Dr. Ying Chang is an Adjunct Professor of the Department of Chemical Engineering, an affiliate member of Precision Health and Integrated Diagnostic Center of the School of Medicine, and a Co-Director of the Taiwan-Stanford Partnership program, LEAP, at Stanford University. She is also a Research Fellow at the Genomics Research Center, Academia Sinica, and an Adjunct Professor at the Center of Liquid Biopsy at Kaohsiung Medical University, Taiwan. Formerly, she was an Assistant Professor in the Department of Chemical Engineering and Materials Science, and the Department of Biomedical Engineering at the University of California-Irvine, Irvine, CA. Prior to her academic appointments, Dr. Chang had worked in various industrial R&D laboratories including as a Senior Engineer for the hard drive media at Maxmedia California, San Jose, CA (now Seagate), a Postdoctoral Scientist for the materials design of GeneChip at Affymetrix Corp, Santa Clara, CA (now Thermal Fisher Scientific). Her recent invention in circulating tumor cells platform has led to a startup company, Cellmax Life in 2013. Highlights of her research include integrated nanomaterials, microfluidics, and bioreactors to control stem cell fates for tissue engineering and liquid biopsy for cancer diagnostics and precision medicine. Dr. Chang received her BS from National Taiwan University and PhD from Stanford University in Chemical Engineering.
Adjunct Professor, Chemical Engineering
Co-Director, Taiwan LEAP@Stanford Program (2017 - Present)
BS, National Taiwan University
PhD, Stanford University, Chemical Engineering
Early Detection and Dynamic Changes of Circulating Tumor Cells in Transgenic NeuN Transgenic (NTTg) Mice with Spontaneous Breast Tumor Development.
2021; 13 (13)
BACKGROUND: This study used NeuN transgenic (NTTg) mice with spontaneous breast tumor development to evaluate the dynamic changes of circulating tumor cells (CTCs) prior to and during tumor development.METHODS: In this longitudinal, clinically uninterrupted study, we collected 75 muL of peripheral blood at the age of 8, 12, 16, and 20 weeks in the first group of five mice, and at the age of 32 weeks, the time of tumor palpability, and one week after tumor palpability in the second group of four mice. Diluted blood samples were run through a modified mouse-CMx chip to isolate the CTCs.RESULTS: The CTC counts of the first group of mice were low (1 ± 1.6) initially. The average CTC counts were 16 ± 9.5, 29.0 ± 18.2, and 70.0 ± 30.3 cells per 75 muL blood at the age of 32 weeks, the time of tumor palpability, and one week after tumor palpability, respectively. There was a significant positive correlation between an increase in CTC levels and tumor vascular density (p-value < 0.01). This correlation was stronger than that between CTC levels and tumor size (p-value = 0.076). The captured CTCs were implanted into a non-tumor-bearing NTTg mouse for xenografting, confirming their viability and tumorigenesis.CONCLUSION: Serial CTCs during an early stage of tumor progression were quantified and found to be positively correlated with the later tumor vascular density and size. Furthermore, the successful generation of CTC-derived xenografts indicates the tumorigenicity of this early onset CTC population.
View details for DOI 10.3390/cancers13133294
View details for PubMedID 34209279
Anticlogging Hemofiltration Device for Mass Collection of Circulating Tumor Cells by Ligand-Free Size Selection.
Langmuir : the ACS journal of surfaces and colloids
A new hemofiltration system was developed to continuously capture circulating tumor cells (CTCs) from a large volume of whole blood using a column that was packed with antifouling zwitterionized silica microspheres. The silica microspheres were modified with sulfobetaine silane (SBSi) to inhibit fouling, resist clogging, and give a high surface wettability and prolonged operation time. Packed microspheres with different diameters formed size-controllable interstitial pores that effectively captured CTCs by ligand-free size selection. For optimized performance of the hemofiltration system, operational factors, including the size of microspheres, flow rate, and cross-sectional area of the column, were considered with respect to the removal rate for colorectal cancer cells and the retention rate for white blood cells and red blood cells. The captured CTCs were collected from the column by density sedimentation. A large quantity of colorectal cancer cells was spiked into sheep blood, and the sample was circulated for 5 h with a total operational volume of 2 L followed by collection and culture in vitro. The results showed that the proposed hemofiltration device selectively removed abundant CTCs from in vitro circulatory blood. The viable cells were harvested for amplification and potential applications for precision medicine.
