Weiming Xia

Three Dimensional Human Neuro-Spheroid Model of Alzheimer's Disease Based on Differentiated Induced Pluripotent Stem Cells.

The testing of candidate drugs to slow progression of Alzheimer’s disease (AD) requires clinical trials that are lengthy and expensive. Efforts to model the biochemical milieu of the AD brain may be greatly facilitated by combining two cutting edge technologies to generate three-dimensional (3D) human neuro-spheroid from induced pluripotent stem cells (iPSC) derived from AD subjects. We created iPSC from blood cells of five AD patients and differentiated them into 3D human neuronal culture. We characterized neuronal markers of our 3D neurons by immunocytochemical staining to validate the differentiation status. To block the generation of pathologic amyloid β peptides (Aβ), the 3D-differentiated AD neurons were treated with inhibitors targeting β-secretase (BACE1) and γ-secretases. As predicted, both BACE1 and γ-secretase inhibitors dramatically decreased Aβ generation in iPSC-derived neural cells derived from all five AD patients, under standard two-dimensional (2D) differentiation conditions. However, BACE1 and γ-secretase inhibitors showed less potency in decreasing Aβ levels in neural cells differentiated under 3D culture conditions. Interestingly, in a single subject AD1, we found that BACE1 inhibitor treatment was not able to significantly reduce Aβ42 levels. To investigate underlying molecular mechanisms, we performed proteomic analysis of 3D AD human neuronal cultures including AD1. Proteomic analysis revealed specific reduction of several proteins that might contribute to a poor inhibition of BACE1 in subject AD1. To our knowledge, this is the first iPSC-differentiated 3D neuro-spheroid model derived from AD patients’ blood. Our results demonstrate that our 3D human neuro-spheroid model can be a physiologically relevant and valid model for testing efficacy of AD drug.

Potential of tocotrienols in the prevention and therapy of Alzheimer's disease

Currently there is no cure for Alzheimers disease (AD); clinical trials are underway to reduce amyloid generation and deposition, a neuropathological hallmark in brains of AD patients. While genetic factors and neuroinflammation contribute significantly to AD pathogenesis, whether increased cholesterol level is a causative factor or a result of AD is equivocal. Prenylation of proteins regulating neuronal functions requires mevalonate-derived farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP). The observation that the levels of FPP and GGPP, but not that of cholesterol, are elevated in AD patients is consistent with the finding that statins, competitive inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, reduce FPP and GGPP levels and amyloid β protein production in preclinical studies. Retrospective studies show inverse correlations between incidence of AD and the intake and serum levels of the HMG CoA reductase-suppressive tocotrienols; tocopherols show mixed results. Tocotrienols, but not tocopherols, block the processing and nuclear localization of sterol regulatory element binding protein-2, the transcriptional factor for HMG CoA reductase and FPP synthase, and enhance the degradation of HMG CoA reductase. Consequently, tocotrienols deplete the pool of FPP and GGPP and potentially blunt prenylation-dependent AD pathogenesis. The antiinflammatory activity of tocotrienols further contributes to their protection against AD. The mevalonate- and inflammation-suppressive activities of tocotrienols may represent those of an estimated 23,000 mevalonate-derived plant secondary metabolites called isoprenoids, many of which are neuroprotective. Tocotrienol-containing plant foods and tocotrienol derivatives and formulations with enhanced bioavailability may offer a novel approach in AD prevention and treatment.

Induced pluripotent stem cells (iPSCs) derived from frontotemporal dementia patient's peripheral blood mononuclear cells

Peripheral blood mononuclear cells (PBMC) were donated by a patient with clinically diagnosed frontotemporal dementia (FTD). Induced pluripotent stem cells (iPSCs) were developed using integration-free CytoTune-iPS Sendai Reprogramming factors which include Sendai virus particles of the four Yamanaka factors Oct, Sox2, Klf4, and c-Myc.

Natural Product and Natural Product-Derived Gamma Secretase Modulators from Actaea Racemosa Extracts

Alzheimer’s disease is characterized by pathogenic oligomerization, aggregation, and deposition of amyloid beta peptide (Aβ), resulting in severe neuronal toxicity and associated cognitive dysfunction. In particular, increases in the absolute or relative level of the major long form of Aβ, Aβ42, are associated with increased cellular toxicity and rapidity of disease progression. As a result of this observation, screening to identify potential drugs to reduce the level of Aβ42 have been undertaken by way of modulating the proteolytic activity of the gamma secretase complex without compromising its action on other essential substrates such as Notch. In this review we summarize results from a program that sought to develop such gamma secretase modulators based on novel natural products identified in the extract of Actaea racemosa, the well-known botanical black cohosh. Following isolation of compound 1 (SPI-014), an extensive medicinal chemistry effort was undertaken to define the SAR of 1 and related semisynthetic compounds. Major metabolic and physicochemical liabilities in 1 were overcome including replacement of both the sugar and acetate moieties with more stable alternatives that improved drug-like properties and resulted in development candidate 25 (SPI-1865). Unanticipated off-target adrenal toxicity, however, precluded advancement of this series of compounds into clinical development.

