CCR5 Decorated Rilpivirine Lipid Nanoparticles Build Myeloid Drug Depots Which Sustains Antiretroviral Activities.

Antiretroviral therapy (ART) improves the quality of life for those living with the human immunodeficiency virus type one (HIV-1). However, poor compliance reduces ART effectiveness and leads to immune compromise, viral mutations, and disease co-morbidities. A novel drug formulation is made whereby a lipid nanoparticle (LNP) carrying rilpivirine (RPV) is decorated with the C-C chemokine receptor type 5 (CCR5). This facilitates myeloid drug depot deposition. Particle delivery to viral reservoirs is tracked by positron emission tomography. The CCR5-mediated RPV LNP cell uptake and retention reduce HIV-1 replication in human monocyte-derived macrophages and infected humanized mice. Focused ultrasound allows the decorated LNP to penetrate the blood-brain barrier and reach brain myeloid cells. These findings offer a role for CCR5-targeted therapeutics in antiretroviral delivery to optimize HIV suppression.

Amyloid-ß specific regulatory T cells attenuate Alzheimer's disease pathobiology in APP/PS1 mice.

BACKGROUND: Regulatory T cells (Tregs) maintain immune tolerance. While Treg-mediated neuroprotective activities are now well-accepted, the lack of defined antigen specificity limits their therapeutic potential. This is notable for neurodegenerative diseases where cell access to injured brain regions is required for disease-specific therapeutic targeting and improved outcomes. To address this need, amyloid-beta (Aß) antigen specificity was conferred to Treg responses by engineering the T cell receptor (TCR) specific for Aß (TCRAß). The TCRAb were developed from disease-specific T cell effector (Teff) clones. The ability of Tregs expressing a transgenic TCRAß (TCRAß -Tregs) to reduce Aß burden, transform effector to regulatory cells, and reverse disease-associated neurotoxicity proved beneficial in an animal model of Alzheimer's disease. METHODS: TCRAß -Tregs were generated by CRISPR-Cas9 knockout of endogenous TCR and consequent incorporation of the transgenic TCRAb identified from Aß reactive Teff monoclones. Antigen specificity was confirmed by MHC-Aß-tetramer staining. Adoptive transfer of TCRAß-Tregs to mice expressing a chimeric mouse-human amyloid precursor protein and a mutant human presenilin-1 followed measured behavior, immune, and immunohistochemical outcomes. RESULTS: TCRAß-Tregs expressed an Aß-specific TCR. Adoptive transfer of TCRAß-Tregs led to sustained immune suppression, reduced microglial reaction, and amyloid loads. 18F-fluorodeoxyglucose radiolabeled TCRAß-Treg homed to the brain facilitating antigen specificity. Reduction in amyloid load was associated with improved cognitive functions. CONCLUSIONS: TCRAß-Tregs reduced amyloid burden, restored brain homeostasis, and improved learning and memory, supporting the increased therapeutic benefit of antigen specific Treg immunotherapy for AD.

Clinical biomarkers for Lewy body diseases.

Synucleinopathies are a group of neurodegenerative disorders characterized by pathologic aggregates of neural and glial α-synuclein (α-syn) in the form of Lewy bodies (LBs), Lewy neurites, and cytoplasmic inclusions in both neurons and glia. Two major classes of synucleinopathies are LB disease and multiple system atrophy. LB diseases include Parkinson's disease (PD), PD with dementia, and dementia with LBs. All are increasing in prevalence. Effective diagnostics, disease-modifying therapies, and therapeutic monitoring are urgently needed. Diagnostics capable of differentiating LB diseases are based on signs and symptoms which might overlap. To date, no specific diagnostic test exists despite disease-specific pathologies. Diagnostics are aided by brain imaging and cerebrospinal fluid evaluations, but more accessible biomarkers remain in need. Mechanisms of α-syn evolution to pathologic oligomers and insoluble fibrils can provide one of a spectrum of biomarkers to link complex neural pathways to effective therapies. With these in mind, we review promising biomarkers linked to effective disease-modifying interventions.

Immune senescence in aged APP/PS1 mice.