View details for DOI 10.1021/acs.langmuir.0c03613
View details for PubMedID 33689353
Scalable Multilayer Cell Collector to Capture Circulating Tumor Cells with an Unlimited Volume Capacity.
ACS biomaterials science & engineering
2019; 5 (6): 2725-2731
Circulating tumor cells (CTCs) have been suggested as the precursors of metastatic cancer. CTC-based characterization has thus been used to monitor tumor status before the onset of metastasis and has shown to be an independent factor. The low abundance of CTCs, however, makes it challenging to employ CTC as a clinical routine, thus making it impossible to address tumor heterogeneity. Here, we present a cell collection prototype for an efficient capture of CTCs from a large volume of body fluids such as blood. An antibody-PEG modified multilayer matrix column is engineered and connected to an apheresis-based circulation system. This setup allows us to capture CTCs repetitively from an unlimited sample volume through the circulation system, thereby increasing the capture count. Compared to conventional CTC capturing devices where the sample handling is generally limited to 1-10 mL, our collector is able to handle a wide range of fluidic sample (40-2000 mL) at a high flow rate (400 mL/min). By processing 90 min in circulation, we obtained an average capture efficiency of at least 75% for the colorectal cancer cell line HCT116 spiked in either 40-200 mL of buffer solution or 40 mL of a whole blood sample. This result highlights a possibility to construct personalized CTC libraries through high-throughput CTC collection for the study of tumor heterogeneity in precision medicine.
View details for DOI 10.1021/acsbiomaterials.9b00315
View details for PubMedID 33405604
- Scalable Multilayer Cell Collector to Capture Circulating Tumor Cells with an Unlimited Volume Capacity ACS BIOMATERIALS SCIENCE & ENGINEERING 2019; 5 (6): 2725–31
- Construction of Cell-Extracellular Matrix Microenvironments by Conjugating ECM Proteins on Supported Lipid Bilayers FRONTIERS IN MATERIALS 2019; 6
Random and aligned electrospun PLGA nanofibers embedded in microfluidic chips for cancer cell isolation and integration with air foam technology for cell release
JOURNAL OF NANOBIOTECHNOLOGY
2019; 17: 31
Circulating tumor cells (CTCs) comprise the high metastatic potential population of cancer cells in the blood circulation of humans; they have become the established biomarkers for cancer diagnosis, individualized cancer therapy, and cancer development. Technologies for the isolation and recovery of CTCs can be powerful cancer diagnostic tools for liquid biopsies, allowing the identification of malignancies and guiding cancer treatments for precision medicine.We have used an electrospinning process to prepare poly(lactic-co-glycolic acid) (PLGA) nanofibrous arrays in random or aligned orientations on glass slips. We then fabricated poly(methyl methacrylate) (PMMA)-based microfluidic chips embedding the PLGA nanofiber arrays and modified their surfaces through sequential coating with using biotin-(PEG)7-amine through EDC/NHS activation, streptavidin (SA), and biotinylated epithelial-cell adhesion-molecule antibody (biotin-anti-EpCAM) to achieve highly efficient CTC capture. When combined with an air foam technology that induced a high shear stress and, thereby, nondestructive release of the captured cells from the PLGA surfaces, the proposed device system operated with a high cell recovery rate.The morphologies and average diameters of the electrospun PLGA nanofibers were characterized using scanning electron microscopy (SEM) and confocal Raman imaging. The surface chemistry of the PLGA nanofibers conjugated with the biotin-(PEG)7-amine was confirmed through time-of-flight secondary ion mass spectrometry (ToF-SIMS) imaging. The chip system was studied for the effects of the surface modification density of biotin-(PEG)7-amine, the flow rates, and the diameters of the PLGA nanofibers on the capture efficiency of EpCAM-positive HCT116 cells from the spiked liquid samples. To assess their CTC capture efficiencies in whole blood samples, the aligned and random PLGA nanofiber arrays were tested for their abilities to capture HCT116 cells, providing cancer cell capture efficiencies of 66 and 80%, respectively. With the continuous injection of air foam into the microfluidic devices, the cell release efficiency on the aligned PLGA fibers was 74% (recovery rate: 49%), while it was 90% (recovery rate: 73%) on the random PLGA fibers, from tests of 200 spiked cells in 2 mL of whole blood from healthy individuals. Our study suggests that integrated PMMA microfluidic chips embedding random PLGA nanofiber arrays may be suitable devices for the efficient capture and recovery of CTCs from whole blood samples.
