Vitamin D3 increases the Caspase-3 p12, MTHFR, and P-glycoprotein reducing amyloid-ß42 in the kidney of a mouse model for Down syndrome.

AIMS: Renal dysfunction has been reported in individuals with Down syndrome (DS); however, the causes and mechanisms involved remain unknown. Here, we present a proposal for how the triplication of the amyloid beta precursor protein (APP) and, mainly the amyloid ß peptide 1-42 (Aß42) can favor the development of renal abnormalities in DS. We evaluated the effects of vitamin D3 (VD3) supplementation on morphofunctional aspects and the repercussions on the presence and localization of Aß42, methylenetetrahydrofolate reductase (MTHFR), caspase-3 p12, and P-glycoprotein (Pgp) in the renal tissue of DS mouse model. MAIN METHODS: Twenty female mice (14-week-old) belonging to the B6EiC3Sn-Rb(12.Ts171665Dn)2Cje/CjeDnJ lineage were divided into four experimental groups (n = 5/group): common diet; trisomy (Ts) and wild-type (Wt); and high doses VD3, Ts(VD3), and Wt(VD3). All the groups were treated for 10 weeks. At 24 weeks, the protocol experimental was interrupted. The kidney was weighed, collected, and processed for immunochemical analysis for Aß42, Caspase-3 p12, MTHFR, and Pgp proteins. All data were analyzed statistically. KEY FINDINGS: Our results showed that VD3 promoted an increase in caspase-3 p12, MTHFR, and Pgp, and consequently contributed to reduced Aß42 in the renal tissue of a mouse model of DS. Furthermore, VD3 treatment affected the plasma creatinine and urea levels and contributed to the attenuation of the dilation of Bowman's space observed in trisomic mice. SIGNIFICANCE: Finally, the results showed that VD3 may activate specific mechanisms involved in reduced Aß42 and tissue repair in the kidneys of a mouse model for Down syndrome.

Combined treatment with the phenolics (-)-epigallocatechin-3-gallate and ferulic acid improves cognition and reduces Alzheimer-like pathology in mice.

"Nutraceuticals" are well-tolerated natural dietary compounds with drug-like properties that make them attractive as Alzheimer's disease (AD) therapeutics. Combination therapy for AD has garnered attention following a recent National Institute on Aging mandate, but this approach has not yet been fully validated. In this report, we combined the two most promising nutraceuticals with complementary anti-amyloidogenic properties: the plant-derived phenolics (-)-epigallocatechin-3-gallate (EGCG, an α-secretase activator) and ferulic acid (FA, a ß-secretase modulator). We used transgenic mice expressing mutant human amyloid ß-protein precursor and presenilin 1 (APP/PS1) to model cerebral amyloidosis. At 12 months of age, we orally administered EGCG and/or FA (30 mg/kg each) or vehicle once daily for 3 months. At 15 months, combined EGCG-FA treatment reversed cognitive impairment in most tests of learning and memory, including novel object recognition and maze tasks. Moreover, EGCG- and FA-treated APP/PS1 mice exhibited amelioration of brain parenchymal and cerebral vascular ß-amyloid deposits and decreased abundance of amyloid ß-proteins compared with either EGCG or FA single treatment. Combined treatment elevated nonamyloidogenic soluble APP-α and α-secretase candidate and down-regulated amyloidogenic soluble APP-ß, ß-C-terminal APP fragment, and ß-secretase protein expression, providing evidence for a shift toward nonamyloidogenic APP processing. Additional beneficial co-treatment effects included amelioration of neuroinflammation, oxidative stress, and synaptotoxicity. Our findings offer preclinical evidence that combined treatment with EGCG and FA is a promising AD therapeutic approach.

Chiral penicillamine-modified selenium nanoparticles enantioselectively inhibit metal-induced amyloid ß aggregation for treating Alzheimer's disease.

Nanometer-scale chirality has gained significant interest from different research fields due to its fundamental importance in nature and living matter. In this study, we design and synthesize chiral penicillamine-capped selenium nanoparticles (l-/d-Pen@Se NPs) that can act as a novel class of chiral amyloid-ß (Aß) inhibitors. In this work, d-Pen@Se NPs demonstrate higher inhibition efficiency, as well as ameliorate cognition and memory impairments. We used rat pheochromocytoma (PC12) cells to perform real-time cell analysis assay (RTCA) to probe the potential cytotoxicity of l-/d-Pen@Se NPs. At any given time point, the cell index decreases as d-Pen@Se NPs concentration increases, demonstrating a concentration-dependent cytotoxic effect on PC12 cells. In addition, d-Pen@Se NPs also reduced Zn2+-induced intracellular Aß40 fibrillation, while l-Pen@Se NPs did not. The histological analysis demonstrates that mice treated with d-Pen@Se NPs did not exhibit signs of in vivo systemic toxicity in major organs. Our findings are highly encouraging in terms of providing substantial evidence of the safety of chiral d-Pen@Se NPs for biomedical application. We expect that these results will be relevant for other chiral NPs for treatment of Alzheimer's disease and have broad implications in NP-based studies and applications.

