Synthetic Human Embryology

Mechanobiology of Human Pluripotent Stem Cells

Cell Mechanics and Mechanotransduction

 

Thrust 1: Synthetic Human Embryology

Human peri- and post-implantation embryonic development remains mysterious, given the scarce human embryo specimens and limited numbers of non-human primate embryos. Recent advances in the generation of human embryo-like structures from human pluripotent stem cells (hPSCs) have sparked great interest in using such synthetic human tissues for advancing human embryology, embryo toxicology, and reproductive medicine.  In this research, we have leveraged the developmental potential and self-organizing properties of hPSCs in conjunction with biomimetic culture systems for developing different synthetic human embryo-like structures.

Specifically, we have developed the first hPSC-based, synthetic embryological model of human post-implantation development that recapitulates multiple embryogenic events including amniotic cavity formation, amnion-epiblast patterning, and primitive streak formation.  We have further shown BMP/SMAD signaling as an autonomous developmental mechanism to provide an early asymmetrically distributed patterning signal to the developing embryoid. Together, our findings provide insight into previously inaccessible but critical embryogenic events in post-implantation human development. Going forward, continuous development of the synthetic human development model will provide a synthetic embryological platform to complement scarce in vivo and ex vivo work that uses live human and non-human primate embryos, thereby opening previously undescribed avenues to advance human embryology, embryo toxicology, and reproductive medicine.


During the first three weeks of human embryo development, a few key embryogenic events occurs, including blastocyst formation, implantation and gastrulation. Gastrulation is the fundamental organizational event that generates the basic body plan and provides the building blocks for all the tissues in the human embryo.

Current students: Yi Zheng, Xufeng Xue, Sajedeh Nasr Esfahani
Collaborators: Deborah L. Gumucio.

References:

  1. Yue Shao, Kenichiro Taniguchi, Ryan F. Townshend, Toshio Miki, Deborah L. Gumucio, and Jianping Fu. A pluripotent stem cell-based model for post-implantation human amniotic sac development. Nature Communications, vol. 8, 208, 2017. [PDF | Supplemental Materials]
  2. Yue Shao, Kenichiro Taniguchi, Katherine Gurdziel, Ryan F. Townshend, Xufeng Xue, Koh Meng Aw Yong, Jianming Sang, Jason R. Spence, Deborah L. Gumucio, and Jianping Fu. Self-organized amniogenesis by human pluripotent stem cells in a biomimetic implantation-like niche. Nature Materials, vol. 16, pp. 419-425, 2017. [PDF | Supplemental Materials]
  3. Kenichiro Taniguchi, Yue Shao, Ryan F. Townshend, Yu-Hwai Tsai, Cynthia J. DeLong, Shawn A. Lopez, Srimonta Gayen, Andrew M. Freddo, Deming J. Chue, Dennis J. Thomas, Jason R. Spence, Benjamin Margolis, Sundeep Kalantry, Jianping Fu, K. Sue O’Shea, and Deborah L. Gumucio. Lumen formation is an intrinsic property of isolated human pluripotent stem cells. Stem Cell Reports, vol. 5, pp. 954-962, 2015. [PDF | Supplemental Materials]

Thrust 2: Mechanobiology of Human Pluripotent Stem Cells

Research on human pluripotent stem cells (hPSCs) has expanded rapidly over the last two decades, owing to the promise of hPSCs for applications in regenerative medicine, disease modeling, and developmental biology studies. Most hPSC studies have so far focused on identifying extrinsic soluble factors, intracellular signaling pathways, and transcriptional networks involved in regulating hPSC behaviors. In our research, we have uniquely focused on a high-risk, high-payoff concept to investigate an emerging functional connection between mechanobiology and some critical questions in the field of hPSCs, including pluripotency, directed differentiation, cell reprogramming and transdifferentiation, functional maturation, and aging. We are also exploring intracellular molecular mechanisms underlying mechanosensitive properties of hPSCs using cell biology and systems biology methods in conjunction with our bioengineering and synthetic micromechanical tools. Our research on mechanobiology of hPSCs will potentially enable drastic advances in large-scale production of hPSCs and their derivatives and contribute significantly to future cell-based regenerative therapies and disease modeling. 

