Direct Conversion of Fibroblast into Neurons for Alzheimer's Disease Research: A Systematic Review.

BACKGROUND: Alzheimer's disease (AD) is a prevalent neurodegenerative disorder without a cure. Innovative disease models, such as induced neurons (iNs), could enhance our understanding of AD mechanisms and accelerate treatment development. However, a review of AD human iN studies is necessary to consolidate knowledge. OBJECTIVE: The objective of this review is to examine the current body of literature on AD human iN cells and provide an overview of the findings to date. METHODS: We searched two databases for relevant studies published between 2010 and 2023, identifying nine studies meeting our criteria. RESULTS: Reviewed studies indicate the feasibility of generating iNs directly from AD patients' fibroblasts using chemical induction or viral vectors. These cells express mature neuronal markers, including MAP-2, NeuN, synapsin, and tau. However, most studies were limited in sample size and primarily focused on autosomal dominant familial AD (FAD) rather than the more common sporadic forms of AD. Several studies indicated that iNs derived from FAD fibroblasts exhibited abnormal amyloid-ß metabolism, a characteristic feature of AD in humans. Additionally, elevated levels of hyperphosphorylated tau, another hallmark of AD, were reported in some studies. CONCLUSION: Although only a limited number of small-scale studies are currently available, AD patient-derived iNs hold promise as a valuable model for investigating AD pathogenesis. Future research should aim to conduct larger studies, particularly focusing on sporadic AD cases, to enhance the clinical relevance of the findings for the broader AD patient population. Moreover, these cells can be utilized in screening potential novel treatments for AD.

Healthcare costs of dementia diseases before, during and after diagnosis: Longitudinal analysis of 17 years of Swedish register data.

INTRODUCTION: This study examines health-care costs attributed to dementia diseases in the 10 years prior to, during, and 6 years after diagnosis. METHODS: Using administrative register data for people diagnosed with dementia (2010-2016) in southern Sweden (n = 21,184), and a comparison group without dementia, health-care costs over 17 years were examined using longitudinal regression analysis. RESULTS: Average annual health-care costs per person were consistently higher before diagnosis in the dementia group (10 years before: Swedish krona (SEK) 2063, P < .005 and 1 year before: SEK8166, P < .005). At diagnosis, health-care costs were more than twice as high (SEK44,410, P < .005). Four to 6 years after diagnosis, there was no significant different in costs compared to comparators. DISCUSSION: Excess health-care cost arise as early as 10 years before a formal diagnosis of dementia, and while there is a spike in cost after diagnosis, health-care costs are no different 4 years after. These findings question currently accepted assumptions on costs of dementia.

REST suppression mediates neural conversion of adult human fibroblasts via microRNA-dependent and -independent pathways.

Direct conversion of human fibroblasts into mature and functional neurons, termed induced neurons (iNs), was achieved for the first time 6 years ago. This technology offers a promising shortcut for obtaining patient- and disease-specific neurons for disease modeling, drug screening, and other biomedical applications. However, fibroblasts from adult donors do not reprogram as easily as fetal donors, and no current reprogramming approach is sufficiently efficient to allow the use of this technology using patient-derived material for large-scale applications. Here, we investigate the difference in reprogramming requirements between fetal and adult human fibroblasts and identify REST as a major reprogramming barrier in adult fibroblasts. Via functional experiments where we overexpress and knockdown the REST-controlled neuron-specific microRNAs miR-9 and miR-124, we show that the effect of REST inhibition is only partially mediated via microRNA up-regulation. Transcriptional analysis confirmed that REST knockdown activates an overlapping subset of neuronal genes as microRNA overexpression and also a distinct set of neuronal genes that are not activated via microRNA overexpression. Based on this, we developed an optimized one-step method to efficiently reprogram dermal fibroblasts from elderly individuals using a single-vector system and demonstrate that it is possible to obtain iNs of high yield and purity from aged individuals with a range of familial and sporadic neurodegenerative disorders including Parkinson's, Huntington's, as well as Alzheimer's disease.

Potassium-induced structural changes of the endoplasmic reticulum in pyramidal neurons in murine organotypic hippocampal slices.

