scholarly journals Immune cells lacking Y chromosome show dysregulation of autosomal gene expression

2021 ◽  
Author(s):  
Jan P. Dumanski ◽  
Jonatan Halvardson ◽  
Hanna Davies ◽  
Edyta Rychlicka-Buniowska ◽  
Jonas Mattisson ◽  
...  
Keyword(s):  
Single Cells ◽  
Cell Types ◽  
Autosomal Gene ◽  
Specific Cell ◽  
Immune Functions ◽  
Chromosome Y ◽  
Late Genes ◽  
Increased Risk

AbstractEpidemiological investigations show that mosaic loss of chromosome Y (LOY) in leukocytes is associated with earlier mortality and morbidity from many diseases in men. LOY is the most common acquired mutation and is associated with aberrant clonal expansion of cells, yet it remains unclear whether this mosaicism exerts a direct physiological effect. We studied DNA and RNA from leukocytes in sorted- and single-cells in vivo and in vitro. DNA analyses of sorted cells showed that men diagnosed with Alzheimer’s disease was primarily affected with LOY in NK cells whereas prostate cancer patients more frequently displayed LOY in CD4 + T cells and granulocytes. Moreover, bulk and single-cell RNA sequencing in leukocytes allowed scoring of LOY from mRNA data and confirmed considerable variation in the rate of LOY across individuals and cell types. LOY-associated transcriptional effect (LATE) was observed in ~ 500 autosomal genes showing dysregulation in leukocytes with LOY. The fraction of LATE genes within specific cell types was substantially larger than the fraction of LATE genes shared between different subsets of leukocytes, suggesting that LOY might have pleiotropic effects. LATE genes are involved in immune functions but also encode proteins with roles in other diverse biological processes. Our findings highlight a surprisingly broad role for chromosome Y, challenging the view of it as a “genetic wasteland”, and support the hypothesis that altered immune function in leukocytes could be a mechanism linking LOY to increased risk for disease.

Epithelial Organotypic Cultures: A Viable Model to Address Mechanisms of Carcinogenesis by Epitheliotropic Viruses

2018 ◽  
Vol 18 (4) ◽  
pp. 246-255 ◽  
Author(s):  
Lara Termini ◽  
Enrique Boccardo
Keyword(s):  
Three Dimensional ◽  
Cell Types ◽  
Experimental Models ◽  
Biological Research ◽  
Specific Gene ◽  
Specific Cell ◽  
Organotypic Cultures ◽  
Skin Physiology

In vitro culture of primary or established cell lines is one of the leading techniques in many areas of basic biological research. The use of pure or highly enriched cultures of specific cell types obtained from different tissues and genetics backgrounds has greatly contributed to our current understanding of normal and pathological cellular processes. Cells in culture are easily propagated generating an almost endless source of material for experimentation. Besides, they can be manipulated to achieve gene silencing, gene overexpression and genome editing turning possible the dissection of specific gene functions and signaling pathways. However, monolayer and suspension cultures of cells do not reproduce the cell type diversity, cell-cell contacts, cell-matrix interactions and differentiation pathways typical of the three-dimensional environment of tissues and organs from where they were originated. Therefore, different experimental animal models have been developed and applied to address these and other complex issues in vivo. However, these systems are costly and time consuming. Most importantly the use of animals in scientific research poses moral and ethical concerns facing a steadily increasing opposition from different sectors of the society. Therefore, there is an urgent need for the development of alternative in vitro experimental models that accurately reproduce the events observed in vivo to reduce the use of animals. Organotypic cultures combine the flexibility of traditional culture systems with the possibility of culturing different cell types in a 3D environment that reproduces both the structure and the physiology of the parental organ. Here we present a summarized description of the use of epithelial organotypic for the study of skin physiology, human papillomavirus biology and associated tumorigenesis.

scholarly journals Rho GTPases as Key Molecular Players within Intestinal Mucosa and GI Diseases

