Transplantation of Human Glial Progenitors to Immunodeficient Neonatal Mice with Amyotrophic Lateral Sclerosis (SOD1/rag2).

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal disease with no effective therapy. The neurodegenerative character of ALS was an appealing target for stem cell-based regenerative approaches. Different types of stem cells have been transplanted in both preclinical and clinical settings, but no convincing outcomes have been noted. Human glial restricted precursors (hGRPs) transplanted intraventricularly to neonatal, immunodeficient mice rescued lifespan of dysmyelinated mice. Intraspinal injection of hGRPs also provided benefits in the mouse model of ALS. Therefore, we have recently developed an immunodeficient model of ALS (double mutant SOD1/rag2), and, in this study, we tested the strategy previously used in dysmyelinated mice of intraventricular transplantation of hGRPs to immunodeficient mice. To maximize potential therapeutic benefits, the cells were implanted into neonates. We used magnetic resonance imaging to investigate the progression of neurodegeneration and therapeutic responses. A cohort of animals was devoted to survival assessment. Postmortem analysis included immunohistochemistry, Nissl staining, and Western blots. Cell transplantation was not associated with improved animal survival, slowing neurodegeneration, or accumulation of misfolded superoxide dismutase 1. Postmortem analysis did not reveal any surviving hGRPs. Grafting into neonatal immunodeficient recipients did not prevent ALS-induced cell loss, which might explain the lack of positive therapeutic effects. The results of this study are in line with the modest effects of clinical neurotransplantations. Therefore, we urge stem cell and ALS communities to develop and implement cell tracking methods to better understand cell fates in the clinic.

Alternative Splicing Regulation of Glycine-Rich Proteins via Target of Rapamycin-Reactive Oxygen Species Pathway in Arabidopsis Seedlings Upon Glucose Stress.

Glucose can serve as both the source of energy and regulatory signaling molecule in plant. Due to the environmental and metabolic change, sugar levels could affect various developmental processes. High glucose environment is hardly conductive to the plant growth but cause development arrest. Increasing evidence indicate that alternative splicing (AS) plays a pivotal role in sugar signaling. However, the regulatory mechanism upon glucose stress remains unclear. The full-length transcriptomes were obtained from the samples of Arabidopsis seedlings with 3% glucose and mock treatment, using Oxford Nanopore sequencing technologies. Further analysis indicated that many genes involved in photosynthesis were significantly repressed and many genes involved in glycolysis, mitochondrial function, and the response to oxidative stress were activated. In total, 1,220 significantly differential alternative splicing (DAS) events related to 619 genes were identified, among which 75.74% belong to intron retention (IR). Notably, more than 20% of DAS events come from a large set of glycine-rich protein (GRP) family genes, such as GRP7, whose AS types mostly belong to IR. Besides the known productive GRP transcript isoforms, we identified a lot of splicing variants with diverse introns spliced in messenger RNA (mRNA) region coding the glycine-rich (GR) domain. The AS pattern of GRPs changed and particularly, the productive GRPs increased upon glucose stress. These ASs of GRP pre-mRNAs triggered by glucose stress could be abolished by AZD-8055, which is an ATP competitive inhibitor for the target of rapamycin (TOR) kinase but could be mimicked by H2O2. Additionally, AS pattern change of arginine/serine-rich splicing factor 31(RS31) via TOR pathway, which was previously described in response to light and sucrose signaling, was also induced in a similar manner by both glucose stress and reactive oxygen species (ROS). Here we conclude that (i) glucose stress suppresses photosynthesis and activates the glycolysis-mitochondria energy relay and ROS scavenging system; (ii) glucose stress triggers transcriptome-wide AS pattern changes including a large set of splicing factors, such as GRPs and RS31; (iii) high sugars regulate AS pattern change of both GRPs and RS31 via TOR-ROS pathway. The results from this study will deepen our understanding of the AS regulation mechanism in sugar signaling.

Genome-wide identification, phylogenetic analysis, and expression profiling of glycine-rich RNA-binding protein (GRPs) genes in seeded and seedless grapes (Vitis vinifera).

