25 research outputs found
Human-Induced Pluripotent Stem Cell-Derived Engineered Cardiac Tissues
By combining tissue engineering techniques with human-induced pluripotent stem cell (hiPSC) technology, human-derived engineered cardiac tissues (ECTs) have been developed using several cell lineage compositions and 3-dimensional geometries. Although hiPSC ECTs are relatively immature compared with native adult heart tissues, they have promising potential as a platform technology for drug-screening and disease modeling, and as grafts for hiPSC-based regenerative heart therapy. This chapter provides the focused overview of the current status of cardiac tissue engineering technology and its possible application
Autoclave Antigen Retrieval Technique for Immunohistochemical Staining of Androgen Receptor in Formalin-Fixed Paraffin Sections of Human Prostate
Although information on the states of androgen receptor (AR) expression at the individual cell level is essential in understanding the human diseased prostate, histological approaches are often hampered by the artifactual loss of AR or AR antigenicity. Here we examined the effects of various antigen retrieval methods, including microwave irradiation and autoclave treatment, for immunohistochemical detection of AR in cultured LNCaP cells and paraffin embedded sections of human prostate, and found beneficial effects of autoclave treatment over microwave treatment. Staining results were consistent with that of AR messenger RNA expression assessed by the reverse transcriptase-polymerase chain reaction method. The procedures described in this article provided intense and reproducible immunostaining for AR, estrogen receptor and progesterone receptor in paraffin sections of the human diseased prostate. Finally, 0.01M EDTA (pH7.4) gave us the most intense nuclear signal for the steroid hormone receptors, though the best soaking solution was 0.01M citrate buffer (pH6.0) considering the lost of tissue morphology
Autoclave Antigen Retrieval Technique for Immunohistochemical Staining of Androgen Receptor in Formalin-Fixed Paraffin Sections of Human Prostate
Although information on the states of androgen receptor (AR) expression at the individual cell level is essential in understanding the human diseased prostate, histological approaches are often hampered by the artifactual loss of AR or AR antigenicity. Here we examined the effects of various antigen retrieval methods, including microwave irradiation and autoclave treatment, for immunohistochemical detection of AR in cultured LNCaP cells and paraffin embedded sections of human prostate, and found beneficial effects of autoclave treatment over microwave treatment. Staining results were consistent with that of AR messenger RNA expression assessed by the reverse transcriptase-polymerase chain reaction method. The procedures described in this article provided intense and reproducible immunostaining for AR, estrogen receptor and progesterone receptor in paraffin sections of the human diseased prostate. Finally, 0.01M EDTA (pH7.4) gave us the most intense nuclear signal for the steroid hormone receptors, though the best soaking solution was 0.01M citrate buffer (pH6.0) considering the lost of tissue morphology
Autoclave Antigen Retrieval Technique for Immunohistochemical Staining of Androgen Receptor in Formalin-Fixed Paraffin Sections of Human Prostate
The myocardial regenerative potential of three-dimensional engineered cardiac tissues composed of multiple human iPS cell-derived cardiovascular cell lineages
Human induced pluripotent stem cells (hiPSCs) are a robust source for cardiac regenerative therapy due to their potential to support autologous and allogeneic transplant paradigms. The in vitro generation of three-dimensional myocardial tissue constructs using biomaterials as an implantable hiPSC-derived myocardium provides a path to realize sustainable myocardial regeneration. We generated engineered cardiac tissues (ECTs) from three cellular compositions of cardiomyocytes (CMs), endothelial cells (ECs), and vascular mural cells (MCs) differentiated from hiPSCs. We then determined the impact of cell composition on ECT structural and functional properties. In vitro force measurement showed that CM+EC+MC