A modified formulation of Huanglian-Jie-Du-Tang reduces memory impairments and ß-amyloid plaques in a triple transgenic mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a degenerative disorder typified by progressive deterioration of memory and the appearance of ß-amyloid peptide (Aß)-rich senile plaques. Recently we have identified a novel function of a patented formulation of modified Huanglian-Jie-Tu-Tang (HLJDT-M), a Chinese herbal medicine, in treating AD in in vitro studies (US patent No. 9,375,457). HLJDT-M is a formulation composed of Rhizoma Coptitis, Cortex Phellodendri and Fructus Gardeniae without Radix Scutellariae. Here, we assessed the efficacy of HLJDT-M on a triple transgenic mouse model of AD (3XTg-AD). Oral administration of HLJDT-M ameliorated the cognitive dysfunction of 3XTg-AD mice and lessened the plaque burden. In addition, biochemical assays revealed a significant decrease in levels of detergent-soluble and acid-soluble Aß via decreasing the levels of full length amyloid-ß precursor protein (FL-APP) and C-terminal fragments of APP (CTFs) in brain lysates of HLJDT-M-treated mice. HLJDT-M treatment also significantly reduced the levels of FL-APP and CTFs in N2a/SweAPP cells. In contrast, treatment using the classical formula HLJDT did not reduce the memory impairment of 3XTg-AD mice and, rather, increased the Aß/Fl-APP/CTFs in both animal and cell culture studies. Altogether, our study indicates that HLJDT-M is a promising herbal formulation to prevent and/or cure AD.

Transgenic mice overexpressing amyloid precursor protein exhibit early metabolic deficits and a pathologically low leptin state associated with hypothalamic dysfunction in arcuate neuropeptide Y neurons.

Weight loss is a prominent early feature of Alzheimer's disease (AD) that often precedes the cognitive decline and clinical diagnosis. While the exact pathogenesis of AD remains unclear, accumulation of amyloid-ß (Aß) derived from the amyloid precursor protein (APP) in the brain is thought to lead to the neuronal dysfunction and death underlying the dementia. In this study, we examined whether transgenic mice overexpressing the Swedish mutation of APP (Tg2576), recapitulating selected features of AD, have hypothalamic leptin signaling dysfunction leading to early body weight deficits. We found that 3-month-old Tg2576 mice, before amyloid plaque formation, exhibit decreased weight with markedly decreased adiposity, low plasma leptin levels, and increased energy expenditure without alterations in feeding behavior. The expression of the orexigenic neuropeptide Y (NPY) in the hypothalamus to the low leptin state was abnormal at basal and fasting conditions. In addition, arcuate NPY neurons exhibited abnormal electrophysiological responses to leptin in Tg2576 hypothalamic slices or wild-type slices treated with Aß. Finally, the metabolic deficits worsened as Tg2576 mice aged and amyloid burden increased in the brain. These results indicate that excess Aß can potentially disrupt hypothalamic arcuate NPY neurons leading to weight loss and a pathologically low leptin state early in the disease process that progressively worsens as the amyloid burden increases. Collectively, these findings suggest that weight loss is an intrinsic pathological feature of Aß accumulation and identify hypothalamic leptin signaling as a previously unrecognized pathogenic site of action for Aß.

Baicalein reduces ß-amyloid and promotes nonamyloidogenic amyloid precursor protein processing in an Alzheimer's disease transgenic mouse model.

Baicalein, a flavonoid isolated from the roots of Scutellaria baicalensis, is known to modulate γ-aminobutyric acid (GABA) type A receptors. Given prior reports demonstrating benefits of GABAA modulation for Alzheimer's disease (AD) treatment, we wished to determine whether this agent might be beneficial for AD. CHO cells engineered to overexpress wild-type amyloid precursor protein (APP), primary culture neuronal cells from AD mice (Tg2576) and AD mice were treated with baicalein. In the cell cultures, baicalein significantly reduced the production of ß-amyloid (Aß) by increasing APP α-processing. These effects were blocked by the GABAA antagonist bicuculline. Likewise, AD mice treated daily with i.p. baicalein for 8 weeks showed enhanced APP α-secretase processing, reduced Aß production, and reduced AD-like pathology together with improved cognitive performance. Our findings suggest that baicalein promotes nonamyloidogenic processing of APP, thereby reducing Aß production and improving cognitive performance, by activating GABAA receptors.

