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Exosomes are emerging as potent and safe delivery carriers for use in vaccinology and therapeutics. A better vaccine for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is needed to provide improved, broader, longer lasting neutralization of SARS-CoV-2, a more robust T cell response, enable widespread global usage, and further enhance the safety profile of vaccines given the likelihood of repeated booster vaccinations. Here, we use Capricor's StealthXTM platform to engineer exosomes to express native SARS-CoV-2 spike Delta variant (STX-S) protein on the surface for the delivery of a protein-based vaccine for immunization against SARS-CoV-2 infection. The STX-S vaccine induced a strong immunization with the production of a potent humoral immune response as demonstrated by high levels of neutralizing antibody not only against the delta SARS-CoV-2 virus but also two Omicron variants (BA.1 and BA.5), providing broader protection than current mRNA vaccines. Additionally, both CD4+ and CD8+ T cell responses were increased significantly after treatment. Quantification of spike protein by ELISA showed that only nanograms of protein were needed to induce a potent immune response. This is a significantly lower dose than traditional recombinant protein vaccines with no adjuvant required, which makes the StealthXTM exosome platform ideal for the development of multivalent vaccines with a better safety profile. Importantly, our exosome platform allows novel proteins, or variants in the case of SARS-CoV-2, to be engineered onto the surface of exosomes in a matter of weeks, comparable with mRNA vaccine technology, but without the cold storage requirements necessary for mRNA vaccines. The ability to utilize exosomes for cellular delivery of proteins, as demonstrated by STX-S, has enormous potential to revolutionize vaccinology by rapidly facilitating antigen presentation at an extremely low dose resulting in a potent, broad antibody response. [ABSTRACT FROM AUTHOR]

Background: Air pollution is widely associated with accelerated cognitive decline at later ages and risk of Alzheimer's disease (AD). Correspondingly, rodent models demonstrate the neurotoxicity of ambient air pollution and its components. Our studies with nano-sized particulate matter (nPM) from urban Los Angeles collected since 2009 have shown pro-amyloidogenic and pro-inflammatory responses. However, recent batches of nPM have diminished induction of the glutamate receptor GluA1 subunit, Iba1, TNFα, Aβ42 peptide, and white matter damage. The same methods, materials, and mouse genotypes were used throughout.Objective: Expand the nPM batch comparisons and evaluate archived brain samples to identify the earliest change in nPM potency.Methods: Batches of nPM were analyzed by in vitro cell assays for NF-κB and Nrf2 induction for comparison with in vivo responses of mouse brain regions from mice exposed to these batches, analyzed by PCR and western blot.Results: Five older nPM batches (2009-2017) and four recent nPM batches (2018, 2019) for NF-κB and Nrf2 induction showed declines in nPM potency after 2017 that paralleled declines of in vivo activity from independent exposures in different years.Conclusion: Transcription-based in vitro assays of nPM corresponded to the loss of in vivo potency for inflammatory and oxidative responses. These recent decreases of nPM neurotoxicity give a rationale for evaluating possible benefits to the risk of dementia and stroke in Los Angeles populations. [ABSTRACT FROM AUTHOR]

Intra-muscle fiber accumulation of ubiquitinated protein aggregates containing several conformationally modified proteins, including amyloid-β and phosphorylated tau, is characteristic of the pathologic phenotype of sporadic inclusion-body myositis (s-IBM), the most common progressive degenerative myopathy of older persons. Abnormalities of protein-degradation, involving both the 26S proteasome and autophagic-lysosomal pathways, were previously demonstrated in s-IBM muscle. NBR1 is a ubiquitin-binding scaffold protein importantly participating in autophagic degradation of ubiquitinated proteins. Whereas abnormalities of p62, a ubiquitin-binding protein, were previously described in s-IBM, abnormalities of NBR1 have not been reported in s-IBM. We have now identified in s-IBM muscle biopsies that NBR1, by: (a) immunohistochemistry, was strongly accumulated within s-IBM muscle-fiber aggregates, where it closely co-localized with p62, ubiquitin, and phosphorylated tau; (b) immunoblots, was increased threefold ( p < 0.001); and (c) immunoprecipitation, was associated with p62 and LC3. By real-time PCR, NBR1 mRNA was increased twofold ( p < 0.01). None of the various disease- and normal-control muscle biopsies had any NBR1 abnormality. In cultured human muscle fibers, NBR1 also physically associated with both p62 and LC3, and experimental inhibition of either the 26S proteasome or the lysosomal activity resulted in NBR1 increase. Our demonstration of NBR1 abnormalities in s-IBM provides further evidence that altered protein degradation pathways may be critically involved in the s-IBM pathogenesis. Accordingly, attempts to unblock defective protein degradation might be a therapeutic strategy for s-IBM patients. [ABSTRACT FROM AUTHOR]

