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Achieving optimal alignment in total knee arthroplasty (TKA) is a critical factor in ensuring optimal outcomes and long-term implant survival. Traditionally, mechanical alignment has been favored to achieve neutral post-operative joint alignment. However, contemporary approaches, such as kinematic alignments and hybrid techniques including adjusted mechanical, restricted kinematic, inverse kinematic, and functional alignments, are gaining attention for their ability to restore native joint kinematics and anatomical alignment, potentially leading to enhanced functional outcomes and greater patient satisfaction. The ongoing debate on optimal alignment strategies considers the following factors: long-term implant durability, functional improvement, and resolution of individual anatomical variations. Furthermore, advancements of computer-navigated and robotic-assisted surgery have augmented the precision in implant positioning and objective measurements of soft tissue balance. Despite ongoing debates on balancing implant longevity and functional outcomes, there is an increasing advocacy for personalized alignment strategies that are tailored to individual anatomical variations. This review evaluates the spectrum of various alignment techniques in TKA, including mechanical alignment, patient-specific kinematic approaches, and emerging hybrid methods. Each technique is scrutinized based on its fundamental principles, procedural techniques, inherent advantages, and potential limitations, while identifying significant clinical gaps that underscore the need for further investigation.

Background: Patients with Alzheimer's disease (AD) frequently present with cerebral amyloid angiopathy (CAA), characterized by the accumulation of beta-amyloid (Aβ) within the cerebral blood vessels, leading to cerebrovascular dysfunction. Pericytes, which wrap around vascular capillaries, are crucial for regulating cerebral blood flow, angiogenesis, and vessel stability. Despite the known impact of vascular dysfunction on the progression of neurodegenerative diseases, the specific role of pericytes in AD pathology remains to be elucidated., Methods: To explore this, we generated pericyte-like cells from human induced pluripotent stem cells (iPSCs) harboring the Swedish mutation in the amyloid precursor protein (APPswe) along with cells from healthy controls. We initially verified the expression of classic pericyte markers in these cells. Subsequent functional assessments, including permeability, tube formation, and contraction assays, were conducted to evaluate the functionality of both the APPswe and control cells. Additionally, bulk RNA sequencing was utilized to compare the transcriptional profiles between the two groups., Results: Our study reveals that iPSC-derived pericyte-like cells (iPLCs) can produce Aβ peptides. Notably, cells with the APPswe mutation secreted Aβ1-42 at levels ten-fold higher than those of control cells. The APPswe iPLCs also demonstrated a reduced ability to support angiogenesis and maintain barrier integrity, exhibited a prolonged contractile response, and produced elevated levels of pro-inflammatory cytokines following inflammatory stimulation. These functional changes in APPswe iPLCs correspond with transcriptional upregulation in genes related to actin cytoskeleton and extracellular matrix organization., Conclusions: Our findings indicate that the APPswe mutation in iPLCs mimics several aspects of CAA pathology in vitro, suggesting that our iPSC-based vascular cell model could serve as an effective platform for drug discovery aimed to ameliorate vascular dysfunction in AD., (© 2024. The Author(s).)

Stroke is a leading cause of permanent disability worldwide. Despite intensive research over the last decades, key anti-inflammatory strategies that have proven beneficial in pre-clinical animal models have often failed in translation. The importance of neutrophils as pro- and anti-inflammatory peripheral immune cells has often been overlooked in ischemic stroke. However, neutrophils rapidly infiltrate into the brain parenchyma after stroke and secrete an array of pro-inflammatory factors including reactive oxygen species, proteases, cytokines, and chemokines exacerbating damage. In this study, we demonstrate the neuroprotective and anti-inflammatory effect of benserazide, a clinically used DOPA decarboxylase inhibitor, using both in vitro models of inflammation and in vivo mouse models of focal cerebral ischemia. Benserazide significantly attenuated PMA-induced NETosis in isolated human neutrophils. Furthermore, benserazide was able to protect both SH-SY5Y and iPSC-derived human cortical neurons when challenged with activated neutrophils demonstrating the clinical relevance of this study. Additional in vitro data suggest the ability of benserazide to polarize macrophages towards M2-phenotypes following LPS stimulation. Neuroprotective effects of benserazide are further demonstrated by in vivo studies where peripheral administration of benserazide significantly attenuated neutrophil infiltration into the brain, altered microglia/macrophage phenotypes, and improved the behavioral outcome post-stroke. Overall, our data suggest that benserazide could serve as a drug candidate for the treatment of ischemic stroke. The importance of our results for future clinical trials is further underlined as benserazide has been approved by the European Medicines Agency as a safe and effective treatment in Parkinson's disease when combined with levodopa., (© 2024. The Author(s).)