Cells ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 66
Author(s):  
Rashmita Pradhan ◽  
Phuong A. Ngo ◽  
Luz d. C. Martínez-Sánchez ◽  
Markus F. Neurath ◽  
Rocío López-Posadas
Keyword(s):  
Intestinal Mucosa ◽  
Rho Gtpases ◽  
Current Knowledge ◽  
Cell Types ◽  
Effector Proteins ◽  
Special Focus ◽  
Specific Cell ◽  
Rho Proteins

Rho proteins operate as key regulators of the cytoskeleton, cell morphology and trafficking. Acting as molecular switches, the function of Rho GTPases is determined by guanosine triphosphate (GTP)/guanosine diphosphate (GDP) exchange and their lipidation via prenylation, allowing their binding to cellular membranes and the interaction with downstream effector proteins in close proximity to the membrane. A plethora of in vitro studies demonstrate the indispensable function of Rho proteins for cytoskeleton dynamics within different cell types. However, only in the last decades we have got access to genetically modified mouse models to decipher the intricate regulation between members of the Rho family within specific cell types in the complex in vivo situation. Translationally, alterations of the expression and/or function of Rho GTPases have been associated with several pathological conditions, such as inflammation and cancer. In the context of the GI tract, the continuous crosstalk between the host and the intestinal microbiota requires a tight regulation of the complex interaction between cellular components within the intestinal tissue. Recent studies demonstrate that Rho GTPases play important roles for the maintenance of tissue homeostasis in the gut. We will summarize the current knowledge on Rho protein function within individual cell types in the intestinal mucosa in vivo, with special focus on intestinal epithelial cells and T cells.

Vertebrate retinal ganglion cells are selected from competent progenitors by the action of Notch

Development ◽  
1995 ◽  
Vol 121 (11) ◽  
pp. 3637-3650 ◽  
Author(s):  
C.P. Austin ◽  
D.E. Feldman ◽  
J.A. Ida ◽  
C.L. Cepko
Keyword(s):  
Cell Fate ◽  
Ganglion Cells ◽  
Notch Pathway ◽  
Cell Types ◽  
Projection Neurons ◽  
Specific Cell ◽  
Vitro Assay ◽  
Notch Activity

The first cells generated during development of the vertebrate retina are the ganglion cells, the projection neurons of the retina. Although they are one of the most intensively studied cell types within the central nervous system, little is known of the mechanisms that determine ganglion cell fate. We demonstrate that ganglion cells are selected from a large group of competent progenitors that comprise the majority of the early embryonic retina and that differentiation within this group is regulated by Notch. Notch activity in vivo was diminished using antisense oligonucleotides or augmented using a retrovirally transduced constitutively active allele of Notch. The number of ganglion cells produced was inversely related to the level of Notch activity. In addition, the Notch ligand Delta inhibited retinal progenitors from differentiating as ganglion cells to the same degree as did activated Notch in an in vitro assay. These results suggest a conserved strategy for neurogenesis in the retina and describe a versatile in vitro and in vivo system with which to examine the action of the Notch pathway in a specific cell fate decision in a vertebrate.

scholarly journals HDAC4 Promotes Growth of Colon Cancer Cells via Repression of p21

2008 ◽  
Vol 19 (10) ◽  
pp. 4062-4075 ◽  
Author(s):  
Andrew J. Wilson ◽  
Do-Sun Byun ◽  
Shannon Nasser ◽  
Lucas B. Murray ◽  
Kanyalakshmi Ayyanar ◽  
...  
Keyword(s):  
Growth Promotion ◽  
Cell Types ◽  
Specific Cell ◽  
Physical Interaction ◽  
Down Regulation ◽  
Proliferative Zone ◽  
P21 Promoter ◽  
Hct116 Cells

