Cathepsin B as a key regulator of ferroptosis in microglia following intracerebral hemorrhage.

Intracerebral hemorrhage (ICH) is a subtype of stroke marked by elevated mortality and disability rates. Recently, mounting evidence suggests a significant role of ferroptosis in the pathogenesis of ICH. Through a combination of bioinformatics analysis and basic experiments, our goal is to identify the primary cell types and key molecules implicated in ferroptosis post-ICH. This aims to propel the advancement of ferroptosis research, offering potential therapeutic targets for ICH treatment. Our study reveals pronounced ferroptosis in microglia and identifies the target gene, cathepsin B (Ctsb), by analyzing differentially expressed genes following ICH. Ctsb, a cysteine protease primarily located in lysosomes, becomes a focal point in our investigation. Utilizing in vitro and in vivo models, we explore the correlation between Ctsb and ferroptosis in microglia post-ICH. Results demonstrate that ICH and hemin-induced ferroptosis in microglia coincide with elevated levels and activity of Ctsb protein. Effective alleviation of ferroptosis in microglia after ICH is achieved through the inhibition of Ctsb protease activity and protein levels using inhibitors and shRNA. Additionally, a notable increase in m6A methylation levels of Ctsb mRNA post-ICH is observed, suggesting a pivotal role of m6A methylation in regulating Ctsb translation. These research insights deepen our comprehension of the molecular pathways involved in ferroptosis after ICH, underscoring the potential of Ctsb as a promising target for mitigating brain damage resulting from ICH.

Geniposide ameliorates brain injury in mice with intracerebral hemorrhage by inhibiting NF-κB signaling.

BACKGROUND: Neuroinflammation and oxidative stress are critical players in intracerebral hemorrhage (ICH). Geniposide is an active component of Gardenia that has anti-inflammatory effects. This study focused on the roles and mechanisms of geniposide in ICH. METHODS: ICH was established by injecting collagenase IV into C57BL/6 mice. To determine the functions of geniposide and NF-κB inhibition in ICH model mice, geniposide (1, 25, or 50 mg/kg) or PDTC (a NF-κB inhibitor) was administered. Neurological functions were assessed with the modified neurological severity score (mNSS) test. Hematoxylin and eosin staining were performed to identify pathological changes. IL-1ß and TNF-α levels were estimated with ELISA kits. NF-κB p65 localization was determined by immunofluorescence staining. Oxidative stress was analyzed by measuring ROS levels. RESULTS: Geniposide alleviated cerebral edema and neurological deficits. Geniposide inhibited neuroinflammation and oxidative stress after ICH, and the inhibitory effects were enhanced by NF-κB inhibition. Additionally, geniposide inhibited NF-κB signaling. CONCLUSION: Geniposide alleviates brain injury by suppressing inflammation and oxidative stress damage in experimental ICH models by inhibiting NF-κB signaling.

Consistency of mouse models with human intracerebral hemorrhage: core targets and non-coding RNA regulatory axis.

Intracerebral hemorrhage (ICH) has a high mortality and disability rate. Numerous basic studies on pathogenesis and therapeutics have been performed in mice. However, the consistency of the experimental mouse model and the human ICH patient remains unclear. This has slowed progress in translational medicine. Furthermore, effective therapeutic targets and reliable regulatory networks for ICH are needed. Therefore, we determined the differentially expressed (DE) messenger RNAs (mRNAs), microRNAs (miRNAs) and circular RNAs (circRNAs) before and after murine ICH and analyzed their regulatory relationships. Subsequently, data on mRNAs from human peripheral blood after ICH were obtained from the Gene Expression Omnibus database. The DE mRNAs after human ICH were compared with those of the mouse. Finally, we obtained seven genes with translational medicine research value and verified them in mice. Then the regulatory network of these genes was analyzed in humans. Similarly, species homologies of these regulatory pathways were identified. In conclusion, we found that the mouse ICH model mimics the human disease mainly in terms of chemokines and inflammatory factors. This has important implications for future research into the mechanisms of ICH injury and repair.

A dispensable role of oligodendrocyte-derived laminin-α5 in brain homeostasis and intracerebral hemorrhage.

