Expression and localization of nuclear proteins in autosomal-dominant Emery-Dreifuss muscular dystrophy with LMNA R377H mutation

BACKGROUND: The autosomal dominant form of Emery-Dreifuss muscular dystrophy (AD-EDMD) is caused by mutations in the gene encoding for the lamins A and C (LMNA). Lamins are intermediate filament proteins which form the nuclear lamina underlying the inner nuclear membrane. We have studied the expression and the localization of nuclear envelope proteins in three different cell types and muscle tissue of an AD-EDMD patient carrying a point mutation R377H in the lamin A/C gene. RESULTS: Lymphoblastoid cells, skin fibroblasts, primary myoblasts and muscle thin sections were studied by immunocytochemistry and electron microscopy. Cellular levels of A-type lamins were reduced compared to control cells. In contrast, the amount of emerin and lamin B appeared unaltered. Cell synchronization experiments showed that the reduction of the cellular level of A-type lamin was due to instability of lamin A. By electron microscopy, we identified a proportion of nuclei with morphological alterations in lymphoblastoid cells, fibroblasts and mature muscle fibres. Immunofluorescence microscopy showed that a major population of the lamin B receptor (LBR), an inner nuclear membrane protein, was recovered in the cytoplasm in association with the ER. In addition, the intranuclear organization of the active form of RNA polymerase II was markedly different in cells of this AD-EDMD patient. This aberrant intranuclear distribution was specifically observed in muscle cells where the pathology of EDMD predominates. CONCLUSIONS: From our results we conclude: Firstly, that structural alterations of the nuclei which are found only in a minor fraction of lymphoblastoid cells and mature muscle fibres are not sufficient to explain the clinical pathology of EDMD; Secondly, that wild type lamin A is required not only for the retention of LBR in the inner nuclear membrane but also for a correct localization of the transcriptionally active RNA pol II in muscle cells. We speculate that a rearrangement of the internal chromatin could lead to muscle-specific disease symptoms by interference with proper mRNA transcription

Sexual differentiation and primordial germ cell distribution in the early horse fetus

It was the aim of this study to characterize the development of the gonads and genital ducts in the equine fetus around the time of sexual differentiation. This included the identification and localization of the primordial germ cell population. Equine fetuses between 45 and 60 days of gestation were evaluated using a combination of micro-computed tomography scanning, immunohistochemistry, and multiplex immunofluorescence. Fetal gonads increased in size 23-fold from 45 to 60 days of gestation, and an even greater increase was observed in the metanephros volume. Signs of mesonephros atrophy were detected during this time. Tubular structures of the fetal testes were present from day 50 onwards, whereas cell clusters dominated in the fetal ovary. The genital ducts were well-differentiated and presented a lumen in all samples. No sign of mesonephric or paramesonephric duct degeneration was detected. Expression of AMH was strong in the fetal testes but absent in ovaries. Irrespective of sex, primordial germ cells selectively expressed LIN28. Migration of primordial germ cells from the mesonephros to the gonad was detected at 45 days, but not at 60 days of development. Their number and distribution within the gonad were influenced (p < 0.05) by fetal sex. Most primordial germ cells (86.8 ± 3.2% in females and 84.6 ± 4.7% in males) were characterized as pluripotent according to co-localization with CD117. However, only a very small percentage of primordial germ cells were proliferating (7.5 ± 1.7% in females and 3.2 ± 1.2% in males) based on co-localization with Ki67. It can be concluded that gonadal sexual differentiation in the horse occurs asynchronously with regard to sex but already before 45 days of gestation.Fil: Scarlet, Dragos. University of Veterinary Medicine Vienna; Suiza. Universitat Zurich; SuizaFil: Handschuh, Stephan. University of Veterinary Medicine Vienna; SuizaFil: Reichart, Ursula. University of Veterinary Medicine Vienna; SuizaFil: Podico, Giorgia. University of Illinois. Urbana - Champaign; Estados UnidosFil: Ellerbrock, Robyn E.. University of Illinois. Urbana - Champaign; Estados UnidosFil: Demyda Peyrás, Sebastián. Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; ArgentinaFil: Canisso, Igor F.. University of Illinois. Urbana - Champaign; Estados UnidosFil: Walter, Ingrid. University of Veterinary Medicine Vienna; SuizaFil: Aurich, Christine. University of Veterinary Medicine Vienna; Suiz