View details for DOI 10.1186/s12951-019-0466-2
View details for Web of Science ID 000459387900003
View details for PubMedID 30782169
View details for PubMedCentralID PMC6379968
Snail-induced claudin-11 prompts collective migration for tumour progression
NATURE CELL BIOLOGY
2019; 21 (2): 251-+
Epithelial-mesenchymal transition (EMT) is a pivotal mechanism for cancer dissemination. However, EMT-regulated individual cancer cell invasion is difficult to detect in clinical samples. Emerging evidence implies that EMT is correlated to collective cell migration and invasion with unknown mechanisms. We show that the EMT transcription factor Snail elicits collective migration in squamous cell carcinoma by inducing the expression of a tight junctional protein, claudin-11. Mechanistically, tyrosine-phosphorylated claudin-11 activates Src, which suppresses RhoA activity at intercellular junctions through p190RhoGAP, maintaining stable cell-cell contacts. In head and neck cancer patients, the Snail-claudin-11 axis prompts the formation of circulating tumour cell clusters, which correlate with tumour progression. Overexpression of snail correlates with increased claudin-11, and both are associated with a worse outcome. This finding extends the current understanding of EMT-mediated cellular migration via a non-individual type of movement to prompt cancer progression.
View details for DOI 10.1038/s41556-018-0268-z
View details for Web of Science ID 000457468300018
View details for PubMedID 30664792
A two-dimensional immunomagnetic nano-net for the efficient isolation of circulating tumor cells in whole blood.
An immunomagnetic "nano-net" was designed and synthesized for specifically capturing rare cells of interest from mixtures. The nano-net, Ab@Lipo-MNP-GO, consists of conjugated antibody molecules on a lipid coated magnetic nanoparticle-graphene oxide sheet complex. The magnetism, chemical composition, and the morphology of the construct and its precursors were characterized by SQUID, FTIR, TGA, DLS and SEM, to confirm the feasibility of the synthetic steps and the resulting properties suitable for solution phase immuno-recognition for cell capture. When applied to capturing circulating tumor cells (CTCs) in oral, colon and lung cancer clinical patients' blood samples, the nano-net construct exhibited far superior ability whereas conventional immunomagnetic beads in some cases were unable to capture any CTCs, even by increasing the bead concentration. Confocal images showed that the nano-net wrapped around the CTCs while the immunomagnetic beads attached them with point contacts. A stable, patch-like multivalent matrix nano-net was demonstrated to tackle the shortcomings of single point contact of immunomagnetic beads to the target cell. This strategy is universal for any cell separation in complex fluids.
View details for DOI 10.1039/c9nr06256d
View details for PubMedID 31538997
Promoting Multivalent Antibody-Antigen Interactions by Tethering Antibody Molecules on a PEGylated Dendrimer-Supported Lipid Bilayer
2018; 19 (2): 426–37
To efficiently isolate maximal quantity of circulating tumor cells (CTCs) and circulating tumor cell microembolis (CTMs) from patient blood by antibody coated microfluidics, a multifunctional, pegylated polyamidoamine-dendrimers conjugated supported lipid bilayer surface construct was proposed to enhance accessibility of antibody molecules to the antigen molecules on target CTCs. The combination of a hydrated, stretchable dendrimer and a laterally mobile supported lipid bilayer (SLB) provide attached antibody molecules with 2.5-dimensional chain movement, achieving multivalency between the surface antibody and cell antigen molecules. An over 170% enhancement is distinctive for Panc-1 cells that expresses low antigen level. Of seven pancreatic ductal adenocarcinoma patients, an average 440 single CTCs and 90 CTMs were collected in 2 mL of peripheral blood, which were 1.6 times and 2.3 times more, than those captured by the SLB-only microfluidics. In summary, we have demonstrated a material design to enhance multivalent antibody-antigen interaction, which is useful for rare cell enrichment and cancer detection.