The class II Histone deacetylase (HDAC), HDAC4, is expressed in a tissue-specific manner, and it represses differentiation of specific cell types. We demonstrate here that HDAC4 is expressed in the proliferative zone in small intestine and colon and that its expression is down-regulated during intestinal differentiation in vivo and in vitro. Subcellular localization studies demonstrated HDAC4 expression was predominantly nuclear in proliferating HCT116 cells and relocalized to the cytoplasm after cell cycle arrest. Down-regulating HDAC4 expression by small interfering RNA (siRNA) in HCT116 cells induced growth inhibition and apoptosis in vitro, reduced xenograft tumor growth, and increased p21 transcription. Conversely, overexpression of HDAC4 repressed p21 promoter activity. p21 was likely a direct target of HDAC4, because HDAC4 down-regulation increased p21 mRNA when protein synthesis was inhibited by cycloheximide. The importance of p21 repression in HDAC4-mediated growth promotion was demonstrated by the failure of HDAC4 down-regulation to induce growth arrest in HCT116 p21-null cells. HDAC4 down-regulation failed to induce p21 when Sp1 was functionally inhibited by mithramycin or siRNA-mediated down-regulation. HDAC4 expression overlapped with that of Sp1, and a physical interaction was demonstrated by coimmunoprecipitation. Chromatin immunoprecipitation (ChIP) and sequential ChIP analyses demonstrated Sp1-dependent binding of HDAC4 to the proximal p21 promoter, likely directed through the HDAC4–HDAC3–N-CoR/SMRT corepressor complex. Consistent with increased transcription, HDAC4 or SMRT down-regulation resulted in increased histone H3 acetylation at the proximal p21 promoter locus. These studies identify HDAC4 as a novel regulator of colon cell proliferation through repression of p21.

scholarly journals Human Organoids for Predictive Toxicology Research and Drug Development

2021 ◽  
Vol 12 ◽  
Author(s):  
Toshikatsu Matsui ◽  
Tadahiro Shinozawa
Keyword(s):  
Stem Cells ◽  
Drug Development ◽  
Disease Modeling ◽  
Cell Types ◽  
Underlying Disease ◽  
Future Research ◽  
Specific Cell ◽  
Predictive Toxicology

Organoids are three-dimensional structures fabricated in vitro from pluripotent stem cells or adult tissue stem cells via a process of self-organization that results in the formation of organ-specific cell types. Human organoids are expected to mimic complex microenvironments and many of the in vivo physiological functions of relevant tissues, thus filling the translational gap between animals and humans and increasing our understanding of the mechanisms underlying disease and developmental processes. In the last decade, organoid research has attracted increasing attention in areas such as disease modeling, drug development, regenerative medicine, toxicology research, and personalized medicine. In particular, in the field of toxicology, where there are various traditional models, human organoids are expected to blaze a new path in future research by overcoming the current limitations, such as those related to differences in drug responses among species. Here, we discuss the potential usefulness, limitations, and future prospects of human liver, heart, kidney, gut, and brain organoids from the viewpoints of predictive toxicology research and drug development, providing cutting edge information on their fabrication methods and functional characteristics.

scholarly journals Perfused Platforms to Mimic Bone Microenvironment at the Macro/Milli/Microscale: Pros and Cons

2022 ◽  
Vol 9 ◽  
Author(s):  
Maria Veronica Lipreri ◽  
Nicola Baldini ◽  
Gabriela Graziani ◽  
Sofia Avnet
Keyword(s):  
Extracellular Matrix ◽  
Bone Structure ◽  
Dynamic Network ◽  
Cell Types ◽  
New Drugs ◽  
Culture Methods ◽  
Increased Risk ◽  
Multiple Cell

As life expectancy increases, the population experiences progressive ageing. Ageing, in turn, is connected to an increase in bone-related diseases (i.e., osteoporosis and increased risk of fractures). Hence, the search for new approaches to study the occurrence of bone-related diseases and to develop new drugs for their prevention and treatment becomes more pressing. However, to date, a reliable in vitro model that can fully recapitulate the characteristics of bone tissue, either in physiological or altered conditions, is not available. Indeed, current methods for modelling normal and pathological bone are poor predictors of treatment outcomes in humans, as they fail to mimic the in vivo cellular microenvironment and tissue complexity. Bone, in fact, is a dynamic network including differently specialized cells and the extracellular matrix, constantly subjected to external and internal stimuli. To this regard, perfused vascularized models are a novel field of investigation that can offer a new technological approach to overcome the limitations of traditional cell culture methods. It allows the combination of perfusion, mechanical and biochemical stimuli, biological cues, biomaterials (mimicking the extracellular matrix of bone), and multiple cell types. This review will discuss macro, milli, and microscale perfused devices designed to model bone structure and microenvironment, focusing on the role of perfusion and encompassing different degrees of complexity. These devices are a very first, though promising, step for the development of 3D in vitro platforms for preclinical screening of novel anabolic or anti-catabolic therapeutic approaches to improve bone health.

