Posts in Category: Stem Cells & Regenerative Medicine

Engineering Cardiac Muscle Tissue: A Maturating Field of Research

Engineering Cardiac Muscle Tissue: A Maturating Field of Research

Florian Weinberger, Ingra Mannhardt, Thomas Eschenhagen

Histological structure of native mouse and rat heart (longitudinal mouse heart in A; cross section of rat heart in B), engineered heart tissue (EHT) from neonatal rat heart cells (longitudinal in C, E, cross section in D), or human pluripotent stem cell–derived cardiomyocytes (F–J). A, staining of actinin (green). B, Staining of sarcolemma with wheat germ agglutinin (white in B). C and D, Staining of actinin (red in C and I), F-actin (green in D and J), or actinin and troponin T (green/red in H) and nuclei in blue (all). E, Hematoxylin and eosin–stained paraffin section. A, Reprinted from Mearini et al135 with permission of the publisher. Copyright © 2014, Macmillan Publishers Limited. B, Reprinted from Bensley et al136 with permission. Copyright © 2016, The Authors. C, D, I, and J, Reprinted from Jackman et al35 with permission of the publisher. Copyright © 2016, Elsevier; and from Eschenhagen et al134 (E) with permission of the publisher. Copyright © 2012, the American Physiological Society. F, Reprinted from Hirt et al36 with permission of the publisher. Copyright © 2014, Elsevier. G, Reprinted from Mannhardt et al78 with permission. Copyright © 2016, The Authors. H, Riegler et al93 with permission of the publisher. Copyright © 2015, American Heart Association. [Powerpoint File]

Engineering Cardiac Muscle Tissue: A Maturating Field of Research

Engineering Cardiac Muscle Tissue: A Maturating Field of Research

Florian Weinberger, Ingra Mannhardt, Thomas Eschenhagen

Plane engineered heart tissue (EHT) on Velcro-covered rods (Eschenhagen et al2, A), ring EHTs (Zimmermann et al24, B), fibrin-based mini-EHT on PDMS (polydimethylsiloxane) racks (Hansen et al25, C), cardiac micro tissues (CMT) on fluorescent pillars (Boudou et al34, D), cardiobundles on PDMS frame (Jackman et al35, Copyright © 2016, Elsevier; E), micro heart muscle (Huebsch et al81, Copyright © 2016, The Authors; F), cardiac biowires (Nunes et al131, Copyright © 2013, Nature Publishing Group; G), cardiac patch (Bian et al133, H). Please note that scale bars are only representative, and sketches might not match the exact dimensions. Graphics in A, B, and D were modified from Eschenhagen et al134 with permission of the publisher. Copyright © 2012, the American Physiological Society. [Powerpoint File]

