Alzheimer's Disease Pathophysiology in the Retina.

The retina is an emerging CNS target for potential noninvasive diagnosis and tracking of Alzheimer's disease (AD). Studies have identified the pathological hallmarks of AD, including amyloid ß-protein (Aß) deposits and abnormal tau protein isoforms, in the retinas of AD patients and animal models. Moreover, structural and functional vascular abnormalities such as reduced blood flow, vascular Aß deposition, and blood-retinal barrier damage, along with inflammation and neurodegeneration, have been described in retinas of patients with mild cognitive impairment and AD dementia. Histological, biochemical, and clinical studies have demonstrated that the nature and severity of AD pathologies in the retina and brain correspond. Proteomics analysis revealed a similar pattern of dysregulated proteins and biological pathways in the retina and brain of AD patients, with enhanced inflammatory and neurodegenerative processes, impaired oxidative-phosphorylation, and mitochondrial dysfunction. Notably, investigational imaging technologies can now detect AD-specific amyloid deposits, as well as vasculopathy and neurodegeneration in the retina of living AD patients, suggesting alterations at different disease stages and links to brain pathology. Current and exploratory ophthalmic imaging modalities, such as optical coherence tomography (OCT), OCT-angiography, confocal scanning laser ophthalmoscopy, and hyperspectral imaging, may offer promise in the clinical assessment of AD. However, further research is needed to deepen our understanding of AD's impact on the retina and its progression. To advance this field, future studies require replication in larger and diverse cohorts with confirmed AD biomarkers and standardized retinal imaging techniques. This will validate potential retinal biomarkers for AD, aiding in early screening and monitoring.

Methoxyacetone revisited.

Conformational space of methoxyacetone (MA) was studied at the MP2/6-311++G(d,p) and DFT(B3LYP)/6-311++G(d,p) levels of theory. Computations predict MA to adopt four conformations, resulting from internal rotations around the O=C-C-O (Trans, Cis) and C-C-O-C (trans, gauche) dihedral angles. The Tt (Trans-trans) conformer is the most stable. The computed energies of two gauche (Tg and Cg) conformers fall in the 3-8 kJ mol-1 range above Tt and should account for 1/3 of the room-temperature gas-phase equilibrium. The energy of Ct form is 11 kJ mol-1 above Tt, and its expected population is negligible (below 1 %). In our earlier work, MA monomers were isolated in cryogenic argon matrices and characterized by infrared spectroscopy. In the experiment, only the most stable Tt conformer was detected in the sample. Signatures of the other conformers were not detected, either in freshly deposited samples, or in samples subjected to different UV irradiations. We rationalize those observations in terms of computed barriers for intramolecular torsions, indicating occurrence of conformational cooling during deposition. The experimental infrared spectrum of the Tt form is now assigned with the aid of anharmonic DFT computations. Exposure of MA to UV irradiation in the 300-260 nm range led to photolysis, according to the Norrish type II mechanism, resulting in dimer between enol acetone and formaldehyde observed as a cage-confined intermediate photoproduct. The subsequent photolysis resulted in the formation of carbon monoxide as the dominating photoproduct, formed in the Norrish type I photoreaction. Mechanistic interpretation of this photo decarbonylation reaction is presented.

An efficient one-pot synthesis and docking studies of bioactive new antiproliferative dispiro[oxindole/acenaphthylenone‒benzofuranone] pyrrolidine scaffolds.

A new and efficient method has been developed to synthesize dispiro[oxindole/acenaphthylenone-benzofuranone]pyrrolidine compounds. This is done by triggering the 1,3-dipolar cycloaddition reaction of azomethine ylides by reacting isatin/acenaphthoquinone with L-picolinic acid/L-proline/sarcosine/L-thioproline/tetrahydroisoquinolines, in a highly regioselective manner in an ionic liquid [DBU][Ac] with 4'-chloro-auron[2-(4-chlorobenzylidene)benzofuran-3(2H)-one]. Single-crystal X-ray diffraction data support the proposed structures of the new compounds. The heterocycles derived from amino acids such as L-picolinic acid, L-proline, and L-thioproline showed significant inhibitory effects against six human solid tumors, including lung, breast, cervix, colon, and others. These new structures were also tested in the active sites of the MDM2 receptor to further study their antiproliferative effects.

