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Amyloid beta (Abeta) 1-42 oligomers accumulate in brains of patients with Mild Cognitive Impairment (MCI) and disrupt synaptic plasticity processes that underlie memory formation. Synaptic binding of Abeta oligomers to several putative receptor proteins is reported to inhibit long-term potentiation, affect membrane trafficking and induce reversible spine loss in neurons, leading to impaired cognitive performance and ultimately to anterograde amnesia in the early stages of Alzheimer's disease (AD). We have identified a receptor not previously associated with AD that mediates the binding of Abeta oligomers to neurons, and describe novel therapeutic antagonists of this receptor capable of blocking Abeta toxic effects on synapses in vitro and cognitive deficits in vivo. Knockdown of sigma-2/PGRMC1 (progesterone receptor membrane component 1) protein expression in vitro using siRNA results in a highly correlated reduction in binding of exogenous Abeta oligomers to neurons of more than 90%. Expression of sigma-2/PGRMC1 is upregulated in vitro by treatment with Abeta oligomers, and is dysregulated in Alzheimer's disease patients' brain compared to age-matched, normal individuals. Specific, high affinity small molecule receptor antagonists and antibodies raised against specific regions on this receptor can displace synthetic Abeta oligomer binding to synaptic puncta in vitro and displace endogenous human AD patient oligomers from brain tissue sections in a dose-dependent manner. These receptor antagonists prevent and reverse the effects of Abeta oligomers on membrane trafficking and synapse loss in vitro and cognitive deficits in AD mouse models. These findings suggest sigma-2/PGRMC1 receptors mediate saturable oligomer binding to synaptic puncta on neurons and that brain penetrant, small molecules can displace endogenous and synthetic oligomers and improve cognitive deficits in AD models. We propose that sigma-2/PGRMC1 is a key mediator of the pathological effects of Abeta oligomers in AD and is a tractable target for small molecule disease-modifying therapeutics.

Studies on agonist-mediated down regulation of alpha1B-adrenergic receptors (AAR) reveals that, in rabbit aortic smooth muscle cells, the norepinephrine-induced downregulation of AAR is accomplished without reducing the receptor half-life (t1/2). A short-lived protein possibly regulates the receptor degradation. The t1/2 in AAR is 172 plus or minus 20 h while norepinephrine treated cells have a t1/2 of 11.8 plus or minus 0.3 h.

Several mutations conferring protection against Alzheimer's disease (AD) have been described, none as profound as the A673T mutation, where carriers are four times less likely to get AD compared to noncarriers. This mutation results in reduced amyloid beta (Aβ) protein production in vitro and lower lifetime Aβ concentration in carriers. Better understanding of the protective mechanisms of the mutation may provide important insights into AD pathophysiology and identify productive therapeutic intervention strategies for disease modification. Aβ(1‐42) protein forms oligomers that bind saturably to a single receptor site on neuronal synapses, initiating the downstream toxicities observed in AD. Decreased formation, toxicity, or stability of soluble Aβ oligomers, or reduction of synaptic binding of these oligomers, may combine with overall lower Aβ concentration to underlie A673T's disease protecting mechanism. To investigate these possibilities, we compared the formation rate of soluble oligomers made from Icelandic A673T mutant and wild type (wt) Aβ(1‐42) synthetic protein, the amount and intensity of oligomer bound to mature primary rat hippocampal/cortical neuronal synapses, and the potency of bound oligomers to impact trafficking rate in neurons in vitro using a physiologically relevant oligomer preparation method. At equal protein concentrations, mutant protein forms approximately 50% or fewer oligomers of high molecular weight (>50 kDa) compared to wt protein. Mutant oligomers are twice as potent at altering the cellular vesicle trafficking rate as wt at equivalent concentrations, however, mutant oligomers have a >4‐fold lower binding affinity to synaptic receptors (Kd = 1,950 vs. 442 nM). The net effect of these differences is a lower overall toxicity at a given concentration. This study demonstrates for the first time that mutant A673T Aβ oligomers prepared with this method have fundamentally different assembly characteristics and biological impact from wt protein and indicates that its disease protecting mechanism may result primarily from the mutant protein's much lower binding affinity to synaptic receptors. This suggests that therapeutics that effectively reduce oligomer binding to synapses in the brain may be beneficial in AD. [ABSTRACT FROM AUTHOR]

