Yacoub, M. H., A. Afifi, H. Hosny, ahmed mahgoub, M. Nagy, A. Elsawy, NairoShehata, A. M. Ibrahim, M. Azer, A. Elafifi, et al., "Structure and function of the aortic and pulmonary outflows in a patient, 12 years after Nikaidoh operation, A cautionary Note", International Journal of Cardiology Congenital Heart Disease, vol. 12, 2023. Abstract

Background
Nikaidoh translocation operation is increasingly being used for patients with TGA, VSD and LVOTO. The early results of this operation are excellent and possibly better than the Rastelli repair. However, the long-term results remain inadequately defined.
Methods and results
Detailed follow up data of a patient, 12 years after Nikaidoh operation, using computerized image analysis are reported. The patient complained of chest pain on exertion. Imaging showed severe dilatation of the non-coronary sinus of the aortic root, compressing the RVOT and pulmonary artery branches resulting in RV dilatation. The dilated aortic sinus showed marked pulsatility with expansion during systole and diastole. As the child was symptomatic, with a risk of aneurysm rupture, excision was performed. This complication is thought to be due to interruption of the innervation of the root as well as cutting vasa vasorum.
Conclusion
Nikaidoh operation is a viable solution for the repair of TGA, VSD, PS. Dilatation of the neo-aortic root should be carefully followed and causes of the dilatation investigated.

WalyEldeen, A. A., S. Sabet, S. E. Anis, T. Stein, and A. M. Ibrahim, "FBLN2 is associated with basal cell markers Krt14 and ITGB1 in mouse mammary epithelial cells and has a preferential expression in molecular subtypes of human breast cancer.", Breast cancer research and treatment, vol. 208, issue 3, pp. 673-686, 2024. Abstract

BACKGROUND: Fibulin-2 (FBLN2) is a secreted extracellular matrix (ECM) glycoprotein and has been identified in the mouse mammary gland, in cap cells of terminal end buds (TEBs) during puberty, and around myoepithelial cells during early pregnancy. It is required for basement membrane (BM) integrity in mammary epithelium, and its loss has been associated with human breast cancer invasion. Herein, we attempted to confirm the relevance of FBLN2 to myoepithelial phenotype in mammary epithelium and to assess its expression in molecular subtypes of human breast cancer.

METHODS: The relationship between FBLN2 expression and epithelial markers was investigated in pubertal mouse mammary glands and the EpH4 mouse mammary epithelial cell line using immunohistochemistry, immunocytochemistry, and immunoblotting. Human breast cancer mRNA data from the METABRIC and TCGA datasets from Bioportal were analyzed to assess the association of Fbln2 expression with epithelial markers, and with molecular subtypes. Survival curves were generated using data from the METABRIC dataset and the KM databases.

RESULTS: FBLN2 knockdown in mouse mammary epithelial cells was associated with a reduction in KRT14 and an increase in KRT18. Further, TGFβ3 treatment resulted in the upregulation of FBLN2 in vitro. Meta-analyses of human breast cancer datasets from Bioportal showed a higher expression of Fbln2 mRNA in claudin-low, LumA, and normal-like breast cancers compared to LumB, Her2 +, and Basal-like subgroups. Fbln2 mRNA levels were positively associated with mesenchymal markers, myoepithelial markers, and markers of epithelial-mesenchymal transition. Higher expression of Fbln2 mRNA was associated with better prognosis in less advanced breast cancer and this pattern was reversed in more advanced lesions.

CONCLUSION: With further validation, these observations may offer a molecular prognostic tool for human breast cancer for more personalized therapeutic approaches.