Timely relief of edema and clearance of waste products, as well as promotion of anti-inflammatory immune responses, reduce ischemic stroke pathology, and attenuate harmful long-term effects post-stroke. The discovery of an extensive and functional lymphatic vessel system in the outermost meningeal layer, dura mater, has opened up new possibilities to facilitate post-stroke recovery by inducing dural lymphatic vessel (dLV) growth via a single injection of a vector encoding vascular endothelial growth factor C (VEGF-C). In the present study, we aimed to improve post-stroke outcomes by inducing dLV growth in mice. We injected mice with a single intracerebroventricular dose of adeno-associated viral particles encoding VEGF-C before subjecting them to transient middle cerebral artery occlusion (tMCAo). Behavioral testing, Gadolinium (Gd) contrast agent-enhanced magnetic resonance imaging (MRI), and immunohistochemical analysis were performed to define the impact of VEGF-C on the post-stroke outcome. VEGF-C improved stroke-induced behavioral deficits, such as gait disturbances and neurological deficits, ameliorated post-stroke inflammation, and enhanced an alternative glial immune response. Importantly, VEGF-C treatment increased the drainage of brain interstitial fluid (ISF) and cerebrospinal fluid (CSF), as shown by Gd-enhanced MRI. These outcomes were closely associated with an increase in the growth of dLVs around the region where we observed increased vefgc mRNA expression within the brain, including the olfactory bulb, cortex, and cerebellum. Strikingly, VEGF-C-treated ischemic mice exhibited a faster and stronger Gd-signal accumulation in ischemic core area and an enhanced fluid outflow via the cribriform plate. In conclusion, the VEGF-C-induced dLV growth improved the overall outcome post-stroke, indicating that VEGF-C has potential to be included in the treatment strategies of post-ischemic stroke. However, to maximize the therapeutic potential of VEGF-C treatment, further studies on the impact of an enhanced dural lymphatic system at clinically relevant time points are essential., (© 2024. The Author(s).)

Brain functionality relies on finely tuned regulation of gene expression by networks of non-coding RNAs (ncRNAs) such as the one composed by the circular RNA ciRS-7 (also known as CDR1as), the microRNA miR-7, and the long ncRNA Cyrano. We describe ischemia-induced alterations in the ncRNA network both in vitro and in vivo and in transgenic mice lacking ciRS-7 or miR-7. Our data show that cortical neurons downregulate ciRS-7 and Cyrano and upregulate miR-7 expression during ischemia. Mice lacking ciRS-7 exhibit reduced lesion size and motor impairment, while the absence of miR-7 alone results in increased ischemia-induced neuronal death. Moreover, miR-7 levels in pyramidal excitatory neurons regulate neurite morphology and glutamatergic signaling, suggesting a potential molecular link to the in vivo phenotype. Our data reveal the role of ciRS-7 and miR-7 in modulating ischemic stroke outcome, shedding light on the pathophysiological function of intracellular ncRNA networks in the brain., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The PSEN1 ΔE9 mutation causes a familial form of Alzheimer's disease (AD) by shifting the processing of amyloid precursor protein (APP) towards the generation of highly amyloidogenic Aβ42 peptide. We have previously shown that the PSEN1 ΔE9 mutation in human-induced pluripotent stem cell (iPSC)-derived astrocytes increases Aβ42 production and impairs cellular responses. Here, we injected PSEN1 ΔE9 mutant astrosphere-derived glial progenitors into newborn mice and investigated mouse behavior at the ages of 8, 12, and 16 months. While we did not find significant behavioral changes in younger mice, spatial learning and memory were paradoxically improved in 16-month-old PSEN1 ΔE9 glia-transplanted male mice as compared to age-matched isogenic control-transplanted animals. Memory improvement was associated with lower levels of soluble, but not insoluble, human Aβ42 in the mouse brain. We also found a decreased engraftment of PSEN1 ΔE9 mutant cells in the cingulate cortex and significant transcriptional changes in both human and mouse genes in the hippocampus, including the extracellular matrix-related genes. Overall, the presence of PSEN1 ΔE9 mutant glia exerted a more beneficial effect on aged mouse brain than the isogenic control human cells likely as a combination of several factors.