Objectives: To evaluate the linkage between age and deficits in innate and adaptive immunity which heralds both Alzheimer's disease (AD) onset and progression. The pathobiological events which underlie and tie these outcomes remain not fully understood. Methods: To investigate age-dependent immunity in AD, we evaluated innate and adaptive immunity in coordinate studies of regulatory T cell (Treg) function, T cell frequencies, and microglial integrity. These were assessed in blood, peripheral lymphoid tissues, and the hippocampus of transgenic (Tg) amyloid precursor protein/presenilin 1 (APP/PS1) against non-Tg mice. Additionally, immune arrays of hippocampal tissue were performed at 4, 6, 12, and 20 months of age. Results: APP/PS1 mice showed progressive impairment of Treg immunosuppressive function with age. There was partial restoration of Treg function in 20-month-old mice. Ingenuity pathway analyses of hippocampal tissues were enriched in inflammatory, oxidative, and cellular activation pathways that paralleled advancing age and AD-pathobiology. Operative genes in those pathways included, but were not limited to triggering receptor on myeloid cells 1 (TREM1), T helper type 1 (Th1), and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathways. Interleukin-17 (IL-17), nitric oxide, acute phase, and T cell receptor signaling pathways were also perturbed. Significant inflammation was observed at 6- and 12-months. However, at 20-months, age associated partial restoration of Treg function reduced inflammatory phenotype. Conclusions: Impaired Treg function, inflammation and oxidative stress were associated with AD pathology. Age associated partial restoration of Treg function in old mice reduced the hippocampal inflammatory phenotype. Restoring Treg suppressive function can be a therapeutic modality for AD.

A Novel Sphingosine Kinase Inhibitor Suppresses Chikungunya Virus Infection.

Chikungunya virus (CHIKV) is a re-emerging arbovirus in the alphavirus genus. Upon infection, it can cause severe joint pain that can last years in some patients, significantly affecting their quality of life. Currently, there are no vaccines or anti-viral therapies available against CHIKV. Its spread to the Americas from the eastern continents has substantially increased the count of the infected by millions. Thus, there is an urgent need to identify therapeutic targets for CHIKV treatment. A potential point of intervention is the sphingosine-1-phosphate (S1P) pathway. Conversion of sphingosine to S1P is catalyzed by Sphingosine kinases (SKs), which we previously showed to be crucial pro-viral host factor during CHIKV infection. In this study, we screened inhibitors of SKs and identified a novel potent inhibitor of CHIKV infection-SLL3071511. We showed that the pre-treatment of cells with SLL3071511 in vitro effectively inhibited CHIKV infection with an EC50 value of 2.91 µM under both prophylactic and therapeutic modes, significantly decreasing the viral gene expression and release of viral particles. Our studies suggest that targeting SKs is a viable approach for controlling CHIKV replication.

Bovine rhinitis B virus is highly prevalent in acute bovine respiratory disease and causes upper respiratory tract infection in calves.

Bovine respiratory disease (BRD) is the most significant cause of cattle morbidity and mortality worldwide. This multifactorial disease has a complex aetiology. Dogma posits a primary viral infection followed by secondary bacterial pneumonia. Bovine rhinitis B virus (BRBV) is an established aetiological agent of BRD, but little is known regarding its pathogenesis. Here, a BRD PCR panel identified 18/153 (11.8 %) lung samples and 20/49 (40.8 %) nasal swabs collected from cattle with respiratory signs as positive for BRBV, which was the most prevalent virus in nasal swabs. Primary bovine tracheal epithelial cells were used to isolate BRBV that was phylogenetically related to contemporary sequences from the USA and Mexico and genetically divergent from the previous sole BRBV isolate. To investigate virus pathogenesis, 1-week-old colostrum-deprived dairy calves were inoculated intranasally with 7.0 log10 TCID50 BRBV. Virus was isolated from nasal swabs, nasal turbinates, trachea and the brain of the challenged animals. Neutralizing antibodies were detected beginning 7 days post-inoculation and peaked at day 14. In situ hybridization (ISH) localized BRBV infection in the upper respiratory ciliated epithelial and goblet cells, occasionally associated with small defects of the superficial cilia lining. Sporadically, pinpoint ISH signals were also detected in cells resembling glial cells in the cerebrum in one calf. Together, these results demonstrate the BRBV infection is highly prevalent in acute BRD samples and while the pathogenicity of BRBV is minimal with infection largely limited to the upper respiratory tract, further research is needed to elucidate a possible initiatory role in BRD.

Isolation and development of bovine primary respiratory cells as model to study influenza D virus infection.

Influenza D virus (IDV) is a novel type of influenza virus that infects and causes respiratory illness in bovines. Lack of host-specific in vitro model that can recapitulate morphology and physiology of in vivo airway epithelial cells has impeded the study of IDV infection. Here, we established and characterized bovine primary respiratory epithelial cells from nasal turbinate, soft palate, and trachea of the same calf. All three cell types showed characteristics peculiar of epithelial cells, polarized into apical-basolateral membrane, and formed tight junctions. Furthermore, these cells expressed both α-2,3- and α-2,6-linked sialic acids with α-2,3 linkage being more abundant. IDV strains replicated to high titers in these cells, while influenza A and B viruses exhibited moderate to low titers, with influenza C virus replication not detected. These findings suggest that bovine primary airway epithelial cells can be utilized to model infection biology and pathophysiology of IDV and other respiratory pathogens.