The Berkeley Human Embryonic Stem Cell Shared Research Laboratory

The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic and other stem cells. Our facility is designed to support human embryonic stem cell (hESC) culture, including high quality cell culture space equipped with biosafety cabinets, incubators, cryogenic storage, and a standard microscope. In addition, we have developed and focused state-of-the-art resources and expertise to meet a growing need for our users, imaging. In particular, we have a two photon and visible confocal fluorescence microscope and a high throughput fluorescence imager that are being increasingly used. In addition, our analytical and sorting flow cytometry capabilities complement the imaging equipment by providing high throughput cell fluorescence measurements. Finally, the facility is equipped with a laminar flow hood to conduct chemistry to create biomaterials and micropatterned surfaces for stem cell culture, as well as subsequent analysis by imaging and flow. We are very thankful to CIRM for enabling the construction and development of this state-of-the-art stem cell facility. Also, the generous resources of CIRM have been additionally leveraged, as the facility director has obtained additional campus funds to purchase additional equipment and further enhance the capabilities of the existing equipment. Our facility is directed by Prof. David Schaffer in collaboration with the management oversight committee. In addition, it is managed by Dr. Mary West, who has successfully overseen the installation of the equipment and resources described above, provides rigorous training to our users, and aids in the development of imaging strategies to enable numerous experiments. In greater scientific detail, 57 students and postdoctoral fellows from the laboratories of 11 PIs have approved access and are using the facility to date. Therefore, these resources are enabling and enhancing a large and growing number of research projects, which are described in greater detail in the full scientific progress report. As one example, the Robey and Winoto labs are developing and optimizing in vitro culture systems to induce T cell development from human embryonic stem cells and iPS cell lines. An important parameter is the choice of the starting cell line, and a number of non-registry human ES cells lines are being compared. Therefore the availability of lab space that is separate from NIH funded labs is key to being able to carry out these studies. As another example, the Healy lab is developing new synthetic substrates for culturing human embryonic stem cells under defined conditions. They have made significant progress with several surface formulations, and quantitative measurements on the maintenance of hESC proliferation and pluripotency are being aided by the imaging equipment in the CIRM facility. As another example, the Schaffer lab has engineered new gene delivery vehicles that can augment and edit the genome of hESCs and neural stem cells. Quantification of gene delivery efficiency and gene targeting has been greatly aided with the flow cytometry and high throughput imaging capabilities of the CIRM facility. As an additional example, the Li lab has been utilizing micropatterned surfaces of variable mechanical properties to study the effects of substrate mechanics on stem cell function, and these efforts have been enabled by the high throughput imager. Finally, the Kumar lab studies how the molecular and mechanical properties of the cellular cytoskeleton regulate cell function. For example, he uses laser ablation of subcellular features such as cytoskeletal filaments to provide real time information on the mechanical properties of the cytoskeleton. He is establishing laser ablation using the multiphoton microscope in the CIRM facility, a capability that will be applied to study cytoskeletal mechanics of stem cells. We thus anticipate that this valuable facility will thus be increasingly utilized to advance a growing number of projects.