Background: Synapse damage and loss are fundamental to the pathophysiology of Alzheimer's disease (AD) and lead to reduced cognitive function. The goal of this review is to address the challenges of forging new clinical development approaches for AD therapeutics that can demonstrate reduction of synapse damage or loss. The key points of this review include the following: Synapse loss is a downstream effect of amyloidosis, tauopathy, inflammation, and other mechanisms occurring in AD. Synapse loss correlates most strongly with cognitive decline in AD because synaptic function underlies cognitive performance. Compounds that halt or reduce synapse damage or loss have a strong rationale as treatments of AD. Biomarkers that measure synapse degeneration or loss in patients will facilitate clinical development of such drugs. The ability of methods to sensitively measure synapse density in the brain of a living patient through synaptic vesicle glycoprotein 2A (SV2A) positron emission tomography (PET) imaging, concentrations of synaptic proteins (e.g., neurogranin or synaptotagmin) in the cerebrospinal fluid (CSF), or functional imaging techniques such as quantitative electroencephalography (qEEG) provides a compelling case to use these types of measurements as biomarkers that quantify synapse damage or loss in clinical trials in AD. Conclusion: A number of emerging biomarkers are able to measure synapse injury and loss in the brain and may correlate with cognitive function in AD. These biomarkers hold promise both for use in diagnostics and in the measurement of therapeutic successes. [ABSTRACT FROM AUTHOR]

Fresh osteochondral allografts are an important treatment option for the repair of full-thickness articular cartilage defects. Viable chondrocytes within the transplanted tissue are considered important to maintaining matrix integrity. The purpose of this study is to determine whether an increase in pH decreases chondrocyte viability during cold storage and whether equilibration of Dulbecco's modified Eagle's medium (DMEM) in 5% CO2 normalizes pH and increases chondrocyte survival during storage at 4°C. Freshly isolated bovine articular chondrocytes cultured in alginate beads were stored for up to 5 days at 4°C or 37°C in DMEM exposed to ambient air or in DMEM equilibrated with 5% CO2. Chondrocyte viability was determined by flow cytometry. Physiologic pH was maintained when DMEM was equilibrated with 5% CO2, while pH increased in ambient air. After 5 days of storage at 4°C, chondrocyte necrosis was higher when stored in ambient air than if equilibrated with 5% CO2. No decrease in chondrocyte viability was observed with storage at 37°C. In addition, chondrocyte viability in bovine cartilage osteochondral cores was examined after storage for 14 days at 4°C in DMEM with and without HEPES, and with and without 5% CO2. Under these conditions, the superficial layer of chondrocytes was more viable when stored in DMEM with HEPES or DMEM equilibrated with 5% CO2 than when stored in DMEM in ambient air. This data shows that an increase in pH decreased bovine chondrocyte viability when refrigerated at 4°C in DMEM, and that optimization of CO2 normalized pH and improved chondrocyte viability during cold storage in DMEM. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:643-650, 2008 [ABSTRACT FROM AUTHOR]