Laminin, a major component of the basal lamina in the CNS, is also expressed in oligodendrocytes (OLs). However, the function of OL-derived laminin remains largely unknown. Here, we performed loss-of-function studies using two OL-specific laminin-α5 conditional knockout mouse lines. Both mutants were grossly normal and displayed intact blood-brain barrier (BBB) integrity. In a mouse model of intracerebral hemorrhage (ICH), control mice and both mutants exhibited comparable hematoma size and neurological dysfunction. In addition, similar levels of hemoglobin and IgG leakage were detected in the mutant brains compared to the controls, indicating comparable BBB damage. Consistent with this finding, subsequent studies revealed no differences in tight junction protein (TJP) and caveolin-1 expression among control and knockout mice, suggesting that neither paracellular nor transcellular mechanism was affected in the mutants. Furthermore, compared to the controls, both mutant lines showed comparable oligodendrocyte number, oligodendrocyte proliferation rate, MBP/MAG levels, and SMI-32 expression, highlighting a minimal role of OL-derived laminin-α5 in OL biology. Together, these findings highlight a dispensable role of OL-derived laminin-α5 in both brain homeostasis and ICH pathogenesis.

Azithromycin reduces hemoglobin-induced innate neuroimmune activation.

Neonatal intraventricular hemorrhage (IVH) releases blood products into the lateral ventricles and brain parenchyma. There are currently no medical treatments for IVH and surgery is used to treat a delayed effect of IVH, post-hemorrhagic hydrocephalus. However, surgery is not a cure for intrinsic brain injury from IVH, and is performed in a subacute time frame. Like many neurological diseases and injuries, innate immune activation is implicated in the pathogenesis of IVH. Innate immune activation is a pharmaceutically targetable mechanism to reduce brain injury and post-hemorrhagic hydrocephalus after IVH. Here, we tested the macrolide antibiotic azithromycin, which has immunomodulatory properties, to reduce innate immune activation in an in vitro model of microglial activation using the blood product hemoglobin (Hgb). We then utilized azithromycin in our in vivo model of IVH, using intraventricular blood injection into the lateral ventricle of post-natal day 5 rat pups. In both models, azithromycin modulated innate immune activation by several outcome measures including mitochondrial bioenergetic analysis, cytokine expression and flow cytometric analysis. This suggests that azithromycin, which is safe for neonates, could hold promise for modulating innate immune activation after IVH.

Group 2 innate lymphoid cells suppress neuroinflammation and brain injury following intracerebral hemorrhage.

Intracerebral hemorrhage (ICH) mobilizes circulating leukocytes that contribute to neuroinflammation and neural injury. However, little is known about the endogenous regulatory immune mechanisms to restrict neuroinflammation following ICH. We examined the role of group 2 innate lymphoid cells (ILC2) that are a specialized subset of innate immune modulators in a mouse model of ICH. We found accumulation of ILC2 in the brain following acute ICH and a concomitant increase of ILC2 within the peripheral lymph nodes. Depletion of ILC2 exacerbated neurodeficits and brain edema after ICH in male and female mice. This aggravated ICH injury was accompanied by augmented microglia activity and leukocyte infiltration. In contrast, expansion of ILC2 using IL-33 led to reduced ICH injury, microglia activity and leukocyte infiltration. Notably, elimination of microglia using a colony stimulating factor 1 receptor inhibitor diminished the exacerbation of ICH injury induced by depletion of ILC2. Brain-infiltrating ILC2 had upregulation of IL-13 after ICH. Results from in vitro assays revealed that ILC2 suppressed thrombin-induced inflammatory activity in microglia-like BV2 cells. Thus, our findings demonstrate that ILC2 suppress neuroinflammation and acute ICH injury.