Background: Species-specific differences in astrocytes and their Alzheimer disease-associated pathology may influence cellular responses to other insults. Herein, human glial chimeric mice were generated to evaluate how Alzheimer disease predisposing genetic background in human astrocytes contributes to behavioral outcome and brain pathology after cortical photothrombotic ischemia., Methods: Neonatal (P0) immunodeficient mice of both sexes were transplanted with induced pluripotent stem cell-derived astrocyte progenitors from Alzheimer disease patients carrying PSEN1 exon 9 deletion (PSEN1 ΔE9), with isogenic controls, with cells from a healthy donor, or with mouse astrocytes or vehicle. After 14 months, a photothrombotic lesion was produced with Rose Bengal in the motor cortex. Behavior was assessed before ischemia and 1 and 4 weeks after the induction of stroke, followed by tissue perfusion for histology., Results: Open field, cylinder, and grid-walking tests showed a persistent locomotor and sensorimotor impairment after ischemia and female mice had larger infarct sizes; yet, these were not affected by astrocytes with PSEN1 ΔE9 background. Staining for human nuclear antigen confirmed that human cells successfully engrafted throughout the mouse brain. However, only a small number of human cells were positive for astrocytic marker GFAP (glial fibrillary acidic protein), mostly located in the corpus callosum and retaining complex human-specific morphology with longer processes compared with host counterparts. While host astrocytes formed the glial scar, human astrocytes were scattered in small numbers close to the lesion boundary. Aβ (beta-amyloid) deposits were not present in PSEN1 ΔE9 astrocyte-transplanted mice., Conclusions: Transplanted human cells survived and distributed widely in the host brain but had no impact on severity of ischemic damage after cortical photothrombosis in chimeric mice. Only a small number of transplanted human astrocytes acquired GFAP-positive glial phenotype or migrated toward the ischemic lesion forming glial scar. PSEN1 ΔE9 astrocytes did not impair behavioral recovery after experimental stroke.

Previous studies have implicated several brain cell types in schizophrenia (SCZ), but the genetic impact of astrocytes is unknown. Considering their high complexity in humans, astrocytes are likely key determinants of neurodevelopmental diseases, such as SCZ. Human induced pluripotent stem cell (hiPSC)-derived astrocytes differentiated from five monozygotic twin pairs discordant for SCZ and five healthy subjects were studied for alterations related to high genetic risk and clinical manifestation of SCZ in astrocyte transcriptomics, neuron-astrocyte co-cultures, and in humanized mice. We found gene expression and signaling pathway alterations related to synaptic dysfunction, inflammation, and extracellular matrix components in SCZ astrocytes, and demyelination in SCZ astrocyte transplanted mice. While Ingenuity Pathway Analysis identified SCZ disease and synaptic transmission pathway changes in SCZ astrocytes, the most consistent findings were related to collagen and cell adhesion associated pathways. Neuronal responses to glutamate and GABA differed between astrocytes from control persons, affected twins, and their unaffected co-twins and were normalized by clozapine treatment. SCZ astrocyte cell transplantation to the mouse forebrain caused gene expression changes in synaptic dysfunction and inflammation pathways of mouse brain cells and resulted in behavioral changes in cognitive and olfactory functions. Differentially expressed transcriptomes and signaling pathways related to synaptic functions, inflammation, and especially collagen and glycoprotein 6 pathways indicate abnormal extracellular matrix composition in the brain as one of the key characteristics in the etiology of SCZ., (© 2021 The Authors. GLIA published by Wiley Periodicals LLC.)

Lipid peroxidation-initiated ferroptosis is an iron-dependent mechanism of programmed cell death taking place in neurological diseases. Here we show that a condensed benzo[b]thiazine derivative small molecule with an arylthiazine backbone (ADA-409-052) inhibits tert-Butyl hydroperoxide (TBHP)-induced lipid peroxidation (LP) and protects against ferroptotic cell death triggered by glutathione (GSH) depletion or glutathione peroxidase 4 (GPx4) inhibition in neuronal cell lines. In addition, ADA-409-052 suppresses pro-inflammatory activation of BV2 microglia and protects N2a neuronal cells from cell death induced by pro-inflammatory RAW 264.7 macrophages. Moreover, ADA-409-052 efficiently reduces infarct volume, edema and expression of pro-inflammatory genes in a mouse model of thromboembolic stroke. Targeting ferroptosis may be a promising therapeutic strategy in neurological diseases involving severe neuronal death and neuroinflammation.