Ahmad, F. S., Y. Jin, A. Grassam-Rowe, Y. Zhou, M. Yuan, X. Fan, R. Zhou, R. Mu-U-Min, C. O'Shea, A. M. Ibrahim, et al., "Generation of cardiomyocytes from human-induced pluripotent stem cells resembling atrial cells with ability to respond to adrenoceptor agonists.", Philosophical transactions of the Royal Society of London. Series B, Biological sciences, vol. 378, issue 1879, pp. 20220312, 2023. Abstract

Atrial fibrillation (AF) is the most common chronic arrhythmia presenting a heavy disease burden. We report a new approach for generating cardiomyocytes (CMs) resembling atrial cells from human-induced pluripotent stem cells (hiPSCs) using a combination of Gremlin 2 and retinoic acid treatment. More than 40% of myocytes showed rod-shaped morphology, expression of CM proteins (including ryanodine receptor 2, -actinin-2 and F-actin) and striated appearance, all of which were broadly similar to the characteristics of adult atrial myocytes (AMs). Isolated myocytes were electrically quiescent until stimulated to fire action potentials with an AM profile and an amplitude of approximately 100 mV, arising from a resting potential of approximately -70 mV. Single-cell RNA sequence analysis showed a high level of expression of several atrial-specific transcripts including , , , , , and . Amplitudes of calcium transients recorded from spontaneously beating cultures were increased by the stimulation of -adrenoceptors (activated by phenylephrine and blocked by prazosin) or -adrenoceptors (activated by isoproterenol and blocked by CGP20712A). Our new approach provides human AMs with mature characteristics from hiPSCs which will facilitate drug discovery by enabling the study of human atrial cell signalling pathways and AF. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Medali, T., D. Couchie, N. Mougenot, M. Mihoc, O. Bergmann, W. Derks, L. I. Szweda, M. Yacoub, S. Soliman, Y. Aguib, et al., "Thioredoxin-1 and its mimetic peptide improve systolic cardiac function and remodeling after myocardial infarction.", FASEB journal : official publication of the Federation of American Societies for Experimental Biology, vol. 38, issue 1, pp. e23291, 2024. Abstract

Myocardial infarction (MI) is characterized by a significant loss of cardiomyocytes (CMs), and it is suggested that reactive oxygen species (ROS) are involved in cell cycle arrest, leading to impaired CM renewal. Thioredoxin-1 (Trx-1) scavenges ROS and may play a role in restoring CM renewal. However, the truncated form of Trx-1, Trx-80, can compromise its efficacy by exerting antagonistic effects. Therefore, a Trx-1 mimetic peptide called CB3 was tested as an alternative way to restore CMs. This study aimed to investigate the effects of Trx-1, Trx-80, and CB3 on mice with experimental MI and study the underlying mechanism of CB3 on CMs. Mouse cardiac parameters were quantified by echocardiography, and infarction size and fibrosis determined using Trichrome and Picro-Sirius Red staining. The study found that Trx-1 and CB3 improved mouse cardiac function, reduced the size of cardiac infarct and fibrosis, and decreased the expression of cardiac inflammatory markers. Furthermore, CB3 polarized macrophages into M2 phenotype, reduced apoptosis and oxidative stress after MI, and increased CM proliferation in cell culture and in vivo. CB3 effectively protected against myocardial infarction and could represent a new class of compounds for treating MI.

Halawa, S., N. Latif, Y. - T. Tseng, A. M. Ibrahim, A. H. Chester, A. Moustafa, Y. Aguib, and M. H. Yacoub, "Profiling Genome-Wide DNA Methylation Patterns in Human Aortic and Mitral Valves.", Frontiers in cardiovascular medicine, vol. 9, pp. 840647, 2022. Abstract