Equity, diversity, and inclusion at the Global Alliance for Genomics and Health.

A lack of diversity in genomics for health continues to hinder equitable leadership and access to precision medicine approaches for underrepresented populations. To avoid perpetuating biases within the genomics workforce and genomic data collection practices, equity, diversity, and inclusion (EDI) must be addressed. This paper documents the journey taken by the Global Alliance for Genomics and Health (a genomics-based standard-setting and policy-framing organization) to create a more equitable, diverse, and inclusive environment for its standards and members. Initial steps include the creation of two groups: the Equity, Diversity, and Inclusion Advisory Group and the Regulatory and Ethics Diversity Group. Following a framework that we call "Reflected in our Teams, Reflected in our Standards," both groups address EDI at different stages in their policy development process.

Thermal Adaptation Alters Response to Thermal Stress and Expression of Virulent Genes (eae, stx1, stx2, and hlyA) in Pathogenic Escherichia coli Isolated from Pork.

Escherichia coli encounter variety of environmental and processing stresses during their growth, survival, and infection. Herein, the thermotolerance behavior and transcription of virulent genes responsible for the pathogenicity in isolated strains of pathogenic E. coli were evaluated. Among 176 E. coli isolates, 4 isolates (2.27%) were confirmed to be pathogenic E. coli, out of which 2 isolates were positive for EHEC and 2 were positive for EPEC based on their virulence factors. Thermotolerance was induced under thermal adaptation at higher temperature, regardless of the pathotypes. Cells grown and adapted at 42 °C, exhibited highest transcription of genes associated with adhesion (eae), hemolysis (hlyA), and shiga toxin production (stx1). However, expression of these genes was downregulated in cells adapted at lower temperature of 4 °C and 25 °C compared to control. Further, transcription of stx2 was upregulated by 70% and 17% at 4 °C and 25 °C, respectively, while the transcription level was reduced by 44% relative to control at 42 °C. The findings indicate that expression of virulent genes in pathogenic E. coli at elevated temperature do not be depend on thermotolerance of the strain harboring these genes.

Retinal arterial Aß40 deposition is linked with tight junction loss and cerebral amyloid angiopathy in MCI and AD patients.

INTRODUCTION: Vascular amyloid beta (Aß) protein deposits were detected in retinas of mild cognitively impaired (MCI) and Alzheimer's disease (AD) patients. We tested the hypothesis that the retinal vascular tight junctions (TJs) were compromised and linked to disease status. METHODS: TJ components and Aß expression in capillaries and larger blood vessels were determined in post mortem retinas from 34 MCI or AD patients and 27 cognitively normal controls and correlated with neuropathology. RESULTS: Severe decreases in retinal vascular zonula occludens-1 (ZO-1) and claudin-5 correlating with abundant arteriolar Aß40 deposition were identified in MCI and AD patients. Retinal claudin-5 deficiency was closely associated with cerebral amyloid angiopathy, whereas ZO-1 defects correlated with cerebral pathology and cognitive deficits. DISCUSSION: We uncovered deficiencies in blood-retinal barrier markers for potential retinal imaging targets of AD screening and monitoring. Intense retinal arteriolar Aß40 deposition suggests a common pathogenic mechanism of failed Aß clearance via intramural periarterial drainage.

Neuroserpin gene therapy inhibits retinal ganglion cell apoptosis and promotes functional preservation in glaucoma.

Our research has proven that the inhibitory activity of the serine protease inhibitor neuroserpin (NS) is impaired because of its oxidation deactivation in glaucoma. Using genetic NS knockout (NS-/-) and NS overexpression (NS+/+ Tg) animal models and antibody-based neutralization approaches, we demonstrate that NS loss is detrimental to retinal structure and function. NS ablation was associated with perturbations in autophagy and microglial and synaptic markers, leading to significantly enhanced IBA1, PSD95, beclin-1, and LC3-II/LC3-I ratio and reduced phosphorylated neurofilament heavy chain (pNFH) levels. On the other hand, NS upregulation promoted retinal ganglion cell (RGC) survival in wild-type and NS-/- glaucomatous mice and increased pNFH expression. NS+/+Tg mice demonstrated decreased PSD95, beclin-1, LC3-II/LC3-I ratio, and IBA1 following glaucoma induction, highlighting its protective role. We generated a novel reactive site NS variant (M363R-NS) resistant to oxidative deactivation. Intravitreal administration of M363R-NS was observed to rescue the RGC degenerative phenotype in NS-/- mice. These findings demonstrate that NS dysfunction plays a key role in the glaucoma inner retinal degenerative phenotype and that modulating NS imparts significant protection to the retina. NS upregulation protected RGC function and restored biochemical networks associated with autophagy and microglial and synaptic function in glaucoma.

