Integrative metabolomics-genomics analysis identifies key networks in a stem cell-based model of schizophrenia.

Schizophrenia (SCZ) is a neuropsychiatric disorder, caused by a combination of genetic and environmental factors. The etiology behind the disorder remains elusive although it is hypothesized to be associated with the aberrant response to neurotransmitters, such as dopamine and glutamate. Therefore, investigating the link between dysregulated metabolites and distorted neurodevelopment holds promise to offer valuable insights into the underlying mechanism of this complex disorder. In this study, we aimed to explore a presumed correlation between the transcriptome and the metabolome in a SCZ model based on patient-derived induced pluripotent stem cells (iPSCs). For this, iPSCs were differentiated towards cortical neurons and samples were collected longitudinally at various developmental stages, reflecting neuroepithelial-like cells, radial glia, young and mature neurons. The samples were analyzed by both RNA-sequencing and targeted metabolomics and the two modalities were used to construct integrative networks in silico. This multi-omics analysis revealed significant perturbations in the polyamine and gamma-aminobutyric acid (GABA) biosynthetic pathways during rosette maturation in SCZ lines. We particularly observed the downregulation of the glutamate decarboxylase encoding genes GAD1 and GAD2, as well as their protein product GAD65/67 and their biochemical product GABA in SCZ samples. Inhibition of ornithine decarboxylase resulted in further decrease of GABA levels suggesting a compensatory activation of the ornithine/putrescine pathway as an alternative route for GABA production. These findings indicate an imbalance of cortical excitatory/inhibitory dynamics occurring during early neurodevelopmental stages in SCZ. Our study supports the hypothesis of disruption of inhibitory circuits to be causative for SCZ and establishes a novel in silico approach that enables for integrative correlation of metabolic and transcriptomic data of psychiatric disease models.

ExerG: adapting an exergame training solution to the needs of older adults using focus group and expert interviews.

BACKGROUND: Exergames are playful technology-based exercise programs. They train physical and cognitive functions to preserve independence in older adults (OAs) with disabilities in daily activities and may reduce their risk of falling. This study gathered in-depth knowledge and understanding of three different user groups' experiences in and relevant needs, worries, preferences, and expectations of technology-based training, to develop an exergame training device for OAs. METHODS: We conducted a qualitative study using semi-structured focus group interviews of primary (OAs in geriatric or neurological rehabilitation) and secondary (health professionals) end users, as well as expert interviews of tertiary end users (health insurance experts or similar), exploring user perspectives on adjusting an existing exergame to OAs' needs. Voice-recorded interviews were transcribed by researchers and analyzed using thematic analysis (TA) following an inductive, data-driven, iterative approach. RESULTS: We interviewed 24 primary, 18 secondary, and 9 tertiary end users at two rehabilitation centers in Austria and Switzerland. Our TA approach identified five to six themes per user group. Themes in the primary end user group reflected aspects of safety, training goals, individuality, game environment, social interactions, and physical and technical overload. Themes in the secondary end user group comprised facets of meaningfulness, distraction through the game environment, safety, gamification elements, the availability and accessibility of the exergame. Tertiary end users' themes addressed aspects of financial reimbursement, suitable target populations, professional training for the handling of exergame devices, training goals, and concerns about the use of exergames in geriatric rehabilitation. CONCLUSIONS: In conclusion, an exergame for OAs must be safe, motivating and fully adaptable to the target group while promoting the return to or preservation of autonomy and independence in daily life. Our findings contribute to developing hard- and software extensions for the ExerG training device. Further research is needed to expand the validity of our findings to larger populations.

Pronounced synergistic neuroprotective effect of GDNF and CNTF on axotomized retinal ganglion cells in the adult mouse.

