Exogenous pyruvate and recombinant human basic fibroblast growth factor maintain pluripotency and enhance global metabolic activity of bovine embryonic stem cells grown on low-density feeder layers.

A suitable microenvironment or niche is essential for self-renewal and pluripotency of stem cells cultured in vitro, including bovine embryonic stem cells (bESCs). Feeder cells participate in the construction of stem cell niche by secreting growth factors and extracellular matrix proteins. In this study, metabolomics and transcriptomics analyses were used to investigate the effects of low-density feeder cells on bESCs. The results showed that bESCs co-cultured with low-density feeder cells experienced a decrease in pluripotent gene expression, cell differentiation, and a reduction of central carbon metabolic activity. When cell-permeable pyruvate (Pyr) and recombinant human basic fibroblast growth factor (rhbFGF) were added to the culture system, the pluripotency of bESCs on low-density feeder layers was rescued, and acetyl-coenzyme A (AcCoA) synthesis and fatty acid de novo synthesis increased. In addition, rhbFGF enhances the effects of Pyr and activates the overall metabolic level of bESCs grown on low-density feeder layers. This study explored the rescue effects of exogenous Pyr and rhbFGF on bESCs cultured on low-density feeder layers, which will provide a reference for improvement of the PSC culture system through the supplementation of energy metabolites and growth factors.

Methanol fixed feeder layers altered the pluripotency and metabolism of bovine pluripotent stem cells.

The pluripotency maintenance of pluripotent stem cells (PSCs) requires the suitable microenvironment, which commonly provided by feeder layers. However, the preparation of feeder layers is time consuming and labor exhaustive, and the feeder cells treated with mitomycin C or γ-ray irradiation bring heterologous contamination. In this study, mouse embryonic fibroblasts (MEFs) were treated by methanol to generate chemical fixed feeder cells, and bovine embryonic stem cells F7 (bESC-F7) cultured on this feeder layer. Then the pluripotency and metabolism of bESC-F7 cultured on methanol-fixed MEFs (MT-MEFs) named MT-F7 was compared with mitomycin C treated MEFs (MC-MEFs). The results showed that bESC-F7 formed alkaline phosphatase positive colonies on MT-MEFs, the relative expression of pluripotent markers of these cells was different from the bESCs cultured on the MC-MEFs (MC-F7). The long-term cultured MT-F7 formed embryoid bodies, showed the ability to differentiate into three germ layers similar to MC-F7. The analyses of RNA-seq data showed that MT-MEFs lead bESCs to novel steady expression patterns of genes regulating pluripotency and metabolism. Furthermore, the bovine expanded pluripotent stem cells (bEPSCs) cultured on MT-MEFs formed classical colonies, maintained pluripotency, and elevated metabolism. In conclusion, MT-MEFs were efficient feeder layer that maintain the distinctive pluripotency and metabolism of PSCs.

Constitutional 763.3 Kb chromosome 1q43 duplication encompassing only CHRM3 gene identified by next generation sequencing (NGS) in a child with intellectual disability.

BACKGROUND: Deletion or duplication on the distal portion of the long arm of chromosome 1 result in complex and highly variable clinical phenotype including.intellectual disability and autism. CASE PRESENTATION: We report on a patient with intellectual disability and a 763.3 Kb duplication on 1q43 that includes only CHRM3, which was detected by next generation sequencing (NGS). The patient presented with intellectual disability, developmental delay, autistic behavior, limited or no speech, social withdrawal, self-injurious, feeding difficulties, strabismus, short stature, hand anomalie, and no seizures, anxiety, or mood swings, and clinodactyly. CONCLUSIONS: We propose that CHRM3 is the critical gene responsible for the common characteristics in the cases with 1q43 duplication and deletion.