Cell-specific and targeted delivery of RNA moieties

2020 ◽  
Author(s):  
Aditi Bhargava ◽  
Peter Ohara ◽  
Luc Jasmin
Keyword(s):  
Nucleic Acids ◽  
Targeted Delivery ◽  
Immune Cell ◽  
Neuronal Cell ◽  
Cell Types ◽  
Specific Cell ◽  
Chemically Modified ◽  
Covalently Linked

AbstractDelivery of therapeutic moieties to specific cell types, such as neurons remains a challenge. Genes present in neurons are also expressed in non-neuronal cell types such as glia where they mediate non-targeted related functions. Thus, non-specific targeting of these proteins/channels has numerous unwanted side effects, as is the case with current small molecules or drug therapies. Current methodologies that use nanoparticles, lipid-mediated uptake, or mannitol in conjunction with lipids to deliver double-stranded RNA (dsRNA) have yielded mixed and unreliable results. We used a neuroanatomical tracer (B subunit of Cholera Toxin (CTB)) that binds to the ganglioside receptors (GM1) expressed on cells, including primary sensory neurons to deliver encapsulated dsRNA. This approach greatly improved delivery of dsRNA to the desired cells by enhancing uptake, reducing vehicle-mediated toxicity and protecting nucleotides from degradation by endonucleases. The delivery complex is internalized, and once inside the cell, the dsRNA naturally dissociates itself from the carrier complex and is very effective in knocking down cognate targets, both in vivo and in vitro. Past methods have used CTB-fusion proteins or chemically modified oligos or DNA moieties that have been covalently conjugated to CTB. Furthermore, CTB conjugated to an antigen, protein, or chemically modified nucleic acid is a potent activator of immune cell (T and B cells, macrophages) response, whereas CTB admixed with antigens or unmodified nucleic acids does not evoke this immune response. Importantly, in our method, the nucleic acids are not covalently linked to the carrier molecules. Thus, our method holds strong potential for targeted delivery of therapeutic moieties for cell types expressing GM1 receptors, including neuronal cell types.

scholarly journals In vivo paracrine interaction between urokinase and its receptor: effect on tumor cell invasion.

1991 ◽  
Vol 115 (4) ◽  
pp. 1107-1112 ◽  
Author(s):  
L Ossowski ◽  
G Clunie ◽  
M T Masucci ◽  
F Blasi
Keyword(s):  
Chorioallantoic Membrane ◽  
Fold Increase ◽  
Cell Types ◽  
Cell Surface Receptor ◽  
In Vivo Model ◽  
Specific Cell ◽  
Upa Receptor ◽  
Surface Receptor

Numerous studies have linked the production of increased levels of urokinase type plasminogen activator (uPA) with the malignant phenotype. It has also been shown that a specific cell surface receptor can bind uPA through a domain distinct and distant from the proteolytic domain. In an in vivo model of invasion, consisting of experimentally modified chorioallantoic membrane (CAM) of a chick embryo, only cells that concurrently expressed both uPA and a receptor for uPA, and in which the receptor was saturated with uPA, were efficient in invasion. To test whether uPA produced by one cell can, in a paracrine fashion, affect the invasive capacity of a receptor-expressing cell, we transfected LB6 mouse cells with human uPA (LB6[uPA]), or human uPA-receptor cDNA (LB6[uPAR]). LB6(uPA) cells released into the medium 1-2 Ploug units of human uPA per 10(6) cells in 24 h. The LB6(uPAR) cells expressed on their surface approximately 12,000 high affinity (Kd 1.7 x 10(-10) M uPA binding sites per cell. Unlabeled LB6(uPA) and 125-IUdR-labeled LB6(uPAR) cells were coinoculated onto experimentally wounded and resealed CAMs and their invasion was compared to that of homologous mixtures of labeled and unlabeled LB6(uPAR) or LB6(uPA) cells. Concurrent presence of both cell types in the CAMs resulted in a 1.8-fold increase of invasion of the uPA-receptor expressing cells. A four-fold stimulation of invasion was observed when cells were cocultured in vitro, prior to in vivo inoculation. Enhancement of invasion was prevented in both sets of experiments by treatment with specific antihuman uPA antibodies, indicating that uPA was the main mediator of the invasion-enhancing, paracrine effect on the receptor-expressing cells.