Cell Therapy Trials in Congenital Heart Disease

Cell Therapy Trials in Congenital Heart Disease

Hidemasa Oh

Preclinical study of intracoronary cardiosphere-derived cell (CDC) infusion in a rat model of right heart failure. A–K, Pulmonary artery (PA) banding was created to induce pressure overload right heart failure in rats (weighing 250–300 g). The left thorax was opened to expose the pulmonary artery. A silk suture was tied tightly around an 18-gauge needle alongside the pulmonary artery, followed by a rapid removal of the needle to leave the pulmonary artery constricted in the lumen equal to the diameter of the needle. Intracoronary infusion was performed 4 weeks after pulmonary artery banding. The ascending aorta and the pulmonary artery were occluded with a snare twice for a 20-s interval, 10 min apart, during which time rats received an infusion of CDCs or vehicle into the aortic root directly. Animals were euthanized at 4 weeks after treatment to obtain immuno-histological data. Masson-trichrome staining and hematoxylin and eosin (H&E) staining are shown. One month after pulmonary artery banding, significant right ventricular hypertrophy and fibrosis were observed. Note that CDC treatment reduced cardiac fibrosis, but not hypertrophy, one month after infusion (F and I). Bars, 2 mm in A to C; 20 μm in D to I. J, Animals were separated into 4 groups: (1) sham-operated animals (n=12), (2) rats subjected to pulmonary artery banding for 4 weeks with vehicle treatment (n=12), and (3 and 4) rats subjected to banding for 4 weeks with 2 doses of CDC infusion (0.5×105 or 1×105 cells: n=12 in each). Cardiac fibrosis induced by right ventricle pressure overload was measured. CDC infusion significantly reduced the fibrotic area in a cell dose-dependent manner. K, CDC treatment did not affect the diameter of myocytes, which are mechanically enlarged by pressure overload. L, Rat CDCs were infected by lentiviral vectors harboring human cytomegalovirus promoter-driven LacZ reporter gene and were subjected to intracoronary transfer into rats 4 weeks after pulmonary artery banding. Clear CDC engraftment could be detected along the endocardium and surrounding capillary vessels where the cells had been injected. M, Cardiomyocytes were stained with α-sarcomeric actin (red). Newly regenerated LacZ-positive cardiac muscle cells could be detected by β-galactosidase staining (green). Substantial cardiomyocyte regeneration was verified 4 weeks after CDC delivery. Bars, 50 μm. N, Engrafted LacZ-positive CDCs were evaluated by X-gal staining and corrected by the number of total nuclei appreciated within the respective area. Fibrotic and nonfibrotic areas were determined by Masson-trichrome staining derived from serial sections. O, Differentiated cardiomyocytes after CDC infusion were verified by the cells coexpressing both α-sarcomeric actin (red) and β-galactosidase (LacZ, green). The frequency of the cells coexpressing α-sarcomeric actin among the β-galactosidase–positive cells is shown. Data are expressed as the means (SD). [Powerpoint File]

Heart Failure in Pediatric Patients With Congenital Heart Disease

Heart Failure in Pediatric Patients With Congenital Heart Disease

Robert B. Hinton, Stephanie M. Ware

Genes causing congenital heart disease (CHD) and cardiomyopathy form a complex network. The network diagrams were generated using ToppCluster (www.toppcluster.cchmc.org) software. Gene lists were derived from clinically available next-generation sequencing panels for cardiomyopathy genes (n=50) and CHD genes (n=44). A, Abstracted cluster network showing a selected subset of features from the following categories: Gene ontology (GO): molecular function; GO: biological processes; GO: cellular component; human phenotype; mouse phenotype; pathway; disease (cardiomyopathy and CHD). The features are color coded by category and connected to cardiomyopathy (white circle) and CHD gene nodes. B, Gene level network with nodes (blue) selected from features in GO: biological processes category. The network illustrates that regulation of cellular component of movement, actin filament-based processes, and cardiac chamber morphogenesis are shared in common between cardiomyopathy and CHD genes, whereas regulation of force of heart contraction and tube development are unshared. [Powerpoint File]

Cardiac Regeneration: Lessons From Development

 

Cardiac Regeneration: Lessons From Development

Francisco X. Galdos, Yuxuan Guo, Sharon L. Paige, Nathan J. VanDusen, Sean M. Wu, William T. Pu

Model of the trabeculation process. During trabeulation, a small fraction of cardiomyocytes (CMs) in the compact myocardium (pink) are first specified as trabeculating CMs (brown). These cells delaminate from the compact myocardium and migrate inward to form the first trabecular CMs. CMs in both compacted and trabecular myocardium further proliferate. This proliferation, together with CM migration and rearrangement, results in protrusion and expansion of the trabecular myocardium (illustration credit: Ben Smith). [Powerpoint File]

Cardiac Regeneration: Lessons From Development

Cardiac Regeneration: Lessons From Development

Francisco X. Galdos, Yuxuan Guo, Sharon L. Paige, Nathan J. VanDusen, Sean M. Wu, William T. Pu

