BioNanotechnology Seminar Series - Spring 2012

Nanotechnology Approaches For Regulating Stem/Cancer Cell Fate

Dr. KiBum Lee, Rutgers
Department of Chemistry and Chemical Biology

Tuesday, April 24, 2012
1000 MNTL, 12:00 - 1:00 PM

Abstract: This talk will focus on the interface of micro-/nano science and cell biology. Even though cell fate (e.g. stem cell differentiation and cancer cell apoptosis) is regulated by interactions with microenvironment cues and intrinsic cellular programs, understanding the functions of microenvironments and manipulating gene expression in stem/cancer cells are hampered by limitations of conventional methods and the lack of extensive knowledge of multiple regulatory signals. If the complex cell behaviors are to be fully investigated, both approaches from nanotechnology—the "top-down" patterning of extracellular matrix (ECM) and signal molecules in combinatorial ways (e.g. ECM compositions, pattern geometry, pattern density and gradient patterns), and the "bottom-up" synthesis of multifunctional nanoparticles and their surface modification with specific signal molecules—should be combined synergistically. To address the aforementioned challenge, our research mainly focuses on three approaches: i) development of combinatorial arrays of microenvironmental signal molecules for investigating cell behaviors; ii) synthesis and utilization of multifunctional nanoparticles as chemotherapeutic reagents against glioblastoma multiforme (GBM); and iii) development of a microfluidic assay platform to identify the optimal conditions for stem cell differentiation and self-renewal.

More specifically, we have applied the combinatorial signal arrays to study the temporal/spatial effect of microenvironmental cues on adhesion, growth, differentiation of functional cells (e.g. neural stem cells and glioblastoma cells). Furthermore, novel synthetic approaches for anti-cancer drugs [e.g. Erlotinib and Histone deacetylase inhibitors (HDAC inhibitors)] and modified siRNA to be linked with nanoparticles have been developed. In parallel research efforts, we have developed a high throughput screening method based on microfluidics to study human embryonic stem cell (hESCs) responses toward multiple microenvironmental cues at the single cell level. In this talk, a summary of the most updated results from these efforts and future directions will be discussed.

 

 

Previous Spring 2012 Seminars

 

Nanostructured Surface for Enhanced Fluorescence Cell Imaging

Austin Hsiao, Bioengineering

Tuesday, April 17, 2012
1000 MNTL, 12:30 - 1:00 PM

Abstract: We have created a nanostructure substrate on which three-dimensional confocal fluorescence cell-imaging sensitivity is amplified for cell membrane and cytoplasm. The nanostructure substrate was fabricated using a plasma etching technique to create a randomized array of nano-pillars. Confocal fluorescence imaging of Chinese Hamster Ovarian cells showed at least 10x enhancement of fluorescence signal from labeled cell membrane and cell cytoplasm compared to traditional glass slide. The observed enhancement may be due to enhanced scattering from the nanostructures and the excitation of localized surface plasmon. Additional fluorescence labels and cell types are under investigation for fluorescence enhancement

 

Human Colon Carcinoma Cells on Gels: Spatial Confine-ment and Mechanosensing

M. Yakut Ali, Mechanical Science and Engineering

Tuesday, April 17, 2012
1000 MNTL, 12:00 - 12:30 PM

Abstract: New Mounting experimental evidences suggest that cells can sense and respond to mechanical cues (i.e. substrate stiffness) and geometric cues in 2D and 3D culture models. Hence, engineered mechanical microenvironment can enable new in vitro mechano-biology studies, which are otherwise complicated to realize. In the first part of my presentation, I will talk about a novel approach to micropattern extracellular matrix (ECM) proteins on 2D polyacrylamide (PA) hydrogels and consequently obtain spatially defined cell culture with precision [1]. This method provides an excellent and robust tool to study the coupled effect of mechanical cues and geometric cues on 2D polyacrylamide hydrogels.