View details for PubMedID 29281787
Site-specific antibody modification and immobilization on a microfluidic chip to promote the capture of circulating tumor cells and microemboli
2017; 53 (29): 4152-4155
We design and synthesize EpCAM antibodies with Fc-domain site-specific linkers that allow preferential alignment when coated on microfluidic devices for capturing circulating tumor cells (CTCs) from colorectal cancer patients. The aligned coating is shown to increase the capture efficiency of CTCs and microemboli by 1.6 and 3.0-fold, respectively (both P < 0.05).
View details for DOI 10.1039/c7cc00247e
View details for Web of Science ID 000399728000024
View details for PubMedID 28352905
Microfluidic Capture and Multiplex Immunofluorescence of Circulating Tumor Cells to Identify Cancer of Origin
CIRCULATING TUMOR CELLS: METHODS AND PROTOCOLS
2017; 1634: 1–19
Circulating tumor cells (CTCs) are an important biomarker and their analysis can be considered a form of "liquid biopsy." The purpose of this book chapter is to describe the use of the 4-channel CMx (cells captured in maximum) microfluidic chip, containing special micropatterns coated with an antibody-conjugated supported lipid bilayer (SLB) on its surface, to capture and isolate CTCs from the blood of cancer patients. Captured CTCs are subsequently released by an air foam to an immunofluorescence (IF) staining panel that enables further analysis, including the identification of the primary cancer source of the CTCs.
View details for DOI 10.1007/978-1-4939-7144-2_1
View details for Web of Science ID 000449398600003
View details for PubMedID 28819837
Strategies for Isolation and Molecular Profiling of Circulating Tumor Cells.
Advances in experimental medicine and biology
2017; 994: 43–66
Cancer is the leading cause of death by disease worldwide, and metastasis is responsible for more than 90% of the mortality of cancer patients. Metastasis occurs when tumor cells leave the primary tumor, travel through the blood stream as circulating tumor cells (CTCs), and then colonize secondary tumors at sites distant from the primary tumor. The capture, identification, and analysis of CTCs offer both scientific and clinical benefits. On the scientific side, the analysis of CTCs could help elucidate possible genetic alterations and signaling pathway aberrations during cancer progression, which could then be used to find new methods to stop cancer progression. On the clinical side, non-invasive testing of a patient's blood for CTCs can be used for patient diagnosis and prognosis, as well as subsequent monitoring of treatment efficacy in routine clinical practice. Additionally, investigation of CTCs early in the progression of cancer may reveal targets for initial cancer detection and for anti-cancer treatment. This chapter will evaluate strategies and devices used for the isolation and identification of CTCs directly from clinical samples of blood. Recent progress in the understanding of the significance of both single CTCs and circulating tumor microemboli will be discussed. Also, advancements in the use of CTC-based liquid biopsy in clinical diagnosis and the potential of CTC-based molecular characterization for use in clinical applications will be summarized.
View details for DOI 10.1007/978-3-319-55947-6_2
View details for PubMedID 28560667
Circulating Tumor Cell Count Correlates with Colorectal Neoplasm Progression and Is a Prognostic Marker for Distant Metastasis in Non-Metastatic Patients
2016; 6: 24517
Enumeration of circulating tumor cells (CTCs) has been proven as a prognostic marker for metastatic colorectal cancer (m-CRC) patients. However, the currently available techniques for capturing and enumerating CTCs lack of required sensitivity to be applicable as a prognostic marker for non-metastatic patients as CTCs are even more rare. We have developed a microfluidic device utilizing antibody-conjugated non-fouling coating to eliminate nonspecific binding and to promote the multivalent binding of target cells. We then established the correlation of CTC counts and neoplasm progression through applying this platform to capture and enumerate CTCs in 2 mL of peripheral blood from healthy (n = 27), benign (n = 21), non-metastatic (n = 95), and m-CRC (n = 15) patients. The results showed that the CTC counts progressed from 0, 1, 5, to 36. Importantly, after 2-year follow-up on the non-metastatic CRC patients, we found that those who had ≥5 CTCs were 8 times more likely to develop distant metastasis within one year after curable surgery than those who had <5. In conclusion, by employing a sensitive device, CTC counts show good correlation with colorectal neoplasm, thus CTC may be as a simple, independent prognostic marker for the non-metastatic CRC patients who are at high risk of early recurrence.