Glycine-rich RNA-binding proteins (GRPs) are essential for many physiological and biochemical processes in plants, especially the response to environmental stresses. GRPs exist widely in angiosperms and gymnosperms plant species; however, their roles in Vitis vinifera are still poorly understood. To characterize VviGRP gene family, we performed a genomic survey, bioinformatics and expression analysis of VviGRPs in grape. We identified nineteen VviGRPs gene family members. The result of bioinformatics analysis showed their motif distribution, gene structure characteristics and chromosomal locations. Then we carried out synteny and phylogenetic analysis to study the origin and evolutionary relationship of GRPs. Tissue-specific expression analysis showed that VviGRPs have different expression patterns. Meanwhile, we studied expression profiles of seventeen ovule-expressed genes during seed development of stenospermocarpic seedless and seeded grapes, and the result showed that most of them have much higher relative expression levels in stenospermocarpic seedless grapes than that of seeded one before 25 days after full bloom (DAFB). It is suggested that VviGRPs may involve in the seed development process. Taken together, our research indicated that VviGRPs are related to seed development and will be beneficial for further investigations into the seed abortion mechanism under stenospermocarpic grapes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01082-3.

Myelin-Independent Therapeutic Potential of Canine Glial-Restricted Progenitors Transplanted in Mouse Model of Dysmyelinating Disease.

BACKGROUND: Dysfunction of glia contributes to the deterioration of the central nervous system in a wide array of neurological disorders, thus global replacement of glia is very attractive. Human glial-restricted precursors (hGRPs) transplanted intraventricularly into neonatal mice extensively migrated and rescued the lifespan in half of the studied mice, whereas mouse GRPs (mGRPs) presented no therapeutic benefit. We studied in the same experimental setting canine GRPs (cGRP) to determine whether their therapeutic potential falls between hGRPs and mGRPs. Additional motivation for the selection of cGRPs was a potential for use in veterinary medicine. METHODS: cGRPs were extracted from the brain of dog fetuses. The cells were transplanted into the anterior or posterior aspect of the lateral ventricle (LV) of neonatal, immunodeficient, dysmyelinated mice (Mbpshi, Rag2 KO; shiv/rag2). Outcome measures included early cell biodistribution, animal survival and myelination assessed with MRI, immunohistochemistry and electron microscopy. RESULTS: Grafting of cGRP into posterior LV significantly extended animal survival, whereas no benefit was observed after anterior LV transplantation. In contrast, myelination of the corpus callosum was more prominent in anteriorly transplanted animals. CONCLUSIONS: The extended survival of animals after transplantation of cGRPs could be explained by the vicinity of the transplant near the brain stem.

Italian Adolescent Young Caregivers of Grandparents: Difficulties Experienced and Support Needed in Intergenerational Caregiving-Qualitative Findings from a European Union Funded Project.

The article aims to describe the experiences of 87 Italian adolescent young caregivers (AYCs) of grandparents (GrPs), with reference to the caregiving stress appraisal model (CSA) that provides a theoretical lens to explore the difficulties encountered and support needed in their caring role. Qualitative data were drawn from an online survey conducted within an EU Horizon 2020 funded project. An inductive thematic analysis was carried out, and the findings were critically interpreted within the conceptual framework of the CSA model. The analysis highlighted three categories of difficulties: material, communication and emotional/psychological. The most common material difficulty was the physical strain associated with moving "uncooperative" disabled older adults. The types of support needed concerned both emotional and material support. The study provides a deeper understanding of the under-studied experiences of AYCs of GrPs. Based on these findings, policies and support measures targeted at AYCs of GrPs should include early needs detection, emotional support and training on intergenerational caring in order to mitigate the stress drivers. Moreover, the study advances the conceptualisation of the CSA model by considering the above-mentioned aspects related to intergenerational caregiving.

Plant Glycine-Rich Proteins in Stress Response: An Emerging, Still Prospective Story.