In the second part, I will discuss on the in-vitro metastasis of human colon carcinoma cells driven solely by the elasticity of the substrate [2]. In the work, our group has shown that human colon carcinoma (HCT-8) cells can exhibit a dissociative, metastasis-like phenotype (MLP) in vitro when cultured on ECM coated substrates with appropriate mechanical stiffness (physio-logically relevant 21–47 kPa), but not on very soft (1 kPa) and very stiff (3.6 GPa) substrates [2]. However, the role of cell-ECM adhesions (integrins) or cell-cell adhesions (E-cadherin) in the mechanosensing process remain elusive. Hence, we cultured the HCT-8 cells on E-cadherin coated soft substrates of appropriate mechanical stiffness (21 kPa) to mimic cell-cell interactions. Interestingly, HCT-8 cells show the distinct dissociative phenotype on E-cadherin coated substrates as well. These results suggest that the E-cadherin, but not the integrins, is the dominant mechanosensor for the MLP on soft substrates. The inhibition of the MLP on E-cadherin coated substrates by blebbistatin, a potent inhibitor of non-muscle myosin II ATPase, indicates that the intracellular forces are involved in the initiation of the MLP. In addition, the actin cytoskeletal structure and nuclei deformation on both fibronectin (ECM) and E-cadherin coated substrates are also investigated before and after the MLP using laser scanning confocal microscopy [3].

 

Multilayered Metal Dielectric nanoLAMPS for Highly Sensitive Multiplexed Detection of Pre-metastatic Cancer Cells

Sean Sivapalan, Materials Science and Engineering

Tuesday, April 10, 2012
1000 MNTL, 12:30 - 1:00 PM

Abstract: We fabricate a new class of nanoLAMPS based on recent theoretical designs by the Bhargava group. A variety of methods are adapted to create alternating metal and dielectric layered nanoparticles with embedded Raman reporter molecules. These reporter molecules are then subject to the intense electromagnetic field within each individual nanoparticle. Raman spectroscopy is used to characterize these electromagnetic field enhancements.

 

Fabrication and Characterization of Multi-Compartment Collagen-Glycosaminoglycan Scaffolds for Tissue Regeneration

Daniel Weisgerber, Materials Science and Engineering

Tuesday, April 10, 2012
1000 MNTL, 12:00 - 12:30 PM

Abstract: We are working to create multi-compartment scaffolds as regenerative templates for multi-tissue interfaces such as the tendon to bone junction (TBJ). Collagen-glycosaminoglycan (CG) scaffolds have previously been successfully employed in a variety of soft tissue regeneration applications. Recently strategies for creating mineralized CG (CGCaP) scaffold variants along with fabrication methods to create layered osteochondral scaffolds have been demonstrated. Here I will discuss methods for the fabrication and characterization of both single and multi-compartment scaffolds targeted for the TBJ. Following production, both histology and microCT imaging approaches were employed to investigate scaffold microstructural features. In the case of the mineralized CGCaP scaffolds, mineral content as well as phase were determined via DMMB and hydroxyproline assays as well as x-ray diffraction approaches, respectively. Bulk mechanical and permeability characterization methods were applied and then compared with ultrasound elastography approaches targeting non-destructive analysis of individual scaffold compartments within the heterogeneous multi-compartment scaffolds. Finally, the biocompatibility of hMSCs within CG and CGCaP scaffold compartments was investigated using bioactivity, gene expression, and functional metrics as a precursor to both in vivo testing as well as subsequent work creating fully interpenetrating TBJ grafts that can drive multi-lineage MSC differentiation in a spatially-selective manner.

 

CYP17: Gatekeeper to Androgen Biosynthesis

Michael Gregory, Biochemistry

Wednesday, March 28, 2012
1000 MNTL, 12:30 - 1:00 PM

Abstract: Human Cytochrome P45017A1 (CYP17) catalyzes the 17 alpha-hydroxylation of pregnenolone and progesterone as well as the subsequent 17,20 carbon-carbon lyase chemistry of its hydroxylated products. CYP17 function plays a central role in human steroid hormone biosynthesis, and its activity is absolutely essential for the formation of androgens. Thus, inhibition of CYP17 has recently been exploited in the treatment of androgen dependent malignancies. Through application of nanotechnology and biophysical tools, we have identified novel characteristics of CYP17 chemistry that may guide development of the next generation of mechanism-based inhibitors.