View details for DOI 10.1038/srep24517
View details for Web of Science ID 000374042500001
View details for PubMedID 27075165
View details for PubMedCentralID PMC4830949
A High Circulating Tumor Cell Count in Portal Vein Predicts Liver Metastasis From Periampullary or Pancreatic Cancer A High Portal Venous CTC Count Predicts Liver Metastases
2016; 95 (16): e3407
Circulating tumor cells (CTCs) released from a periampullary or pancreatic cancer can be more frequently detected in the portal than the systemic circulation and potentially can be used to identify patients with liver micrometastases. Aims of this study is to determine if CTCs count in portal venous blood of patients with nonmetastatic periampullary or pancreatic adenocarcinoma can be used as a predictor for subsequent liver metastases. CTCs were quantified in portal and peripheral venous blood samples collected simultaneously during pancreaticoduodenectomy in patients with presumed periampullary or pancreatic adenocarcinoma without image-discernible metastasis. Postoperatively patients were monitored for liver metastasis by abdominal magnetic resonance imaging or computed tomography every 3 months for 1 year. Sixty patients with a pathological diagnosis of periampullary or pancreatic adenocarcinoma were included in the study. Multivariate analysis indicated that portal CTC count was a significant predictor for liver metastases within 6 months after surgery. Eleven of 13 patients with a high portal CTCs count (defined as >112 CMx Platform estimated CTCs in 2 mL blood) developed liver metastases within 6 months after surgery. In contrast, only 6 of 47 patients with a low portal CTC count developed liver metastases (P < 0.0001). A value of 112 CMx Platform estimated CTCs had 64.7% sensitivity and 95.4% specificity to predict liver metastases within 6 months after surgery. We concluded that a high CTC count in portal venous blood collected during pancreaticoduodenectomy in patients with periampullary or pancreatic adenocarcinoma without metastases detected by currently available imaging tools is a significant predictor for liver metastases within 6 months after surgery.
View details for DOI 10.1097/MD.0000000000003407
View details for Web of Science ID 000376924800022
View details for PubMedID 27100430
View details for PubMedCentralID PMC4845834
Sensitive and Specific Biomimetic Lipid Coated Microfluidics to Isolate Viable Circulating Tumor Cells and Microemboli for Cancer Detection
2016; 11 (3): e0149633
Here we presented a simple and effective membrane mimetic microfluidic device with antibody conjugated supported lipid bilayer (SLB) "smart coating" to capture viable circulating tumor cells (CTCs) and circulating tumor microemboli (CTM) directly from whole blood of all stage clinical cancer patients. The non-covalently bound SLB was able to promote dynamic clustering of lipid-tethered antibodies to CTC antigens and minimized non-specific blood cells retention through its non-fouling nature. A gentle flow further flushed away loosely-bound blood cells to achieve high purity of CTCs, and a stream of air foam injected disintegrate the SLB assemblies to release intact and viable CTCs from the chip. Human blood spiked cancer cell line test showed the ~95% overall efficiency to recover both CTCs and CTMs. Live/dead assay showed that at least 86% of recovered cells maintain viability. By using 2 mL of peripheral blood, the CTCs and CTMs counts of 63 healthy and colorectal cancer donors were positively correlated with the cancer progression. In summary, a simple and effective strategy utilizing biomimetic principle was developed to retrieve viable CTCs for enumeration, molecular analysis, as well as ex vivo culture over weeks. Due to the high sensitivity and specificity, it is the first time to show the high detection rates and quantity of CTCs in non-metastatic cancer patients. This work offers the values in both early cancer detection and prognosis of CTC and provides an accurate non-invasive strategy for routine clinical investigation on CTCs.