The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic and other stem cells. Our facility is designed to support human embryonic stem cell (hESC) culture, including high quality cell culture space equipped with biosafety cabinets, incubators, cryogenic storage, and a standard microscope. In addition, we have developed and focused state-of-the-art resources and expertise to meet a growing need for our users, imaging. In particular, we have a two photon and visible confocal fluorescence microscope and a high throughput fluorescence imager (ImageXpress) that are being increasingly used. Our analytical and sorting flow cytometry capabilities complement the imaging equipment by providing high throughput cell fluorescence measurements. Finally, the facility is equipped with a laminar flow hood to conduct chemistry to create biomaterials and micropatterned surfaces for stem cell culture, as well as subsequent analysis by imaging and flow. We are very thankful to CIRM for enabling the construction and development of this state-of-the-art stem cell facility. Also, the generous resources of CIRM have been additionally leveraged, as the facility director has obtained additional campus funds to purchase additional equipment and further enhance the capabilities of the existing equipment. Our facility is directed by Prof. David Schaffer in collaboration with the management oversight committee. In addition, it is managed by Dr. Mary West, who has successfully overseen the installation of the equipment and resources described above, provides rigorous training to our users, and aids in the development of imaging strategies to enable numerous experiments. In greater scientific detail, 75 students and postdoctoral fellows from the laboratories of 21 PIs have approved access and are using the facility to date. Therefore, these resources are enabling and enhancing a large and growing number of research projects, which are described in greater detail in the full scientific progress report. The goal of a project in the Song Li lab that has made great progress this year is to derive functional neural crest stem cells (NCSCs) from embryonic stem cells and induced pluripotent stem cells. The derived NCSCs were used in nerve and blood vessel regeneration. In addition, other projects aimed at identification and characterization of adult stem cells in blood vessels. The marker expression and multipotency were determined by using ImageXpress and flow cytometry at CIRM SSCF. The next phase of this study focuses on the function of vascular stem cells in other diseases including arthrosclerosis. As another example, the Healy lab is developing new synthetic substrates for culturing human embryonic stem cells under defined conditions. Progress was made and published by the Healy lab in creating synthetic culture surfaces and materials for long-term hESC propagation, a project that has involved heavy usage of the ImageXpress. As a final example, the Schaffer lab has engineered new gene delivery vehicles that can augment and edit the genome of hESCs and neural stem cells. Quantification of gene delivery efficiency and gene targeting has been greatly aided with the flow cytometry and high throughput imaging capabilities of the CIRM facility. Finally, the Kumar lab studies how the molecular and mechanical properties of the cellular cytoskeleton regulate cell function. For example, he uses laser ablation of subcellular features such as cytoskeletal filaments to provide real time information on the mechanical properties of the cytoskeleton. He is establishing laser ablation using the multiphoton microscope in the CIRM facility, a capability that will be applied to study cytoskeletal mechanics of stem cells. We anticipate in the upcoming year that this valuable facility will thus be increasingly utilized to advance a growing number of projects.