Novel behavioural characteristics of the APP(Swe)/PS1ΔE9 transgenic mouse model of Alzheimer's disease.

In order to better understand animal models of Alzheimer's disease, novel phenotyping strategies have been established for transgenic mouse models. In line with this, the current study characterised male APPxPS1 transgenic mice on mixed C57BL/6JxC3H/HeJ background for the first time for social recognition memory, sensorimotor gating, and spatial memory using the cheeseboard test as an alternative to the Morris water maze. Furthermore, locomotion, anxiety, and fear conditioning were evaluated in transgenic and wild type-like animals. APPxPS1 males displayed task-dependent hyperlocomotion and anxiety behaviours and exhibited social recognition memory impairments compared to wild type-like littermates. Spatial learning and memory, fear conditioning, and sensorimotor gating were unaffected in APPxPS1 transgenic mice. In conclusion, this study describes for the first time social recognition memory deficits in male APPxPS1 mice and suggests that spatial learning and memory deficits reported in earlier studies are dependent on the sex and genetic background of the APPxPS1 mouse line used. Furthermore, particular test conditions of anxiety and spatial memory paradigms appear to impact on the behavioural response of this transgenic mouse model for Alzheimer's disease.

A dual mechanism linking NGF/proNGF imbalance and early inflammation to Alzheimer's disease neurodegeneration in the AD11 anti-NGF mouse model.

The neurotrophin Nerve Growth Factor (NGF) is essential for the maintenance and differentiation of basal forebrain cholinergic neurons. Since basal forebrain cholinergic neurons represent one major neuronal population affected and progressively degenerating in Alzheimer's disease (AD), interest has grown for NGF as a potential therapeutic agent in neurodegenerative disorders linked to aging, particularly for AD. However, no evidence was available, to link, in a cause-effect manner, deficits in NGF signalling to the broader activation in the Alzheimer's cascade, besides cholinergic deficits. The phenotypic analysis of the AD11 anti-NGF transgenic mouse, obtained by the "neuroantibodies" phenotypic protein knock out strategy, allowed demonstrating a direct causal link between NGF deprivation and AD pathology. Since then, extensive mechanistic studies on the AD11 model provided a new twist to the concept that alterations in NGF transport and signalling play a crucial role in sporadic Alzheimer's neurodegeneration, leading to the hypothesis of "Neurotrophic imbalance" as an upstream driver for sporadic AD. The results obtained with the AD11 anti-NGF mice highlight the fact that the particular mode of NGF neutralization, with an NGF antibody expressed in the brain, selectively interfering with mature NGF versus unprocessed proNGF, plays a major role in the mechanism of neurodegeneration, and could lead to new insights into the mechanisms of human sporadic AD. Here, we will review (1) the renewed neurotrophic imbalance hypothesis for AD and (2) the mechanisms underlying the neurodegenerative phenotype of AD11 anti-NGF mice.

L-3-n-butylphthalide regulates amyloid precursor protein processing by PKC and MAPK pathways in SK-N-SH cells over-expressing wild type human APP695.

Amyloid precursor protein (APP) is cleaved by α-secretase, within the amyloid-ß (Aß) sequence, resulting in the release of a secreted fragment (αAPPs) and precluding Aß production. We investigated the effects of a promising anti-AD new drug, l-3-n-butylphthalide (L-NBP), on APP processing and Aß generation in neuroblastoma SK-N-SH cells overexpressing wild-type human APP695. L-NBP significantly increased αAPPs release, and reduced Aß generation. The steady-state full-length APP levels were unaffected by L-NBP. It suggested that L-NBP regulated APP processing towards to the non-amyloidogenic α-secretase pathway. Protein kinase C (PKC) and mitogen activated protein (MAP) kinase might be involved in L-NBP-induced αAPPs secretion. L-NBP significantly increased PKCα and ɛ activations, lowered PKCγ activation and increased the phosphorylation of p44/p42 MAPK. Furthermore, PKC and MAPK inhibitors partially reduced L-NBP-induced αAPPs secretion. The results suggested alternative pharmacological mechanisms of L-NBP regarding the treatment of Alzheimer's disease (AD).