The endoplasmic reticulum (ER) structure is of central importance for the regulation of cellular anabolism, stress response, and signal transduction. Generally continuous, the ER can temporarily undergo dramatic structural rearrangements resulting in a fragmented appearance. In this study we assess the dynamic nature of ER fission in pyramidal neurons in organotypic hippocampal slice cultures stimulated by depolarizing concentration of potassium (50 mM). The slices were obtained from transgenic mice expressing fluorescent ER-targeted DsRed2 protein. We employed live tissue confocal microscopy imaging with fluorescence recovery after photobleaching (FRAP) to monitor the extent of structural rearrangements of the ER. In control slices, the ER structure was continuous. Potassium stimulation resulted in extensive fragmentation (fission), whereas return to basal potassium levels (2.5 mM) led to ER fusion and normalization of ER structure. This ER fission/fusion could be repeated several times in the same neuron, demonstrating the reversibility of the process. Blockade of the N-methyl-D-aspartate receptor (NMDAR) with the antagonist D-AP5 or removal of extracellular Ca(2+) prevented depolarization-induced ER fission. ER fission is sensitive to temperature, and decreasing temperature from 35°C to 30°C augments fission, implying that the altering of ER continuity may be a protective response against damage. We conclude that events that generate membrane depolarisation in brain tissue lead to the release of endogenous glutamate that may regulate neuronal ER continuity. The rapid and reversible NMDAR-mediated changes in ER structure reflect an adaptive, innate property of the ER for synaptic activation as well as response to tissue stress, injury, and disease.

Dendritic EGFP-STIM1 activation after type I metabotropic glutamate and muscarinic acetylcholine receptor stimulation in hippocampal neuron.

Several signaling pathways in neurons engage the endoplasmic reticulum (ER) calcium store by triggering calcium release. After release, ER calcium levels must be restored. In many non-neuronal cell types, this is mediated by store-operated calcium entry (SOCE), a cellular homeostatic mechanism that activates specialized store-operated calcium channels (SOC). Although much evidence supports the existence of SOCE in neurons, its importance has been difficult to determine because of the abundance of calcium channels expressed and the lack of SOC-specific pharmacological agents. We have explored the function of the SOCE-inducing protein STIM1 in neurons. In EGFP-STIM1-expressing hippocampal neurons, the sarco- and endoplasmic reticulum calcium ATPase (SERCA) inhibitor thapsigargin caused rapid aggregation (i.e., activation) of STIM1 in soma and dendrites. Upon STIM1 activation by thapsigargin, a dramatic reduction in STIM1 mobility was detected by fluorescence recovery after photobleaching (FRAP). By triggering release of ER calcium with 3,5-dihydroxyphenylglycine (DHPG) or carbachol (Cch), agonists of type I metabotropic glutamate receptors (mGluR) and muscarinic acetylcholine receptors (mAChR), respectively, STIM1 was activated, and calcium entry (likely to represent SOCE) occurred in dendrites. It is therefore possible that neuronal SOCE is activated by physiological stimuli, some of which may alter the postsynaptic calcium signaling properties.

Rapid fragmentation of the endoplasmic reticulum in cortical neurons of the mouse brain in situ following cardiac arrest.

Neuronal endoplasmic reticulum (ER), continuous from soma to dendritic spines, undergoes rapid fragmentation in response to N-methyl-D-aspartate (NMDA) receptor stimulation in hippocampal slices and neuronal primary cultures. Here, we show that ER fragments in the mouse brain following cardiac arrest (CA) induced brain ischemia. The ER structure was assessed in vivo in cortical pyramidal neurons in transgenic mice expressing ER-targeted GFP using two-photon laser scanning microscopy with fluorescence recovery after photobleaching (FRAP). Endoplasmic reticulum fragmentation occurred 1 to 2 minutes after CA and once induced, fragmentation was rapid (<15 seconds). We propose that acute ER fragmentation may be a protective response against severe ischemic stress.

Endoplasmic reticulum dynamics in hippocampal dendritic spines induced by agonists of type I metabotropic glutamate but not by muscarinic acetylcholine receptors.

Neurons in the hippocampus exhibit subpopulations of dendritic spines that contain endoplasmic reticulum (ER). ER in spines is important for synaptic activity and its associated Ca(2+) signaling. The dynamic distribution of ER to spines is regulated by diacylglycerol and partly mediated by protein kinase C, metalloproteinases and γ-secretase. In this study, we explored whether pharmacological activation of type I metabotropic glutamate receptors (mGluRs) and muscarinic acetylcholine receptors (mAChRs) known to activate phospholipase C would have any effect on spine ER content. We found that DHPG (100 µM) but not carbachol (10 µM) caused a reduction in the number of spines with ER. We further found that ER Ca(2+) depletion triggered by thapsigargin (200 nM) had no effect on ER localization in spines.