Grant Number: CL1-00519 PI Name: David V. Schaffer Email: schaffer@berkeley.edu Project Title: The Berkeley Human Embryonic Stem Cell Shared Research Laboratory a. Scope of Research Taking Place in the Facility The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic stem cells, as well as other stem cells. Our facility focuses on providing imaging, flow cytometry, and sorting. To date, 90 students and postdoctoral fellows from the laboratories of 21 PIs use the facility. We list some examples here: Irina Conboy (BioE). The Conboy lab studies the intersection of aging and stem cell science. If age-imposed decline in the regenerative capacity of stem cells was understood, the debilitating lack of organ maintenance in the old could be ameliorated and perhaps, even reversed. They used the SSCF confocal microscope and flow cytometer in their studies. Paliwal P, et al. (2012) Age dependent increase in the levels of osteopontin inhibits skeletal muscle regeneration. Aging 4(8):553. Paliwal P, Conboy IM. (2011) Inhibitors of tyrosine phosphatases and apoptosis reprogram lineage-marked differentiated muscle to myogenic progenitor cells. Chem Biol 18(9):1153. Sanjay Kumar (BioE). The Kumar Lab’s interests center around the macromolecular basis of cell shape, mechanics, and adhesion, with special emphasis on the nervous system. Specifically, they seek to understand how elements of the cytoskeleton and adhesion machinery physically interact to form a three-dimensional architecture that drives cell shape and shape-dependent behavior and transduces biochemical signals. The SSCF swept field confocal was used to document changes in soft ECMs (MacKay). This microscope was also used by researcher Amit Pathik, who developed paradigm for investigating matrix regulation of invasion (Pathak). J. L. MacKay, et al. (2012). A genetic strategy for the dynamic and graded control of cell mechanics, motility, and matrix remodeling. Biophysical Journal 102: 434. A. Pathak and S. Kumar (2012). Independent regulation of tumor cell migration by matrix stiffness and confinement. PNAS 109 (26): 10334. David V. Schaffer (ChemE, BioE, and Helen Wills Neuroscience Institute). One recently-completed CIRM-funded project involved directed evolution and engineering of new viral gene delivery vehicles capable of highly efficient delivery to human embryonic and induced pluripotent stem cells. This Adeno-associated virus (AAV) variant study involved heavy usage of the flow cytometer and the ImageXpress Micro to quantitate the number of successfully infected (and gene-targeted) cells and colonies (Asuri). In a current CIRM-funded project, we are engineering synthetic materials for the scaleable expansion and dopaminergic differentiation of human ESCs and iPSCs (Keung). Also, we are developing optogenetic tools to investigate signaling pathways involved in stem cell fate decisions (Bugaj). Finally, we discovered a novel signaling pathway that regulates the differentiation of adult neural stem cells into neurons (Ashton). Asuri, P., et al. (2012) “Directed Evolution of Adeno-Associated Virus for Enhanced Gene Delivery and Gene Targeting in Human Pluripotent Stem Cells.” Molecular Therapy 20:329. Bugaj, L.J., et al. (2013) “A Modular Optogenetic Platform for Inducible Protein Clustering and Signaling Activation in Mammalian Cells.” Nature Methods 10:249. Keung, A.J., et al. (2012) “Soft Microenvironments Promote the Early Neurogenic Differentiation but not Self-renewal of Human Pluripotent Stem Cells.” Integrative Biology 4:1049. Ashton, R.S., et al. (2012) “Astrocytes Regulate Adult Hippocampal Neurogenesis Through Ephrin-B Signaling.” Nature Neuroscience 15:1399. b. Management and Use of the Laboratory The overall management of the CIRM Shared Stem Cell Facility CIRM SSCF is coordinated by the management oversight committee.CIRM SSCF is managed by Dr. Mary West. The facility is open to qualified researchers from 9:30 am to 5 pm with controlled access afterhours. c. Activities of Hired Laboratory Personnel The CIRM Shared Stem Cell Facility is managed by Dr. Mary West who is responsible for training, equipment function and maintenance. CIRM is also funding an operator, Alma Faleros, for the BD FACS instrument, who works from 12:30 to 9:30pm. d. Activities of the Oversight Committee The oversight committee is composed of Facility Director David Schaffer and Profs. Kevin Healy, Song Li, Sanjay Kumar, Irina Conboy, and Astar Winoto. The committee meets regularly with the Facility Manager to obtain updates on the facility, discuss prioritization for new equipment purchases, facility finances, and policies for access. e. Plans and Any Expected Changes for the Next Reporting Period In the next year we anticipate busy operation of the facility with no major changes.