Cardiac valves exhibit highly complex structures and specialized functions that include dynamic interactions between cells, extracellular matrix (ECM) and their hemodynamic environment. Valvular gene expression is tightly regulated by a variety of mechanisms including epigenetic factors such as histone modifications, RNA-based mechanisms and DNA methylation. To date, methylation fingerprints of non-diseased human aortic and mitral valves have not been studied. In this work we analyzed the differential methylation profiles of 12 non-diseased aortic and mitral valve tissue samples (in matched pairs). Analysis of methylation data [reduced representation bisulfite sequencing (RRBS)] of 16,101 promoters genome-wide revealed 584 differentially methylated (DM) promoters, of which 13 were reported in endothelial mesenchymal trans-differentiation (EMT), 37 in aortic and mitral valve disease and 7 in ECM remodeling. Both functional classification as well as network analysis showed that the genes associated with the DM promoters were enriched for WNT-, Cadherin-, Endothelin-, PDGF-, HIF-1 and VEGF- signaling implicated in valvular physiology and pathophysiology. Additional enrichment was detected for TGFB-, NOTCH- and Integrin- signaling involved in EMT as well as ECM remodeling. This data provides the first insight into differential regulation of human aortic and mitral valve tissue and identifies candidate genes linked to DM promoters. Our work will improve the understanding of valve biology, valve tissue engineering approaches and contributes to the identification of relevant drug targets.

Ibrahim, A. M., A. Bilsland, S. Rickelt, J. S. Morris, and T. Stein, "A matrisome RNA signature from early-pregnancy mouse mammary fibroblasts predicts distant metastasis-free breast cancer survival in humans.", Breast cancer research : BCR, vol. 23, issue 1, pp. 90, 2021. Abstract

BACKGROUND: During pregnancy, the mouse mammary ductal epithelium branches and grows into the surrounding stroma, requiring extensive extracellular matrix (ECM) and tissue remodelling. It therefore shows parallels to cancer invasion. We hypothesised that similar molecular mechanisms may be utilised in both processes, and that assessment of the stromal changes during pregnancy-associated branching may depict the stromal involvement during human breast cancer progression.

METHODS: Immunohistochemistry (IHC) was employed to assess the alterations within the mouse mammary gland extracellular matrix during early pregnancy when lateral branching of the primary ductal epithelium is initiated. Primary mouse mammary fibroblasts from three-day pregnant and age-matched non-pregnant control mice, respectively, were 3D co-cultured with mammary epithelial cells to assess differences in their abilities to induce branching morphogenesis in vitro. Transcriptome analysis was performed to identify the underlying molecular changes. A signature of the human orthologues of the differentially expressed matrisome RNAs was analysed by Kaplan-Meier and multi-variate analysis in two large breast cancer RNA datasets (Gene expression-based Outcome for Breast cancer Online (GOBO) und Kaplan-Meier Plotter), respectively, to test for similarities in expression between early-pregnancy mouse mammary gland development and breast cancer progression.

RESULTS: The ECM surrounding the primary ductal network showed significant differences in collagen and basement membrane protein distribution early during pregnancy. Pregnancy-associated fibroblasts (PAFs) significantly enhanced branching initiation compared to age-matched control fibroblast. A combined signature of 64 differentially expressed RNAs, encoding matrisome proteins, was a strong prognostic indicator of distant metastasis-free survival (DMFS) independent of other clinical parameters. The prognostic power could be significantly strengthened by using only a subset of 18 RNAs (LogRank P ≤ 1.00e-13; Hazard ratio (HR) = 2.42 (1.8-3.26); p = 5.61e-09). The prognostic power was confirmed in a second breast cancer dataset, as well as in datasets from ovarian and lung cancer patients.

CONCLUSIONS: Our results describe for the first time the early stromal changes that accompany pregnancy-associated branching morphogenesis in mice, specify the early pregnancy-associated molecular alterations in mouse mammary fibroblasts, and identify a matrisome signature as a strong prognostic indicator of human breast cancer progression, with particular strength in oestrogen receptor (ER)-negative breast cancers.

