012-38-1)on February 20,2012.The protocol was registered with the Chinese Clinical Trial Registry(Registration No.ChiCTR-COC-17013559).The animal study was approved by the Institutional Animal Care and Use Committee of China Medical University(approval No.2017008)on March 8,2017.

Relationship between different surgical methods,hemorrhage position,hemorrhage volume,surgical timing,and treatment outcome of hypertensive intracerebral hemorrhage

BACKGROUND:The present study aimed to explore the relationship between surgical methods,hemorrhage position,hemorrhage volume,surgical timing and treatment outcome of hypertensive intracerebral hemorrhage(HICH).METHODS:A total of 1 310 patients,who had been admitted to six hospitals from January 2004 to January 2008,were divided into six groups according to different surgical methods:craniotomy through bone fl ap(group A),craniotomy through a small bone window(group B),stereotactic drilling drainage(group C1 and group C2),neuron-endoscopy operation(group D) and external ventricular drainage(group E) in consideration of hemorrhage position,hemorrhage volume and clinical practice. A retrospective analysis was made of surgical timing and curative effect of the surgical methods.RESULTS:The effectiveness rate of the methods was 74.12% for 1 310 patients after onemonth follow-up. In this series,the disability rate was 44.82% 3–6 months after the operation. Among the 1 310 patients,241(18.40%) patients died after the operation. If hematoma volume was >80 mL and the operation was performed within 3 hours,the mortality rate of group A was signifi cantly lower than that of groups B,C,D,and E(P<0.05). If hematoma volume was 50–80 mL and the operation was performed within 6–12 hours,the mortality rate of groups B and D was lower than that of groups A,C and E(P<0.05). If hematoma volume was 20–50 mL and the operation was performed within 6–24 hours,the mortality rate of group C was lower than that of groups A,B and D(P<0.05).CONCLUSIONS:Craniotomy through a bone f lap is suitable for patients with a large hematoma and hernia of the brain. Stereotactic drilling drainage is suggested for patients with hematoma volume less than 80 mL. The curative effect of HICH individualized treatment would be improved via the suitable selection of operation time and surgical method according to the position and volume of hemorrhage.

Intranasal insulin ameliorates neurological impairment after intracerebral hemorrhage in mice

In Alzheimer’s disease and ischemic stroke,intranasal insulin can act as a neuroprotective agent.However,whether intranasal insulin has a neuroprotective effect in intracerebral hemorrhage and its potential mechanisms remain poorly understood.In this study,a mouse model of autologous blood-induced intracerebral hemorrhage was treated with 0.5,1,or 2 IU insulin via intranasal delivery,twice per day,until 24 or 72 hours after surgery.Compared with saline treatment,1 IU intranasal insulin treatment significantly reduced hematoma volume and brain edema after cerebral hemorrhage,decreased blood-brain barrier permeability and neuronal degeneration damage,reduced neurobehavioral deficits,and improved the survival rate of mice.Expression levels of p-AKT and p-GSK3βwere significantly increased in the perihematoma tissues after intranasal insulin therapy.Our findings suggest that intranasal insulin therapy can protect the neurological function of mice after intracerebral hemorrhage through the AKT/GSK3βsignaling pathway.The study was approved by the Ethics Committee of the North Sichuan Medical College of China(approval No.NSMC(A)2019(01))on January 7,2019.

Neuroprotective Effect of Chrysophanol as a PI3K/AKT/mTOR Signaling Inhibitor in an Experimental Model of Autologous Blood-induced Intracerebral Hemorrhage