ECTs possessed preferential electromechanical properties versus ECTs without vascular cells indicating that incorporation of vascular cells augmented tissue maturation and function. The inclusion of MCs facilitated more mature CM sarcomeric structure, preferential alignment, and activated multiple tissue maturation pathways. The CM+EC+MC ECTs implanted onto infarcted, immune tolerant rat hearts engrafted, displayed both host and graft-derived vasculature, and ameliorated myocardial dysfunction. Thus, a composition of CMs and multiple vascular lineages derived from hiPSCs and incorporated into ECTs promotes functional maturation and demonstrates myocardial replacement and perfusion relevant for clinical translation
Impact of Cell Composition and Geometry on Human Induced Pluripotent Stem Cells-Derived Engineered Cardiac Tissue
The current study describes a scalable, porous large-format engineered cardiac tissue (LF-ECT) composed of human induced pluripotent stem cells (hiPSCs) derived multiple lineage cardiac cells with varied 3D geometries and cell densities developed towards the goal of scale-up for large animal pre-clinical studies. We explored multiple 15 × 15 mm ECT geometries using molds with rectangular internal staggered posts (mesh, ME), without posts (plain sheet, PS), or long parallel posts (multiple linear bundles, ML) and a gel matrix containing hiPSC-derived cardiomyocytes, endothelial, and vascular mural cells matured in vitro for 14 days. ME-ECTs displayed the lowest dead cell ratio (p < 0.001) and matured into 0.5 mm diameter myofiber bundles with greater 3D cell alignment and higher active stress than PS-ECTs. Increased initial ECT cell number beyond 6 M per construct resulted in reduced cell survival and lower active stress. The 6M-ME-ECTs implanted onto 1 week post-infarct immune tolerant rat hearts engrafted, displayed evidence for host vascular coupling, and recovered myocardial structure and function with reduced scar area. We generated a larger (30 × 30 mm) ME-ECT to confirm scalability. Thus, large-format ECTs generated from hiPSC-derived cardiac cells may be feasible for large animal preclinical cardiac regeneration paradigms
Chronic optical pacing conditioning of h-iPSC engineered cardiac tissues
The immaturity of human induced pluripotent stem cell derived engineered cardiac tissues limits their ability to regenerate damaged myocardium and to serve as robust in vitro models for human disease and drug toxicity studies. Several chronic biomimetic conditioning protocols, including mechanical stretch, perfusion, and/or electrical stimulation promote engineered cardiac tissue maturation but have significant technical limitations. Non-contacting chronic optical stimulation using heterologously expressed channelrhodopsin light-gated ion channels, termed optogenetics, may be an advantageous alternative to chronic invasive electrical stimulation for engineered cardiac tissue conditioning. We designed proof-of-principle experiments to successfully transfect human induced pluripotent stem cell derived engineered cardiac tissues with a desensitization resistant, chimeric channelrhodopsin protein, and then optically paced engineered cardiac tissues to accelerate maturation. We transfected human induced pluripotent stem cell engineered cardiac tissues using an adeno-associated virus packaged chimeric channelrhodopsin and then verified optically paced by whole cell patch clamp. Engineered cardiac tissues were then chronically optically paced above their intrinsic beat rates in vitro from day 7 to 14. Chronically optically paced resulted in improved engineered cardiac tissue electrophysiological properties and subtle changes in the expression of some cardiac relevant genes, though active force generation and histology were unchanged. These results validate the feasibility of a novel chronically optically paced paradigm to explore non-invasive and scalable optically paced–induced engineered cardiac tissue maturation strategies
【無菌性膿疱の診断と治療】 膿疱症におけるinvolucrinとelafin
雑誌掲載版Involucrinは正常表皮の辺縁帯の構成成分で細胞外に形成される脂質膜と結合し、表皮のバリア機能に重要な働きをしており、尋常性乾癖表皮では発現が冗進している。Elafinは正常表皮では発現しないが、尋常性乾癖表皮では豊富に発現され、好中球の蛋白分解酵素を阻害するとともに、辺縁帯の成分ともなる。膿庖性乾癖ではinvolucrinの発現は冗進しているが、elafinは過剰な蛋白分解酵素と複合体を形成して相対的に不足することがある