The Flavonol Quercitrin Hinders GSK3 Activity and Potentiates the Wnt/β-Catenin Signaling Pathway

The Wnt/β-catenin signaling pathway dictates cell proliferation and differentiation during embryonic development and tissue homeostasis. Its deregulation is associated with many pathological conditions, including neurodegenerative disease, frequently downregulated. The lack of efficient treatment for these diseases, including Alzheimer’s disease (AD), makes Wnt signaling an attractive target for therapies. Interestingly, novel Wnt signaling activating compounds are less frequently described than inhibitors, turning the quest for novel positive modulators even more appealing. In that sense, natural compounds are an outstanding source of potential drug leads. Here, we combine different experimental models, cell-based approaches, neuronal culture assays, and rodent behavior tests with Xenopus laevis phenotypic analysis to characterize quercitrin, a natural compound, as a novel Wnt signaling potentiator. We find that quercitrin potentiates the signaling in a concentration-dependent manner and increases the occurrence of the Xenopus secondary axis phenotype mediated by Xwnt8 injection. Using a GSK3 biosensor, we describe that quercitrin impairs GSK3 activity and increases phosphorylated GSK3β S9 levels. Treatment with XAV939, an inhibitor downstream of GSK3, impairs the quercitrin-mediated effect. Next, we show that quercitrin potentiates the Wnt3a-synaptogenic effect in hippocampal neurons in culture, which is blocked by XAV939. Quercitrin treatment also rescues the hippocampal synapse loss induced by intracerebroventricular injection of amyloid-β oligomers (AβO) in mice. Finally, quercitrin rescues AβO-mediated memory impairment, which is prevented by XAV939. Thus, our study uncovers a novel function for quercitrin as a Wnt/β-catenin signaling potentiator, describes its mechanism of action, and opens new avenues for AD treatments.     

Quantitative Analysis of Plant Cytosolic Calcium Signals in Response to Water Activated by Low-Power Non-Thermal Plasma

Non-thermal plasma technology is increasingly being applied in the plant biology field. Despite the variety of beneficial effects of plasma-activated water (PAW) on plants, information about the mechanisms of PAW sensing by plants is still limited. In this study, in order to link PAW perception to the positive downstream responses of plants, transgenic Arabidopsis thaliana seedlings expressing the Ca²⁺-sensitive photoprotein aequorin in the cytosol were challenged with water activated by low-power non-thermal plasma generated by a dielectric barrier discharge (DBD) source. PAW sensing by plants resulted in the occurrence of cytosolic Ca²⁺ signals, whose kinetic parameters were found to strictly depend on the operational conditions of the plasma device and thus on the corresponding mixture of chemical species contained in the PAW. In particular, we highlighted the effect on the intracellular Ca²⁺ signals of low doses of DBD-PAW chemicals and also presented the effects of consecutive plant treatments. The results were discussed in terms of the possibility of using PAW-triggered Ca²⁺ signatures as benchmarks to accurately modulate the chemical composition of PAW in order to induce environmental stress resilience in plants, thus paving the way for further applications in agriculture.     

Tolerance,Adaptation, and Cell Response Elicited by Micromonospora sp. Facing Tellurite Toxicity: A Biological and Physical-Chemical Characterization

The intense use of tellurium (Te) in industrial applications, along with the improper disposal of Te-derivatives, is causing their accumulation in the environment, where oxyanion tellurite (TeO₃²⁻) is the most soluble, bioavailable, and toxic Te-species. On the other hand, tellurium is a rare metalloid element whose natural supply will end shortly with possible economic and technological effects. Thus, Te-containing waste represents the source from which Te should be recycled and recovered. Among the explored strategies, the microbial TeO₃²⁻ biotransformation into less toxic Te-species is the most appropriate concerning the circular economy. Actinomycetes are ideal candidates in environmental biotechnology. However, their exploration in TeO₃²⁻ biotransformation is scarce due to limited knowledge regarding oxyanion microbial processing. Here, this gap was filled by investigating the cell tolerance, adaptation, and response to TeO₃²⁻ of a Micromonospora strain isolated from a metal(loid)-rich environment. To this aim, an integrated biological, physical-chemical, and statistical approach combining physiological and biochemical assays with confocal or scanning electron (SEM) microscopy and Fourier-transform infrared spectroscopy in attenuated total reflectance mode (ATR-FTIR) was designed. Micromonospora cells exposed to TeO₃²⁻ under different physiological states revealed a series of striking cell responses, such as cell morphology changes, extracellular polymeric substance production, cell membrane damages and modifications, oxidative stress burst, protein aggregation and phosphorylation, and superoxide dismutase induction. These results highlight this Micromonospora strain as an asset for biotechnological purposes.     