Amyloid beta (Abeta) 1–42 oligomers accumulate in brains of patients with Mild Cognitive Impairment (MCI) and disrupt synaptic plasticity processes that underlie memory formation. Synaptic binding of Abeta oligomers to several putative receptor proteins is reported to inhibit long-term potentiation, affect membrane trafficking and induce reversible spine loss in neurons, leading to impaired cognitive performance and ultimately to anterograde amnesia in the early stages of Alzheimer's disease (AD). We have identified a receptor not previously associated with AD that mediates the binding of Abeta oligomers to neurons, and describe novel therapeutic antagonists of this receptor capable of blocking Abeta toxic effects on synapses in vitro and cognitive deficits in vivo. Knockdown of sigma-2/PGRMC1 (progesterone receptor membrane component 1) protein expression in vitro using siRNA results in a highly correlated reduction in binding of exogenous Abeta oligomers to neurons of more than 90%. Expression of sigma-2/PGRMC1 is upregulated in vitro by treatment with Abeta oligomers, and is dysregulated in Alzheimer's disease patients' brain compared to age-matched, normal individuals. Specific, high affinity small molecule receptor antagonists and antibodies raised against specific regions on this receptor can displace synthetic Abeta oligomer binding to synaptic puncta in vitro and displace endogenous human AD patient oligomers from brain tissue sections in a dose-dependent manner. These receptor antagonists prevent and reverse the effects of Abeta oligomers on membrane trafficking and synapse loss in vitro and cognitive deficits in AD mouse models. These findings suggest sigma-2/PGRMC1 receptors mediate saturable oligomer binding to synaptic puncta on neurons and that brain penetrant, small molecules can displace endogenous and synthetic oligomers and improve cognitive deficits in AD models. We propose that sigma-2/PGRMC1 is a key mediator of the pathological effects of Abeta oligomers in AD and is a tractable target for small molecule disease-modifying therapeutics.

In this study, a panel of 46 compounds containing five different scaffolds known to have high σ 2 receptor affinity were screened. 6,7-Dimethoxy-2-[4-(4-methoxyphenyl)butan-2-yl]-1,2,3,4-tetrahydroisoquinoline [(±)- 7 ] ( K i for σ 1 = 48.4 ± 7.7 nM, and K i for σ 2 = 0.59 ± 0.02 nM) and its desmethyl analogue, (±)- 8 ( K i for σ 1 = 108 ± 35 nM, and K i for σ 2 = 4.92 ± 0.59 nM), showed excellent binding affinity and subtype selectivity for σ 2 receptors. In vitro cell binding indicated that σ 2 receptor binding of [ 11 C]-(±)- 7 and [ 11 C]-(±)- 8 was dependent on TMEM97 protein expression. In PET studies, the peak brain uptake of [ 11 C]-(±)- 7 (8.28 ± 2.52%ID/cc) was higher than that of [ 11 C]-(±)- 8 (4.25 ± 0.97%ID/cc) with specific distribution in the cortex and hypothalamus. Brain uptake or tissue binding was selectively inhibited by ligands with different σ 2 receptor binding affinities. The results suggest [ 11 C]-(±)- 7 can be used as a PET radiotracer for imaging the function of σ 2 receptors in central nervous system disorders.

Objective: The objective of the present study was to investigate the potential of application of growth factor genes to induce chondrogenic differentiation of human-derived mesenchymal stem cells (MSCs). The growth factor genes evaluated in the present study were transforming growth factor 1 (TGF-beta1) and insulin-like growth factor 1 (IGF-1)., Methods: Human MSCs were transduced with the adenoviral vectors carrying either TGF-beta1 or IGF-1 (AdTGF-beta1 and AdIGF-1 respectively) or a combination of both growth factor genes at different multiplicities of infection (MOI) and were then made into pellets. Pellets were also made from nontransduced cells and maintained in culture medium supplemented with 10 ng/mL of TGF-beta1. At specified time points, histological analysis, cartilage matrix gene expression, and immunofluorescence were performed to determine the extent of chondrogenic differentiation., Results: MSCs transduced with the AdTGF-beta1 demonstrated robust chondrogenic differentiation, while those made from AdIGF-1 did not. AdTGF-beta1 pellets demonstrated aggrecan gene expression as early as day 3 of pellet culture, while type II collagen gene expression was detected by day 10 of culture. The AdIGF-1, alone or in combination with TGF-beta1 pellets, did not show any type II collagen gene expression at any time point. By immunofluoresecence, type X collagen was distributed throughout the matrix in TGF-beta1 protein pellets while the growth factor gene pellets displayed scant staining., Conclusion: The results suggest that sustained administration of TGF-beta1 may be more effective in suppressing terminal differentiation than intermittent dosing and thus effective for cartilage repair.