Ischemic stroke, the third leading cause of death in the Western world, affects mainly the elderly and is strongly associated with comorbid conditions such as atherosclerosis or diabetes, which are pathologically characterized by increased inflammation and are known to influence the outcome of stroke. Stroke incidence peaks during influenza seasons, and patients suffering from infections such as pneumonia prior to stroke exhibit a worse stroke outcome. Earlier studies have shown that comorbidities aggravate the outcome of stroke, yet the mediators of this phenomenon remain obscure. Here, we show that acute peripheral inflammation aggravates stroke-induced neuronal damage and motor deficits specifically in aged mice. This is associated with increased levels of plasma proinflammatory cytokines, rather than with an increase of inflammatory mediators in the affected brain parenchyma. Nascent transcriptomics data with mature microRNA sequencing were used to identify the neuron-specific miRNome, in order to decipher dysregulated miRNAs in the brains of aged animals with stroke and co-existing inflammation. We pinpoint a previously uninvestigated miRNA in the brain, miR-127, that is highly neuronal, to be associated with increased cell death in the aged, LPS-injected ischemic mice. Target prediction tools indicate that miR-127 interacts with several basally expressed neuronal genes, and of these we verify miR-127 binding to Psmd3. Finally, we report reduced expression of miR-127 in human stroke brains. Our results underline the impact of peripheral inflammation on the outcome of stroke in aged subjects and pinpoint molecular targets for restoring endogenous neuronal capacity to combat ischemic stroke., (© 2020 A.I. Virtanen Institute. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Background: Ischemic stroke is a devastating disease without a cure. The available treatments for ischemic stroke, thrombolysis by tissue plasminogen activator, and thrombectomy are suitable only to a fraction of patients and thus novel therapeutic approaches are urgently needed. The neuroinflammatory responses elicited secondary to the ischemic attack further aggravate the stroke-induced neuronal damage. It has been demonstrated that these responses are regulated at the level of non-coding RNAs, especially miRNAs., Methods: We utilized lentiviral vectors to overexpress miR-669c in BV2 microglial cells in order to modulate their polarization. To detect whether the modulation of microglial activation by miR-669c provides protection in a mouse model of transient focal ischemic stroke, miR-669c overexpression was driven by a lentiviral vector injected into the striatum prior to induction of ischemic stroke., Results: Here, we demonstrate that miR-669c-3p, a member of chromosome 2 miRNA cluster (C2MC), is induced upon hypoxic and excitotoxic conditions in vitro and in two different in vivo models of stroke. Rather than directly regulating the neuronal survival in vitro, miR-669c is capable of attenuating the microglial proinflammatory activation in vitro and inducing the expression of microglial alternative activation markers arginase 1 (Arg1), chitinase-like 3 (Ym1), and peroxisome proliferator-activated receptor gamma (PPAR-γ). Intracerebral overexpression of miR-669c significantly decreased the ischemia-induced cell death and ameliorated the stroke-induced neurological deficits both at 1 and 3 days post injury (dpi). Albeit miR-669c overexpression failed to alter the overall Iba1 protein immunoreactivity, it significantly elevated Arg1 levels in the ischemic brain and increased colocalization of Arg1 and Iba1. Moreover, miR-669c overexpression under cerebral ischemia influenced several morphological characteristics of Iba1 positive cells. We further demonstrate the myeloid differentiation primary response gene 88 (MyD88) transcript as a direct target for miR-669c-3p in vitro and show reduced levels of MyD88 in miR-669c overexpressing ischemic brains in vivo., Conclusions: Collectively, our data provide the evidence that miR-669c-3p is protective in a mouse model of ischemic stroke through enhancement of the alternative microglial/macrophage activation and inhibition of MyD88 signaling. Our results accentuate the importance of controlling miRNA-regulated responses for the therapeutic benefit in conditions of stroke and neuroinflammation.

Community forestry (CF) is increasingly recognized as one of the key solutions to forest management and governance challenges in the developing world. At its heart is the participation of local communities in forest management, bringing multiple benefits in forest health as well as the wellbeing of the local community and the wider society. However, CF in Nepal is felt by many to not be delivering on its potential. This paper presents illustrative examples from a capacity development needs assessment (CDNA), which was conducted to understand challenges CF in Nepal faces, focusing on two districts: Rupandehi and Sindhupalchok. The application of the CDNA framework revealed the multiple challenges facing the CF program. They range from lack of multi-stakeholder platform at high level for discussion and resolution of CF-related issues, lack of system to review the program as well as systematic capacity development opportunities for stakeholders and weak governance, altogether contributing to costly and complex process for local community to participate and benefit from the program. We argue that behind these challenges underlie the distrust to local communities and tendency to retain power inequality that is tilted toward techno-bureaucracy, limitation of the program within sectoral boundary of forestry as well as lack of political commitment and overall weak capacity of the status apparatus. We highlight that revitalizing the CF program requires utilization of state restructuring process to expand its scope and improve its governance. Similarly, we emphasize the need for coordinated efforts among stakeholders to relax regulatory burdens, to engage local communities in policy processes, and to set up a capacity development system for supporting community rights mainly in the commercial management of community forests-to ensure that CF delivers.

Neuroinflammation is strongly induced by cerebral ischemia. The early phase after the onset of ischemic stroke is characterized by acute neuronal injury, microglial activation, and subsequent infiltration of blood-derived inflammatory cells, including macrophages. Therefore, modulation of the microglial/macrophage responses has increasingly gained interest as a potential therapeutic approach for the ischemic stroke. In our study, we investigated the effects of peripherally administered interleukin 13 (IL-13) in a mouse model of permanent middle cerebral artery occlusion (pMCAo). Systemic administration of IL-13 immediately after the ischemic insult significantly reduced the lesion volume, alleviated the infiltration of CD45 + leukocytes, and promoted the microglia/macrophage alternative activation within the ischemic region, as determined by arginase 1 (Arg1) immunoreactivity at 3 days post-ischemia (dpi). Moreover, IL-13 enhanced the expression of M2a alternative activation markers Arg1 and Ym1 in the peri-ischemic (PI) area, as well as increased plasma IL-6 and IL-10 levels at 3 dpi. Furthermore, IL-13 treatment ameliorated gait disturbances at day 7 and 14 and sensorimotor deficits at day 14 post-ischemia, as analyzed by the CatWalk gait analysis system and adhesive removal test, respectively. Finally, IL-13 treatment decreased neuronal cell death in a coculture model of neuroinflammation with RAW 264.7 macrophages. Taken together, delivery of IL-13 enhances microglial/macrophage anti-inflammatory responses in vivo and in vitro, decreases ischemia-induced brain cell death, and improves sensory and motor functions in the pMCAo mouse model of cerebral ischemia.