Neuroprotection is among the potential treatment options for glaucoma and other retinal pathologies characterized by the loss of retinal ganglion cells (RGCs). Here, we examined the impact of a neural stem (NS) cell-based intravitreal co-administration of two neuroprotective factors on the survival of axotomized RGCs. To this aim we used lentiviral vectors to establish clonal NS cell lines ectopically expressing either glial cell line-derived neurotrophic factor (GDNF) or ciliary neurotrophic factor (CNTF). The modified NS cell lines were intravitreally injected either separately or as a 1:1 mixture into adult mice one day after an optic nerve lesion, and the number of surviving RGCs was determined in retinal flat-mounts two, four and eight weeks after the lesion. For the transplantation experiments, we selected a GDNF- and a CNTF-expressing NS cell line that promoted the survival of axotomized RGCs with a similar efficacy. Eight weeks after the lesion, GDNF-treated retinas contained 3.8- and CNTF-treated retinas 3.7-fold more RGCs than control retinas. Of note, the number of surviving RGCs was markedly increased when both factors were administered simultaneously, with 14.3-fold more RGCs than in control retinas eight weeks after the lesion. GDNF and CNTF thus potently and synergistically rescued RGCs from axotomy-induced cell death, indicating that combinatorial neuroprotective approaches represent a promising strategy to effectively promote the survival of RGCs under pathological conditions.

Degeneration of Photoreceptor Cells in Arylsulfatase G-Deficient Mice.

PURPOSE: Retinal degeneration is a common feature of several lysosomal storage disorders, including the mucopolysaccharidoses, a group of metabolic disorders that is characterized by widespread accumulation of glycosaminoglycans due to lysosomal enzyme dysfunction. We used a new mouse model of mucopolysaccharidosis IIIE to study the effect of Arylsulfatase G (ARSG) deficiency on retina integrity. METHODS: The retina of Arsg knockout mice aged 1 to 24 months was studied by immunohistochemistry and Western blot analysis. Electron microscopic analyses were performed on retinas from 15- and 22-month-old animals. Photoreceptor and microglia cell numbers and retina thickness were determined to quantitatively characterize retinal degeneration in ARSG-deficient mice. RESULTS: Arsg knockout mice showed a progressive degeneration of photoreceptor cells starting between 1 and 6 months of age, resulting in the loss of more than 50% of photoreceptor cells in 24-month-old mice. Photoreceptor loss was accompanied by reactive astrogliosis, reactive microgliosis that was evident in the outer but not inner retina, and elevated expression levels of some lysosomal proteins. Electron microscopic analyses of retinas revealed no evidence for the presence of storage vacuoles. Of note, expression of ARSG protein in wild-type mice was detectable only in the RPE which, however, appeared morphologically unaffected in knockout mice at the electron microscopic level. CONCLUSIONS: To our knowledge, this is the first study demonstrating that ARSG deficiency results in progressive photoreceptor degeneration and dysregulation of various lysosomal proteins.

Sustained Neural Stem Cell-Based Intraocular Delivery of CNTF Attenuates Photoreceptor Loss in the nclf Mouse Model of Neuronal Ceroid Lipofuscinosis.

A sustained intraocular administration of neurotrophic factors is among the strategies aimed at establishing treatments for currently untreatable degenerative retinal disorders. In the present study we have analyzed the neuroprotective effects of a continuous neural stem (NS) cell-based intraocular delivery of ciliary neurotrophic factor (CNTF) on photoreceptor cells in the nclf mouse, an animal model of the neurodegenerative lysosomal storage disorder variant late infantile neuronal ceroid lipofuscinosis (vLINCL). To this aim, we genetically modified adherently cultivated NS cells with a polycistronic lentiviral vector encoding a secretable variant of CNTF together with a Venus reporter gene (CNTF-NS cells). NS cells for control experiments (control-NS cells) were modified with a vector encoding the reporter gene tdTomato. Clonal CNTF-NS and control-NS cell lines were established using fluorescent activated cell sorting and intravitreally grafted into 14 days old nclf mice at the onset of retinal degeneration. The grafted cells preferentially differentiated into astrocytes that were attached to the posterior side of the lenses and the vitreal side of the retinas and stably expressed the transgenes for at least six weeks, the latest post-transplantation time point analyzed. Integration of donor cells into host retinas, ongoing proliferation of grafted cells or adverse effects of the donor cells on the morphology of the host eyes were not observed. Quantitative analyses of host retinas two, four and six weeks after cell transplantation revealed the presence of significantly more photoreceptor cells in eyes with grafted CNTF-NS cells than in eyes with grafted control-NS cells. This is the first demonstration that a continuous intraocular administration of a neurotrophic factor attenuates retinal degeneration in an animal model of neuronal ceroid lipofuscinosis.

The sheddase ADAM10 is a potent modulator of prion disease.