Seed plants are sessile organisms that have developed a plethora of strategies for sensing, avoiding, and responding to stress. Several proteins, including the glycine-rich protein (GRP) superfamily, are involved in cellular stress responses and signaling. GRPs are characterized by high glycine content and the presence of conserved segments including glycine-containing structural motifs composed of repetitive amino acid residues. The general structure of this superfamily facilitates division of GRPs into five main subclasses. Although the participation of GRPs in plant stress response has been indicated in numerous model and non-model plant species, relatively little is known about the key physiological processes and molecular mechanisms in which those proteins are engaged. Class I, II, and IV members are known to be involved in hormone signaling, stress acclimation, and floral development, and are crucial for regulation of plant cells growth. GRPs of class IV [RNA-binding proteins (RBPs)] are involved in alternative splicing or regulation of transcription and stomatal movement, seed, pollen, and stamen development; their accumulation is regulated by the circadian clock. Owing to the fact that the overexpression of GRPs can confer tolerance to stress (e.g., some are involved in cold acclimation and may improve growth at low temperatures), these proteins could play a promising role in agriculture through plant genetic engineering. Consequently, isolation, cloning, characterization, and functional validation of novel GRPs expressed in response to the diverse stress conditions are expected to be growing areas of research in the coming years. According to our knowledge, this is the first comprehensive review on participation of plant GRPs in the response to diverse stress stimuli.

Marked Differences in the Submandibular Salivary Proteome between Sardinian Alcohol-Preferring and Sardinian Alcohol-Non Preferring Rats Revealed by an Integrated Top-Down-Bottom-Up Proteomic Platform.

Sardinian alcohol-preferring (sP) and Sardinian alcohol-non preferring (sNP) rats have been selectively bred for opposite alcohol preference and consumption. Aiming to verify possible differences at the proteomics level between sP and sNP rats, we investigated the salivary proteome by a a liquid chromatography-mass spectrometry top-down-bottom-up integrated approach. For this purpose, submandibular saliva was collected from alcohol-naive sP and sNP rats under isoprenaline stimulation. A total of 200 peptides and proteins were detected and quantified in the two rat lines, 149 of which were characterized in their naturally occurring structure. The data are available via ProteomeXchange with identifier PXD006997. Surprisingly, sP rats exhibited marked quantitative and qualitative differences with respect to sNP rats, namely higher levels of proteoforms originating from submandibular gland protein C, and from submandibular rat protein 2, as well as those of several unidentified peptides and proteins. sP rats expressed some proteins not detectable in sNP rats such as the glutamine and glutamic acid-rich protein (GRP)-CB. The isoform GRP-B, detectable in both rat lines, was more abundant in sNP rats. The submandibular saliva of sNP rats was also characterized by very high levels of GRP-B proteolytic peptides and rat salivary protein 1. Whether these differences could contribute to the opposite alcohol preference and consumption of sP and sNP rats is currently unknown and requires further investigation.

Heterologous expression of a novel Zoysia japonica salt-induced glycine-rich RNA-binding protein gene, ZjGRP, caused salt sensitivity in Arabidopsis.

KEY MESSAGE: A novel Zoysia japonica salt-induced glycine-rich RNA-binding protein gene was cloned in this study and its overexpression caused salt sensitivity in transgenic Arabidopsis. Glycine-rich RNA-binding proteins (GRPs) play crucial roles in diverse plant developmental processes. However, the mechanisms and functions of GRPs in salinity stress responses remain largely unknown. In this study, rapid amplification of cDNA end (RACE) PCR methods was adopted to isolate ZjGRP from Zosyia japonica, a salt-tolerant grass species. ZjGRP cDNA was 456 bp in length, corresponding to 151 amino acids. ZjGRP was localized in the nucleus and cytoplasm, and was found particularly abundantly in stomatal guard cells. Quantitative real-time PCR showed that ZjGRP was expressed in the roots, stems, and leaves of Zoysia japonica, with the greatest expression seen in the fast-growing leaves. Furthermore, expression of ZjGRP was strongly induced by treatment with NaCl, ABA, MeJA, and SA. Overexpression of ZjGRP in Arabidopsis reduced the rate of germination and retarded seedling growth. ZjGRP-overexpressing Arabidopsis thaliana exhibited weakened salinity tolerance, likely as a result of effects on ion transportation, osmosis, and antioxidation. This study indicates that ZjGRP plays an essential role in inducing salt sensitivity in transgenic plants.

Diversification of the plant-specific hybrid glycine-rich protein (HyGRP) genes in cereals.