Background: Ischemic stroke is one of the main causes of death and disability worldwide. It is caused by the cessation of cerebral blood flow resulting in the insufficient delivery of glucose and oxygen to the neural tissue. The inflammatory response initiated by ischemic stroke in order to restore tissue homeostasis in the acute phase of stroke contributes to delayed brain damage., Methods: By using in vitro models of neuroinflammation and in vivo model of permanent middle cerebral artery occlusion, we demonstrate the neuroprotective and anti-inflammatory effects of sulfosuccinimidyl oleate sodium (SSO)., Results: SSO significantly reduced the lipopolysaccharide/interferon-γ-induced production of nitric oxide, interleukin-6 and tumor necrosis factor-α, and the protein levels of inflammatory enzymes including nitric oxide synthase 2, cyclooxygenase-2 (COX-2), and p38 mitogen-activated protein kinase (MAPK) in microglia, without causing cell toxicity. Although SSO failed to directly alleviate glutamate-induced excitotoxicity in murine cortical neurons, it prevented inflammation-induced neuronal death in microglia-neuron co-cultures. Importantly, oral administration of SSO in Balb/c mice subjected to permanent occlusion of the middle cerebral artery reduced microglial activation in the peri-ischemic area and attenuated brain damage. This in vivo neuroprotective effect of SSO was associated with a reduction in the COX-2 and heme oxygenase-1 immunoreactivities., Conclusions: Our results suggest that SSO is an anti-inflammatory and a possible therapeutic candidate in diseases such as stroke where inflammation is a central hallmark.

Background: DHCR24, involved in the de novo synthesis of cholesterol and protection of neuronal cells against different stress conditions, has been shown to be selectively downregulated in neurons of the affected brain areas in Alzheimer's disease., Methods: Here, we investigated whether the overexpression of DHCR24 protects neurons against inflammation-induced neuronal death using co-cultures of mouse embryonic primary cortical neurons and BV2 microglial cells upon acute neuroinflammation. Moreover, the effects of DHCR24 overexpression on dendritic spine density and morphology in cultured mature mouse hippocampal neurons and on the outcome measures of ischemia-induced brain damage in vivo in mice were assessed., Results: Overexpression of DHCR24 reduced the loss of neurons under inflammation elicited by LPS and IFN-γ treatment in co-cultures of mouse neurons and BV2 microglial cells but did not affect the production of neuroinflammatory mediators, total cellular cholesterol levels, or the activity of proteins linked with neuroprotective signaling. Conversely, the levels of post-synaptic cell adhesion protein neuroligin-1 were significantly increased upon the overexpression of DHCR24 in basal growth conditions. Augmentation of DHCR24 also increased the total number of dendritic spines and the proportion of mushroom spines in mature mouse hippocampal neurons. In vivo, overexpression of DHCR24 in striatum reduced the lesion size measured by MRI in a mouse model of transient focal ischemia., Conclusions: These results suggest that the augmentation of DHCR24 levels provides neuroprotection in acute stress conditions, which lead to neuronal loss in vitro and in vivo.

Developing new therapies for stroke is urgently needed, as this disease is the leading cause of death and disability worldwide, and the existing treatment is only available for a small subset of patients. The interruption of blood flow to the brain during ischemic stroke launches multiple immune responses, characterized by infiltration of peripheral immune cells, the activation of brain microglial cells, and the accumulation of immune mediators. Copper is an essential trace element that is required for many critical processes in the brain. Copper homeostasis is disturbed in chronic neurodegenerative diseases and altered in stroke patients, and targeted copper delivery has been shown to be protective against chronic neurodegeneration. This study was undertaken to assess whether the copper bis(thiosemicarbazone) complex, Cu II (atsm), is beneficial in acute brain injury, in preclinical mouse models of ischemic stroke. We demonstrate that the copper complex Cu II (atsm) protects neurons from excitotoxicity and N2a cells from OGD in vitro, and is protective in permanent and transient ischemia models in mice as measured by functional outcome and lesion size. Copper delivery in the ischemic brains modulates the inflammatory response, specifically affecting the myeloid cells. It reduces CD45 and Iba1 immunoreactivity, and alters the morphology of Iba1 positive cells in the ischemic brain. Cu II (atsm) also protects endogenous microglia against ischemic insult and reduces the proportion of invading monocytes. These results demonstrate that the copper complex Cu II (atsm) is an inflammation-modulating compound with high therapeutic potential in stroke and is a strong candidate for the development of therapies for acute brain injury.