Current Status of Cardiac Xenotransplantation: Report of a Workshop of the German Heart Transplant Centers, Martinsried, March 3, 2023

This report comprises the contents of the presentations and following discussions of a workshop of the German Heart Transplant Centers in Martinsried, Germany on cardiac xenotransplantation. The production and current availability of genetically modified donor pigs, preservation techniques during organ harvesting, and immunosuppressive regimens in the recipient are described. Selection criteria for suitable patients and possible solutions to the problem of overgrowth of the xenotransplant are discussed. Obviously microbiological safety for the recipient and close contacts is essential, and ethical considerations to gain public acceptance for clinical applications are addressed. The first clinical trial will be regulated and supervised by the Paul-Ehrlich-Institute as the National Competent Authority for Germany, and the German Heart Transplant Centers agreed to cooperatively select the first patients for cardiac xenotransplantation

Temperature calibration of Mg/Ca ratios in the intermediate water benthic foraminifer Hyalinea balthica

Author Posting. © American Geophysical Union, 2011. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geochemistry Geophysics Geosystems 12 (2011): Q04003, doi:10.1029/2010GC003333.Core top samples from Indonesian and northeast Atlantic depth transects were used to calibrate Mg/Ca and δ18O in tests of the calcitic benthic foraminifer Hyalinea balthica to bottom water temperature between 4°C and 13°C. This shallow infaunal species is primarily abundant in neritic to upper bathyal sediments (<600 m). Both linear and exponential calibrations suggest a temperature sensitivity of ~12% per °C that is ~4 times higher than observed in other species of deep-sea benthic foraminifera. Culture experiments support the core top calibration. We find no discernible effect of salinity and saturation on Mg/Ca. Comparison between the measured benthic foraminiferal δ18O and predicted equilibrium values suggests that on average H. balthica δ18O is 0.64‰ ± 0.13‰ lower than predicted from the equilibrium composition. To test the reliability of using paired H. balthica Mg/Ca and δ18O measurements for reconstructing seawater δ18Osw and salinity, we apply this calibration to another depth transect from Cape Ghir off NW Africa, which was not included in the calibration. Based on error analysis of the calibration data and this validation test, we show that the uncertainty of reconstructing bottom water temperature and salinity from paired Mg/Ca and δ18O measurements of H. balthica is better than ±0.7°C and ±0.69 practical salinity scale, respectively. The small uncertainties allow for the reconstruction of seawater density to better than 0.3σθ units, which is precise enough for the identification of specific water masses and reconstruction of changes in their properties. We propose that the relatively high Mg content and temperature sensitivity of H. balthica might be due to minor, biologically mediated contribution of high-Mg calcite to the primarily low Mg calcite test, which is influenced by the ambient temperature. This hypothesis, if correct, suggests that benthic species with relatively high Mg/Ca may be better suited for deepwater temperature reconstructions than species that have thus far been more commonly used.This project was funded by NSF Awards OCE 02‐20922 and 09‐02977 to YR, OCE 09‐28607 to MK, OCE02‐20776 to DWO, and DFG priority program INTERDYNAMIK to AM

Conditional Stat1 Ablation Reveals the Importance of Interferon Signaling for Immunity to Listeria monocytogenes Infection

