Biology subjects

Leinenga, G.

Publications and source records attributed to Leinenga, G..

5 recordsLinked to original sources

bioRxiv2023

Scanning ultrasound improves memory and enhances functional outcomes in Alzheimer mice in the absence of amyloid-beta reduction

A prevalent view in treating age-dependent disorders including Alzheimers disease (AD) is that the underlying amyloid plaque pathology must be targeted for cognitive improvements. In contrast, we report here that repeated scanning ultrasound (SUS) treatment at 1 MHz frequency can ameliorate memory deficits in the APP23 mouse model of AD without reducing amyloid-{beta} (A{beta}) burden. Different from previous studies that had shown A{beta} clearance as a consequence of blood-brain barrier (BBB) opening, here, the BBB was not opened as no microbubbles were used. Quantitative proteomics and functional magnetic resonance imaging revealed that ultrasound induced long-lasting functional changes that correlate with the improvement in memory. Intriguingly, the treatment was more effective at a higher frequency (1MHz) than at a frequency within the range currently explored in clinical trials in AD patients (286 kHz). Together, our data suggest frequency-dependent bio-effects of ultrasound and a dissociation of cognitive improvement and A{beta} clearance, with important implications for the design of trials for AD therapies. SummaryThe therapeutic effect of ultrasound on memory in AD mice leads to altered protein expression and improved functional connectivity in the absence of amyloid-{beta} removal. Of two frequencies explored, the higher ultrasound frequency (1 MHz) is more effective.

neuroscience↗

bioRxiv2022

A sporadic Alzheimer's blood-brain barrier model for developing ultrasound-mediated delivery of Aducanumab and anti-Tau antibodies.

RationaleThe blood-brain barrier (BBB) is a major impediment to therapeutic intracranial drug delivery for the treatment of neurodegenerative diseases, including Alzheimers disease (AD). Focused ultrasound applied together with microbubbles (FUS+MB) is a novel technique to transiently open the BBB and increase drug delivery. Evidence suggests that FUS+MB is safe, however the effects of FUS+MB on human BBB cells, especially in the context of AD, remain sparsely investigated. MethodsHere we generated BBB cells (induced brain endothelial cells (iBECs) and astrocytes (iAstrocytes)) from apolipoprotein E gene allele E4 (APOE4, high AD risk) and allele E3 (APOE3, lower AD risk) carrying patient-derived induced pluripotent stem cells (iPSCs). We then developed a human sporadic AD BBB cell platform to investigate the effects of FUS+MB on BBB cells and screen for the delivery of two potentially therapeutic AD antibodies. ResultsWe utilized this robust and reproducible human BBB model to demonstrate increased delivery of therapeutic AD antibodies across the BBB following FUS+MB treatment, including an analogue of Aducanumab (AduhelmTM; anti-amyloid-{beta}) and a novel anti-Tau antibody RNF5. Our results also demonstrate the safety of FUS+MB indicated by minimal changes in the cell transcriptome as well as little or no changes in cell viability and inflammatory responses within the first 24 h post FUS+MB. Finally, we report a more physiologically relevant hydrogel-based 2.5D BBB model as a key development for FUS+MB-mediated drug delivery screening, with potentially higher translational utility. ConclusionOur results demonstrate an important translatable patient BBB cell model for identifying FUS+MB-deliverable drugs and screening for cell- and patient-specific effects of FUS+MB, accelerating the use of FUS+MB as a therapeutic modality in AD. One Sentence SummaryFocused ultrasound increases the in vitro delivery of therapeutic antibodies Aducanumab and anti-Tau in a sporadic Alzheimers disease patient-derived blood-brain barrier cell model.

neuroscience↗

bioRxiv2021

Transcriptional signature in microglia isolated from an Alzheimers disease mouse model treated with scanning ultrasound

RationaleIntracranial scanning ultrasound combined with intravenously injected microbubbles (SUS+MB) has been shown to transiently open the blood-brain barrier and reduce amyloid-{beta} (A{beta}) pathology in the APP23 mouse model of Alzheimers disease (AD). This has been accomplished, at least in part, through the activation of microglial cells; however, their response to the SUS treatment is only incompletely understood. MethodsWild-type (WT) and APP23 mice were subjected to SUS+MB, using non-SUS+MB-treated mice as sham controls. After 48 hours, the APP23 mice were injected with methoxy-XO4 to label A{beta} aggregates, followed by microglial isolation into XO4+ and XO4- populations using flow cytometry. Both XO4+ and XO4- cells were subjected to RNA sequencing and their transcriptome was analyzed through a bioinformatics pipeline. ResultsThe transcriptomic analysis of the microglial cells revealed a clear segregation depending on genotype (AD model versus WT mice), as well as treatment (SUS+MB versus sham) and A{beta} internalization (XO4+ versus XO4- microglia). Differential gene expression analysis detected 278 genes that were significantly changed by SUS+MB in the XO4+ cells (248 up/30 down) and 242 in XO- cells (225 up/17 down). Not surprisingly given previous findings of increased phagocytosis of plaques following SUS+MB, the pathway analysis highlighted that the treatment induced an enrichment in genes related to the phagosome pathway in XO4+ microglia; however, when comparing SUS+MB to sham, the analysis revealed an enrichment in genes involved in the cell cycle in both the XO4+ and XO4- microglial population. ConclusionOur data provide a comprehensive analysis of microglia in an AD mouse model subjected to ultrasound treatment as a function of A{beta} internalization, one of the defining hallmarks of AD. Several differentially expressed genes are highlighted, pointing to an ultrasound-induced activation of cell cycle mechanisms in microglial cells isolated from APP23 mice treated with SUS+MB. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=86 SRC="FIGDIR/small/473590v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@9ae94org.highwire.dtl.DTLVardef@1a48d22org.highwire.dtl.DTLVardef@2dcb9dorg.highwire.dtl.DTLVardef@92dfb2_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2021