Collagens are key structural components of basement membranes, providing a scaffold for other components or adhering cells. Collagens and collagen-derived active fragments contribute to biological activities such as cell growth, differentiation and migration. Here, we report that collagen XV knock-out (ColXV KO) mice are resistant to experimental ischemic stroke. Interestingly, the infarcts of ColXV KO mice were as small as those of wild-type (WT) mice thrombolysed with recombinant tissue plasminogen activator (rtPA), the actual treatment for ischemic stroke. Importantly, there were no differences in the architecture of cerebrovascular anatomy between WT and ColXV KO mice. We found a twofold increase of the most potent pro-angiogenic factor, type A vascular growth endothelial factor (VEGF-A) in the ipsilateral cortex of rtPA-treated ischemic WT mice compared with untreated ischemic and sham-operated counterparts. A similar increase of VEGF-A was also found in both rtPA and untreated ischemic ColXV KO mice compared with sham ColXV KO mice. Finally, we evidenced that the levels of ColXV were increased in the plasma of WT mice treated with rtPA compared with untreated ischemic counterparts. Altogether, this study indicates that the lack ColXV is protective after stroke and that the degradation of endothelial ColXV may contribute to the beneficial effect of rtPA after ischemic stroke. The neuroprotection observed in ColXV KO mice may be attributed to the increased VEGF-A production following stroke in the ischemic territory.

Stroke is a highly debilitating, often fatal disorder for which current therapies are suitable for only a minor fraction of patients. Discovery of novel, effective therapies is hampered by the fact that advanced age, primary age-related tauopathy or comorbidities typical to several types of dementing diseases are usually not taken into account in preclinical studies, which predominantly use young, healthy rodents. Here we investigated for the first time the neuroprotective potential of bexarotene, an FDA-approved agent, in a co-morbidity model of stroke that combines high age and tauopathy with thromboembolic cerebral ischemia. Following thromboembolic stroke bexarotene enhanced autophagy in the ischemic brain concomitantly with a reduction in lesion volume and amelioration of behavioral deficits in aged transgenic mice expressing the human P301L-Tau mutation. In in vitro studies bexarotene increased the expression of autophagy markers and reduced autophagic flux in neuronal cells expressing P301L-Tau. Bexarotene also restored mitochondrial respiration deficits in P301L-Tau neurons. These newly described actions of bexarotene add to the growing amount of compelling data showing that bexarotene is a potent neuroprotective agent, and identify a novel autophagy-modulating effect of bexarotene.

Stroke represents a global challenge and is a leading cause of permanent disability worldwide. Despite much effort, translation of research findings to clinical benefit has not yet been successful. Failure of neuroprotection trials is considered, in part, due to the low quality of preclinical studies, low level of reproducibility across different laboratories and that stroke co-morbidities have not been fully considered in experimental models. More rigorous testing of new drug candidates in different experimental models of stroke and initiation of preclinical cross-laboratory studies have been suggested as ways to improve translation. However, to our knowledge, no drugs currently in clinical stroke trials have been investigated in preclinical cross-laboratory studies. The cytokine interleukin 1 is a key mediator of neuronal injury, and the naturally occurring interleukin 1 receptor antagonist has been reported as beneficial in experimental studies of stroke. In the present paper, we report on a preclinical cross-laboratory stroke trial designed to investigate the efficacy of interleukin 1 receptor antagonist in different research laboratories across Europe. Our results strongly support the therapeutic potential of interleukin 1 receptor antagonist in experimental stroke and provide further evidence that interleukin 1 receptor antagonist should be evaluated in more extensive clinical stroke trials., (© The Author(s) 2015.)

Transient forebrain ischemia induces delayed death of the hippocampal pyramidal neurons, particularly in the CA2 and medial CA1 area. Early pharmacological inhibition of inflammatory response can ameliorate neuronal death, but it also inhibits processes leading to tissue regeneration. Therefore, research efforts are now directed to modulation of post-ischemic inflammation, with the aim to promote beneficial effects of inflammation and limit adverse effects. Transcription factor NF-κB plays a key role in the inflammation and cell survival/apoptosis pathways. In the brain, NF-κB is predominantly found in the form of a heterodimer of p65 (RelA) and p50 subunit, where p65 has a transactivation domain while p50 is chiefly involved in DNA binding. In this study, we subjected middle-aged Nfkb1 knockout mice (lacking p50 subunit) and wild-type controls of both sexs to 17 min of transient forebrain ischemia and assessed mouse performance in a panel of behavioral tests after two weeks of post-operative recovery. We found that ischemia failed to induce clear memory and motor deficits, but affected spontaneous locomotion in genotype- and sex-specific way. We also show that both the lack of the NF-κB p50 subunit and female sex independently protected CA2 hippocampal neurons from ischemia-induced cell death. Additionally, the NF-κB p50 subunit deficiency significantly reduced ischemia-induced microgliosis, astrogliosis, and neurogenesis. Lower levels of hippocampal microgliosis significantly correlated with faster spatial learning. We conclude that NF-κB regulates the outcome of transient forebrain ischemia in middle-aged subjects in a sex-specific way, having an impact not only on neuronal death but also specific inflammatory responses and neurogenesis.