Retinal pathological features and proteome signatures of Alzheimer's disease.

Alzheimer's disease (AD) pathologies were discovered in the accessible neurosensory retina. However, their exact nature and topographical distribution, particularly in the early stages of functional impairment, and how they relate to disease progression in the brain remain largely unknown. To better understand the pathological features of AD in the retina, we conducted an extensive histopathological and biochemical investigation of postmortem retina and brain tissues from 86 human donors. Quantitative examination of superior and inferior temporal retinas from mild cognitive impairment (MCI) and AD patients compared to those with normal cognition (NC) revealed significant increases in amyloid ß-protein (Aß42) forms and novel intraneuronal Aß oligomers (AßOi), which were closely associated with exacerbated retinal macrogliosis, microgliosis, and tissue atrophy. These pathologies were unevenly distributed across retinal layers and geometrical areas, with the inner layers and peripheral subregions exhibiting most pronounced accumulations in the MCI and AD versus NC retinas. While microgliosis was increased in the retina of these patients, the proportion of microglial cells engaging in Aß uptake was reduced. Female AD patients exhibited higher levels of retinal microgliosis than males. Notably, retinal Aß42, S100 calcium-binding protein B+ macrogliosis, and atrophy correlated with severity of brain Aß pathology, tauopathy, and atrophy, and most retinal pathologies reflected Braak staging. All retinal biomarkers correlated with the cognitive scores, with retinal Aß42, far-peripheral AßOi and microgliosis displaying the strongest correlations. Proteomic analysis of AD retinas revealed activation of specific inflammatory and neurodegenerative processes and inhibition of oxidative phosphorylation/mitochondrial, and photoreceptor-related pathways. This study identifies and maps retinopathy in MCI and AD patients, demonstrating the quantitative relationship with brain pathology and cognition, and may lead to reliable retinal biomarkers for noninvasive retinal screening and monitoring of AD.

Computational refinement identifies functional destructive single nucleotide polymorphisms associated with human retinoid X receptor gene.

Alterations in the nuclear retinoid X receptor (RXRs) signalling have been implicated in neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease, stroke, multiple sclerosis and glaucoma. Single nucleotide polymorphisms (SNPs) are the main cause underlying single nucleic acid variations which in turn determine heterogeneity within various populations. These genetic polymorphisms have been suggested to associate with various degenerative disorders in population-wide analysis. This bioinformatics study was designed to investigate, search, retrieve and identify deleterious SNPs which may affect the structure and function of various RXR isoforms through a computational and molecular modelling approach. Amongst the 1,813 retrieved SNPs several were found to be deleterious with rs140464195_G139R, rs368400425_R358W and rs368586400_L383F RXRα mutant variants being the most detrimental ones causing changes in the interatomic interactions and decreasing the flexibility of the mutant proteins. Molecular genetics analysis identified seven missense mutations in RXRα/ß/γ isoforms. Two novel mutations SNP IDs (rs1588299621 and rs1057519958) were identified in RXRα isoform. We used several in silico prediction tools such as SIFT, PolyPhen, I-Mutant, Protein Variation Effect Analyzer (PROVEAN), PANTHER, SNP&Go, PhD-SNP and SNPeffect to predict pathogenicity and protein stability associated with RXR mutations. The structural assessment by DynaMut tool revealed that hydrogen bonds were affected along with hydrophobic and carbonyl interactions resulting in reduced flexibility at the mutated residue positions but ultimately stabilizing the molecule as a whole. Summarizing, analysis of the missense mutations in RXR isoforms showed a mix of conclusive and inconclusive genotype-phenotype correlations suggesting the use of sophisticated computational analysis tools for studying RXR variants.Communicated by Ramaswamy H. Sarma.