Regulation of cardiac progenitor proliferation and differentiation. A, Schematic showing the anterolateral position of first heart field (FHF) progenitors and dorsomedial position of second heart field (SHF) progenitors at embryonic day (E) 7.5. Canonical wingless-type MMTV integration site family member (WNTs), Sonic Hedgehog (SHH), and fibroblast growth factors (FGFs) are expressed dorsally in the region encompassed by the SHF, whereas noncanonical WNTs, BMP2, and FGF8 are expressed ventrally, where the FHF is present. FHF progenitors make up the cardiac crescent and differentiate before the SHF to form the developing heart tube at E8.0. SHF maintains their proliferative state and elongate the heart tube by migrating and differentiating at the inflow and outflow poles of the heart. B, Noncanonical WNTs, BMP2/4, and FGF8 signaling drives FHF progenitors to differentiates toward the myocyte lineage. Meanwhile, canonical WNT/β-catenin, SHH, and FGFs maintain SHF progenitor proliferation. SHF progenitor migration to the outflow and inflow poles of the heart tube exposes them to BMP2/4 and noncanonical WNTs, which drives SHF progenitors to exit their proliferative state and differentiate. C, Canonical WNT/ β-catenin signaling inhibits the differentiation of cardiac progenitors to the myocytes. BMP signaling activates SMAD4 that binds to the transcription factor HOPX to directly inhibit canonical WNT/β-catenin. Moreover, noncanonical WNTs such as WNT5a and WNT11 also inhibit canonical WNT/β-catenin to drive cardiac progenitor differentiation (illustration credit: Ben Smith). [Powerpoint File]

Cardiac Regeneration: Lessons From Development

Cardiac Regeneration: Lessons From Development

Francisco X. Galdos, Yuxuan Guo, Sharon L. Paige, Nathan J. VanDusen, Sean M. Wu, William T. Pu

Specification of mesodermal precursors. Schematic representing the signaling events leading to mesodermal specification during early development. NODAL (nodal growth differentiation factor) is first expressed proximally at embryonic day (E) 5.0. Through an autoregulatory loop, NODAL activates its own expression throughout the epiblast (shown in light purple) and goes on to induce the expression of NODAL antagonists, LEFTY1 and CER1, in the distal visceral endoderm at E5.5 (DVE). The DVE migrates anteriorly where it specifies the anterior portion of the embryo as shown in the yellow hues at E6.5 to 7.5. The anterior visceral endoderm (AVE, yellow) limits NODAL signaling to the posterior of the embryo. Along with wingless-type MMTV integration site family member 3 (WNT3) and bone morphogenic protein (BMP) signaling, NODAL specifies early primitive streak progenitors to the mesoderm fate. [Powerpoint File]

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Ji-Hye Jung, Xuebin Fu, Phillip C. Yang

Biogenesis of exosome. Exosomes are compartmentalized into the multivesicular bodies (MVBs), which fuse with the cell membrane and released to the extracellular space. This process prevents exosomes from degradation by the lysosomes. Exosomes comprise of various transmembrane and cytosolic proteins, such as integrins, CD9, CD63, and CD81. Furthermore, exosomes retain donor cells’ proteins, DNA fragments, miRNAs, and noncoding RNAs within the bilipid membrane. As such, exosomes preserve the genetic information of donor cells from enzymatic degradation while enabling targeted delivery of specific cargo. [Powerpoint File]

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Ji-Hye Jung, Xuebin Fu, Phillip C. Yang

Exosomes mediated intercellular communication in heart. Exosomes facilitate communication among cardiomyocytes, endothelial cells, and vascular smooth muscle cells in the infarcted area of the heart. Exosomes transfer signaling molecules, such as miRNAs, mRNAs, and proteins to confer paracrine effects on the neighboring cells. In addition, pathological and physiological influence on the heart stimulates exosome secretion. Therefore, cardiac exosomes under pathological conditions could be used as ideal markers for diagnostic tools in the clinic. [Powerpoint FIle]

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Exosomes Generated From iPSC-Derivatives: New Direction for Stem Cell Therapy in Human Heart Diseases

