Biology subjects

Massaad, C.

Publications and source records attributed to Massaad, C..

4 recordsLinked to original sources

bioRxiv2025

Role of oligodendroglial ADAM10 in oligodendrocyte maturation, myelination and myelin maintenance

In the central nervous system, oligodendrocytes extend their processes to wrap axons to form myelin sheaths, ensuring saltatory conduction of action potentials and providing trophic support for the axon. Our previous work demonstrated that pharmacological activation of -secretase promotes myelin protection and remyelination after both ex vivo and in vivo demyelination. Based on these findings, the present study aims to investigate the role of oligodendroglial ADAM10 (OLA10; a member of the a-secretase family) in myelin development and maintenance. Using an inducible knockout mouse model for OLA10 (KOOL-A10), we demonstrated that OLA10 deficiency results in delayed maturation of OPC in primary culture, and altered myelination in primary neuron/glia co-culture. Furthermore, an induction of OLA10 deficiency in adult mice was associated with an alteration of myelin sheath thickness and long-term motor and cognitive deficit in males but not in females. Overall, our study highlights the subtle but significant role of ADAM10 expressed by oligodendrocytes in the formation and maintenance of the myelin over time, while emphasizing a sexual dimorphism that carries important relevance for current research.

neuroscience↗

bioRxiv2021

RNA-Seq is not required to determine stable reference genes for qPCR normalization

Assessment of differential gene expression by qPCR is heavily influenced by the choice of reference genes. Although numerous statistical approaches have been proposed to determine the best reference genes, they can give rise to conflicting results depending on experimental conditions. Hence, recent studies propose the use of RNA-Seq to identify stable genes followed by the application of different statistical approaches to determine the best set of reference genes for qPCR data normalization. In this study, we demonstrate that the statistical approach to determine the best reference genes from randomly selected candidates is more important than the preselection of stable candidates from RNA-Seq data. Using a qPCR data normalization workflow that we have previously established; we show that qPCR data normalization using randomly chosen conventional reference genes renders the same results as stable reference genes selected from RNA-Seq data. We validated these observations in two distinct cross-sectional experimental conditions involving human iPSC derived microglial cells and mouse sciatic nerves. These results taken together show that given a robust statistical approach for reference gene selection, stable genes selected from RNA-Seq data do not offer any significant advantage over commonly used reference genes for normalizing qPCR assays.

molecular biology↗

bioRxiv2020

Retracing Schwann cell developmental transitions in embryonic dissociated DRG cultures

Embryonic Dissociated Dorsal Root Ganglia cultures are often used to investigate the role of novel molecular pathways or drugs in Schwann cell development and myelination. These cultures largely recapitulate the order of cellular and molecular events that occur in Schwann cells of embryonic nerves. However, the timing of Schwann cell developmental transitions, notably the transition from Schwann Cell Precursors to immature Schwann cells and then to myelinating Schwann cells, has not been estimated so far in this culture system. In this study, we determined the expression profiles of Schwann cell developmental genes during the first week of culture and then compared our data to the expression profiles of these genes in developing spinal nerves. This helped in identifying that Schwann Cell Precursors transition into immature Schwann Cells between the 5th and 7th day in vitro. Furthermore, we also investigated the transition of immature cells into pro-myelinating and myelinating Schwann cells upon the induction of myelination in vitro. Our results suggest that Schwann cell differentiation beyond the immature stage can be observed as early as 4 days post the induction of myelination in cocultures. Finally, we compared the myelinating potential of coculture-derived Schwann cell monocultures to cultures established from neonatal sciatic nerves and found that both these cultures system exhibit similar myelinating phenotypes. In effect, our results allow for a better understanding and interpretation of coculture experiments especially in studies that aim to elucidate the role of a novel actor in Schwann Cell development and myelination.

developmental biology↗

bioRxiv2019

Optimal use of statistical methods to validate reference gene stability in longitudinal studies

Multiple statistical approaches have been proposed to validate reference genes in qPCR assays. However, conflicting results from these statistical methods pose a major hurdle in the choice of the best reference genes. Indeed, as their respective approaches to calculating reference gene stability is different, their suitability has to be tested for a given experimental setting. In this study, the stability of 10 candidate reference genes (Actb, Gapdh, Tbp, Sdha, Pgk1, Ppia, Rpl13a, Hsp60, Mrpl10, Rps26) was assessed using four common statistical approaches (GeNorm, NormFinder, Coefficient of Variation analysis and Pairwise {Delta}Ct method) in a longitudinal setting. We used the development of the cerebellum and the spinal cord of mice as a model to assess the suitability of these statistical methods for reference gene validation. GeNorm and the Pairwise {Delta}Ct were found to be ill suited due to a fundamental assumption in their stability calculations. Whereas, NormFinder and Coefficient of Variation analysis fare better provided they are used complementarily. We therefore devised a workflow combining these two methods for validating reference genes in developmental studies. This workflow proves to be more robust than any of the methods used individually.

molecular biology↗