Structural Brain Injury on Brain Magnetic Resonance Imaging in Acute Respiratory Distress Syndrome.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is an acute inflammatory respiratory failure condition that may be associated with brain injury. We aimed to describe the types of structural brain injuries detected by brain magnetic resonance imaging (MRI) among patients with ARDS. METHODS: We retrospectively reviewed and collected data on brain injuries as detected by brain MRI during index hospitalization of all patients with ARDS at a single tertiary center in the United States from January 2010 to October 2018 (pre-COVID era). Structural brain injuries were classified as cerebral ischemia (ischemic infarct and hypoxic-ischemic brain injury) or cerebral hemorrhage (intraparenchymal hemorrhage, cerebral microbleeds, subarachnoid hemorrhage, and subdural hematoma). Descriptive statistics were conducted. RESULTS: Of the 678 patients with ARDS, 66 (9.7%) underwent brain MRI during their ARDS illness. The most common indication for brain MRI was encephalopathy (45.4%), and the median time from hospital admission to MRI was 10 days (interquartile range 4-17). Of 66 patients, 29 (44%) had MRI evidence of brain injury, including cerebral ischemia in 33% (22 of 66) and cerebral hemorrhage in 21% (14 of 66). Among those with cerebral ischemia, common findings were bilateral globus pallidus infarcts (n = 7, 32%), multifocal infarcts (n = 5, 23%), and diffuse hypoxic-ischemic brain injury (n = 3, 14%). Of those with cerebral hemorrhage, common findings were cerebral microbleeds (n = 12, 86%) and intraparenchymal hemorrhage (n = 2, 14%). Patients with ARDS with cerebral hemorrhage had significantly greater use of rescue therapies, including prone positioning (28.6% vs. 5.8%, p = 0.03), inhaled vasodilator (35.7% vs. 11.5%, p = 0.046), and recruitment maneuver (14.3% vs. 0%, p = 0.04). CONCLUSIONS: Structural brain injury was not uncommon among selected patients with ARDS who underwent brain MRI. The majority of brain injuries seen were bilateral globus pallidus infarcts and cerebral microbleeds.

[Analysis of the prevalence of anticoagulant therapy in patients with cognitive disorders and cerebral amyloid angiopathy (CAA)]. / Analyse zur Häufigkeit einer gerinnungshemmenden Medikation bei Patientinnen mit kognitiven Störungen und zerebraler Amyloidangiopathie (CAA).

OBJECTIVES: To investigate the prevalence of coincident anticoagulation in patients with cognitive disorders and possible or probable cerebral amyloid angiopathy (CAA) as well as the relationship between the presence of oral anticoagulation and CAA-specific lesion load. MATERIALS AND METHODS: Patients with subjective cognitive decline (SCD), amnestic and non-amnestic mild cognitive impairment (aMCI/naMCI), Alzheimer's disease (AD), mixed dementia (MD) and vascular dementia (VD) who presented to our outpatient dementia clinic between February 2016 and October 2020 were included in this retrospective analysis. Patients underwent cranial magnetic resonance imaging (MRI). MRI data sets were analyzed regarding the presence of CAA-related MRI biomarkers to determine CAA prevalence. Presence of anticoagulant therapy was determined by chart review. RESULTS: Within the study period, 458 patients (209 male, 249 female, mean age 73.2 ± 9.9 years) with SCD (n = 44), naMCI (n = 40), aMCI (n = 182), AD (n = 120), MD (n = 68) and VD (n = 4) were analyzed. A total of 109 patients (23.8%) were diagnosed with possible or probable CAA. CAA prevalence was highest in aMCI (39.4%) and MD (28.4%). Of patients with possible or probable CAA, 30.3% were under platelet aggregation inhibition, 12.8% were treated with novel oral anticoagulants and 3.7% received phenprocoumon treatment. Regarding the whole study cohort, patients under oral anticoagulation showed more cerebral microbleeds (p = 0.047). There was no relationship between oral anticoagulation therapy and the frequency of cortical superficial siderosis (p = 0.634). CONCLUSION: CAA is a frequent phenomenon in older patients with cognitive disorders. Almost half of CAA patients receive anticoagulant therapy. Oral anticoagulation is associated with a higher number of cortical and subcortical microbleeds.

Deubiquitylating Enzyme OTUB1 Facilitates Neuronal Survival After Intracerebral Hemorrhage Via Inhibiting NF-κB-triggered Apoptotic Cascades.

