Tau protein misfolding and aggregation are central to Tauopathies, yet the temporal dynamics of Tau interactions in vivo remain poorly understood. Here, we applied quantitative proteomics to demonstrate that the interactome of human Tau in adult Drosophila brains changes dynamically over a 12-day time course, revealing a progressive shift from early cytosolic and ribosomal associations to late enrichment of mitochondrial and synaptic partners. Notably, the mitochondrial pore protein Porin/VDAC1 was identified as a late-stage interactor and functional analyses demonstrated that Tau overexpression impairs mitochondrial respiration, elevates oxidative damage, and disrupts carbohydrate homeostasis. To validate this temporally specific interaction, Porin was downregulated, resulting in reduced Tau mitochondrial association, phosphorylation and aggregation. Paradoxically, however, Porin attenuation exacerbated Tau-induced toxicity, including shortened lifespan, locomotor deficits, and impaired learning. These findings indicate that while Porin facilitates pathological Tau modifications, it is also essential for neuronal resilience, highlighting a complex role in modulating Tau toxicity. Our study provides a temporal map of Tau-associated proteome changes in vivo and identifies mitochondria as critical mediators of Tau-driven neurodegeneration.

Background: Optimal porosity and surface area of additively manufactured Ti-6Al-4V scaffolds for early human mesenchymal stem cell (hMSC) retention remain unclear, despite their importance in reconstruction of critical-sized bone defects. This study evaluated scaffold architectures fulfilling biomechanical criteria for hMSC and osteocyte survival, proliferation, differentiation, cell cycle, and retention. Methods: Primary hMSCs from four healthy donors were characterized by flow cytometry, immunofluorescence, and Western blotting. Cells were seeded onto Ti-6Al-4V scaffolds with graded porosity (P50–P90, 50–90% porosity) and cultured for 120 h. Viability and retention were measured by trypan blue exclusion, apoptosis and cell cycle by flow cytometry, and osteogenic differentiation by collagen I, osteocalcin, and Akt phosphorylation analyses. Collagen-embedded hMSCs and osteoblasts were used to assess migration and phenotype maintenance. Results: The densest scaffold, P50, consistently retained significantly more viable hMSCs and differentiated osteoblasts than P60 and P70 (p < 0.005 in both cases, One-way ANOVA analysis, significance level a = 0.05 followed by Bonferroni correction). Among the scaffold architectures investigated, P50 demonstrated the most favorable early cellular retention under the present experimental conditions. No scaffold-induced apoptosis or proliferation changes were detected. Osteogenic differentiation and Akt phosphorylation increased during culture. In collagen-containing scaffolds, cells migrated from the matrix and preferentially coated titanium struts, with P50 supporting superior attachment. Conclusions: Among the scaffold architectures investigated, the low-porosity/high-surface-area P50 design provided the most favorable early microenvironment for hMSC and osteoblast retention without cytotoxic effects.

Metabolic homeostasis is a key determinant of stress resilience and healthy ageing. Pomegranate juice (PGJ) and its characteristic ellagitannin punicalagin (PUN) have been associated with beneficial metabolic effects, but the mechanisms through which they influence metabolic adaptation and healthspan, and whether they act through shared or distinct pathways, remain poorly defined. We investigated the effects of lyophilized PGJ (50, 100, and 200 μg/mL) and PUN (25, 50, and 100 μM) on metabolic regulation and stress resilience in Caenorhabditis elegans. Nuclear magnetic resonance spectroscopy was employed for metabolic profiling of PGJ, whereas ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry enabled the quantification of PUN at 2.4 ± 0.127 µg/mg lyophilized juice. Under glucose-enriched conditions, PGJ (200 μg/mL) and PUN (100 μM) reduced normalised Nile Red fluorescence to 0.53 and 0.59 of vehicle levels. On day 10 of adulthood, both treatments increased mean survival under oxidative stress by 1.73- and 1.50-fold, respectively. Both treatments increased MDT-15 reporter response (1.13- and 1.44-fold, respectively), whereas PGJ preferentially increased AAK-2- and FAT-6-associated responses and PUN elevated NHR-49- and FAT-7 expression. PUN also improved locomotor performance under amyloid-β-induced proteotoxic stress. Overall, PGJ and PUN elicited partially overlapping metabolic responses in C. elegans, with NHR-49 and MDT-15 emerging as treatment-responsive regulators associated with lipid homeostasis.