Aguib, Y., M. Allouba, R. Walsh, A. M. Ibrahim, S. Halawa, A. Afify, M. Hosny, P. I. Theotokis, A. Galal, S. Elshorbagy, et al., "New Variant With a Previously Unrecognized Mechanism of Pathogenicity in Hypertrophic Cardiomyopathy.", Circulation, vol. 144, issue 9, pp. 754-757, 2021.
Eissa, M. I., M. A. El-Sherbiny, A. M. Ibrahim, A. Abdelsadik, M. M. Mohamed, and M. S. El-Halawany, "Biochemical and Histopathological studies on female and male Wistar rats fed on genetically modified soybean meals (Roundup Ready)", The Journal of Basic and Applied Zoology, 2019.
Al Kindi, H. N., A. M. Ibrahim, M. Roshdy, B. S. Abdelghany, D. Yehia, A. N. Masoud, W. Simry, Y. Aguib, and M. H. Yacoub, "Clinical, cellular, and molecular characterisation of cardiac rhabdomyoma in tuberous sclerosis.", Cardiology in the young, pp. 1-9, 2021. Abstract

BACKGROUND: Rhabdomyoma is the most common cardiac tumour in children. It is usually associated with tuberous sclerosis complex caused by mutations in TSC-1 or TSC-2 genes. This tumour typically regresses by unknown mechanisms; however, it may cause inflow or outflow obstruction that necessitates urgent surgery. Here we investigate the clinical features and the genetic analysis of patients with tuberous sclerosis complex presenting with large rhabdomyoma tumours. We also investigate the potential role of autophagy and apoptosis in the pathogenesis of this tumour.

METHODS: All the patients with cardiac rhabdomyoma referred to Aswan Heart Centre from 2010 to 2018 were included in this study. Sanger sequencing was performed for coding exons and the flanking intronic regions of TSC1 and TSC2 genes. Histopathological evaluation, immunohistochemistry, and western blotting were performed with P62, LC3b, caspase3, and caspase7, to evaluate autophagic and apoptotic signaling.

RESULTS: Five patients were included and had the clinical features of tuberous sclerosis complex. Three patients, who were having obstructive tumours, were found to have pathogenic mutations in TSC-2. The expression of two autophagic markers, P62 and LC3b, and two apoptotic markers, caspase3 and caspase7, were increased in the tumour cells compared to normal surrounding myocardial tissue.

CONCLUSION: All the patients with rhabdomyoma were diagnosed to have tuberous sclerosis complex. The patients who had pathogenic mutations in the TSC-2 gene had a severe disease form necessitating urgent intervention. We also demonstrate the potential role of autophagy and apoptosis as a possible mechanism for tumourigenesis and regression. Future studies will help in designing personalised treatment for cardiac rhabdomyoma.

Ibrahim, A. M., M. Roshdy, S. Elshorbagy, M. Hosny, S. Halawa, D. Yehia, H. A. Elfawy, A. elDessouki, F. Mohamed, A. Ellithy, et al., "An Investigation of Fibulin-2 in Hypertrophic Cardiomyopathy.", International journal of molecular sciences, vol. 21, issue 19, 2020. Abstract

Hypertrophic cardiomyopathy (HCM) is the most common inherited heart muscle disease, with a prevalence of at least 1 in 500 in the general population. The disease is pleiotropic and is characterized by an increased stiffness of the myocardium, partly due to changes in the extracellular matrix (ECM), with elevated levels of interstitial fibrosis. Myocardial fibrosis is linked to impaired diastolic function and possibly phenotypic heterogeneity of HCM. The ECM consists of a very large number of proteins, which actively interact with each other as well as with myocardial cells. The role of other multiple components of the ECM in HCM has not been defined. Fibulin-2 is a glycoprotein component of the ECM, which plays an important role during embryogenesis of the heart; however, its role in adult myocardium has not been adequately studied. We here describe, for the first time, abnormal expression of fibulin-2 in the myocardium in patients with HCM as compared to normal controls. This abnormal expression was localized in the cytoplasm of myocardial cells and in the interstitial fibroblasts. In addition, fibulin-2 levels, measured by ELISA, were significantly elevated in the serum of patients with HCM as compared to normal controls.

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