The deubiquitylase OTU domain-containing ubiquitin aldehyde-binding protein 1 (OTUB1) has been implicated in the pathogenesis of various human diseases. However, the molecular mechanism by which OTUB1 participates in the pathogenesis of intracerebral hemorrhage (ICH) remains elusive. In the present study, we established an autologous whole blood fusion-induced ICH model in C57BL/6 J mice. We showed that the upregulation of OTUB1 contributes to the attenuation of Nuclear factor kappa B (NF-κB) and its downstream apoptotic signaling after ICH. OTUB1 directly associates with NF-κB precursors p105 and p100 after ICH, leading to attenuated polyubiquitylation of p105 and p100. Moreover, we revealed that NF-κB signaling was modestly activated both in ICH tissues and hemin-exposed HT-22 neuronal cells, accompanied with the activation of NF-κB downstream pro-apoptotic signaling. Notably, overexpression of OTUB1 strongly inhibited hemin-induced NF-κB activation, whereas interference of OTUB1 led to the opposite effect. Finally, we revealed that lentiviral transduction of OTUB1 markedly ameliorated hemin-induced apoptotic signaling and HT-22 neuronal death. Collectively, these findings suggest that the upregulation of OTUB1 serves as a neuroprotective mechanism in antagonizing neuroinflammation-induced NF-κB signaling and neuronal death, shed new light on manipulating intracellular deubiquitylating pathways as novel interventive approaches against ICH-induced secondary neuronal damage and death.

Use of MR signal intensity variations to highlight structures at risk along brain biopsy trajectories.

OBJECTIVE: Postoperative intracerebral hemorrhages are significant complications following brain stereotactic biopsy. They can derive from anatomical structure (sulci, vessels) damage that is missed during stereotactic trajectory planning. In this study, the authors investigated the ability to detect contact between structures at risk and stereotactic trajectories using signal analysis from MRI obtained during clinical practice, with the aim to propose a visual tool to highlight areas with anatomical structures at risk of damage along the biopsy trajectory. METHODS: The authors retrospectively analyzed actual stereotactic trajectories using intraoperative imaging (intraoperative 2D radiographs in the exploratory data set and intraoperative 3D scans in the confirmatory data set). The MR signal variation along each biopsy trajectory was matched with the patient's anatomy. RESULTS: In the exploratory data set (n = 154 patients), 32 contacts between the actual biopsy trajectory and an anatomical structure at risk were identified along 28 (18.2%) biopsy trajectories, corresponding to 8 preventable intracerebral hemorrhages. Variations of the mean derivative of the MR signal intensity were significantly different between trajectories with and without contact (the pathological threshold of the mean derivative of the MR signal intensity was defined as ± 0.030 arbitrary units; p < 0.0001), with a sensitivity of 89.3% and specificity of 74.6% to detect a contact. In the confirmatory data set (n = 73 patients), the sensitivity and specificity of the 0.030 threshold to detect a contact between the actual stereotactic trajectory and an anatomical structure at risk were 81.3% and 68.4%, respectively. CONCLUSIONS: Variations of the mean derivative of the MR signal intensity can be converted into a green/red color code along the planned biopsy trajectory to highlight anatomical structures at risk, which can help neurosurgeons during the surgical planning of stereotactic procedures.

FGF21 alleviates endothelial mitochondrial damage and prevents BBB from disruption after intracranial hemorrhage through a mechanism involving SIRT6.

BACKGROUND: Disruption of the BBB is a harmful event after intracranial hemorrhage (ICH), and this disruption contributes to a series of secondary injuries. We hypothesized that FGF21 may have protective effects after intracranial hemorrhage (ICH) and investigated possible underlying molecular mechanisms. METHODS: Blood samples of ICH patients were collected to determine the relationship between the serum level of FGF21 and the [Formula: see text]GCS%. Wild-type mice, SIRT6flox/flox mice, endothelial-specific SIRT6-homozygous-knockout mice (eSIRT6-/- mice) and cultured human brain microvascular endothelial cells (HCMECs) were used to determine the protective effects of FGF21 on the BBB. RESULTS: We obtained original clinical evidence from patient data identifying a positive correlation between the serum level of FGF21 and [Formula: see text]GCS%. In mice, we found that FGF21 treatment is capable of alleviating BBB damage, mitigating brain edema, reducing lesion volume and improving neurofunction after ICH. In vitro, after oxyhemoglobin injury, we further explored the protective effects of FGF21 on endothelial cells (ECs), which are a significant component of the BBB. Mitochondria play crucial roles during various types of stress reactions. FGF21 significantly improved mitochondrial biology and function in ECs, as evidenced by alleviated mitochondrial morphology damage, reduced ROS accumulation, and restored ATP production. Moreover, we found that the crucial regulatory mitochondrial factor deacylase sirtuin 6 (SIRT6) played an irreplaceable role in the effects of FGF21. Using endothelial-specific SIRT6-knockout mice, we found that SIRT6 deficiency largely diminished these neuroprotective effects of FGF21. Then, we revealed that FGF21 might promote the expression of SIRT6 via the AMPK-Foxo3a pathway. CONCLUSIONS: We provide the first evidence that FGF21 is capable of protecting the BBB after ICH by improving SIRT6-mediated mitochondrial homeostasis.