Dysferlin is a 237-kDa transmembrane protein involved in calcium-mediated sarcolemma resealing. Dysferlin gene mutations cause limb-girdle muscular dystrophy (LGMD) 2B, Miyoshi myopathy (MM) and distal myopathy of the anterior tibialis. Considering that a secondary Dysferlin reduction has also been described in other myopathies, our original goal was to identify cases with a Dysferlin deficiency without dysferlin gene mutations. The dysferlin gene is huge, composed of 55 exons that span 233 140 bp of genomic DNA. We performed a thorough mutation analysis in 65 LGMD/MM patients with ≤20% Dysferlin. The screening was exhaustive, as we sequenced both genomic DNA and cDNA. When required, we used other methods, including real-time PCR, long PCR and array CGH. In all patients, we were able to recognize the primary involvement of the dysferlin gene. We identified 38 novel mutation types. Some of these, such as a dysferlin gene duplication, could have been missed by conventional screening strategies. Nonsense-mediated mRNA decay was evident in six cases, in three of which both alleles were only detectable in the genomic DNA but not in the mRNA. Among a wide spectrum of novel gene defects, we found the first example of a 'nonstop' mutation causing a dysferlinopathy. This study presents the first direct and conclusive evidence that an amount of Dysferlin ≤20% is pathogenic and always caused by primary dysferlin gene mutations. This demonstrates the high specificity of a marked reduction of Dysferlin on western blot and the value of a comprehensive molecular approach for LGMD2B/MM diagnosis. [ABSTRACT FROM AUTHOR]

Background: AD is associated with increases in lipid peroxidation and decreased antioxidant defense. Little is known about antioxidant defense of brain lipid rafts which are the site of APP processing. Oxidative damage to lipid rafts increases amyloid production. Lipid hydroperoxidases, glutathione peroxidase 4 (GPx4) and peroxiredoxin 6 (Prdx6) are potential detoxifying enzymes capable of reducing oxidized phospholipids and oxysterols within cellular membranes. We hypothesized that lipid hydroperoxidases would be reduced in ApoE4 carriers, which also have deficient cholesterol transport. To determine this, we investigated the status of the AD lipid raft by ApoE isoform in human frontal cortex. Method: Lipid rafts were isolated (Invent Biotechnologies; LR‐039) from postmortem AD (ApoE3,3, ApoE3,4, and ApoE4,4) and age‐matched controls (ApoE3,3) and were analyzed by western blot and enzymatic assay. Result: Lipid raft ApoE protein level of AD brains showed large decreases: 43% (Ctl vs AD3,3; p<0.01), 53% (Ctl vs AD3,4; p<0.001) and 70% (Ctl vs AD4,4; p<0.0001). Total cholesterol content of lipid rafts decreased 25% (Ctl vs AD; p<0.01). GPx4 protein of lipid rafts decreased 30% (Ctl vs AD3,3; p<0.01), 45% (Ctl vs AD3,4; p<0.0001) and 57% (Ctl vs AD4,4; p<0.001). AD brains from ApoE4,4 carriers had 36%(p<0.01) lower GPx4 localization than ApoE3,3. Prdx6 localization was not detected in the lipid rafts. Conclusion: Reductions in ApoE, total cholesterol, and GPx4 in the AD lipid raft suggests its vulnerability during AD. Furthermore, many of these decreases were ApoE isoform dependent, suggesting that antioxidant defense may differ by ApoE isoform. [ABSTRACT FROM AUTHOR]

