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1.
Cells Tissues Organs ; 209(4-6): 165-176, 2020.
Article in English | MEDLINE | ID: mdl-33423027

ABSTRACT

In the past, the heart muscle was thought to originate from a single source of myocardial progenitor cells. More recently, however, an additional source of myocardial progenitors has been revealed to be the second heart field, and chicken embryos were important for establishing this concept. However, there have been few studies in chicken on how this field contributes to heart muscles in vitro. We have developed an ex vivo experimental system from chicken embryos between stages HH17-20 to investigate how mesodermal progenitors in the second branchial arch (BA2) differentiate into cardiac muscles. Using this method, we presented evidence that the progenitor cells within the BA2 arch differentiated into beating cardiomyocytes in vitro. The beating explant cells were positive for cardiac actin, Nkx2.5, and ventricular myosin heavy chain. In addition, we performed a time course for the expression of second heart field markers (Isl1 and Nkx2.5) in the BA2 from stage HH16 to stage HH21 using in situ hybridization. Accordingly, using EGFP-based cell labeling techniques and quail-chicken cell injection, we demonstrated that mesodermal cells from the BA2 contributed to the outflow tract and ventricular myocardium in vivo. Thus, our findings highlight the cardiogenic potential of chicken BA2 mesodermal cells in vitro and in vivo.


Subject(s)
Branchial Region , Chickens , Myocardium , Animals , Cell Differentiation , Chick Embryo , Heart , Myocytes, Cardiac , Stem Cells
2.
Eur J Cell Biol ; 94(2): 101-13, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25555464

ABSTRACT

The formin homology domain-containing protein1 (FHOD1) suppresses actin polymerization by inhibiting nucleation, but bundles actin filaments and caps filament barbed ends. Two polyclonal antibodies against FHOD1 were generated against (i) its N-terminal sequence (residues 1-339) and (ii) a peptide corresponding the sequence from position 358-371, which is unique for FHOD1 and does not occur in its close relative FHOD3. After affinity purification both antibodies specifically stain purified full length FHOD1 and a band of similar molecular mass in homogenates of cardiac muscle. The antibody against the N-terminus of FHOD1 was used for immunostaining cells of established lines, primary neonatal (NRC) and adult (ARC) rat cardiomyocytes and demonstrated the presence of FHOD1 in HeLa and fibroblastic cells along stress fibers and within presumed lamellipodia and actin arcs. In NRCs and ARCs we observed a prominent staining of presumed intercalated discs (ICD). Immunostaining of sections of hearts with both anti-FHOD1 antibodies confirmed the presence of FHOD1 in ICDs and double immunostaining demonstrated its colocalisation with cadherin, plakoglobin and a probably slightly shifted localization to connexin43. Similarly, immunostaining of isolated mouse or pig ICDs corroborated the presence of FHOD1 and its colocalisation with the mentioned cell junctional components. Anti-FHOD1 immunoblots of isolated ICDs demonstrated the presence of an immunoreactive band comigrating with purified FHOD1. Conversely, an anti-peptide antibody specific for FHOD3 with no cross-reactivity against FHOD1 immunostained on sections of cardiac muscle and ARCs the myofibrils in a cross-striated pattern but not the ICDs. In addition, the anti-peptide-FHOD1 antibody stained the lateral sarcolemma of ARCs in a banded pattern. Double immunostaining with anti-cadherin and -integrin-ß1 indicated the additional localization of FHOD1 in costameres. Immunostaining of cardiac muscle sections or ARCs with antibodies against mDia3-FH2-domain showed colocalisation with cadherin along the lateral border of cardiomyocytes suggesting also its presence in costameres.


Subject(s)
Costameres/metabolism , Fetal Proteins/metabolism , Myocardium/metabolism , Nuclear Proteins/metabolism , Animals , Animals, Newborn , Antibodies/metabolism , Cadherins/metabolism , Cell Line , Connexin 43/metabolism , Formins , Humans , Mice , Microfilament Proteins/metabolism , Myocardium/cytology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/ultrastructure , Rats , Stress Fibers/metabolism , Swine , gamma Catenin/metabolism
3.
Cytoskeleton (Hoboken) ; 71(2): 95-107, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24382810

ABSTRACT

F-actin treadmilling plays a key part in cell locomotion. Because immunofluorescence showed colocalisation of thymosin beta4 (Tß4) with cofilin-1 and Arp2/3 complex in lamellipodia, we analyzed combinations of these proteins on F-actin-adenosine triphosphate (ATP)-hydrolysis, which provides a measure of actin treadmilling. Actin depolymerising factor (ADF)/cofilin stimulated treadmilling, while Tß4 decreased treadmilling, presumably by sequestering monomers. Tß4 added together with ADF/cofilin also inhibited the treadmilling, relative to cofilin alone, but both the rate and extent of depolymerization were markedly enhanced in the presence of both these proteins. Arp2/3 complex reversed the sequestering activity of Tß4 when equimolar to actin, but not in the additional presence of cofilin-1 or ADF. Transfection experiments to explore the effects of changing the intracellular concentration of Tß4 in HeLa cells showed that an increase in Tß4 resulted in reduced actin filaments bundles and narrower lamellipodia, and a conspicuous decrease of cell migration as seen by two different assays. In contrast, cells transfected with a vector leading to Tß4 knockdown by small interfering RNA (siRNA) displayed prominent actin filament networks within the lamellipodia and the leading lamella and enhanced migration. The experiments reported here demonstrate the importance of the interplay of these different classes of actin-binding proteins on cell behaviour.


Subject(s)
Actins/metabolism , Destrin/metabolism , Thymosin/metabolism , Actin Depolymerizing Factors/metabolism , Cell Movement/physiology , HeLa Cells , Humans
4.
Ann Anat ; 193(5): 453-8, 2011 Oct 20.
Article in English | MEDLINE | ID: mdl-21684729

ABSTRACT

Hyperostosis frontalis interna is a morphological pattern characterized by single or multiple bony nodules situated on the inner lamina of the frontal bone. It is seldom found in males, but it is a common phenomenon among post-menopausal females in modern societies but relatively rare in antiquity. The etiopathogenesis of the trait is a matter of debate and ranges from genetic predisposition to epigenetic, while endocrine disturbances, aging, and dietary factors are also listed among the causes. We studied the frequency, characteristic features, and etiopathogenesis of the disease in recent cadaveric and dry skull specimens. The frequency of hyperostosis frontalis interna in cadavers and dry skull materials was almost identical, 12.5% and 12.3%, respectively. In cadavers, 87.5% of severe hyperostosis frontalis interna cases were found in females over 65 years-old. Interestingly, in two cadavers we found hyperostotic lesions spreading onto adjacent tissues such as the dura and falx cerebri. We provide some new aspects that may help in better understanding of the etiopathogenesis of hyperostosis frontalis interna. Thereby, we discuss the various etiopathogenesis models found in the literature.


Subject(s)
Hyperostosis Frontalis Interna/pathology , Adolescent , Adult , Aged , Female , Frontal Bone/pathology , Humans , Hyperostosis Frontalis Interna/epidemiology , Male , Middle Aged , Prevalence , Young Adult
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