E3 ligase Nedd4L promotes macrophage M1 polarization and exacerbates brain damage by TRAF3/TBK1 signaling pathway after ICH in mice.

BACKGROUND: Intracerebral hemorrhage (ICH) is a serious medical problem, and promising strategy is limited. Macrophage initiated brain inflammatory injury following ICH, but the molecular mechanism had not been well identified. E3 ligase Nedd4L is implicated in the pathogenesis of the inflammatory immune response. METHODS: In the present study, we detected the levels of Nedd4L in macrophages following ICH. Furthermore, Macrophage M1 polarization, pro-inflammatory cytokine production, BBB disruption, brain water content and neurological function were examined in ICH mice. RESULTS: Here, we demonstrated that E3 ligase Nedd4L levels of macrophage increased following ICH, promoted M1 polarization inflammation by TRAF3. Nedd4L promoted BBB disruption, as well as neurological deficits. Inhibition of Nedd4L significantly attenuated M1 polarization in vivo. Inhibition of Nedd4L decreased TRAF3 and TBK1 levels, and subsequent phosphorylation of p38 and NF-κB p65 subunit following ICH. CONCLUSIONS: Our data demonstrated that Nedd4L was involved in the pathogenesis of ICH, which promoted inflammatory responses and exacerbated brain damage by TRAF3 following ICH.

Cerebral Microbleeds Are Associated with Widespread Blood-Brain Barrier Leakage.

INTRODUCTION: The pathogenesis of cerebral microbleeds (CMBs) is incompletely understood, but blood-brain barrier (BBB) leakage may play a key role. This study aimed to investigate the relationship between compromised BBB integrity and CMBs as well as cognitive function. METHODS: Ninety-seven participants were enrolled in this cross-sectional study, involving 24 CMB patients. Dynamic contrast-enhanced-magnetic resonance imaging was used to measure BBB permeability, and cognitive function was assessed by Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA). RESULTS: Compared with participants without CMBs, CMB patients had higher volume transfer constant (Ktrans, all p < 0.01) and area under the concentration curve (AUC, all p < 0.05) in normal-appearing white matter (NAWM), white matter hyperintensities (WMH), cortical gray matter (CGM), and deep gray matter (DGM). Multivariable linear regression analyses revealed that CMB patients had significantly higher Ktrans in NAWM and AUC in NAWM, WMH, and CGM after adjustment for age, sex, vascular risk factors, and cognitive scores. MMSE and MoCA scores decreased with increasing Ktrans in WMH and DGM as well as AUC in WMH after adjustment for age, sex, CMB group, and education length. CONCLUSION: This study demonstrated that widespread BBB leakage was prevalent in CMB patients, suggesting that compromised BBB integrity may play a key role in the pathogenesis of CMBs and could lead to cognitive impairment.

Detection rate of brain MR and MR angiography for neuroimaging abnormality in patients with newly diagnosed left-sided infective endocarditis.