So far, our research has unambiguously unraveled the mechanosensitive properties of hPSCs and their roles in directed neural differentiation and subtype specification of hPSCs. Our mechanistic work has further led to the discovery of an intervened regulatory network emerged from converging and reinforcing signal integration of TGF-β, WNT, Hippo, Rho-GTPase, and the actomyosin cytoskeleton that forms a molecular framework required for contextual, integrated responses of hPSCs.

In the future, we will extend our mechanobiology research into the exciting field of cell reprogramming and transdifferentiation related to different types of neural cells including neural precursor cells. We will continue to leverage micro/nanoengineering and systems biology tools in conjunction with new discoveries at the interface of mechanotransduction, epigenetics, and classic signaling networks to engineer and control transcriptional landscapes and thus achieve superior cell reprogramming and transdifferentiation efficiencies toward specific, precise neuronal subtypes most susceptible to disease and traumatic injury.


Micro/nanoengineering ex vivo stem cell niche

Current students: Xufeng Xue
Collaborators: Paul H. Krebsbach, Eva Feldman, Andres J. Garcia.

References:

  1. Yubing Sun and Jianping Fu. Harnessing mechanobiology of human pluripotent stem cells for regenerative medicine. ACS Chemical Neuroscience, vol. 5, pp. 621-623, 2014. [PDF]
  2. Yubing Sun, Koh Meng Aw Yong, Luis G. Villa-Diaz, Xiaoli Zhang, Weiqiang Chen, Renee Philson, Shinuo Weng, Haoxing Xu, Paul H. Krebsbach, and Jianping Fu. Hippo/YAP-mediated rigidity-dependent motor neuron differentiation of human pluripotent stem cells. Nature Materials, vol. 13, pp. 599-604, 2014. [PDF | Supplemental Materials
  3. Yubing Sun and Jianping Fu. Mechanobiology: A new frontier for human pluripotent stem cells. Integrative Biology, vol. 5, pp. 450-457, 2013. [PDF]
  4. Yubing Sun, Luis G Villa-Diaz, Raymond Hiu-Wai Lam, Weiqiang Chen, Pasul H. Krebsbach, and Jianping Fu. Matrix mechanics regulates fate decisions of human embryonic stem cells. PLoS ONE, vol. 7, e37178, 2012. [PDF]
  5. Weiqiang Chen, Luis G Villa-Diaz, Yubing Sun, Shinuo Weng, Raymond Hiu-Wai Lam, Lin Han, Rong Fan, Paul H. Krebsbach, and Jianping Fu. Nanotopography influences adhesion, spreading, and self-renewal of human embryonic stem cells. ACS Nano, vol. 6, pp. 4094-4103, 2012. [PDF | supplemental materials]

Thrust 3: Cell Mechanics and Mechanotransduction

External forces and matrix mechanics play a key role in the regulation of cell function. Cells sense and response to external forces and changes in matrix mechanics by modulating their endogenous cytoskeletal contractility, balanced by external forces or resistant forces generated by the deformation of the extracellular matrix (ECM). Thus, it appears that cells are mechano-sensitive and –responsive to mechanical forces and matrix mechanics through a modulated delicate force balance between the endogenous cytoskeletal contractility and external mechanical forces transmitted across the cell-ECM adhesions. Indeed, such tensional homeostasis in the intracellular cytoskeleton (CSK) has a key role in the regulation of basic cellular functions, such as cell proliferation, apoptosis, adhesion, and migration. Deregulation of the tensional homeostasis in cells contributes to the pathogenesis of several human diseases, such as atherosclerosis, osteoarthritis and osteoporosis, and cancer.

The force balance transmitted across the mechanical continuum of ECM-integrin-CSK can regulate integrin-mediated adhesion signaling (such as FAK and Src signaling) to coordinate downstream integrated cell function. These biophysical signals are sensed at the adhesion sites in which integrins provide the mechanical linkage between the ECM and the actin CSK. Exposure of cells to mechanical strain, fluid shear stress, or plating cells on substrates with varying elastic moduli, will activate integrins, which promote recruitment of scaffold and signaling proteins to strengthen adhesions and to transmit biochemical signals into the cell. These mechanotransduction pathways establish positive feedback loops in which integrin engagement activates acto-myosin CSK contractility, which in turn reinforces adhesions. Thus, the level of CSK contractility generated inside the cell is directly proportional to the adhesion strength and the matrix elastic modulus and dictates the cellular responses of cells.