Regulating microglial miR-155 transcriptional phenotype alleviates Alzheimer's-induced retinal vasculopathy by limiting Clec7a/Galectin-3+ neurodegenerative microglia.

Single cell RNA sequencing studies identified novel neurodegeneration-associated microglial (MGnD/DAM) subtypes activated around cerebral amyloid plaques. Micro-RNA (miR)-155 of the TREM2-APOE pathway was shown to be a key transcriptional regulator of MGnD microglial phenotype. Despite growing interest in studying manifestations of Alzheimer's disease (AD) in the retina, a CNS organ accessible to noninvasive high-resolution imaging, to date MGnD microglia have not been studied in the AD retina. Here, we discovered the presence and increased populations of Clec7a+ and Galectin-3+ MGnD microglia in retinas of transgenic APPSWE/PS1L166P AD-model mice. Conditionally targeting MGnD microglia by miR-155 ablation via the tamoxifen-inducible CreERT2 system in APPSWE/PS1L166P mice diminished retinal Clec7a+ and Galectin-3+ microglial populations while increasing homeostatic P2ry12+ microglia. Retinal MGnD microglia were often adhering to microvessels; their depletion protected the inner blood-retina barrier and reduced vascular amyloidosis. Microglial miR-155 depletion further limits retinal inflammation. Mass spectrometry analysis revealed enhanced retinal PI3K-Akt signaling and predicted IL-8 and Spp1 decreases in mice with microglia-specific miR-155 knockout. Overall, this study identified MGnD microglia in APPSWE/PS1L166P mouse retina. Transcriptional regulation of these dysfunctional microglia mitigated retinal inflammation and vasculopathy. The protective effects of microglial miR-155 ablation should shed light on potential treatments for retinal inflammation and vascular damage during AD and other ocular diseases.

Transcriptomic and proteomic retinal pigment epithelium signatures of age-related macular degeneration.

There are currently no treatments for geographic atrophy, the advanced form of age-related macular degeneration. Hence, innovative studies are needed to model this condition and prevent or delay its progression. Induced pluripotent stem cells generated from patients with geographic atrophy and healthy individuals were differentiated to retinal pigment epithelium. Integrating transcriptional profiles of 127,659 retinal pigment epithelium cells generated from 43 individuals with geographic atrophy and 36 controls with genotype data, we identify 445 expression quantitative trait loci in cis that are asssociated with disease status and specific to retinal pigment epithelium subpopulations. Transcriptomics and proteomics approaches identify molecular pathways significantly upregulated in geographic atrophy, including in mitochondrial functions, metabolic pathways and extracellular cellular matrix reorganization. Five significant protein quantitative trait loci that regulate protein expression in the retinal pigment epithelium and in geographic atrophy are identified - two of which share variants with cis- expression quantitative trait loci, including proteins involved in mitochondrial biology and neurodegeneration. Investigation of mitochondrial metabolism confirms mitochondrial dysfunction as a core constitutive difference of the retinal pigment epithelium from patients with geographic atrophy. This study uncovers important differences in retinal pigment epithelium homeostasis associated with geographic atrophy.

Mechanical regulation of cell-cycle progression and division.

Cell-cycle progression and division are fundamental biological processes in animal cells, and their biochemical regulation has been extensively studied. An emerging body of work has revealed how mechanical interactions of cells with their microenvironment in tissues, including with the extracellular matrix (ECM) and neighboring cells, also plays a crucial role in regulating cell-cycle progression and division. We review recent work on how cells interpret physical cues and alter their mechanics to promote cell-cycle progression and initiate cell division, and then on how dividing cells generate forces on their surrounding microenvironment to successfully divide. Finally, the article ends by discussing how force generation during division potentially contributes to larger tissue-scale processes involved in development and homeostasis.

Vascular access complications in patients undergoing veno-arterial ecmo and their impact on survival in patients with refractory cardiogenic shock: A retrospective 8-year study.