Objective Intracerebral hemorrhage(ICH)refers to predominant,sporadic,and non-traumatic bleeding in the brain parenchyma.The PI3K/AKT/mTOR signaling pathway is an important signal transduction pathway regulated by enzyme-linked receptors and has many biological functions in mammals.It plays a key role in neuronal metabolism,gene expression regulation,and tissue homeostasis in the healthy and diseased brain.Methods In the present study,the role of the PI3K/AKT/mTOR pathway inhibitor chrysophanol(CPH)(10 mg/kg and 20 mg/kg,orally)in the improvement of ICH-associated neurological defects in rats was investigated.Autologous blood(20µL/5 min/unilateral/intracerebroventricular)mimics ICH-like defects involving cellular and molecular dysfunction and neurotransmitter imbalance.The current study also included various behavioral assessments to examine cognition,memory,and motor and neuromuscular coordination.The protein expression levels of PI3K,AKT,and mTOR as well as myelin basic protein and apoptotic markers,such as Bax,Bcl-2,and caspase-3,were examined using ELISA kits.Furthermore,the levels of various neuroinflammatory cytokines and oxidative stress markers were assessed.Additionally,the neurological severity score,brain water content,gross brain pathology,and hematoma size were used to indicate neurological function and brain edema.Results CPH was found to be neuroprotective by restoring neurobehavioral alterations and significantly reducing the elevated PI3K,AKT,and mTOR protein levels,and modulating the apoptotic markers such as Bax,Bcl-2,and caspase-3 in rat brain homogenate.CPH substantially reduced the inflammatory cytokines like interleukin(IL)-1β,IL-6,and tumor necrosis factor-α.CPH administration restored the neurotransmitters GABA,glutamate,acetylcholine,dopamine,and various oxidative stress markers.Conclusion Our results show that CPH may be a promising therapeutic approach for overcoming neuronal damage caused by the overexpression of the PI3K/AKT/mTOR signaling pathway in ICH-induced neurological dysfunctions in rats.

Study of clinical features of amyloid angiopathy hemorrhage and hypertensive intracerebral hemorrhage

Objective: The purpose of this study was to differentiate between cerebral amyloid angiopathy (CAA) and hypertension (HTN) based on hemorrhage pattern interpretation. Methods: From June 1994 to Oct., 2000, 83 patients admitted to our service with acute intracerebral hemorrhage (ICH) were investigated retrospectively; 41 patients with his-tologically proven diagnosis of cerebral amyloid angiography and 42 patients with clear history of hypertension were investigated. Results: Patients with a CAA-related ICH were significantly older than patients with a HTN-related ICH (74.0 years vs 66.5 years, P<0.05). There was a significantly higher number of hematomas> ml in CAA (85.3%) when compared with HTN (59.5%). No basal ganglional hemorrhage was seen in CAA, but in 40.5% in HTN. In CAA-related ICH, su-barachnoid hemorrhage (SAH) was seen in 26 patients (63.4%) compared to only 11 patients (26.2%) in HTN-related ICH. Intraventricular hemorrhage was seen in 24.4% in CAA, and in 26.2% in HTN. Typical features of CAA-related ICH included lobar distribution affecting mainly the lobar superficial areas, lobulated appearance, rupture into the subarachnoid space, and secondary IVH from the lobar hemorrhage. More specifically, multiplicity of hemorrhage, bilaterality, and repeated episodes also strongly suggest the diagnosis of CAA. Multiple hemorrhages, defined as 2 or more separate he-matomas in multiple lobes, accounted for 17.1% in CAA-related ICH. Conclusion: There are certain features in CAA on CT and MRI and in clinical settings. To some extent, these features may contribute to distinguishing CAA from HTN related ICH.

Intraclot Recombinant Tissue-type Plasminogen Activator Reduces Perihematomal Edema and Mortality in Patients with Spontaneous Intracerebral Hemorrhage