Signal transducer and activator of transcription 1 (Stat1) is a key player in responses to interferons (IFN). Mutations of Stat1 cause severe immune deficiencies in humans and mice. Here we investigate the importance of Stat1 signaling for the innate and secondary immune response to the intracellular bacterial pathogen Listeria monocytogenes (Lm). Cell type-restricted ablation of the Stat1 gene in naïve animals revealed unique roles in three cell types: macrophage Stat1 signaling protected against lethal Lm infection, whereas Stat1 ablation in dendritic cells (DC) did not affect survival. T lymphocyte Stat1 reduced survival. Type I IFN (IFN-I) signaling in T lymphocytes reportedly weakens innate resistance to Lm. Surprisingly, the effect of Stat1 signaling was much more pronounced, indicating a contribution of Stat1 to pathways other than the IFN-I pathway. In stark contrast, Stat1 activity in both DC and T cells contributed positively to secondary immune responses against Lm in immunized animals, while macrophage Stat1 was dispensable. Our findings provide the first genetic evidence that Stat1 signaling in different cell types produces antagonistic effects on innate protection against Lm that are obscured in mice with complete Stat1 deficiency. They further demonstrate a drastic change in the cell type-dependent Stat1 requirement for memory responses to Lm infection

Mechanisms Establishing TLR4-Responsive Activation States of Inflammatory Response Genes

Precise control of the innate immune response is required for resistance to microbial infections and maintenance of normal tissue homeostasis. Because this response involves coordinate regulation of hundreds of genes, it provides a powerful biological system to elucidate the molecular strategies that underlie signal- and time-dependent transitions of gene expression. Comprehensive genome-wide analysis of the epigenetic and transcription status of the TLR4-induced transcriptional program in macrophages suggests that Toll-like receptor 4 (TLR4)-dependent activation of nearly all immediate/early- (I/E) and late-response genes results from a sequential process in which signal-independent factors initially establish basal levels of gene expression that are then amplified by signal-dependent transcription factors. Promoters of I/E genes are distinguished from those of late genes by encoding a distinct set of signal-dependent transcription factor elements, including TATA boxes, which lead to preferential binding of TBP and basal enrichment for RNA polymerase II immediately downstream of transcriptional start sites. Global nuclear run-on (GRO) sequencing and total RNA sequencing further indicates that TLR4 signaling markedly increases the overall rates of both transcriptional initiation and the efficiency of transcriptional elongation of nearly all I/E genes, while RNA splicing is largely unaffected. Collectively, these findings reveal broadly utilized mechanisms underlying temporally distinct patterns of TLR4-dependent gene activation required for homeostasis and effective immune responses

Cells of the adult human heart

Abstract: Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and therapeutic strategies require a deeper understanding of the molecular processes involved in the healthy heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavour. Here, using state-of-the-art analyses of large-scale single-cell and single-nucleus transcriptomes, we characterize six anatomical adult heart regions. Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, and reveal distinct atrial and ventricular subsets of cells with diverse developmental origins and specialized properties. We define the complexity of the cardiac vasculature and its changes along the arterio-venous axis. In the immune compartment, we identify cardiac-resident macrophages with inflammatory and protective transcriptional signatures. Furthermore, analyses of cell-to-cell interactions highlight different networks of macrophages, fibroblasts and cardiomyocytes between atria and ventricles that are distinct from those of skeletal muscle. Our human cardiac cell atlas improves our understanding of the human heart and provides a valuable reference for future studies

Cells of the adult human heart

Abstract: Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and therapeutic strategies require a deeper understanding of the molecular processes involved in the healthy heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavour. Here, using state-of-the-art analyses of large-scale single-cell and single-nucleus transcriptomes, we characterize six anatomical adult heart regions. Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, and reveal distinct atrial and ventricular subsets of cells with diverse developmental origins and specialized properties. We define the complexity of the cardiac vasculature and its changes along the arterio-venous axis. In the immune compartment, we identify cardiac-resident macrophages with inflammatory and protective transcriptional signatures. Furthermore, analyses of cell-to-cell interactions highlight different networks of macrophages, fibroblasts and cardiomyocytes between atria and ventricles that are distinct from those of skeletal muscle. Our human cardiac cell atlas improves our understanding of the human heart and provides a valuable reference for future studies