The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic, induced pluripotent, and adult stem cells. We have reached a steady state usage of ~90 students and postdoctoral fellows from the laboratories of 21 PIs at the SSCF, located in Stanley Hall located on the North east end of campus. In addition, cell sorting instrumentation remains in the campus flow facility. Based on the interests of the users, we have continued in Year 4 to provide informational seminars, conduct new instrument demonstrations, offer bulk discounted rates on expensive stem cell medias, and find creative ways to obtain additional instrumentation that will be useful to users. For instance, this year we added considerable automated equipment by incorporating a High-Throughput Screening Facility (HTSF) into our management and administrative structure. This facility includes cell seeding equipment within a BSL-2 cell culture hood, two automated liquid handlers with complementary capabilities, a multi-label plate reader, and automated imager (similar to the SSCF ImageXpress Micro) with a robotic servicing arm for screening of multi-well plates. Most automation equipment was retained at its originally location in Li Ka Shing, in the CIRM Center of Excellence facility, and some CIRM-funded faculty are currently are using the facility to screen for protein partners in pathways that are unique to stem cell biology. In Stanley Hall, the SSCF recently added a new room, B203 Stanley, for a state-of-the-art instrument that helps to detect levels of multiple proteins or growth factors simultaneously from a small sample of tissue or serum. In addition, the HTSF automated imaging equipment and accompanying computer server was moved to B203 Stanley so the facility could have direct management over training and use. These additions this year will further enhance the offerings to stem cell researchers to widen the breadth of the instrumentation that we could offer to researchers, who are grateful that we have these instruments in our facility because of the shared aspect of maintenance and training. The previously highlighted PIs (detailed in Years 2 and 3) continue to have success using SSCF instrumentation and we would like to highlight publications by researchers who have thus made significant advancements to their research. Publications: Downing T., J. Soto, C. Morez, T. Houssin, A. Fritz, F. Yuan, J. Chu, S. Patel, D.V. Schaffer, S. Li, (2013), "Biophysical regulation of epigenetic state and cell reprogramming". Nature Materials, 12:1154-62 (PMID: 24141451). Cousin, W., M. Ho, R. Desai, A. Tham, R. Chen, S. Kung, C. Elab, I. Conboy, (2013) "Regenerative Capacity of Old Muscle Stem Cells Declines without Significant Accumulation of DNA Damage" Public Library of Science (PLoS), 8:e63528 (PMID: 23704914). Yousef, H., M.J. Conboy, J. Li, M. Zeiderman, T. Vazin, C. Schlesinger, D.V. Schaffer, and I.M. Conboy (2013) “hESC-Secreted Proteins can be Enriched for Multiple Regenerative Therapies by Heparin-Binding.” Aging, 5:357-372 (PMID 23793469). Elabd, C., W. Cousin, R. Chen, M. Chooljian, J. Pham, I. Conboy, M. Conboy (2013) "DNA methyltransferase-3-dependent nonrandom template segregation in differentiating embryonic stem cells." The Journal of Cell Biology, 203:73-85 (PMID: 24127215). A. Pathak and S. Kumar (2013). “Transforming potential and matrix stiffness co-regulate confinement sensitivity of tumor cell migration.” Integrative Biology 5: 1067-1075 (PMID: 23832051). Ma, Z., S. Koo, M. Finnegan, P. Loskill, N. Huebsch, N. Marks, B. Conklin, C. Grigoropoulos, K. Healy K. (2013) “Three-dimensional filamentous human diseased cardiac tissue model”, Biomaterials. (5):1367-1377 (PMID: 24268663). Halkias, J., H. Melichar, K. Taylor, J. Ross, B. Yen, S. Cooper, A. Winoto, and E. Robey (2013) "Opposing chemokine gradients control human thymocyte migration in situ" The Journal of Clinical Investigation, (5):2131-42 (PMID: 23585474). Bugaj, L.J., A.T. Choksi, C.K. Mesuda, R.S. Kane, and D.V. Schaffer (2013) “A Modular Optogenetic Platform for Inducible Protein Clustering and Signaling Activation in Mammalian Cells.” Nature Methods, 10:249-252 (PMID 23377377). Conway, A., T. Vazin, D.P. Spelke, N.A. Rode, K.E. Healy, R.S. Kane, and D.V. Schaffer (2013) “Multivalent Ligands to Control Stem Cell Behaviour in Vitro and in Vivo.” Nature Nanotechnology, 8:831-838 (PMID 24141540). Keung, A.J., M. Dong, D.V. Schaffer (co-corresponding author), and S. Kumar (2013) “Pan-neuronal Maturation But Not Neuronal Subtype Differentiation of Adult Neural Stem Cells is Mechanosensitive.” Scientific Reports, 3:1817 (PMID 23660869). Lei, Y. and D.V. Schaffer “A Fully Defined and Scalable 3D Culture System for Human Pluripotent Stem Cell Expansion and Differentiation.” Proceedings of the National Academy of Sciences USA (in press).

