Biology subjects

Mohamed, T. M. A.

Publications and source records attributed to Mohamed, T. M. A..

2 recordsLinked to original sources

bioRxiv2020

Heart Slice Culture System Reliably Demonstrates Clinical Drug-Related Cardiotoxicity

The limited availability of human heart tissue and its complex cell composition are major limiting factors for reliable testing drug efficacy, toxicity and understanding mechanism. Recently, we developed a functional human and pig heart slice biomimetic culture system that fully preserves the viability and functionality of 300 {micro}m heart slices for 6 days. Here, we tested the reliability of this culture system in delineating the mechanisms of known anti-cancer drugs that cause cardiomyopathy. We tested three anti-cancer drugs (doxorubicin, trastuzumab, and sunitinib) associated with different mechanisms leading to cardiotoxicity at three concentrations and assessed the effect of these drugs on heart slice viability, structure, function and transcriptome. Slices incubated with any of these drugs for 48 h showed significant loss in viability, cardiomyocyte structure and functionality. Mechanistically, RNA sequencing demonstrated a significant downregulation of cardiac genes and upregulation of oxidative response in doxorubicin-treated tissues. Trastuzumab treatment caused major downregulation in cardiac muscle contraction-related genes, consistent with its clinically known direct effect on cardiomyocytes. Interestingly, sunitinib treatment resulted in significant downregulation of angiogenesis-related genes in line with its mechanism of action. Heart slices are not only able to demonstrate the expected toxicity of doxorubicin and trastuzumab similar to hiPS-derived-cardiomyocytes; they are superior in detecting sunitinib cardiotoxicity phenotypes and mechanism in the clinically relevant concentration range, 100 nM - 1 {micro}M. These results indicate that heart slice tissue culture models have the potential to become a reliable platform for testing drug toxicity and mechanism of action.

cell biology↗

bioRxiv2019

Unique Transcription Factor Functions Regulate Epigenetic and Transcriptional Dynamics During Cardiac Reprogramming

Direct lineage conversion, whereby a somatic cell assumes a new cellular identity, can be driven by ectopic expression of combinations of lineage-enriched transcription factors. To determine the molecular mechanisms by which expression of Gata4, Mef2c, and Tbx5 (GMT) induces direct reprogramming from a cardiac fibroblast toward an induced cardiomyocyte, we performed a comprehensive transcriptomic and epigenomic interrogation of the reprogramming process. Single cell RNA sequencing indicated that a reprogramming trajectory was acquired within 48 hours of GMT introduction, did not require cell division, and was limited mainly by successful expression of GMT. Evaluation of chromatin accessibility by ATAC-seq supported the expression dynamics and revealed widespread chromatin remodeling at early stages of the reprogramming process. Chromatin immunoprecipitation followed by sequencing of each factor alone or in combinations revealed that GMT bind DNA individually and in combination, and that ectopic expression of either Mef2c or Tbx5 is sufficient in some contexts to increase accessibility. We also find evidence for cooperative facilitation and refinement of each factors binding in a combinatorial setting. A random-forest classifier that integrated the observed gene expression dynamics with regions of dynamic chromatin accessibility suggested Tbx5 binding is a primary driver of gene expression changes and revealed additional transcription factor motifs co-segregating with reprogramming factor motifs, suggesting new factors that may be involved in the reprogramming process. These results begin to explain the mechanisms by which transcription factors normally expressed in multiple germ layers can function combinatorially to direct lineage conversion.

molecular biology↗