Cerebral stroke induces massive Th1-shifted inflammation both in the brain and the periphery, contributing to the outcome of stroke. A Th1-type response is neurotoxic whereas a Th2-type response is accompanied by secretion of anti-inflammatory cytokines, such as interleukin-4 (IL-4). Interleukin-33 (IL-33) is a cytokine known to induce a shift towards the Th2-type immune response, polarize macrophages/microglia towards the M2-type, and induce production of anti-inflammatory cytokines. We found that the plasma levels of the inhibitory IL-33 receptor, sST2, are increased in human stroke and correlate with a worsened stroke outcome, suggesting an insufficient IL-33-driven Th2-type response. In mouse, peripheral administration of IL-33 reduced stroke-induced cell death and improved the sensitivity of the contralateral front paw at 5days post injury. The IL-33-treated mice had increased levels of IL-4 in the spleen and in the peri-ischemic area of the cortex. Neutralization of IL-4 by administration of an IL-4 antibody partially prevented the IL-33-mediated protection. IL-33 treatment also reduced astrocytic activation in the peri-ischemic area and increased the number of Arginase-1 immunopositive microglia/macrophages at the lesion site. In human T-cells, IL-33 treatment induced IL-4 secretion, and the conditioned media from IL-33-exposed T-cells reduced astrocytic activation. This study demonstrates that IL-33 is protective against ischemic insult by induction of IL-4 secretion and may represent a novel therapeutic approach for the treatment of stroke., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Neural stem/progenitor cells (NPCs) proliferate and produce new neurons in neurogenic areas throughout the lifetime. While these cells represent potential therapeutic treatment of neurodegenerative diseases, regulation of neurogenesis is not completely understood. We show that deficiency of nuclear factor erythroid 2-related factor (Nrf2), a transcription factor induced in response to oxidative stress, prevents the ischemia-induced increase in newborn neurons in the subgranular zone of the dentate gyrus. Consistent with this finding, the growth of NPC neurospheres was increased by lentivirus-mediated overexpression of Nrf2 gene or by treatment with pyrrolidine dithiocarbamate (PDTC), an Nrf2 activating compound. Also, neuronal differentiation of NPCs was increased by Nrf2 overexpression or PDTC treatment but reduced by Nrf2 deficiency. To investigate the impact of Nrf2 on NPCs in Alzheimer's disease (AD), we treated NPCs with amyloid beta (Aβ), a toxic peptide associated with neurodegeneration and cognitive abnormalities in AD. We found that Aβ1-42-induced toxicity and reduction in neurosphere proliferation were prevented by Nrf2 overexpression, while Nrf2 deficiency enhanced the Aβ1-42-induced reduction of neuronal differentiation. On the other hand, Aβ1-40 had no effect on neurosphere proliferation in wt NPCs but increased the proliferation of Nrf2 overexpressing neurospheres and reduced it in Nrf2-deficient neurospheres. These results suggest that Nrf2 is essential for neuronal differentiation of NPCs, regulates injury-induced neurogenesis and provides protection against Aβ-induced NPC toxicity., (© 2014 AlphaMed Press.)