Claudin-5 binder enhances focused ultrasound-mediated opening in an in vitro blood-brain barrier model

RationaleThe blood-brain barrier (BBB) while functioning as a gatekeeper of the brain, impedes cerebral drug delivery. An emerging technology to overcome this limitation is focused ultrasound (FUS). When FUS interacts with intravenously injected microbubbles (FUS+MB), the BBB opens, transiently allowing the access of therapeutic agents into the brain. However, the ultrasound parameters need to be tightly tuned: when the acoustic pressure is too low there is no opening, and when it is too high, bleeds can occur. We therefore asked whether BBB permeability can be increased by combining FUS+MB with a second modality such that in a clinical setting lower acoustic pressures could be potentially used. MethodsGiven that FUS achieves BBB opening by the disruption of tight junction (TJ) proteins such as claudin-5 of brain endothelial cells, we generated a stable MDCK II cell line (eGFP-hCldn5-MDCK II) that expresses fluorescently tagged human claudin-5. Two claudin-5 binders, mC5C2 (a peptide) and cCPEm (a truncated form of an enterotoxin), that have been reported previously to weaken the barrier, were synthesized and assessed for their abilities to enhance the permeability of cellular monolayers. We then performed a comparative analysis of single and combination treatments. ResultsWe successfully generated a novel cell line that formed functional monolayers as validated by an increased transendothelial electrical resistance (TEER) reading and a low (< 0.2%) permeability to sodium fluorescein (376 Da). We found that the binders exerted a time- and concentration-dependent effect on BBB opening when incubated over an extended period, whereas FUS+MB caused a rapid barrier opening followed by recovery after 12 hours within the tested pressure range. Importantly, preincubation with cCPEm prior to FUS+MB treatment resulted in greater barrier opening compared to either FUS+MB or cCPEm alone as measured by reduced TEER values and an increased permeability to fluorescently labelled 40 kDa dextran (FD40). ConclusionThe data suggest that pre-incubation with clinically suitable binders to TJ proteins may be a general strategy to facilitate safer and more effective ultrasound-mediated BBB opening in cellular and animal systems and potentially also for the treatment of human diseases of the brain.

neuroscience↗

bioRxiv2021

A comparative study of the effects of Aducanumab and scanning ultrasound on amyloid plaques and behavior in the APP23 mouse model of Alzheimer disease

BackgroundAducanumab is an anti-amyloid-{beta} (A{beta}) antibody that achieved reduced amyloid pathology in Alzheimers disease (AD) trials, but it is controversial whether it also improved cognition. It has been claimed that this would require a sufficiently high cumulative dose of the antibody in the brain. Therapeutic ultrasound, in contrast, has only begun to be investigated in human AD clinical trials. We have previously shown that scanning ultrasound in combination with intravenously injected microbubbles (SUS), that temporarily and safely opens the blood-brain barrier (BBB), removes amyloid and restores cognition in APP23 mice. It has not been directly tested how the effects of SUS compare to immunotherapy or whether a combination therapy is more effective. MethodsIn a study comprising four treatment arms, we tested the efficacy of an Aducanumab analogue, Adu, in comparison to SUS, as well as a combination therapy in APP23 mice, using sham as a control (aged 13-22 months). The active place avoidance (APA) test was used to test spatial memory, and histology and ELISA were used to measure amyloid. Brain antibody levels were also determined. ResultsWe found that both Adu and SUS reduced the total plaque area in the hippocampus to a similar degree, with no additive effect in the combination treatment (SUS+Adu). Whereas there was only a trend towards a reduction for both Adu and SUS in the cortex, the combination trial yielded a statistically significant reduction compared to sham. Only the SUS and SUS+Adu groups included animals that had their plaque load reduced to below 1% from above 10%. There was a robust improvement in spatial memory for SUS+Adu only. In this group, when measured three days post-treatment, Adu levels were still 5-fold increased in the combination therapy compared to delivery of Adu on its own. Together, these findings suggest that SUS should be seriously considered as a treatment option for AD. Alternatively, a combination trial using Aducanumab together with ultrasound to increase brain levels of Aducanumab may be warranted, as the two approaches may engage different (albeit shared) clearance mechanisms.

neuroscience↗