Binding of Gamma-Glutamyl Transferase to TLR4 Signalling Allows Tissue Factor Activation in Monocytes

Gamma-glutamyl transferase (GGT) is involved in the progression of atherosclerosis, since its enzymatic activity promotes the generation of reactive oxygen species (ROS). Besides, GGT may act as a prothrombotic factor by inducing tissue factor (TF) expression, independently of its enzymatic activity. The aim of this study was to assess whether GGT-induced TF stimulation was a consequence of binding to toll-like receptor 4 (TLR4) expressed on monocytes, the precursors of macrophages and foam cells which colocalize with GGT activity within atherosclerotic plaques. Experiments were performed in human peripheral blood mononuclear cells (PBMCs), THP-1 cells (a monocytic cellular model), and HEK293 cells, which were genetically modified to study the activation of TLR4. TF procoagulant activity was assessed by a one-stage clotting time test, and TF protein expression was estimated by western blot. Human recombinant (hr) GGT protein increased TF procoagulant activity and protein expression in both PBMCs and THP-1 cells. The GGT-induced TF stimulation was prevented by cellular pretreatment with TLR4/NF-κB inhibitors (LPS-Rs, CLI-095, and BAY-11-7082), and HEK293 cells lacking TLR4 confirmed that TLR4 is essential for GGT-induced activation of NF-κB. In conclusion, hrGGT induced TF expression in monocytes through a cytokine-like mechanism that involved the activation of TLR4/NF-κB signaling.     

The PINK1 p.Asn521Thr Variant Is Associated with Earlier Disease Onset in GRN/C9orf72 Frontotemporal Lobar Degeneration

Genetic frontotemporal lobar degeneration (FTLD) is characterized by heterogeneous phenotypic expression, with a disease onset highly variable even in patients carrying the same mutation. Herein we investigated if variants in lysosomal genes modulate the age of onset both in FTLD due to GRN null mutations and C9orf72 expansion. In a total of 127 subjects (n = 74 GRN mutations and n = 53 C9orf72 expansion carriers), we performed targeted sequencing of the top 98 genes belonging to the lysosomal pathway, selected based on their high expression in multiple brain regions. We described an earlier disease onset in GRN/C9orf72 pedigrees in subjects carrying the p.Asn521Thr variant (rs1043424) in PTEN-induced kinase 1 (PINK1), a gene that is already known to be involved in neurodegenerative diseases. We found that: (i) the PINK1 rs1043424 C allele is significantly associated with the age of onset; (ii) every risk C allele increases hazard by 2.11%; (iii) the estimated median age of onset in homozygous risk allele carriers is 10–12 years earlier than heterozygous/wild type homozygous subjects. A replication study in GRN/C9orf72 negative FTLD patients confirmed that the rs1043424 C allele was associated with earlier disease onset (−5.5 years in CC versus A carriers). Understanding the potential mechanisms behind the observed modulating effect of the PINK1 gene in FTLD might prove critical for identifying biomarkers and/or designing drugs to modify the age of onset, especially in GRN/C9orf72-driven disease.     

Translational Implications for Radiosensitizing Strategies in Rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood and adolescence that includes FP-RMS, harboring the fusion oncoprotein PAX3/7-FOXO1 and FN-RMS, often mutant in the RAS pathway. Risk stratifications of RMS patients determine different prognostic groups and related therapeutic treatment. Current multimodal therapeutic strategies involve surgery, chemotherapy (CHT) and radiotherapy (RT), but despite the deeper knowledge of response mechanisms underpinning CHT treatment and the technological improvements that characterize RT, local failures and recurrence frequently occur. This review sums up the RMS classification and the management of RMS patients, with special attention to RT treatment and possible radiosensitizing strategies for RMS tumors. Indeed, RMS radioresistance is a clinical problem and further studies aimed at dissecting radioresistant molecular mechanisms are needed to identify specific targets to hit, thus improving RT-induced cytotoxicity.     