The study aimed to investigate the impact of intraclot recombinant tissue-type plasminogen activator （rt-PA） on perihematomal edema （PHE） development in patients with intracerebral hemorrhage （ICH） treated with minimally invasive surgery （MIS） and the effects of intraclot rt-PA on the 30-day survival. We reviewed the medical records of ICH patients undergoing MIS between October 2011 and July 2013. A volumetric analysis was done to assess the change in PHE and ICH volumes at pre-MIS （T1）, post-MIS （T2） and day 10-16 （T3） following diagnostic computed tomographic scans （To）. Forty-three patients aged 52.8±11.1 years with （n=30） or without rt-PA （n=13） were enrolled from our institutional ICH database. The median rt-PA dose was 1.5 （1） mg, with a maximum dose of 4.0 mg. The ratio of clot evacuation was significantly increased by intraclot rt-PA as compared with controls （77.9%±20.4% vs. 64%±15%; P=0.046）. From TI to T2, reduction in PHE volume was strongly associ- ated with the percentage of clot evacuation （p=0.34; P=-0.027）. In addition, PHE volume was positively correlated with residual ICH volume at the same day （p ranging from 0.39-0.56, P〈0.01）. There was no correlation between the cumulative dose of rt-PA and early （T2） PHE volume （p=0.24; P=0.12） or de- layed （T3） PHE volume （p=0.19; P=0.16）. The 30-day mortality was zero in this cohort. In the selected cohort of ICH patients treated with MIS, intraclot rt-PA accelerated clot removal and had no effects on PHE formation. MIS aspiration and low dose of rt-PA seemed to be feasible to reduce the 30-day mor- tality in patients with severe ICH. A large, randomized study addressing dose titration and long-term outcome is needed.

Changes of TXA_2 and PGI_2 during Postoperative Hypertensive Crisis in Patients with Hypertensive Intracerebral Hemorrhage

In order to explore the changes and the roles of TXA2 and PGI2 during postoperative hypertensive crisis in patients with hypertensive intracerebral hemorrhage, 31 cases subject to craniotomy were divided into three groups： group A, 9 patients with postoperative hypertensive crisis; group B, 13 patients without postoperative hypertensive crisis; and group C, 9 patients without history of hypertension and hypertensive intracerebral hemorrhage. TXA2, TXB2, 6-keto-PGF1α and PGI2 were measured after operation in the three groups respectively. The postoperative blood pressure in group A, including SBP and DBP, was elevated more obviously than that in the other two groups. TXA2 and PGI2 in group A were significantly higher than those in other two groups （P〈0.01）. Moreover, the ratio of TXB2 to 6-keto-PGF1α in group A was significantly higher than that in other two groups （P〈0.05）. The increase of TXA2 and the relative inadequacy of prostacyclin, especially 6-keto-PGF1α, may play roles in the postoperative hypertensive crisis. And the increased value of TXB2 to 6-keto-PGF1α could provide the basis for diagnosis of postoperative hypertensive crisis.

Exosomal miR-23b from bone marrow mesenchymal stem cells alleviates oxidative stress and pyroptosis after intracerebral hemorrhage

Our previous studies showed that miR-23b was downregulated in patients with intracerebral hemorrhage(ICH). This indicates that miR-23b may be closely related to the patho-physiological mechanism of ICH, but this hypothesis lacks direct evidence. In this study, we established rat models of ICH by injecting collagenase Ⅶ into the right basal ganglia and treating them with an injection of bone marrow mesenchymal stem cell(BMSC)-derived exosomal miR-23b via the tail vein. We found that edema in the rat brain was markedly reduced and rat behaviors were improved after BMSC exosomal miR-23b injection compared with those in the ICH groups. Additionally, exosomal miR-23b was transported to the microglia/macrophages, thereby reducing oxidative stress and pyroptosis after ICH. We also used hemin to mimic ICH conditions in vitro. We found that phosphatase and tensin homolog deleted on chromosome 10(PTEN) was the downstream target gene of miR-23b, and exosomal miR-23b exhibited antioxidant effects by regulating the PTEN/Nrf2 pathway. Moreover, miR-23b reduced PTEN binding to NOD-like receptor family pyrin domain containing 3(NLRP3) and NLRP3 inflammasome activation, thereby decreasing the NLRP3-dependent pyroptosis level. These findings suggest that BMSC-derived exosomal miR-23b exhibits antioxidant effects through inhibiting PTEN and alleviating NLRP3 inflammasome-mediated pyroptosis, thereby promoting neurologic function recovery in rats with ICH.