The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic, induced pluripotent, and adult stem cells. The Flow Cytometry Facility in 461 Li Ka Shing houses the CIRM funded cell sorter. The B129 Stanley suite contains cell culture hoods, four incubators, and other essential lab equipment. The SSCF also has an automated wide field ImageXpress Micro, and two analytical flow cytometers, and a 2-photon and confocal fluorescence microscope. In B203 Stanley, there is a BioRad BioPlex MAGPIX reader for magnetic bead–based multiplexed immunoassays. We have reached a steady state usage of ~90 students and postdoctoral fellows from the laboratories of 21 PIs. Based on the interests of the users, we have continued in Year 5 to obtain additional instrumentation that will be useful to users. For instance, this year we added two independent hypoxia chambers to fit within one of our standard incubators so that experiments at low O2 conditions (similar to the O2 concentration tissues experience inside the body) can be performed. We also now manage a histology lab with equipment to creating slides from tissue specimens. These additions this year will further enhance the offerings to UCB and extended community stem cell researchers. The previously highlighted PIs continue to have success using SSCF instrumentation and we would like to highlight publications by researchers who have made significant advancements to their research. Publications: Li J, Han S, Cousin W, Conboy IM. (2014) "Age-specific functional epigenetic changes in p21 and p16 in injury-activated satellite cells." Stem Cells, 10:1002 (PMID: 25447026). Yousef, H., M.J. Conboy, H. Mamiya, M. Zeiderman, C. Schlesinger, D.V. Schaffer, and I.M. Conboy (2014) “Mechanisms of action of hESC-secreted proteins that enhance human and mouse myogenesis” Aging, 8:602-20 (PMID 25109702). Elabd C, Cousin W, Upadhyayula P, Chen RY, Chooljian MS, Li J, Kung S, Jiang KP, Conboy IM (2014) "Oxytocin is an age-specific circulating hormone that is necessary for muscle maintenance and regeneration." Nat Commun, 5:4082 (PMID: 24915299). MacKay, J.L. and S. Kumar (2014). “Simultaneous and independent tuning of RhoA and Rac1 activity with orthogonally inducible promoters.” Integrative Biology 6: 885-894 (PMID: 25044255). Kim, Y. and S. Kumar (2014). “CD44-mediated adhesion to hyaluronic acid contributes to mechanosensing and invasive motility.” Molecular Cancer Research 12: 1416-1429 (PMID: 24962319). Umesh, V., A. Rape, T. A. Ulrich, and S. Kumar (2014). “Microenvironmental stiffness enhances glioma cell proliferation by stimulating epidermal growth factor receptor signaling.” PLOS ONE 9: E101771 (PMID: 25000176) Rape, D. and S. Kumar (2014). “A composite hydrogel platform for the dissection of tumor cell migration at tissue interfaces.” Biomaterials 35: 8846-8853 (PMID: 25047626). Jha, A. K., W. M. Jackson, and K. E. Healy (2014) “Controlling osteogenic stem cell differentiation via soft bioinspired hydrogels.” PLoS One 9:e98640 (PMID: 24937602) Huebsch, Nathaniel, Peter Loskill, Mohammad Mandegar, Natalie C. Marks, Alice S. Sheehan, Zhen Ma, Anurag A. Mathur (2014) "Automated video-based analysis of contractility and calcium flux in human iPS–derived cardiomyocytes cultured over different spatial scales." Tissue Engineering Part C Methods (EPub ahead of print) (PMID: 25333967) Ma, Z. Koo S, Finnegan MA, Loskill P, Huebsch N, Marks NC, Conklin BR, Grigoropoulos CP, Healy KE (2014) Three-dimensional filamentous human diseased cardiac tissue model. Biomaterials 35, 1367–77 (PMID: 24268663). Yousef, H., A. Morgenthaler, C. Schlesinger, L. Bugaj, I. Conboy, and D.V. Schaffer “Age-Associated Increase in BMP Signaling inhibits Hippocampal Neurogenesis.” Stem Cells (in press). Fritz, A., S.R. Mao, M.G. West, and D.V. Schaffer (2015) “A Medium-Throughput Analysis of Signaling Pathways Involved in Early Stages of Stem Cell Reprogramming.” Biotechnology & Bioengineering 1:209-19 (PMID: 25065366). Hughes, A.J., D.P. Spelke, Z. Xu, C-C. Kang, D.V. Schaffer (co-corresponding author), and A.E. Herr (2014) “Single-cell Western Blotting.” Nature Methods, 11:749-755 (PMID 24880876). Leo, Y., D. Jeong, J, Xiao, and D.V. Schaffer “Developing Defined and Scalable 3D Culture Systems for Culturing Human Pluripotent Stem Cells at High Densities.” Cell and Molecular Bioengineering 2:172-183 (PMID: 25419247). Conway, A. and D.V. Schaffer (2014) “Biomaterial Microenvironments to Support the Generation of New Neurons in the Adult Brain.” Stem Cells, 32:1220-1229 (PMID 24449485). Vazin, T., R.S. Ashton, A. Conway, N. Rode, S.M. Lee, V. Bravo, K.E. Healy, R.S. Kane, and D.V. Schaffer (2014) “The Effect of Multivalent Sonic Hedgehog on Differentiation of Human Embryonic Stem Cells into Dopaminergic and GABAergic Neurons.” Biomaterials, 35:941-948 (PMID 24172856).