 

Cationic, Helical Polypeptide Based Gene Delivery for IMR90 Fibroblasts and Human Embryonic Stem Cells

Jonathan Yen, Bioengineering

Wednesday, March 28, 2012
1000 MNTL, 12:00 - 12:30 PM

Abstract: New diblock copolymers consisting of poly(ethyleneglycol)-block-poly(?-(4-(piperidin-1-yl)ethanaminomethylbenzyl)-L-glutamate), PEG-b-PVBLG-8 were synthesized for the effective transfection of hard to transfect cells like IMR90, human fetal lung fibroblasts and human embryonic stem cells (hESC). The diblock copolymers maintain a water soluble cationic helical polypeptides block, with the PEG reducing toxicity of the PVBLG-8. It also allows the copolymer to maintain its high helical content, without dramatically compromising its ability to condense the DNA. Polymers with varying degrees of polymerization of PVBLG-8 were synthesized to identify the most effective vehicle for gene delivery.

The diblock polymer with a polymerization degree of 287 demonstrates greater transfection efficiency and lower toxicity in IMR90 cells when compared with the commercial Lipofectamine 2000, giving a transfection efficiency of 21.4% and about 80% cell viability. We also demonstrate the potential for the polymer to be used in hESC. In summary, this polymer demonstrates the effective use of peptide based delivery vehicles for gene delivery

 

Nanotechnology: Ethical Implications

Dr. Christopher Hook, Mayo Clinic

Thursday, March 15, 2012
12:00-1:00 p.m. at Carle Forum Pollard Auditorium

Dr. Christopher Hook is a hematologist/oncologist and ethicist who practices at Mayo Clinic in Rochester, MN. He lectures widely and we have scheduled him to come to Carle on Thursday, March 15th from noon till 1 pm, to lecture at our Bioethics Seminar. Dr. Hook is planning to speak to our seminar series audience on the topic "Nanotechnology" with a focus on cancer.

 

Direct Write Assembly of 3-D Microperiodic Hydrogel Scaffolds For Stem Cell Culture and Tissue Engineering

Lucas Osterbur, Materials Science and Engineering

Tuesday, February 21, 2012
1000 MNTL, 12:30 - 1:00 PM

Abstract: Stem cells have demonstrated remarkable potential for tissue engineering and regenerative medicine through their ability for self-renewal and controlled differentiation. However, the development of optimal three dimensional (3D) scaffolds that overcome the limitations of 2D cell culture remains an elusive challenge in stem cell research. Here, we demonstrate the use of direct-write assembly to create tailored hydrogel scaffolds composed of 3D microperiodic architectures for applications with two types of stem cells, human embryonic stem cells (hESCs) and mesenchymal stem cells (mSCs). Photo-polymerizable poly(hyaluronic acid) (pHA) is synthesized as an ink for scaffold fabrication due to its demonstrated biocompatibility, biodegradability, and tailorable functionality. hESCs are cultured on 3D pHA scaffolds as an alternative to traditional culture methods based on 2D substrates with mouse embryonic fibroblast (MEF) feeder layers. Initial results demonstrate the potential of pHA scaffolds as chemically defined, biomimetic systems for proliferation of hESCs in an undifferentiated state. 3D pHA scaffolds are also under development for repair of articular defects using porcine, adipose-derived mSCs. Cartilage developed from mSCs in vitro on these 3D scaffolds exhibits a morphology that is distinct from that observed on 2D polystyrene controls. Animal protocols are under review for implantation of 3D pHA scaffolds seeded with mSCs.

 

Mesenchymal Stem Cells Contribute to Vascular Growth in Skel-etal Muscle in Response to Eccentric Exercise   |   View Presentation