Ji-Hye Jung, Xuebin Fu, Phillip C. Yang

Personalized therapy with the exosomes generated from induced pluripotent stem cell (iPSC)–derived cardiomyocytes (iCMs). Exosomes from patient-specific iPSC derivatives provide a platform for personalized therapy by simulating endogenous repair. Exosomes can be directed to specific injured site of each individual patient to promote salvage of the existing injured cardiac cells. Exosomes isolated from the supernatant of iCM culture undergo ultracentrifugation methods, as well as advanced characterization and quantification by transmission electronical microscopy, Nanosight, and dynamic light scattering. [Powerpoint File]

From Microscale Devices to 3D Printing: Advances in Fabrication of 3D Cardiovascular Tissues

From Microscale Devices to 3D Printing: Advances in Fabrication of 3D Cardiovascular Tissues

Anton V. Borovjagin, Brenda M. Ogle, Joel L. Berry, Jianyi Zhang

Bioprinting is usually accomplished using a combination of gel and cells. Laser-assisted bioprinting (A) using Laser-induced forward transfer relies on the focused energy of a laser onto an energy absorbing ribbon to induce bioink droplet formation. This technique is advantageous because it avoids the problem of clogging of the bioink nozzle that plagues other bioprinting techniques. Multiphoton excitation-based printing (B) is accomplished via photocrosslinking of proteins or polymers in the focal volume of the laser and excels in its high resolution and ability to polymerize many native proteins that do not form hydrogels spontaneously outside the body. Inkjet printing (C), one of the most common printing techniques, relies on a vapor bubble or a piezoelectric actuator to displace material to extrude the bioink from a nozzle. Robotic dispensing (D) uses other mechanical means of displacing bioink under robotic control. ECM indicates extracellular matrix. [Powerpoint File]

From Microscale Devices to 3D Printing: Advances in Fabrication of 3D Cardiovascular Tissues

From Microscale Devices to 3D Printing: Advances in Fabrication of 3D Cardiovascular Tissues

Anton V. Borovjagin, Brenda M. Ogle, Joel L. Berry, Jianyi Zhang

In vitro testing of cells and tissues may occur in several ways. Microfluidic systems (A) have emerged as a tool for basic science studies of the effect of highly controlled fluid mechanical and solid mechanical forces on single cell types or cocultures. Microfluidic systems are also gaining favor as a diagnostic tool and a platform for drug development. Organoid cultures (B) are described as organ buds grown in culture that feature realistic microanatomy and are useful as cellular models of human disease. These cultures have found utility in the study of basic mechanisms of organ-specific diseases. Spheroid cultures (C) feature sphere-shaped clusters of a single cell type or coculture sustained in a gel or a bioreactor in order to interact with their 3D surroundings and are useful in testing drug efficacy and toxicity. (D) Engineered heart tissues are constructed by polymerizing an extracellular matrix–based gel containing cardiac cell types between 2 elastomeric posts or similar structures allowing auxotonic contraction of cardiomyocytes. This allows approximation of the normal conditions of the heart contracting against the hydrostatic pressure imposed by the circulation. This type of tissue construct has been used for testing toxicity of drugs and basic studies of muscle function and interplay between multiple cardiac cell types. [Powerpoint File]

Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps

Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps

Kathleen M. Broughton, Mark A. Sussman

Cardiac regenerative medicine product to marketplace. Several companies focused on the treatment of heart failure and regenerative cardiac medicine have emerged over the past decade. At this time, no commercially available product is approved by the US FDA and commercially available in the United States. Each company is positioned based on its most current clinical status in ClinicalTrials.gov and the company website. This figure was last updated on February 1, 2016. [Powerpoint File]

Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps

Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps

Kathleen M. Broughton, Mark A. Sussman

Strengths, Weaknesses, Opportunities, and Threats (SWOT) analysis of individual adult stems cells as a cardiovascular therapy. The analysis focus is the individual adult stem cell therapeutic treatment options. The analysis focuses on the internal workings of the individual treatment options based on current clinical trial results and ongoing preclinical research. [Powerpoint File]