scholarly journals Intratumoral platelet aggregate formation in a murine preclinical glioma model depends on podoplanin expression on tumor cells

2019 ◽  
Vol 3 (7) ◽  
pp. 1092-1102 ◽  
Author(s):  
Barbara Costa ◽  
Tanja Eisemann ◽  
Jens Strelau ◽  
Ingrid Spaan ◽  
Andrey Korshunov ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Tumor Cells ◽  
Cell Types ◽  
Platelet Receptor ◽  
Increased Risk ◽  
Platelet Aggregates ◽  
Glioma Model ◽  
Novel Strategy

Abstract Binding of the sialomucin-like transmembrane glycoprotein podoplanin (PDPN) to the platelet receptor C-type lectin-like receptor 2 induces platelet activation and aggregation. In human high-grade gliomas, PDPN is highly expressed both in tumor cells and in tumor-associated astrocytes. In glioma patients, high expression of PDPN is associated with worse prognosis and has been shown to correlate with intratumoral platelet aggregation and an increased risk of venous thromboembolism (VTE). To functionally assess the role of PDPN in platelet aggregation in vivo, we established a syngeneic orthotopic murine glioma model in C57/Bl6 mice, based on transplantation of p53- and Pten-deficient neural stem cells. This model is characterized by the presence of intratumoral platelet aggregates and by the upregulation of PDPN both in glioma cells and in astrocytes, reflecting the characteristics of human gliomas. Deletion of PDPN either in tumor cells or in astrocytes resulted in glioma formation with similar penetrance and grade compared with control mice. Importantly, only the lack of PDPN in tumor cells, but not in astrocytes, caused a significant reduction in intratumoral platelet aggregates, whereas in vitro, both cell types have similar platelet aggregation-inducing capacities. Our results demonstrate a causative link between PDPN and platelet aggregation in gliomas and pinpoint the tumor cells as the major players in PDPN-induced platelet aggregation. Our data indicate that blocking PDPN specifically on tumor cells could represent a novel strategy to prevent platelet aggregation and thereby reduce the risk of VTE in glioma patients.

scholarly journals An alternative approach to produce versatile retinal organoids with accelerated ganglion cell development

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Philip E. Wagstaff ◽  
Anneloor L. M. A. ten Asbroek ◽  
Jacoline B. ten Brink ◽  
Nomdo M. Jansonius ◽  
Arthur A. B. Bergen
Keyword(s):  
Ganglion Cells ◽  
Cell Types ◽  
Retinal Cell ◽  
Specific Cell ◽  
Retinal Ganglion ◽  
In Vivo Models ◽  
Alternative Approach

AbstractGenetically complex ocular neuropathies, such as glaucoma, are a major cause of visual impairment worldwide. There is a growing need to generate suitable human representative in vitro and in vivo models, as there is no effective treatment available once damage has occured. Retinal organoids are increasingly being used for experimental gene therapy, stem cell replacement therapy and small molecule therapy. There are multiple protocols for the development of retinal organoids available, however, one potential drawback of the current methods is that the organoids can take between 6 weeks and 12 months on average to develop and mature, depending on the specific cell type wanted. Here, we describe and characterise a protocol focused on the generation of retinal ganglion cells within an accelerated four week timeframe without any external small molecules or growth factors. Subsequent long term cultures yield fully differentiated organoids displaying all major retinal cell types. RPE, Horizontal, Amacrine and Photoreceptors cells were generated using external factors to maintain lamination.

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