We aimed to investigate the detection rate of brain MR and MR angiography for neuroimaging abnormality in newly diagnosed left-sided infective endocarditis patients with/without neurological symptoms. This retrospective study included consecutive patients with definite or possible left-sided infective endocarditis according to the modified Duke criteria who underwent brain MRI and MR angiography between March 2015 and October 2020. The detection rate for neuroimaging abnormality on MRI was defined as the number of patients with positive brain MRI findings divided by the number of patients with left-sided infective endocarditis. Positive imaging findings included acute ischemic lesions, cerebral microbleeds, hemorrhagic lesions, and infectious aneurysms. In addition, aneurysm rupture rate and median period to aneurysm rupture were evaluated on follow-up studies. A total 115 patients (mean age: 55 years ± 19; 65 men) were included. The detection rate for neuroimaging abnormality was 77% (89/115). The detection rate in patients without neurological symptoms was 70% (56/80). Acute ischemic lesions, cerebral microbleeds, and hemorrhagic lesions including superficial siderosis and intracranial hemorrhage were detected on MRI in 56% (64/115), 57% (66/115), and 20% (23/115) of patients, respectively. In particular, infectious aneurysms were detected on MR angiography in 3% of patients (4/115), but MR angiography in 5 patients (4.3%) was insignificant for infectious aneurysm, which were detected using CT angiography (n = 3) and digital subtraction angiography (n = 2) during follow-up. Among the 9 infectious aneurysm patients, aneurysm rupture occurred in 4 (44%), with a median period of aneurysm rupture of 5 days. The detection rate of brain MRI for neuroimaging abnormality in newly diagnosed left-sided infective endocarditis patients was high (77%), even without neurological symptoms (70%).

Cerebral Microbleed Patterns and Cortical Amyloid-ß: The ARIC-PET Study.

BACKGROUND: Cerebral microbleeds (CMBs) are associated with cognitive decline, but their importance outside of cerebral amyloid angiopathy and the mechanisms of their impact on cognition are poorly understood. We evaluated the cross-sectional association between CMB patterns and cerebral Aß (amyloid-ß) deposition, by florbetapir positron emission tomography. METHODS: The longitudinal ARIC study (Atherosclerosis Risk in Communities) recruited individuals from 4 US communities from 1987 to 1989. From 2012 to 2014, the ARIC-PET (Atherosclerosis Risk in Communities - Positron Emission Tomography) ancillary recruited 322 nondemented ARIC participants who completed 3T brain magnetic resonance imaging with T2*GRE as part of ARIC visit 5 to undergo florbetapir positron emission tomography imaging. Magnetic resonance imaging images were read for CMBs and superficial siderosis; on positron emission tomography, global cortical standardized uptake value ratio >1.2 was considered a positive Aß scan. Multivariable logistic regression models evaluated CMB characteristics in association with Aß positivity. Effect modification by sex, race, APOE status, and cognition was evaluated. RESULTS: CMBs were present in 24% of ARIC-PET participants. No significant associations were found between CMBs and Aß positivity, but a pattern of isolated lobar CMBs or superficial siderosis was associated with over 4-fold higher odds of elevated Aß when compared with those with no CMBs (odds ratio, 4.72 [95% CI, 1.16-19.16]). A similar elevated risk was not observed in those with isolated subcortical or mixed subcortical and either lobar CMBs or superficial siderosis. Although no significant interactions were found, effect estimates for elevated Aß were nonsignificantly lower (P>0.10, odds ratio, 0.4-0.6) for a mixed CMB pattern, and odds ratios were nonsignificantly higher for lobar-only CMBs for 4 subgroups: women (versus men); Black participants (versus White participants), APOE ε4 noncarriers (versus carriers), and cognitively normal (versus mild cognitive impairment). CONCLUSIONS: In this community-based cohort of nondemented adults, lobar-only pattern of CMBs or superficial siderosis is most strongly associated with brain Aß, with no elevated risk for a mixed CMB pattern. Further studies are needed to understand differences in CMB patterns and their meaning across subgroups.

Histopathological Correlates of Lobar Microbleeds in False-Positive Cerebral Amyloid Angiopathy Cases.