The sigma-1 receptor enhances brain plasticity and functional recovery after experimental stroke.

Stroke leads to brain damage with subsequent slow and incomplete recovery of lost brain functions. Enriched housing of stroke-injured rats provides multi-modal sensorimotor stimulation, which improves recovery, although the specific mechanisms involved have not been identified. In rats housed in an enriched environment for two weeks after permanent middle cerebral artery occlusion, we found increased sigma-1 receptor expression in peri-infarct areas. Treatment of rats subjected to permanent or transient middle cerebral artery occlusion with 1-(3,4-dimethoxyphenethyl)-4-(3-phenylpropyl)piperazine dihydrochloride, an agonist of the sigma-1 receptor, starting two days after injury, enhanced the recovery of lost sensorimotor function without decreasing infarct size. The sigma-1 receptor was found in the galactocerebroside enriched membrane microdomains of reactive astrocytes and in neurons. Sigma-1 receptor activation increased the levels of the synaptic protein neurabin and neurexin in membrane rafts in the peri-infarct area, while sigma-1 receptor silencing prevented sigma-1 receptor-mediated neurite outgrowth in primary cortical neuronal cultures. In astrocytic cultures, oxygen and glucose deprivation induced sigma-1 receptor expression and actin dependent membrane raft formation, the latter blocked by sigma-1 receptor small interfering RNA silencing and pharmacological inhibition. We conclude that sigma-1 receptor activation stimulates recovery after stroke by enhancing cellular transport of biomolecules required for brain repair, thereby stimulating brain plasticity. Pharmacological targeting of the sigma-1 receptor provides new opportunities for stroke treatment beyond the therapeutic window of neuroprotection.

Rho kinase inhibition protects CA1 cells in organotypic hippocampal slices during in vitro ischemia.

The actin cytoskeleton is a dynamic superstructure that regulates multiple cellular functions and that has been implicated in cell death regulation. We investigated whether modulating the neuronal actin cytoskeleton polymerization by Rho-GTPase kinase (ROCK) inhibition influences cell death in hippocampal neuronal cultures and in murine organotypic hippocampal slice cultures subjected to in vitro ischemia (IVI). During IVI, spines on vehicle treated hippocampal neurons collapsed and large dendritic actin aggregates were formed. Following ROCK inhibition by Y27632, the actin aggregates were markedly smaller while large filopodia extended from the dendritic trunk. Y27632 also provided strong neuroprotection of hippocampal pyramidal CA1 neurons, which was of similar magnitude as protection by NMDA receptor blockade. Likewise, treatment with the F-actin depolymerizing agent latrunculin during IVI diminished actin aggregation and mitigated cell death following IVI. We propose that ROCK inhibition protects neurons against ischemic damage by disrupting actin polymerization thereby mitigating NMDA receptor induced toxicity and releasing ATP bound to actin for cellular energy use. We conclude that ROCK inhibitors abrogate multiple detrimental processes and could therefore be useful in stroke therapy.

NMDA receptor stimulation induces reversible fission of the neuronal endoplasmic reticulum.

With few exceptions the endoplasmic reticulum (ER) is considered a continuous system of endomembranes within which proteins and ions can move. We have studied dynamic structural changes of the ER in hippocampal neurons in primary culture and organotypic slices. Fluorescence recovery after photobleaching (FRAP) was used to quantify and model ER structural dynamics. Ultrastructure was assessed by electron microscopy. In live cell imaging experiments we found that, under basal conditions, the ER of neuronal soma and dendrites was continuous. The smooth and uninterrupted appearance of the ER changed dramatically after glutamate stimulation. The ER fragmented into isolated vesicles in a rapid fission reaction that occurred prior to overt signs of neuronal damage. ER fission was found to be independent of ER calcium levels. Apart from glutamate, the calcium ionophore ionomycin was able to induce ER fission. The N-methyl, D-aspartate (NMDA) receptor antagonist MK-801 inhibited ER fission induced by glutamate as well as by ionomycin. Fission was not blocked by either ifenprodil or kinase inhibitors. Interestingly, sub-lethal NMDA receptor stimulation caused rapid ER fission followed by fusion. Hence, ER fission is not strictly associated with cellular damage or death. Our results thus demonstrate that neuronal ER structure is dynamically regulated with important consequences for protein mobility and ER luminal calcium tunneling.