The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic, induced pluripotent, and adult stem cells. The Flow Cytometry Facilities in 461 Li Ka Shing and 493 LSA house the CIRM funded cell sorters. The B129 Stanley suite contains cell culture hoods, four incubators, and other essential lab equipment. The SSCF also has an automated wide field ImageXpress Micro, and two analytical flow cytometers, and a 2-photon and confocal fluorescence microscope system. In B203 Stanley, there is a BioRad MAGPIX reader for magnetic bead–based multiplexed immunoassays. We have reached a steady state usage of ~90 students and postdoctoral fellows from the laboratories of 21 PIs. Based on the interests of the users, we have continued in Year 6 to maintain state of the art instrumentation for the research community. This year we upgraded our Image Xpress Micro to have the larger format CMOS camera and LED light source to improve imaging capability and reduce long-term cost to run the instrument. The previously highlighted PIs continue to have success using SSCF instrumentation and we would like to highlight publications by some of these researchers who have made significant advancements to their research by using our facility. Publications: B. Zheng, T. Vazin, P.W. Goodwill, A. Conway, A. Verma, E.U. Saritas, D.V. Schaffer, and S.M. Conolly Magnetic Particle Imaging tracks the long-term fate of in vivo neural cell implants with high image contrast. Scientific Reports, 2015, 10(5)14055 (PMID: 26358296). S. Y. Wong, T. A. Ulrich, L. P. Deleyrolle, J. L. MacKay, J.-M. Lin, R. T. Martuscello, M. A. Jundi, B. A. Reynolds, and S. Kumar (2015). Constitutive activation of myosin-dependent contractility sensitizes glioma tumor-initiating cells to mechanical inputs and reduces tumor invasion. Cancer Research 75: 1113-1122 (PMID: 25634210). A. D. Rape, M. Zibinsky, N. Murthy, and S. Kumar (2015). A synthetic hydrogel for the high-throughput study of cell-ECM interactions. Nature Communications 6: 8129 (PMID: 26350361). Han, Bruce W., Hans Layman, Nikhil A. Rode, Anthony Conway, David V. Schaffer, Nancy J. Boudreau, Wesley M. Jackson, and Kevin E. Healy. Multivalent Conjugates of Sonic Hedgehog Accelerate Diabetic Wound Healing. Tissue Engineering. Part A 21, no. 17–18 (September 2015): 2366–78. doi:10.1089/ten.TEA.2014.0281 (PMID: 26154888). Ma, Zhen, Jason Wang, Peter Loskill, Nathaniel Huebsch, Sangmo Koo, Felicia L. Svedlund, Natalie C. Marks, et al. Self-Organizing Human Cardiac Microchambers Mediated by Geometric Confinement. Nature Communications 6 (July 14, 2015). doi:10.1038/ncomms8413 (PMID: 26172574). Amit K. Jha, Anurag Mathur, Felicia L. Svedlund, Jianqin Ye, Yerem Yeghiazarians, Kevin E. Healy. Molecular weight and concentration of heparin in hyaluronic acid-based matrices modulates growth factor retention kinetics and stem cell fate. Journal of Controlled Release 209 (2015): 308-316 (PMID: 25931306). Mathur, Anurag, Peter Loskill, Kaifeng Shao, Nathaniel Huebsch, SoonGweon Hong, Sivan G. Marcus, Natalie Marks et al. Human iPSC-based Cardiac Microphysiological System For Drug Screening Applications. Scientific reports 5 (2015) (PMID: 25748532). Jha, Amit K., et