OBJECTIVE: A definite diagnosis of cerebral amyloid angiopathy (CAA), characterized by the accumulation of amyloid ß in walls of cerebral small vessels, can only be obtained through pathological examination. A diagnosis of probable CAA during life relies on the presence of hemorrhagic markers, including lobar cerebral microbleeds (CMBs). The aim of this project was to study the histopathological correlates of lobar CMBs in false-positive CAA cases. METHODS: In 3 patients who met criteria for probable CAA during life, but showed no CAA upon neuropathological examination, lobar CMBs were counted on ex vivo 3T magnetic resonance imaging (MRI) and on ex vivo 7T MRI. Areas with lobar CMBs were next sampled and cut into serial sections, on which the CMBs were then identified. RESULTS: Collectively, there were 25 lobar CMBs on in vivo MRI and 22 on ex vivo 3T MRI of the analyzed hemispheres. On ex vivo MRI, we targeted 12 CMBs for sampling, and definite histopathological correlates were retrieved for 9 of them, of which 7 were true CMBs. No CAA was found on any of the serial sections. The "culprit vessels" associated with the true CMBs instead showed moderate to severe arteriolosclerosis. Furthermore, CMBs in false-positive CAA cases tended to be located more often in the juxtacortical or subcortical white matter than in the cortical ribbon. INTERPRETATION: These findings suggest that arteriolosclerosis can generate lobar CMBs and that more detailed investigations into the exact localization of CMBs with respect to the cortical ribbon could potentially aid the diagnosis of CAA during life. ANN NEUROL 2023;94:856-870.

Revisiting the role of the complement system in intracerebral hemorrhage and therapeutic prospects.

Intracerebral hemorrhage (ICH) is a stroke subtype characterized by non-traumatic rupture of blood vessels in the brain, resulting in blood pooling in the brain parenchyma. Despite its lower incidence than ischemic stroke, ICH remains a significant contributor to stroke-related mortality, and most survivors experience poor outcomes that significantly impact their quality of life. ICH has been accompanied by various complex pathological damage, including mechanical damage of brain tissue, hematoma mass effect, and then leads to inflammatory response, thrombin activation, erythrocyte lysis, excitatory amino acid toxicity, complement activation, and other pathological changes. Accumulating evidence has demonstrated that activation of complement cascade occurs in the early stage of brain injury, and the excessive complement activation after ICH will affect the occurrence of secondary brain injury (SBI) through multiple complex pathological processes, aggravating brain edema, and pathological brain injury. Therefore, the review summarized the pathological mechanisms of brain injury after ICH, specifically the complement role in ICH, and its related pathological mechanisms, to comprehensively understand the specific mechanism of different complements at different stages after ICH. Furthermore, we systematically reviewed the current state of complement-targeted therapies for ICH, providing a reference and basis for future clinical transformation of complement-targeted therapy for ICH.

Intracerebral Hemorrhage-Induced Brain Injury: the Role of Lysosomal-Associated Transmembrane Protein 5.

Intracerebral hemorrhage (ICH) is a lethal stroke with high mortality or disability. However, effective therapy for ICH damage is generally lacking. Previous investigations have suggested that lysosomal protein transmembrane 5 (LAPTM5) is involved in various pathological processes, including autophagy, apoptosis, and inflammation. In this study, we aimed to identify the expression and functions of LAPTM5 in collagenase-induced ICH mouse models and hemoglobin-induced cell models. We found that LAPTM5 was highly expressed in brain tissues around the hematoma, and double immunostaining studies showed that LAPTM5 was co-expressed with microglia cells, neurons, and astrocytes. Following ICH, the mice presented increased brain edema, blood-brain barrier permeability, and neurological deficits, while pathological symptoms were alleviated after the LAPTM5 knockdown. Adeno-associated virus 9-mediated downregulation of LAPTM5 also improves ICH-induced secondary cerebral damage, including neuronal degeneration, the polarization of M1-like microglia, and inflammatory cascades. Furthermore, LAPTM5 promoted activation of the nuclear factor kappa-B (NF-κB) pathway in response to neuroinflammation. Further investigations indicated that brain injury improved by LAPTM5 knockdown was further exacerbated after the overexpression of receptor-interacting protein kinase 1 (RIP1), which is revealed to trigger the NF-κB pathway. In vitro experiments demonstrated that LAPTM5 silencing inhibited hemoglobin-induced cell function and confirmed regulation between RIP1 and LAPTM5. In conclusion, the present study indicates that LAPTM5 may act as a positive regulator in the context of ICH by modulating the RIP1/NF-κB pathway. Thus, it may be a candidate gene for further study of molecular or therapeutic targets.

A bibliometric analysis of cerebral microbleeds and cognitive impairment.