Introduction: Veno-arterial extracorporeal membrane oxygenation (ECMO) is well-recognized treatment modality for patients with refractory cardiogenic shock. Uncomplicated cannulation is a prerequisite and basis for achieving a successful outcome in ECMO. Vascular access is obtained either by surgical cut-down. Common vascular access complications are bleeding and limb ischemia. Objective: To evaluate cannulation technique, the incidence of vascular complications, and their impact on the outcome. Methods: A retrospective data analysis conducted on 95 patients receiving ECMO from 2013 to 2020 was done. The patients were divided into two groups: no vascular access complications (non-VAC group) and vascular access complications (VAC group). The groups were compared related to the hospital and ICU stays and blood transfusion. Results: The patients in both groups were demographically and clinically comparable. The Non-VAC group had 75 patients, whereas the VAC group had a total of 20 patients. The main complication observed in the VAC group was bleeding from the cannulation site which required more blood transfusion than the non-VAC group (6.8 ± 1.02 vs 4.2 ± 1.26). Limb ischemia was another complication seen in the VAC group (4.2%, n = 4). Two patients had delayed bleeding after decannulation. The overall average length of stay in the hospital was statistically similar in both the groups (22 days in the VAC group vs 18 days in the non-VAC group), but the average ICU stay was more in the VAC group compared to the non-VAC group (18 days vs 12.06 days). Conclusion: Bleeding and limb ischemia are the important vascular access site complications, which increase blood transfusion requirements, ICU stay, and overall hospital stay.

Disulfiram enhances meropenem activity against NDM- and IMP-producing carbapenem-resistant Acinetobacter baumannii infections.

OBJECTIVES: To evaluate the in vitro and in vivo efficacy of the FDA-approved drug disulfiram in combination with meropenem against MBL-expressing carbapenem-resistant Acinetobacter baumannii. METHODS: Chequerboard and antibiotic resistance reversal analysis were performed using 25 clinical isolates producing different MBLs. Three representative strains harbouring NDM, IMP or non-MBL genes were subjected to a time-kill assay to further evaluate this synergistic interaction. Dose-dependent inhibition by disulfiram was assessed to determine IC50 for NDM-1, IMP-7, VIM-2 and KPC-2. Further, to test the efficacy of meropenem monotherapy and meropenem in combination with disulfiram against NDM- and IMP-harbouring A. baumannii, an experimental model of systemic infection and pneumonia was developed using BALB/c female mice. RESULTS: Chequerboard and antibiotic reversal assay displayed a synergistic interaction against MBL-expressing A. baumannii strains with 4- to 32-fold reduction in MICs of meropenem. In time-kill analysis, meropenem and disulfiram exhibited synergy against NDM- and IMP-producing carbapenem-resistant A. baumannii (CRAb) isolates. In vitro dose-dependent inhibition analysis showed that disulfiram inhibits NDM-1 and IMP-7 with IC50 values of 1.5 ± 0.6 and 16.25 ± 1.6 µM, respectively, with slight or no inhibition of VIM-2 (<20%) and KPC-2. The combination performed better in the clearance of bacterial load from the liver and spleen of mice infected with IMP-expressing CRAb. In the pneumonia model, the combination significantly decreased the bacterial burden of NDM producers compared with monotherapy. CONCLUSIONS: These results strongly suggest that the combination of disulfiram and meropenem represents an effective treatment option for NDM- and IMP-associated CRAb infections.

Evaluating associations of RNFL thickness and multifocal VEP with cognitive assessment and brain MRI volumes in older adults: Optic nerve decline and cognitive change (ONDCC) initiative.

To examine the relationships of retinal structural (optical coherence tomography) and visual functional (multifocal visual evoked potentials, mfVEP) indices with neuropsychological and brain structural measurements in healthy older subjects. 95 participants (mean (SD) age 68.1 (9.0)) years were recruited in the Optic Nerve Decline and Cognitive Change (ONDCC) study in this observational clinical investigation. OCT was conducted for retinal nerve fibre layer (RNFL) and mfVEP for amplitude and latency measurements. Participants undertook neuropsychological tests for cognitive performance and MRI for volumetric evaluation of various brain regions. Generalised estimating equation models were used for association analysis (p < 0.05). The brain volumetric measures including total grey matter (GM), cortex, thalamus, hippocampal and fourth ventricular volumes were significantly associated with global and sectoral RNFL. RNFL thickness correlated with delayed recalls of California verbal learning test (CVLT) and Rey complex figure test (RCFT). The mfVEP amplitudes associated with cerebral white matter (WM) and cingulate GM volumes in MRI and CVLT, RCFT and trail making test outcomes. A significant association of mfVEP latency with logical memory delayed recall and thalamus volume was also observed. Our results suggested significant association of specific RNFL and mfVEP measures with distinctive brain region volumes and cognitive tests reflecting performance in memory, visuospatial and executive functional domains. These findings indicate that the mfVEP and RNFL measurements may parallel brain structural and neuropsychological measures in the older population.