The prion protein (PrP(C)) is highly expressed in the nervous system and critically involved in prion diseases where it misfolds into pathogenic PrP(Sc). Moreover, it has been suggested as a receptor mediating neurotoxicity in common neurodegenerative proteinopathies such as Alzheimer's disease. PrP(C) is shed at the plasma membrane by the metalloprotease ADAM10, yet the impact of this on prion disease remains enigmatic. Employing conditional knockout mice, we show that depletion of ADAM10 in forebrain neurons leads to posttranslational increase of PrP(C) levels. Upon prion infection of these mice, clinical, biochemical, and morphological data reveal that lack of ADAM10 significantly reduces incubation times and increases PrP(Sc) formation. In contrast, spatiotemporal analysis indicates that absence of shedding impairs spread of prion pathology. Our data support a dual role for ADAM10-mediated shedding and highlight the role of proteolytic processing in prion disease.

Neural stem cell-based intraocular administration of ciliary neurotrophic factor attenuates the loss of axotomized ganglion cells in adult mice.

PURPOSE: To analyze the neuroprotective effect of intravitreally grafted neural stem (NS) cells genetically modified to secrete ciliary neurotrophic factor (CNTF) on intraorbitally lesioned retinal ganglion cells (RGCs) in adult mice. METHODS: Adherently cultivated NS cells were genetically modified to express a secretable variant of mouse CNTF together with the fluorescent reporter protein Venus. Clonal CNTF-secreting NS cell lines were established using fluorescence activated cell sorting, and intravitreally grafted into adult mice 1 day after an intraorbital crush of the optic nerve. Brn-3a-positive RGCs were counted in flat-mounted retinas at different postlesion intervals to evaluate the neuroprotective effect of the CNTF-secreting NS cells on the axotomized RGCs. Anterograde axonal tracing experiments were performed to analyze the regrowth of the injured RGC axons in CNTF-treated retinas. RESULTS: Intravitreally grafted NS cells preferentially differentiated into astrocytes that survived in the host eyes, stably expressed CNTF, and significantly attenuated the loss of the axotomized RGCs over a period of at least 4 months, the latest postlesion time point analyzed. Depending on the postlesion interval analyzed, the number of RGCs in eyes with grafted CNTF-secreting NS cells was 2.8-fold to 6.4-fold higher than in eyes with grafted control NS cells. The CNTF-secreting NS cells additionally induced long-distance regrowth of the lesioned RGC axons. CONCLUSIONS: Genetically modified clonal NS cell lines may serve as a useful tool for preclinical studies aimed at evaluating the therapeutic potential of a sustained cell-based intravitreal administration of neuroprotective factors in mouse models of glaucoma.

Genetically modified neural stem cells for a local and sustained delivery of neuroprotective factors to the dystrophic mouse retina.

A continuous intraocular delivery of neurotrophic factors (NFs) is being explored as a strategy to rescue photoreceptor cells and visual functions in degenerative retinal disorders that are currently untreatable. To establish a cell-based intraocular delivery system for a sustained administration of NFs to the dystrophic mouse retina, we used a polycistronic lentiviral vector to genetically modify adherently cultivated murine neural stem (NS) cells. The vector concurrently encoded a gene of interest, a reporter gene, and a resistance gene and thus facilitated the selection, cloning, and in vivo tracking of the modified cells. To evaluate whether modified NS cells permit delivery of functionally relevant quantities of NFs to the dystrophic mouse retina, we expressed a secretable variant of ciliary neurotrophic factor (CNTF) in NS cells and grafted the cells into the vitreous space of Pde6b(rd1) and Pde6b(rd10) mice, two animal models of retinitis pigmentosa. In both mouse lines, grafted cells attached to the retina and lens, where they differentiated into astrocytes and some neurons. Adverse effects of the transplanted cells on the morphology of host retinas were not observed. Importantly, the CNTF-secreting NS cells significantly attenuated photoreceptor degeneration in both mutant mouse lines. The neuroprotective effect was significantly more pronounced when clonally derived NS cell lines selected for high expression levels of CNTF were grafted into Pde6b(rd1) mice. Intravitreal transplantations of modified NS cells may thus represent a useful method for preclinical studies aimed at evaluating the therapeutic potential of a cell-based intraocular delivery of NFs in mouse models of photoreceptor degeneration.