al. Enhanced survival and engraftment of transplanted stem cells using growth factor sequestering hydrogels. Biomaterials 47 (2015): 1-12 (PMID: 25682155). Huebsch, Nathaniel, Peter Loskill, Mohammad A. Mandegar, Natalie C. Marks, Alice S. Sheehan, Zhen Ma, Anurag A. Mathur et al. Automated video-based analysis of contractility and calcium flux in human iPS–derived cardiomyocytes cultured over different spatial scales. Tissue Engineering ja (2015) (PMID: 25333967). Bugaj, L.J., D. P. Spelke, C.K. Mesuda, M. Varedi, R.S. Kane, and D.V. Schaffer (2015) “Regulation of Endogenous Transmembrane Receptors Through Optogenetic Cry2 Clustering.” Nature Communications, 6:6898 (PMID 25902152). Kotterman, M.A., T. Vazin, and D.V. Schaffer (2015) “Adeno-Associated Viral Variant Enhances Selective Gene Delivery to Neural Stem Cells in Vivo.” Development, 142:1885-1892 (PMID 25968319). Fritz, A.L., M.M. Adil, S.R. Mao, D.V. Schaffer (2015) “cAMP and EPAC Signaling Functionally Replace OCT4 During Induced Pluripotent Stem Cell Reprogramming.” Molecular Therapy, 23:952-963 (PMID 25666918). Yousef, H., M.J. Conboy, A. Morgenthaler, C. Schlesinger, L. Bugaj, P. Paliwal, C. Greer, I.M. Conboy, and D.V. Schaffer (2015) “Systemic Attenuation of the TGF-β Pathway by a Single Drug Simultaneously Rejuvenates Hippocampal Neurogenesis and Myogenesis in the Same Old Mammal.” Oncotarget, 6:11959-11978 (PMID: 26003168).

Public Summary of Scientific Progress: The central mission of our Berkeley CIRM Shared Stem Cell Facility (SSCF) is to provide our East Bay users with knowledge, expertise, training, and equipment to advance scientific knowledge of human embryonic, induced pluripotent, and adult stem cells. The Flow Cytometry Facility in 461 Li Ka Shing houses the CIRM funded cell sorter. The B129 Stanley suite contains cell culture hoods, four incubators, and other essential lab equipment. The SSCF also has an automated wide field ImageXpress Micro, and two analytical flow cytometers, and a 2-photon and confocal fluorescence microscope. In B203 Stanley, there is a BioRad BioPlex MAGPIX reader for magnetic bead–based multiplexed immunoassays. We have reached a steady state usage of ~90 students and postdoctoral fellows from the laboratories of 25 PIs. Based on the interests of the users, we have continued in Year 7NCE to maintain state of the art instrumentation for the research community by using some of Year6 funds to help bridge salary and instrument service contract funding from CIRM funding to alternatives. We are committed to continuing operating this facility, as it has become an important resource to the UC Berkeley research community. The previously highlighted PIs continue to have success using SSCF instrumentation and we would like to highlight publications by researchers who have made significant advancements to their research. Publications: Hojeong Jeon, Sangmo Koo, Willie Mae Reese, Peter Loskill, Costas P Grigoropoulos, Kevin E Healy. (2015) “Directing cell migration and organization via nanocrater-patterned cell-repellent interfaces.” Nature Materials 14(9): 918-923 (PMCID: PMC4545687). Kevin M Tharp, Amit K Jha, Judith Kraiczy, Alexandra Yesian, Grigory