Heather Huntsman, Kinesiology and Community Health

Tuesday, February 21, 2012
1000 MNTL, 12:00 - 12:30 PM

Abstract: Tissue health is critically dependent on vascularization to support growth and function following injury. In addition to multiple implications in the regenerative potential of other tissues, mesenchymal stem cells have been shown to promote vessel formation both in vivo and in vitro. Our recent work has established that transgenic overexpression of the α7 integrin in skeletal muscle (α7Tg ) can enhance the presence of Sca-1+CD45- mesenchymal stem cells (mMSCs) which facilitate myogenesis. PURPOSE: The purpose of this study was to determine the extent to which angiogenesis is increased in α7Tg muscle following acute or repeated bouts of eccentric exercise and elucidate a role for mMSCs in this event. METHODS: mMSCs were isolated from α7Tg muscle by fluorescent activated cell sorting (FACS) and pericyte markers were examined by flow cytometry. Wild type (WT) and α7Tg mice (5 wk) were subjected to single or multiple bouts (3x/wk, 4 wks) of downhill running exercise. Additionally, DiI-labeled mMSCs were injected into WT mice. Measures of angiogenesis and vessel growth were evaluated by immunohistochemistry. RESULTS: A large percentage of isolated mMSCs were positive for pericyte markers. DiI-labeled mMSCs injected into WT muscle migrated to the vascular niche and incorporated directly into vessels. Although capillary:fiber ratio, capillary density and tortuosity index did not increase, the number of large vessels was significantly increased in α7Tg muscle following single and repeated bouts of exercise (p<0.05; 3-fold for repeated bouts) and in WT muscle receiving mMSC transplantation (P<0.05; 48%). CONCLUSION: This study demonstrates that mMSCs contribute to vascular growth in skeletal muscle in response to eccentric exercise, and that this adaptation is coordinated with increased myogenesis previously reported. Well-orchestrated responses similar to this may be a key mechanism in the successful regeneration of several tissue types.

 

Analyzing Protein Complexes from Mammalian Cells at the Single Molecular Level  |   View Presentation

Yang Kevin Xiang, Molecular and integrative Physiology

Tuesday, February 7, 2012
1000 MNTL, 12:00 - 1:00 PM

Abstract: Proteins perform most cellular functions in macromolecular complexes. Quantitative information on macromolecular complexes is scarce. The same protein often participates in different complexes to exhibit diverse functionality. Current ensemble approaches of identifying cellular protein interactions cannot reveal physiological permutations of these interactions. I will discuss a single-molecule pull-down (SiMPull) assay that combines the principles of a conventional pull-down assay with single-molecule fluorescence microscopy and enables direct visualization of individual cellular protein complexes. SiMPull can reveal how many proteins and of which kinds  are  present  in  the in vivo complex, as we show using protein kinase A.  We then demonstrate a wide applicability to various signaling proteins found in the cytosol, membrane and cellular organelles, and to endogenous protein complexes from animal tissue extracts. The pulled-down proteins are functional and are used, without further processing, for single-molecule biochemical studies. SiMPull should provide a rapid, sensitive and robust platform for analyzing protein assemblies in biological pathways.

 

Exploring Academic Collaboration with the University of Cape Coast, Ghana with the Global Health Initiative at UIUC   |   View Presentation

Erich Lidstone, Department of Bioengineering

Tuesday, January 24, 2012
1000 MNTL, 12:30 - 1:00 PM

Abstract: The University of Illinois at Urbana-Champaign has a longstanding history of excellent research with regard to science and engineering, and the Global Health Initiative (GHI) at UIUC hopes to apply this research expertise toward the development of low-cost and resource-limited diagnostic tools and therapeutic solutions. In an exploratory two-week visit to Accra and Cape Coast, Ghana, students, faculty, and staff of the GHI were able to learn about and explore many aspects of Ghana's health care system, interacting with clinicians, social health workers, and patients at a number of hospitals, clinics, and homes. Students were able to meet also with a number of research faculty at the University of Cape Coast (UCC) to discuss how UIUC might help to learn from and collaborate with UCC. In visits to hospitals, students were able to learn about necessary and high use equipment directly from the technicians using them, as well as the laboratory facility directors. Finally, the welcoming community at Cape Coast helped students to learn about the state of medical care in Ghana and points which might benefit from improvement through new technology, greater resources, or greater supply. Please join us to learn about developments in Cape Coast as well as about how to get involved with future events coordinated by the Global Health Initiative at UIUC.

 

DNA Mediated Synthesis of Novel Gold Nanoflowers for Diagnostic and Therapeutic Applications   |   View Presentation

Brian N.Y. Wong, Department of Materials Science and Engineering

Tuesday, January 24, 2012
1000 MNTL, 12:00 - 12:30 PM

Abstract: My research is focused at exploring the applications of a new class of gold nanoflower structure that exhibits interesting cellular uptake properties, tunable size and optical properties, and an exceptionally simple synthesis protocol. These properties make it an ideal platform for cancer diagnosis, imaging, and as a platform for therapy. My work has been focused on optimizing the size of nanoflowers while maintaining the functional DNA aptamer that is responsible for cancer recognition and demonstrating selective uptake in vitro. The goal of the project is a complete cancer theragnostic platform that can be used to image, diagnose, and treat a variety of diseases by choosing the appropriate apatmer and nanoflower size.