Quantitative proteome profiling reveals molecular hallmarks of egg quality in Atlantic halibut: impairments of transcription and protein folding impede protein and energy homeostasis during early development.

BACKGROUND: Tandem mass tag spectrometry (TMT labeling-LC-MS/MS) was utilized to examine the global proteomes of Atlantic halibut eggs at the 1-cell-stage post fertilization. Comparisons were made between eggs judged to be of good quality (GQ) versus poor quality (BQ) as evidenced by their subsequent rates of survival for 12 days. Altered abundance of selected proteins in BQ eggs was confirmed by parallel reaction monitoring spectrometry (PRM-LC-MS/MS). Correspondence of protein levels to expression of related gene transcripts was examined via qPCR. Potential mitochondrial differences between GQ and BQ eggs were assessed by transmission electron microscopy (TEM) and measurements of mitochondrial DNA (mtDNA) levels. RESULTS: A total of 115 proteins were found to be differentially abundant between GQ and BQ eggs. Frequency distributions of these proteins indicated higher protein folding activity in GQ eggs compared to higher transcription and protein degradation activities in BQ eggs. BQ eggs were also significantly enriched with proteins related to mitochondrial structure and biogenesis. Quantitative differences in abundance of several proteins with parallel differences in their transcript levels were confirmed in egg samples obtained over three consecutive reproductive seasons. The observed disparities in global proteome profiles suggest impairment of protein and energy homeostasis related to unfolded protein response and mitochondrial stress in BQ eggs. TEM revealed BQ eggs to contain significantly higher numbers of mitochondria, but differences in corresponding genomic mtDNA (mt-nd5 and mt-atp6) levels were not significant. Mitochondria from BQ eggs were significantly smaller with a more irregular shape and a higher number of cristae than those from GQ eggs. CONCLUSION: The results of this study indicate that BQ Atlantic halibut eggs are impaired at both transcription and translation levels leading to endoplasmic reticulum and mitochondrial disorders. Observation of these irregularities over three consecutive reproductive seasons in BQ eggs from females of diverse background, age and reproductive experience indicates that they are a hallmark of poor egg quality. Additional research is needed to discover when in oogenesis and under what circumstances these defects may arise. The prevalence of this suite of markers in BQ eggs of diverse vertebrate species also begs investigation.

Harnessing the Antioxidative Potential of Dental Pulp Stem Cell-Conditioned Medium in Photopolymerized GelMA Hydrogels.

Gelatin methacryloyl (GelMA) stands out for its biocompatibility, tunability, and functionality, being often selected as a scaffolding material. However, the biological modulations induced by its photocrosslinking process on mesenchymal stem cells as well as stress mitigation measures remain insufficiently explored. By using GelMA of Good Manufacturing Practice (GMP) grade, this study aimed (a) to achieve a comprehensive understanding of the biological effects of photocrosslinking process with a specific focus on oxidative stress and (b) to develop a strategy to mitigate the adverse effects by employing conditioned medium (CM) by dental pulp stem cells (DPSCs). Following photocrosslinking, pathways related to oxidative phosphorylation and DNA repair were enriched in the presence of DPSC-CM carrying various antioxidants such as peroxiredoxin (PRDX) 1-6 and superoxide dismutase type 1 (SOD1), while the control samples exhibited enrichment in inflammatory signaling pathways. Incorporating DPSC-CM into the hydrogel notably reduced the degree of cellular oxidation caused by photocrosslinking and stress responses, resulting in improved cell viability, growth, motility, and osteogenic differentiation, as well as fewer apoptotic and senescent cells compared to those without DPSC-CM. The deteriorated biocompatibility of freshly crosslinked GelMA hydrogel was confirmed by the disrupted vasculature of chorioallantoic membranes in chicken embryos after implantation, which was prevented by DPSC-CM. In conclusion, this study demonstrates the robust antioxidative effects of DPSC-CM, mitigating the negative effect of GelMA photocrosslinking processes.