Common proteomic profiles of induced pluripotent stem cell-derived three-dimensional neurons and brain tissue from Alzheimer patients

We established a unique platform for proteomic analysis of cultured three-dimensional (3D) neurons and brain tissue from Alzheimers disease (AD) patients. We collected peripheral blood mononuclear cells (PBMC), converted PBMC to induced pluripotent stem cell (iPSC) lines, and differentiated the iPSC into human 3D neuro-spheroids. The postmortem brain tissue from the superior frontal cortex, inferior frontal cortex and cerebellum area of the AD patients was compared to the same regions from the control subjects. Proteomic analysis of 3D neuro-spheroids derived from AD subjects revealed the alteration of a number of proteins involved in axon growth, mitochondrial function, and antioxidant defense. Similar analysis of post-mortem AD brain tissue revealed significant alteration in proteins involved in oxidative stress, neuro-inflammation, along with proteins related to axonal injury. These results clearly indicate that the dysfunction of 3D neurons from AD patients in our in vitro environment is comparable to the post-mortem AD brain tissue in vivo. In conclusion, our study revealed a number of candidate proteins that have important implications in AD pathogenesis and supports the notion that the iPSC-derived 3D neuronal system functions as a model to examine novel aspects of AD pathology. SIGNIFICANCE In this study, we present a unique platform for proteomic analysis of induced pluripotent stem cell-derived three dimensional (3D) neurons and compare the results to those from three regions of post-mortem brain tissue from Alzheimers disease patients and normal control subjects. Our results show that the dysfunction of 3D neurons from AD patients in our in vitro environment is comparable to the post-mortem AD brain tissue in vivo. Our results revealed several candidate proteins that have important implications in AD pathogenesis.

Peripheral blood mononuclear cell-converted induced pluripotent stem cells (iPSCs) from an early onset Alzheimer's patient

Improvement in transduction efficiency makes it possible to convert blood cells into induced pluripotent stem cells (iPSC). In this study, we generated an iPSC line from peripheral blood mononuclear cells (PBMC) donated by a patient who exhibited memory deficit at age 59; outcome of positron emission tomography scan is consistent with a diagnosis of Alzheimers disease. Integration-free CytoTune-iPS Sendai Reprogramming factors which include Sendai virus particles of the four Yamanaka factors Oct4, Sox2, Klf4, and c-Myc were introduced to PBMC to convert them to iPSCs without retention of virus. Three germ layer differentiation was induced to demonstrate the pluripotency of these iPSCs.

Amyloid β synaptotoxicity is Wnt-PCP dependent and blocked by fasudil

Synapse loss is the structural correlate of the cognitive decline indicative of dementia. In the brains of Alzheimers disease sufferers, amyloid β (Aβ) peptides aggregate to form senile plaques but as soluble peptides are toxic to synapses. We previously demonstrated that Aβ induces Dickkopf‐1 (Dkk1), which in turn activates the Wnt–planar cell polarity (Wnt‐PCP) pathway to drive tau pathology and neuronal death.

An amylin analog used as a challenge test for Alzheimer's disease

Preclinical studies demonstrate the potential of amylin in the diagnosis of Alzheimers disease (AD). We aimed to lay the foundation for repurposing the amylin analog and a diabetes drug, pramlintide, for AD in humans.

Lewy Body Pathology and Chronic Traumatic Encephalopathy Associated With Contact Sports

Traumatic brain injury has been associated with increased risk of Parkinson disease and parkinsonism, and parkinsonism and Lewy body disease (LBD) can occur with chronic traumatic encephalopathy (CTE). To test whether contact sports and CTE are associated with LBD, we compared deceased contact sports athletes (n = 269) to cohorts from the community (n = 164) and the Boston University Alzheimer disease (AD) Center (n = 261). Participants with CTE and LBD were more likely to have β-amyloid deposition, dementia, and parkinsonism than CTE alone (p < 0.05). Traditional and hierarchical clustering showed a similar pattern of LBD distribution in CTE compared to LBD alone that was most frequently neocortical, limbic, or brainstem. In the community-based cohort, years of contact sports play were associated with neocortical LBD (OR = 1.30 per year, p = 0.012), and in a pooled analysis a threshold of >8 years of play best predicted neocortical LBD (ROC analysis, OR = 6.24, 95% CI = 1.5-25, p = 0.011), adjusting for age, sex, and APOE ɛ4 allele status. Clinically, dementia was significantly associated with neocortical LBD, CTE stage, and AD; parkinsonism was associated with LBD pathology but not CTE stage. Contact sports participation may increase risk of developing neocortical LBD, and increased LBD frequency may partially explain extrapyramidal motor symptoms sometimes observed in CTE.

A role for APP in Wnt signalling links synapse loss with β-amyloid production

In Alzheimer’s disease (AD), the canonical Wnt inhibitor Dickkopf-1 (Dkk1) is induced by β-amyloid (Aβ) and shifts the balance from canonical towards non-canonical Wnt signalling. Canonical (Wnt-β-catenin) signalling promotes synapse stability, while non-canonical (Wnt-PCP) signalling favours synapse retraction; thus Aβ-driven synapse loss is mediated by Dkk1. Here we show that the Amyloid Precursor Protein (APP) co-activates both arms of Wnt signalling through physical interactions with Wnt co-receptors LRP6 and Vangl2, to bi-directionally modulate synapse stability. Furthermore, activation of non-canonical Wnt signalling enhances Aβ production, while activation of canonical signalling suppresses Aβ production. Together, these findings identify a pathogenic-positive feedback loop in which Aβ induces Dkk1 expression, thereby activating non-canonical Wnt signalling to promote synapse loss and drive further Aβ production. The Swedish familial AD variant of APP (APPSwe) more readily co-activates non-canonical, at the expense of canonical Wnt activity, indicating that its pathogenicity likely involves direct effects on synapses, in addition to increased Aβ production. Finally, we report that pharmacological inhibition of the Aβ-Dkk1-Aβ positive feedback loop with the drug fasudil can restore the balance between Wnt pathways, prevent dendritic spine withdrawal in vitro, and reduce Aβ load in vivo in mice with advanced amyloid pathology. These results clarify a relationship between Aβ accumulation and synapse loss and provide direction for the development of potential disease-modifying treatments.