To aid in the mechanistic investigation of mechanotransduction, we have established different micromechanical tools and systems that allow for quantitative controls and real-time measurements of mechanical stimuli and cellular biomechanical responses. Our unique technology developments are particularly useful for investigations of mechanotransduction centering on the ECM-integrin-CSK signaling axis to generate quantitative descriptions of the functional relations between matrix mechanics, external forces, cytoskeletal contraction, cell stiffness, and adhesion signaling and morphogenesis.


Micromechanical tools for precise control and measurement of mechanical stimuli and responses

Current students: Xufeng Xue
Collaborators: Krishna Garikipati, Cheri X. Deng.

References:

  1. Shinuo Weng, Yue Shao, Weiqiang Chen, and Jianping Fu. Mechanosensitive subcellular rheostasis drives emergent single-cell mechanical homeostasis. Nature Materials, vol. 15, pp. 961-967, 2016. [PDF | Supplemental Materials]
  2. Di Chen, Yubing Sun, Madhu S. R. Gudur, Yising Hsiao, Ziqi Wu, Jianping Fu, and Cheri X. Deng. Two bubble acoustic tweezing cytometry for biomechanical probing and stimulation of cells. Biophysical Journal, vol. 108, pp. 32-42, 2015. [PDF]
  3. Yue Shao, Jennifer M. Mann, Weiqiang Chen, and Jianping Fu. Global architecture of F-actin cytoskeleton regulates cell shape-dependent endothelial mechanotransduction. Integrative Biology, vol. 6, pp. 300-311, 2014. [PDF | Supplemental Materials]
  4. Zhenzhen Fan, Yubing Sun, Di Chen, Donald Tay, Weiqiang Chen, Cheri X. Deng, and Jianping Fu. Acoustic tweezing cytometry for live-cell subcellular control of intracellular cytoskeleton contractility. Scientific Reports, vol. 3, 2176, 2013. [PDF | Supplemental Materials]
  5. Raymond Hiu-Wai Lam, Shinuo Weng, Wei Lu, and Jianping Fu. Live-cell subcellular measurement of cell stiffness using a microengineered stretchable micropost array membrane. Integrative Biology, vol. 4, pp. 1289-1298, 2012. [PDF]
  6. Raymond Hiu-Wai Lam, Yubing Sun, Weiqiang Chen, and Jianping Fu. Elastomeric microposts integrated into microfluidics for flow-mediated endothelial mechanotransduction analysis. Lab on a Chip, vol. 12, pp. 1865-1873, 2012. [PDF | supplemental materials]
  7. Jennifer M. Mann, Raymond Hiu-Wai Lam, Shinuo Weng, Yubing Sun, and Jianping Fu. A silicone-based stretchable micropost array membrane for monitoring live-cell subcellular cytoskeletal response. Lab on a Chip, vol. 12, pp. 731-740, 2012. [PDF | supplemental materials]
  8. Shang-You Tee, Jianping Fu, Christopher S. Chen, and Paul A. Janmey. Cell shape and substrate rigidity both regulate cell stiffness. Biophysical Journal, vol. 100, pp. L25-27, Mar. 2011. [PDF | supplemental materials]
  9. Michael T. Yang, Jianping Fu, Yang-Kao Wang, Ravi A. Desai, and Christopher S. Chen. Assaying stem cell mechanobiology on microfabricated elastomeric substrates with geometrically modulated rigidity. Nature Protocols, vol. 6, pp. 187-213, 2011. [PDF]
  10. Jianping Fu, Yang-Kao Wang, Michael T. Yang, Ravi A. Desai, Xiang Yu, Zhijun Liu, and Christopher S. Chen. Mechanical regulation of stem cell function using geometrically modulated elastomeric substrates. Nature Methods, vol. 7, pp.733-736, 2010. [PDF | supplemental materials]