Background: Cerebral microbleeds (MBs) are more prevalent in APOE4 carriers and associated with Alzheimer Disease (AD). Loss of cerebrovascular integrity, blood‐brain barrier breakdown, and reduced Aβ clearance are also shown by APOE4 carrier mice. In AD, increased brain amyloid coupled with increased oxidative stress may synergistically contribute to MBs prevalence. EFAD mice carrying familial dominant AD genes in combination with human apoE alleles are a model for sex differences in AD, with greater amyloid accumulation and microbleeds in female E4FAD mice. Independently, Aβ and heme from microbleeds contribute to increased oxidative stress. New data suggest increased lipid peroxidation in E4 mice at baseline as measured by 4‐HNE. Peroxiredoxin 6 (Prdx6) and glutathione peroxidase 4 (GPx4) are the only known antioxidant enzymes capable of detoxifying oxidized phospholipids. Method: Prdx6 and GPx4 were studied by Western blot in cortical lysates of 6mo old EFAD mice. To evaluate Aβ contribution, we analyzed Prdx6 and GPx4 in 2mo old EFAD mice, which show 2 to 10‐fold lower Aβ compared to 6mo EFAD mice, and 6mo old EFAD non carrier siblings (no AD‐transgene). Result: Prdx6 was decreased in E4FAD mice (M: ‐30%; F: ‐50%) compared to E3FAD mice. GPx4 was decreased only n E4FAD females (M: no change; F: ‐50%). Younger mice did not show differences in either lipid peroxidases at 2mo. Interestingly, Prdx6 but not GPx4 was reduced in 6mo old female E4FAD non carrier siblings. Conclusion: Our data suggest that the decreased levels of Prdx6 in 6mo old female E4 carrier mice in absence of Aβ might impair their ability to reduce oxidized lipids and predispose them to membrane alteration. Moreover, aging and increase of Aβ load together might contribute to the reduction of GPx4 levels in APOE4 carriers and the accumulation of MBs. Additional studies are required to clarify the underlying interaction sex‐apoE allele in MBs onset and its contribution to the pathogenesis of AD. [ABSTRACT FROM AUTHOR]

Introduction: Chronic air pollution (AirPoll) is associated with accelerated cognitive decline and risk of Alzheimer's disease (AD). Correspondingly, wild-type and AD-transgenic rodents exposed to AirPoll have increased amyloid peptides and behavioral impairments., Methods: We examined the γ-secretase modulator GSM-15606 for potential AirPoll protection by its attenuating of amyloid beta (Aβ)42 peptide production. Male and female wild-type mice were fed GSM-15606 during an 8-week inhalation exposure to AirPoll subfractions, ambient nanoparticulate matter (nPM), and diesel exhaust particles (DEP)., Results: GSM-15606 decreased Aβ42 during nPM and DEP exposure without changing beta- or gamma-secretase activity or BACE1 and PS1 protein levels. DEP increased lateral ventricle volume by 25%., Discussion: These enzyme responses are relevant to AD drug treatments, as well as to the physiological functions of the Aβ42 peptide. GSM-15606 attenuation of Aβ42 may benefit human exposure to AirPoll., Highlights: Gamma-secretase modulator (GSM-15606) attenuates the amyloidogenic amyloid beta (Aβ)42 peptide during exposure to air pollution, which may be a mechanism by which air pollution increases Alzheimer's disease (AD) risk. AD drug treatments may also consider Aβ homeostasis among the chronic effects of GSM-15606 and other amyloid reduction treatments on secretase enzymes., (© 2024 The Author(s). Alzheimer's & Dementia published by Wiley Periodicals LLC on behalf of Alzheimer's Association.)