Gamma-secretase and metalloproteinase activity regulate the distribution of endoplasmic reticulum to hippocampal neuron dendritic spines.

The neuronal endoplasmic reticulum (ER) contributes to many physiological and pathological processes in the brain. A subset of dendritic spines on hippocampal neurons contains ER that may contribute to synapse-specific intracellular signaling. Distribution of ER to spines is dynamic, but knowledge of the regulatory mechanisms is lacking. In live cell imaging experiments we now show that cultured hippocampal neurons rapidly lost ER from spines after phorbol ester treatment. ER loss was reduced by inhibiting gamma-secretase (DAPT at 2 microM) and metalloproteinase (TAPI-0 and GM6001 at 4 microM) activity. Inhibition of protein kinase C also diminished loss of ER by preventing exit of ER from spines. Furthermore, gamma-secretase and metalloproteinase inhibition, in the absence of phorbol ester, triggered a dramatic increase in spine ER content. Metalloproteinases and gamma-secretase cleave several transmembrane proteins. Many of these substrates are known to localize to adherens junctions, a structural specialization with which spine ER interacts. One interesting possibility is thus that ER content within spines may be regulated by proteolytic activity affecting adherens junctions. Our data demonstrate a hitherto unknown role for these two proteolytic activities in regulating dynamic aspects of cellular ultrastructure, which is potentially important for cellular calcium homeostasis and several intracellular signaling pathways.

Dynamic distribution of endoplasmic reticulum in hippocampal neuron dendritic spines.

The role of the endoplasmic reticulum (ER) localized in dendritic spines has become a subject of intense interest because of its potential functions in local protein synthesis and signal transduction. Although it is recognized from electron microscopic studies that not all spines contain ER, little is know of its dynamic regulation or turnover. Here, we report a surprising degree of turnover of ER within spines. Using confocal microscopy imaging we observed continuity of spine-ER with dendritic ER in hippocampal primary neurons. Over 24 h, less than 50% of spine ER was stable. Despite this high degree of turn over, we identified a significant subset of spines that maintained ER for at least 4 days. These results indicate that within a single neuron, the organelle composition of a spine is unexpectedly dynamic and may explain aspects of the spine-to-spine variation in calcium spike magnitude and localized protein synthesis and trafficking.

Identification of two distinct progenitor populations in the lateral ganglionic eminence: implications for striatal and olfactory bulb neurogenesis.

The lateral ganglionic eminence (LGE) is known to give rise to striatal projection neurons as well as interneurons, which migrate in the rostral migratory stream (RMS) to populate the granule cell and glomerular layers of the olfactory bulb. Because all of these neuronal subtypes express Distalless-related (DLX) homeobox proteins during their differentiation, we set out to further characterize progenitors in the Dlx-positive domain of the LGE. Previous studies have shown that the LIM homeobox protein Islet1 (ISL1) marks the LGE subventricular zone (SVZ) and differentiating striatal projection neurons. However, ISL1 is not expressed in neurons of the developing olfactory bulb or the RMS. We show here that the dorsal-most portion of the Dlx-expressing region of the LGE SVZ lacks ISL1 cells. This dorsal domain, however, contains cells that express the ETS transcription factor Er81, which is also expressed in granule and periglomerular cells of the developing and adult olfactory bulb. Moreover, the adult SVZ and RMS contain numerous Er81-positive cells. Fate-mapping studies using Dlx5/6-cre transgenic mice demonstrate that Er81-positive cells in the granule cell and glomerular layers of the olfactory bulb derive from the Dlx-expressing SVZ region. These findings suggest that the LGE SVZ contains two distinct progenitor populations: a DLX(+);ISL1(+) population representing striatal progenitors and a DLX(+);Er81(+) population comprising olfactory bulb interneuron progenitors. In support of this, mice mutant for the homeobox genes Gsh2 and Gsh1/2, which show olfactory bulb defects, exhibit dramatically reduced numbers of Er81-positive cells in the LGE SVZ as well as in the olfactory bulb mantle.