Inflammation is a major mechanism of acute brain injury and chronic neurodegeneration. This neuroinflammation is known to be substantially regulated by the transcription factor NF-κB, which is predominantly found in the form of heterodimer of p65 (RelA) and p50 subunit, with p50/p50 homodimers being also common. The p65 subunit has a transactivation domain, whereas p50 is chiefly involved in DNA binding. Binding of the p65/p50 heterodimers is thought to induce expression of numerous proinflammatory genes in microglia. Here we show that cultured microglia deficient for the gene (Nfkb1) encoding p50 subunit show reduced induction of proinflammatory mediators, increased expression of anti-inflammatory genes, and increased expression of CD45, an immunoregulatory molecule, in response to lipopolysaccharide (LPS) exposure, but increased capacity to take up β-amyloid (Aβ) which is associated with enhanced release of tumor necrosis factor alpha (TNFα). However, Nfkb1 deficiency strongly increases leukocyte infiltration and the expression of proinflammatory genes in response to intrahippocampal administration of LPS. Also, when crossing Nfkb1 deficient mice with APdE9 transgenic mice the expression of proinflammatory genes was strongly enhanced, whereas Aβ burden was slightly but significantly reduced. These alterations in expression of inflammatory mediators in Nfkb1 deficient mice were associated with reduced expression of CD45. Our data demonstrates a crucial and complex role p50 subunit of NF-κB in brain inflammation, especially in regulating the phenotype of microglia after acute and chronic inflammatory insults relevant to clinical conditions, contributing to both pro-inflammatory and anti-inflammatory responses of microglia, infiltration of leukocytes, and clearance of Aβ in Alzheimer's disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Polyamines, spermidine, spermine and their precursor putrescine, are ubiquitous cell components essential for normal cell growth. Increased polyamine levels and enhanced biosynthesis have been associated with malignant transformation and tumor formation, and thus, the polyamines have been considered to be a meaningful target to cancer therapies. However, clinical cancer treatment trials using inhibitors of polyamine synthesis have been unsuccessful probably due to compensatory uptake of polyamines from extracellular sources. The antizyme proteins regulate both polyamine biosynthesis and transport, and thus, the antizymes could provide an efficient approach to control cellular proliferation compared to the mere inhibition of biosynthesis. To define the role of antizymes in proliferative processes associated with the whole animal, we have generated transgenic mice overexpressing mouse antizyme 1 gene under its own regulatory sequences. Antizyme 1 protein was abundantly expressed in various organs and the expressed antizyme protein was functional as ornithine decarboxylase activity was significantly reduced in all tissues analyzed. However, antizyme 1 overexpression caused only minor changes in tissue polyamine levels demonstrating the challenges in using the "antizyme approach" to deplete polyamines in a living animal. Neither were there any changes in cellular proliferation in the proliferative tissues of transgenic animals. Interestingly though, there was occurrence of abnormally high level of apoptosis in the non-proliferating part of the colon epithelia. Otherwise, the transgenic founder mice appeared healthy and out of seven founders six were fertile. However, none of the founders could transmit the transgene suggesting that the antizyme 1 overexpression may be deleterious to transgenic gametes.

Ischemic stroke is confounded by conditions such as atherosclerosis, diabetes, and infection, all of which alter peripheral inflammatory processes with concomitant impact on stroke outcome. The majority of the stroke patients are elderly, but the impact of interactions between aging and inflammation on stroke remains unknown. We thus investigated the influence of age on the outcome of stroke in animals predisposed to systemic chronic infection. Th1-polarized chronic systemic infection was induced in 18-22 month and 4-month-old C57BL/6j mice by administration of Trichuris muris (gut parasite). One month after infection, mice underwent permanent middle cerebral artery occlusion and infarct size, brain gliosis, and brain and plasma cytokine profiles were analyzed. Chronic infection increased the infarct size in aged but not in young mice at 24 h. Aged, ischemic mice showed altered plasma and brain cytokine responses, while the lesion size correlated with plasma prestroke levels of RANTES. Moreover, the old, infected mice exhibited significantly increased neutrophil recruitment and upregulation of both plasma interleukin-17α and tumor necrosis factor-α levels. Neither age nor infection status alone or in combination altered the ischemia-induced brain microgliosis. Our results show that chronic peripheral infection in aged animals renders the brain more vulnerable to ischemic insults, possibly by increasing the invasion of neutrophils and altering the inflammation status in the blood and brain. Understanding the interactions between age and infections is crucial for developing a better therapeutic regimen for ischemic stroke and when modeling it as a disease of the elderly., (© 2013 The Anatomical Society and John Wiley & Sons Ltd.)

Background: Numerous clinical trials in stroke have failed, most probably partially due to preclinical studies using young, healthy male rodents with little relevance to the heterogenic conditions of human stroke. Co-morbid conditions such as atherosclerosis and infections coupled with advanced age are known to contribute to increased risk of cerebrovascular diseases. Clinical and preclinical studies have shown that the E4 allele of human apolipoprotein (ApoE4) is linked to poorer outcome in various conditions of brain injury and neurodegeneration, including cerebral ischemia. Since ApoE is a known regulator of lipid homeostasis, we studied the impact of a high-cholesterol diet in aged mice in the context of relevant human ApoE isoforms on the outcome of focal brain ischemia., Methods: Aged mice expressing human E3 and E4 isoforms of ApoE in C57BL/6J background and C57BL/6J mice fed on either a high-fat diet or a normal diet underwent permanent middle cerebral artery occlusion. The impact of a high-cholesterol diet was assessed by measuring the serum cholesterol level and the infarction volume was determined by magnetic resonance imaging. Sensorimotor deficits were assessed using an adhesive removal test and the findings were correlated with inflammatory markers., Results: We show that expression of human ApoE4 renders aged mice fed with a western-type diet more susceptible to sensorimotor deficits upon stroke. These deficits are not associated with atherosclerosis but are accompanied with altered astroglial activation, neurogenesis, cyclooxygenase-2 immunoreactivity and increased plasma IL-6., Conclusions: Our results support the hypothesis that ApoE alleles modify the inflammatory responses in the brain and the periphery, thus contributing to altered functional outcome following stroke.