[Effects of extracts of Ginseng and Ginkgo biloba on hippocampal acetylcholine and monoamines in PDAP-pV717I transgenic mice].

OBJECTIVE: To study the effects and its possible mechanism of Naoweikang (NWK), a composite of ginseng and ginkgo extracts, on hippocampal neurotransmitters in APP transgenic mice. METHODS: P-DAPPV717I transgenic mice were taken as the model of Alzheimer's disease (AD) and be treated with different doses of NWK (31 mg/kg and 62 mg/kg) respectively by gastrogavage once per day for 12 weeks. Contents of hippocampal acetylcholine (ACh), monoamine neurotransmitters and their metabolites were determined with high performance liquid chromatography. RESULTS: Compared with nontransgenic mice, the levels of ACh and 5-HIAA in hippocampus of transgenic mice lowered significantly (P < 0.01), while 5-HT increased significantly (P < 0.05), and the levels of norepinephrine and dopamine increased by 14.6% and 17.7%, respectively. After 12-week administration, the ACh level increased significantly in the two NWK treated groups (P<0.05), and the 5-HT level in the high dose NWK treated group decreased (P<0.05), as compared with those in the untreated transgenic mice. CONCLUSION: NWK shows a significant regulatory effect on the activities of hippocampal acetylcholine and monoamine system, especially the cholinergic and 5-HT systems, in APP transgenic mice, which might be one of its mechanisms in improving learning and memory of AD model, and therefore, NWK might exert certain curative effect on AD.

Inhibition of human IAPP fibril formation does not prevent beta-cell death: evidence for distinct actions of oligomers and fibrils of human IAPP.

Type 2 diabetes mellitus (T2DM) is characterized by an approximately 60% deficit in beta-cell mass, increased beta-cell apoptosis, and islet amyloid derived from islet amyloid polypeptide (IAPP). Human IAPP (hIAPP) forms oligomers, leading to either amyloid fibrils or toxic oligomers in an aqueous solution in vitro. Either application of hIAPP on or overexpression of hIAPP in cells induces apoptosis. It remains controversial whether the fibrils or smaller toxic oligomers induce beta-cell apoptosis. Rifampicin prevents hIAPP amyloid fibril formation and has been proposed as a potential target for prevention of T2DM. We examined the actions of rifampicin on hIAPP amyloid fibril and toxic oligomer formation as well as its ability to protect beta-cells from either application of hIAPP or endogenous overexpression of hIAPP (transgenic rats and adenovirus-transduced beta-cells). We report that rifampicin (Acocella G. Clin Pharmacokinet 3: 108-127, 1978) prevents hIAPP fibril formation, but not formation of toxic hIAPP oligomers (Bates G. Lancet 361: 1642-1644, 2003), and does not protect beta-cells from apoptosis induced by either overexpression or application of hIAPP. These data emphasize that toxic hIAPP oligomers, rather than hIAPP fibrils, initiate beta-cell apoptosis and that screening tools to identify inhibitors of amyloid fibril formation are likely to be less useful than those that identify inhibitors of toxic oligomer formation. Finally, rifampicin and related molecules do not appear to be useful as candidates for prevention of T2DM.

Cerebral amyloid induces aberrant axonal sprouting and ectopic terminal formation in amyloid precursor protein transgenic mice.

A characteristic feature of Alzheimer's disease (AD) is the formation of amyloid plaques in the brain. Although this hallmark pathology has been well described, the biological effects of plaques are poorly understood. To study the effect of amyloid plaques on axons and neuronal connectivity, we have examined the axonal projections from the entorhinal cortex in aged amyloid precursor protein (APP) transgenic mice that exhibit cerebral amyloid deposition in plaques and vessels (APP23 mice). Here we report that entorhinal axons form dystrophic boutons around amyloid plaques in the entorhinal termination zone of the hippocampus. More importantly, entorhinal boutons were found associated with amyloid in ectopic locations within the hippocampus, the thalamus, white matter tracts, as well as surrounding vascular amyloid. Many of these ectopic entorhinal boutons were immunopositive for the growth-associated protein GAP-43 and showed light and electron microscopic characteristics of axonal terminals. Our findings suggest that (1) cerebral amyloid deposition has neurotropic effects and is the main cause of aberrant sprouting in AD brain; (2) the magnitude and significance of sprouting in AD have been underestimated; and (3) cerebral amyloid leads to the disruption of neuronal connectivity which, in turn, may significantly contribute to AD dementia.