The nature of cell division forces in epithelial monolayers.

Epithelial cells undergo striking morphological changes during division to ensure proper segregation of genetic and cytoplasmic materials. These morphological changes occur despite dividing cells being mechanically restricted by neighboring cells, indicating the need for extracellular force generation. Beyond driving cell division itself, forces associated with division have been implicated in tissue-scale processes, including development, tissue growth, migration, and epidermal stratification. While forces generated by mitotic rounding are well understood, forces generated after rounding remain unknown. Here, we identify two distinct stages of division force generation that follow rounding: (1) Protrusive forces along the division axis that drive division elongation, and (2) outward forces that facilitate postdivision spreading. Cytokinetic ring contraction of the dividing cell, but not activity of neighboring cells, generates extracellular forces that propel division elongation and contribute to chromosome segregation. Forces from division elongation are observed in epithelia across many model organisms. Thus, division elongation forces represent a universal mechanism that powers cell division in confining epithelia.

Inner retinal injury in experimental glaucoma is prevented upon AAV mediated Shp2 silencing in a caveolin dependent manner.

SH2 domain containing tyrosine phosphatase 2 (Shp2; PTPN11) regulates several intracellular pathways downstream of multiple growth factor receptors. Our studies implicate that Shp2 interacts with Caveolin-1 (Cav-1) protein in retinal ganglion cells (RGCs) and negatively regulates BDNF/TrkB signaling. This study aimed to investigate the mechanisms underlying the protective effects of shp2 silencing in the RGCs in glaucomatous conditions. Methods: Shp2 was silenced in the Cav-1 deficient mice and the age matched wildtype littermates using adeno-associated viral (AAV) constructs. Shp2 expression modulation was performed in an acute and a chronic mouse model of experimental glaucoma. AAV2 expressing Shp2 eGFP-shRNA under a strong synthetic CAG promoter was administered intravitreally in the animals' eyes. The contralateral eye received AAV-eGFP-scramble-shRNA as control. Animals with Shp2 downregulation were subjected to either microbead injections or acute ocular hypertension experimental paradigm. Changes in inner retinal function were evaluated by measuring positive scotopic threshold response (pSTR) while structural and biochemical alterations were evaluated through H&E staining, western blotting and immunohistochemical analysis of the retinal tissues. Results: A greater loss of pSTR amplitudes was observed in the WT mice compared to Cav-1-/- retinas in both the models. Silencing of Shp2 phosphatase imparted protection against inner retinal function loss in chronic glaucoma model in WT mice. The functional rescue also translated to structural preservation of ganglion cell layer in the chronic glaucoma condition in WT mice which was not evident in Cav-1-/- mice retinas. Conclusions: This study indicates that protective effects of Shp2 ablation under chronic experimental glaucoma conditions are dependent on Cav-1 in the retina, suggesting in vivo interactions between the two proteins.

Mouse model of Alzheimer's disease demonstrates differential effects of early disease pathology on various brain regions.

Different parts of the brain are affected distinctively in various stages of the Alzheimer's disease (AD) pathogenesis. Identifying the biochemical changes in specific brain regions is key to comprehend the neuropathological mechanisms in early pre-symptomatic phases of AD. Quantitative proteomics profiling of four distinct areas of the brain of young APP/PS1 mouse model of AD was performed followed by biochemical pathway enrichment analysis. Findings revealed fundamental compositional and functional shifts even in the early stages of the disease. This novel study highlights unique proteome and biochemical pathway alterations in specific regions of the brain that underlie the early stages of AD pathology and will provide a framework for future longitudinal studies. The proteomics data were deposited into the ProteomeXchange Consortium via PRIDE with the identifier PXD019192.