Modeling RTT Syndrome by iPSC-Derived Neurons from Male and Female Patients with Heterogeneously Severe Hot-Spot MECP2 Variants

Rett syndrome caused by MECP2 variants is characterized by a heterogenous clinical spectrum accounted for in 60% of cases by hot-spot variants. Focusing on the most frequent variants, we generated in vitro iPSC-neurons from the blood of RTT girls with p.Arg133Cys and p.Arg255*, associated to mild and severe phenotype, respectively, and of an RTT male harboring the close to p.Arg255*, p.Gly252Argfs*7 variant. Truncated MeCP2 proteins were revealed by Western blot and immunofluorescence analysis. We compared the mutant versus control neurons at 42 days for morphological parameters and at 120 days for electrophysiology recordings, including girls’ isogenic clones. A precocious reduced morphological complexity was evident in neurons with truncating variants, while in p.Arg133Cys neurons any significant differences were observed in comparison with the isogenic wild-type clones. Reduced nuclear size and branch number show up as the most robust biomarkers. Patch clamp recordings on mature neurons allowed the assessment of cell biophysical properties, V-gated currents, and spiking pattern in the mutant and control cells. Immature spiking, altered cell capacitance, and membrane resistance of RTT neurons, were particularly pronounced in the Arg255* and Gly252Argfs*7 mutants. The overall results indicate that the specific markers of in vitro cellular phenotype mirror the clinical severity and may be amenable to drug testing for translational purposes.     

Tomato Sterol 22-desaturase Gene CYP710A11: Its Roles in Meloidogyne incognita Infection and Plant Stigmasterol Alteration

Sterols are isoprenoid-derived lipids that play essential structural and functional roles in eukaryotic cells. Plants produce a complex mixture of sterols, and changes in plant sterol profiles have been linked to plant–pathogen interactions. β-Sitosterol and stigmasterol, in particular, have been associated with plant defense. As nematodes have lost the ability to synthesize sterols de novo, they require sterols from the host. Tomato (Solanum lycopersicum) plants infected by the plant parasitic nematode Meloidogyne incognita show a reduced level of stigmasterol and a repression of the gene CYP710A11, encoding the sterol C-22 desaturase that is responsible for the conversion of β-sitosterol to stigmasterol. In this study, we investigated the role of the tomato sterol C-22 desaturase gene CYP710A11 in the response to infection by M. incognita. We explored the plant–nematode interaction over time by analyzing the plant sterol composition and CYP710A11 gene regulation in S. lycopersicum after M. incognita infection. The temporal gene expression analysis showed that 3 days after inoculation with M. incognita, the CYP710A11 expression was significantly suppressed in the tomato roots, while a significant decrease in the stigmasterol content was observed after 14 days. A cyp710a11 knockout mutant tomato line lacking stigmasterol was analyzed to better understand the role of CYP710A11 in nematode development. M. incognita grown in the mutant line showed reduced egg mass counts, presumably due to the impaired growth of the mutant. However, the nematodes developed as well as they did in the wild-type line. Thus, while the suppression of CYP710A11 expression during nematode development may be a defense response of the plant against the nematode, the lack of stigmasterol did not seem to affect the nematode. This study contributes to the understanding of the role of stigmasterol in the interaction between M. incognita and tomato plants and shows that the sterol C-22 desaturase is not essential for the success of M. incognita.     

AQP4-MOG Double-Positive Neuromyelitis Optica Spectrum Disorder: Case Report with Central and Peripheral Nervous System Involvement and Review of Literature

(1) The co-occurrence of AQP4 and myelin oligodendrocyte glycoprotein (MOG) antibodies in patients with demyelinating disorders is extremely rare. In addition, a concomitant involvement of the peripheral nervous system (PNS) has been described either in association with AQP4 antibodies-positive neuromyelitis optica spectrum disorder (NMOSD), or MOG-associated disease. We report on a case of NMOSD with co-occurrence of AQP4 and MOG antibodies and concomitant central and peripheral nervous system involvement. We also reviewed available cases of AQP4-MOG double-positive patients. (2) Brain and spine MRI, cerebrospinal fluid studies, and electrophysiological test were performed. Serum AQP4 and MOG positivity was assessed with live cell-based assay. (3) A 62-year-old woman presented with recurrent optic neuritis, myelitis, and radiculitis, tested positive for AQP4 and MOG antibodies, and was treated successfully with rituximab. (4) Although few cases of AQP4-MOG double-positive patients were already described mostly affecting females with a concomitant spinal cord and optical nerve involvement, we describe the first case of double-positive NMOSD with the peculiar involvement of both central and peripheral nervous system.