Currently approved vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have focused solely on the spike protein to provide immunity. The first vaccines were developed rapidly using spike mRNA delivered by lipid nanoparticles but required ultralow-temperature storage and have had limited immunity against variations in spike. Subsequently, protein-based vaccines were developed, which offer broader immunity but require significant time for development and the use of an adjuvant to boost the immune response. Here, exosomes were used to deliver a bivalent protein-based vaccine in which two independent viral proteins were used. Exosomes were engineered to express either SARS-CoV-2 delta spike (Stealth X-Spike [STX-S]) or the more conserved nucleocapsid (Stealth X-Nucleocapsid [STX-N]) protein on the surface. When administered as a single product (STX-S or STX-N) or in combination (STX-S+N), both STX-S and STX-N induced strong immunization with the production of potent humoral and cellular immune responses. Interestingly, these results were obtained with the administration of only nanograms of protein and without an adjuvant. In two independent animal models (mouse and rabbit), the administration of nanograms of the STX-S+N vaccine resulted in increased antibody production, potent neutralizing antibodies with cross-reactivity to other variants of spike, and strong T-cell responses. Importantly, no competition of immune responses was observed, allowing the delivery of nucleocapsid with spike to offer improved SARS-CoV-2 immunity. These data show that the StealthX exosome platform has the enormous potential to revolutionize vaccinology by combining the advantages of mRNA and recombinant protein vaccines into a superior, rapidly generated, low-dose vaccine resulting in potent, broader immunity. IMPORTANCE The pandemic emergency has brought to light the need for a new generation of rapidly developed vaccines that induce longer-lasting, potent, and broader immune responses. While the mRNA vaccines played a critical role during the emergency in reducing SARS-CoV-2 hospitalization rates and deaths, more efficient approaches are needed. A multivalent, protein-based vaccine delivered by exosomes could meet this urgent need due to the high speed of development, manufacturability, and the ability to produce a strong antibody response, with neutralizing antibodies and a strong T-cell response able to broadly combat viral infection with a minimum number of injections., Competing Interests: The authors declare a conflict of interest. The authors are employees of Capricor Therapeutics.

The neurotoxicity of air pollution is undefined for sex and APOE alleles. These major risk factors of Alzheimer's disease (AD) were examined in mice given chronic exposure to nPM, a nano-sized subfraction of urban air pollution. In the cerebral cortex, female mice had two-fold more genes responding to nPM than males. Transcriptomic responses to nPM had sex- APOE interactions in AD-relevant pathways. Only APOE 3 mice responded to nPM in genes related to Abeta deposition and clearance ( Vav2 , Vav3 , S1009a ). Other responding genes included axonal guidance, inflammation (AMPK, NFKB, APK/JNK signaling), and antioxidant signaling (NRF2, HIF1A). Genes downstream of NFKB and NRF2 responded in opposite directions to nPM. Nrf2 knockdown in microglia augmented NFKB responses to nPM, suggesting a critical role of NRF2 in air pollution neurotoxicity. These findings give a rationale for epidemiologic studies of air pollution to consider sex interactions with APOE alleles and other AD-risk genes., Competing Interests: AH, MC, KD, CD, MT, NS, ML, CS, TT, HF, HZ, TM, CF No competing interests declared, (© 2020, Haghani et al.)

Background: Ambient particulate matter (PM) smaller than 2.5 µm in diameter (PM 2.5 ) undergoes diurnal changes in chemical composition due to photochemical oxidation. In this study we examine the relationships between oxidative activity and inflammatory responses associated with these diurnal chemical changes. Because secondary PM contains a higher fraction of oxidized PM species, we hypothesized that PM 2.5 collected during afternoon hours would induce a greater inflammatory response than primary, morning PM 2.5 . Methods: Time-integrated aqueous slurry samples of ambient PM 2.5 were collected using a direct aerosol-into-liquid collection system during defined morning and afternoon time periods. PM 2.5 samples were collected for 5 weeks in the late summer (August-September) of 2016 at a central Los Angeles site. Morning samples, largely consisting of fresh primary traffic emissions (primary PM), were collected from 6-9am (am-PM 2.5 ), and afternoon samples were collected from 12-4pm (pm-PM 2.5 ), when PM composition is dominated by products of photochemical oxidation (secondary PM). The two diurnally phased PM 2.5 slurries (am- and pm-PM 2.5 ) were characterized for chemical composition and BV-2 microglia were assayed in vitro for oxidative and inflammatory gene responses. Results: Contrary to expectations, the am-PM 2.5 slurry had more proinflammatory activity than the pm-PM 2.5 slurry as revealed by nitric oxide (NO) induction, as well as the upregulation of proinflammatory cytokines IL-1β, IL-6, and CCL2 (MCP-1), as assessed by messenger RNA production. Conclusions: The diurnal differences observed in this study may be in part attributed to the greater content of transition metals and water-insoluble organic carbon (WIOC) of am-PM 2.5 (primary PM) vs. pm-PM 2.5 (secondary PM), as these two classes of compounds can increase PM 2.5 toxicity., Competing Interests: No competing interests were disclosed.