BACKGROUND AND OBJECTIVES: Cerebral microbleeds (CMBs) are imaging markers for small cerebral vascular diseases, which can accumulate and impact the corresponding brain networks. CMBs can affect cognitive function, including executive function, information processing speed, and visuospatial memory. Bibliometrics is a scientific and innovative method that can analyze and visualize the scientific field quantitatively. In this study, we aimed to use bibliometric analysis to demonstrate the relationship and mechanisms between CMBs and cognitive impairment. Furthermore, we reviewed the relationship between CMBs and different cognitive disorders. The use of bibliometrics can help further clarify this relationship. METHODS: We retrieved articles on CMBs and cognitive impairment from the Web of Science Core Collection. The keywords (such as stroke, dementia, and cerebral amyloid angiopathy), authors, countries, institutions and journals, in the field were visually analyzed using VOSviewer software and bibliometric websites. RESULTS: This bibliometric analysis reveals the related trends of CMBs in the field of cognitive impairment. CMBs, along with other small vascular lesions, constitute the basis of cognitive impairment, and studying CMBs is essential to understand the mechanisms underlying cognitive impairment. CONCLUSION: This bibliometric analysis reveals a strong link between CMBs and cognitive impairment-related diseases and that specific brain networks were affected by CMBs. This provides further insights into the possible mechanisms and causes of CMBs and cognitive impairment. The direct and indirect damage (such as oxidative stress and neuroinflammation) to the brain caused by CMBs, destruction of the frontal-subcortical circuits, elevated Cystatin C levels, and iron deposition are involved in the occurrence and development of cognitive impairment. CMBs may be a potential marker for detecting, quantifying, and predicting cognitive impairment.

A role for P-selectin and complement in the pathological sequelae of germinal matrix hemorrhage.

BACKGROUND: Germinal matrix hemorrhage is a devastating disease of pre-term infancy commonly resulting in post-hemorrhagic hydrocephalus, periventricular leukomalacia, and subsequent neurocognitive deficits. We demonstrate vascular expression of the adhesion molecule P-selectin after GMH and investigate a strategy to specifically target complement inhibition to sites of P-selectin expression to mitigate the pathological sequelae of GMH. METHODS: We prepared two fusion proteins consisting of different anti-P-selectin single chain antibodies (scFv's) linked to the complement inhibitor Crry. One scFv targeting vehicle (2.12scFv) blocked the binding of P-selectin to its PSGL-1 ligand expressed on leukocytes, whereas the other targeting vehicle (2.3scFv) bound P-selectin without blocking ligand binding. Post-natal C57BL/6 J mice on day 4 (P4) were subjected to collagenase induced-intraventricular hemorrhage and treated with 2.3Psel-Crry, 2.12Psel-Crry, or vehicle. RESULTS: Compared to vehicle treatment, 2.3Psel-Crry treatment after induction of GMH resulted in reduced lesion size and mortality, reduced hydrocephalus development, and improved neurological deficit measurements in adolescence. In contrast, 2.12Psel-Crry treatment resulted in worse outcomes compared to vehicle. Improved outcomes with 2.3Psel-Crry were accompanied by decreased P-selectin expression, and decreased complement activation and microgliosis. Microglia from 2.3Psel-Crry treated mice displayed a ramified morphology, similar to naïve mice, whereas microglia in vehicle treated animals displayed a more ameboid morphology that is associated with a more activated status. Consistent with these morphological characteristics, there was increased microglial internalization of complement deposits in vehicle compared to 2.3Psel-Crry treated animals, reminiscent of aberrant C3-dependent microglial phagocytosis that occurs in other (adult) types of brain injury. In addition, following systemic injection, 2.3Psel-Crry specifically targeted to the post-GMH brain. Likely accounting for the unexpected finding that 2.12Psel-Crry worsens outcome following GMH was the finding that this construct interfered with coagulation in this hemorrhagic condition, and specifically with heterotypic platelet-leukocyte aggregation, which express P-selectin and PSGL-1, respectively. CONCLUSIONS: GMH induces expression of P-selectin, the targeting of which with a complement inhibitor protects against pathogenic sequelae of GMH. A dual functioning construct with both P-selectin and complement blocking activity interferes with coagulation and worsens outcomes following GMH, but has potential for treatment of conditions that incorporate pathological thrombotic events, such as ischemic stroke.