Plant-specific hybrid proline- or glycine-rich proteins (HyP/GRPs) are involved in diverse gene functions including plant development and responses to biotic and abiotic stresses. The quantitative trait locus, qLTG3-1, enhances seed germination in rice under low-temperature conditions and encodes a member with a glycine-rich motif of the HyP/GRP family. The function of this gene may be related to the weakening of tissue covering the embryo during seed germination. In the present study, the diversification of the HyP/GRP gene family was elucidated in rice based on phylogenetic relationships and gene expression levels. At least 21 members of the HyP/GRP family have been identified in the rice genome and clustered in five regions on four chromosomes by tandem and chromosomal duplications. Of these, OsHyPRP05 (qLTG3-1) and its paralogous gene, OsHyPRP21, had a glycine-rich motif. Furthermore, orthologous genes with a glycine-rich motif and the HyP/GRP gene family were detected in four genome-sequenced monocots: 12 in barley, 10 in Brachypodium, 20 in maize, and 28 in sorghum, using a BLAST search of qLTG3-1 as the query. All members of the HyP/GRP family in these five species were classified into seven main groups, which were clustered together in these species. These results suggested that the HyP/GRP gene family was formed in the ancestral genome before the divergence of these species. The collinearity of chromosomal regions around qLTG3-1 and its orthologous genes were conserved among rice, Brachypodium, sorghum, and maize, indicating that qLTG3-1 and orthologous genes conserve gene function during seed germination.

Effectiveness of tobacco control television advertising in changing tobacco use in England: a population-based cross-sectional study.

AIM: To examine whether government-funded tobacco control television advertising shown in England between 2002 and 2010 reduced adult smoking prevalence and cigarette consumption. DESIGN: Analysis of monthly cross-sectional surveys using generalised additive models. SETTING: England. PARTICIPANTS: More than 80 000 adults aged 18 years or over living in England and interviewed in the Opinions and Lifestyle Survey. MEASUREMENTS: Current smoking status, smokers' daily cigarette consumption, tobacco control gross rating points (GRPs-a measure of per capita advertising exposure combining reach and frequency), cigarette costliness, tobacco control activity, socio-demographic variables. FINDINGS: After adjusting for other tobacco control policies, cigarette costliness and individual characteristics, we found that a 400-point increase in tobacco control GRPs per month, equivalent to all adults in the population seeing four advertisements per month (although actual individual-level exposure varies according to TV exposure), was associated with 3% lower odds of smoking 2 months later [odds ratio (OR) = 0.97, 95% confidence interval (CI) = 0.95, 0.999] and accounted for 13.5% of the decline in smoking prevalence seen over this period. In smokers, a 400-point increase in GRPs was associated with a 1.80% (95%CI = 0.47, 3.11) reduction in average cigarette consumption in the following month and accounted for 11.2% of the total decline in consumption over the period 2002-09. CONCLUSION: Government-funded tobacco control television advertising shown in England between 2002 and 2010 was associated with reductions in smoking prevalence and smokers' cigarette consumption.

Classification and comparative analysis of Curcuma longa L. expressed sequences tags (ESTs) encoding glycine-rich proteins (GRPs).

Glycine-rich proteins (GRPs) are a group of proteins characterized by their high content of glycine residues often occurring in repetitive blocs. The diverse expression pattern and sub cellular localization of various GRPs suggest their implication in different physiological processes. Several GRPs has been isolated and characterized from different monocots and dicots. However, little or no information is available about the structure and function of GRPs in asexually reproducing plants. In this study, in-silico analysis of expressed sequence tag database resulted in the isolation of fifty-one GRPs from Curcuma longa L., an asexually reproducible plant of great medicinal and economic significance. Phylogenetic analysis grouped the GRPs into four distinct classes based on conserved motifs and nature of glycine-rich repeats. Majority of the isolated GRPs exhibited high homology with known GRPs from other plants that are expressed in response to various stresses. The presence of high structural diversity and signal peptide in some GRPs suggest their diverse physiological role and tissue specific localization. The isolated sequences can be used as a framework for cloning, characterization and expressional analysis of GRPs in response to various biotic and abiotic stresses in Curcuma longa as well as other asexually reproducing plants.