Karateev, Riccardo Sinisi, Elena A Dubikovskaya, Kevin E Healy, Andreas Stahl. (2015) “Matrix-Assisted Transplantation of Functional Beige Adipose Tissue.” Diabetes 64(11): 3713-3724 (PMID: 26293504). Peter Loskill, Sivan G Marcus, Anurag Mathur, Willie Mae Reese, Kevin E Healy. (2015) “μOrgano: A Lego®-Like Plug & Play System for Modular Multi-Organ-Chips.” PLoS One 10(10): e0139587 (PMID: 26440672). Amit K Jha, Kevin M Tharp, Shane Browne, Jianqin Ye, Andreas Stahl, Yerem Yeghiazarians, Kevin E Healy. (2016) “Matrix metalloproteinase-13 mediated degradation of hyaluronic acid-based matrices orchestrates stem cell engraftment through vascular integration.” Biomaterials 89: 136-147 (PMID: 26967648). Eda I Altiok, Jorge L Santiago-Ortiz, Felicia L Svedlund, Aline Zbinden, Amit K Jha, Deepika Bhatnagar, Peter Loskill, Wesley M Jackson, David V Schaffer, Kevin E Healy. (2016) “Multivalent hyaluronic acid bioconjugates improve sFlt-1 activity in vitro.” Biomaterials 93: 95-105 (PMID: 27086270). Zheng. B., M.P. von See, E. Yu, B. Gunel, K. Lu, T. Vazin, D.V. Schaffer, P.W. Goodwill, and S.M. Conolly (2016) “Quantitative Magnetic Particle Imaging Monitors the Transplantation, Biodistribution, and Clearance of Stem Cells in Vivo.” Theranostics, 6:291-301 (PMID 26909106). Luque, T., M.S. Kang, D.V. Schaffer, and S. Kumar (2016) “Microelastic Mapping of the Rat Dentate Gyrus.” Royal Society Open Science, 3:150702 (PMID 27152213). Chen, S. Y.S. Na, M.E. Todhunger, S. Hsiao, A.W. Bremer, O.J. Scheideler, P.R. Bomdica, M.M. Haharbiz, Z.J. Gartner, D.V. Schaffer (2016) “Interrogating Stem Cell Fate Decisions with High-throughput Arrays of Cellular Communities.” Nature Communications, 7:10309 (PMID 26754526). Zheng, V., T. Vazin, P.W. Goodwill, A. Conway, A. Verma, E. Ulku Saritas, D.V. Schaffer, and S.M. Conolly (2015) “Magnetic Particle Imaging Tracks the Long-term Fate of in Vivo Neural Cell Implants with High Image Contrast.” Scientific Reports, 5:14055 (PMID: 26358296). Han, B.W., H. Layman, N.A. Rode, A. Conway, D.V. Schaffer, N.J. Boudreau, W.M. Jackson, and K.E. Healy (2015) “Multivalent Conjugates of Sonic Hedgehog Accelerate Diabetic Wound Healing.” Tissue Engineering, 21:2366-2378 (PMID 26154888). Sia J, Sun R, Chu J, Li S. (2016) “Dynamic culture improves cell reprogramming efficiency.” Biomaterials 92:36-45 (PMID: 27031931). Lee K, Rafi M, Feng X, Tang R, Lingampalli N, Aran K, Murthy N. (2015) “In vivo delivery of transcription factors with multifunctional oligonucleotides.” Nature Materials 14(7), 701-706 (PMID: 25915034). Lee K, Yu P, Lingampalli N, Tang R, Kim H, Murthy N. (2015) “Peptide-enhanced mRNA transfection in cultured mouse cardiac fibroblasts and direct reprogramming towards cardiomyocyte-like cells.” Int J Nanomedicine 10, 1841-1854 (PMID: 25834424). Kunwoo Lee, Michael Conboy, Fuguo Jiang, Hyun Jin Kim, Mark A. Dewitt, Vanessa A. Mackley, Wen-chin Huang, Kevin Chang, Anirudh Rao, Colin Skinner, Tamanna Shobha, Freeman Lan, Nicolas L. Bray, Song Li, Jacob E. Corn, Kazunori Kataoka, Jennifer A. Doudna, Irina Conboy, Niren Murthy “Gold nanoparticle-mediated in vivo delivery of CRISPR/Cas9 ribonucleoprotein and donor DNA.” (Submitted).