View details for DOI 10.1371/journal.pone.0149633
View details for Web of Science ID 000371735200030
View details for PubMedID 26938471
View details for PubMedCentralID PMC4777486
Clinical Significance of Circulating Tumor Microemboli as a Prognostic Marker in Patients with Pancreatic Ductal Adenocarcinoma
2016; 62 (3): 505–13
Characterization of circulating tumor cells (CTCs) has been used to provide prognostic, predictive, and pharmacodynamic information in many different cancers. However, the clinical significance of CTCs and circulating tumor microemboli (CTM) in patients with pancreatic ductal adenocarcinoma (PDAC) has yet to be determined.In this prospective study, CTCs and CTM were enumerated in the peripheral blood of 63 patients with PDAC before treatment using anti-EpCAM (epithelial cell adhesion molecule)-conjugated supported lipid bilayer-coated microfluidic chips. Associations of CTCs and CTM with patients' clinical factors and prognosis were determined.CTCs were abundant [mean (SD), 70.2 (107.6)] and present in 81% (51 of 63) of patients with PDAC. CTM were present in 81% (51 of 63) of patients with mean (SD) 29.7 (1101.4). CTM was an independent prognostic factor of overall survival (OS) and progression free survival (PFS). Patients were stratified into unfavorable and favorable CTM groups on the basis of CTM more or less than 30 per 2 mL blood, respectively. Patients with baseline unfavorable CTM, compared with patients with favorable CTM, had shorter PFS (2.7 vs 12.1 months; P < 0.0001) and OS (6.4 vs 19.8 months; P < 0.0001). Differences persisted if we stratified patients into early and advanced diseases. The number of CTM before treatment was an independent predictor of PFS and OS after adjustment for clinically significant factors.The number of CTM, instead of CTCs, before treatment is an independent predictor of PFS and OS in patients with PDAC.
View details for DOI 10.1373/clinchem.2015.248260
View details for Web of Science ID 000371225200015
View details for PubMedID 26861552
Identifying cancer origin using circulating tumor cells
CANCER BIOLOGY & THERAPY
2016; 17 (4): 430–38
Circulating tumor cells (CTCs) have become an established clinical evaluation biomarker. CTC count provides a good correlation with the prognosis of cancer patients, but has only been used with known cancer patients, and has been unable to predict the origin of the CTCs. This study demonstrates the analysis of CTCs for the identification of their primary cancer source. Twelve mL blood samples were equally dispensed on 6 CMx chips, microfluidic chips coated with an anti-EpCAM-conjugated supported lipid bilayer, for CTC capture and isolation. Captured CTCs were eluted to an immunofluorescence (IF) staining panel consisting of 6 groups of antibodies: anti-panCK, anti-CK18, anti-CK7, anti-TTF-1, anti-CK20/anti-CDX2, and anti-PSA/anti-PSMA. Cancer cell lines of lung (H1975), colorectal (DLD-1, HCT-116), and prostate (PC3, DU145, LNCaP) were selected to establish the sensitivity and specificity for distinguishing CTCs from lung, colorectal, and prostate cancer. Spiking experiments performed in 2mL of culture medium or whole blood proved the CMx platform can enumerate cancer cells of lung, colorectal, and prostate. The IF panel was tested on blood samples from lung cancer patients (n = 3), colorectal cancer patients (n = 5), prostate cancer patients (n = 5), and healthy individuals (n = 12). Peripheral blood samples found panCK(+) and CK18(+) CTCs in lung, colorectal, and prostate cancers. CTCs expressing CK7(+) or TTF-1(+), (CK20/ CDX2)(+), or (PSA/ PSMA)(+) corresponded to lung, colorectal, or prostate cancer, respectively. In conclusion, we have designed an immunofluorescence staining panel to identify CTCs in peripheral blood to correctly identify cancer cell origin.
View details for DOI 10.1080/15384047.2016.1141839
View details for Web of Science ID 000375583500013
View details for PubMedID 26828696
View details for PubMedCentralID PMC4910938
- Chemoresponsive surface-tethered polypeptide brushes based on switchable secondary conformations RSC ADVANCES 2015; 5 (105): 86113-86119
Efficient elusion of viable adhesive cells from a microfluidic system by air foam
AMER INST PHYSICS. 2014: 052001
We developed a new method for releasing viable cells from affinity-based microfluidic devices. The lumen of a microchannel with a U-shape and user-designed microstructures was coated with supported lipid bilayers functionalized by epithelial cell adhesion molecule antibodies to capture circulating epithelial cells of influx solution. After the capturing process, air foam was introduced into channels for releasing target cells and then carrying them to a small area of membrane. The results show that when the air foam is driven at linear velocity of 4.2 mm/s for more than 20 min or at linear velocity of 8.4 mm/s for more than 10 min, the cell releasing efficiency approaches 100%. This flow-induced shear stress is much less than the physiological level (15 dyn/cm(2)), which is necessary to maintain the intactness of released cells. Combining the design of microstructures of the microfluidic system, the cell recovery on the membrane exceeds 90%. Importantly, we demonstrate that the cells released by air foam are viable and could be cultured in vitro. This novel method for releasing cells could power the microfluidic platform for isolating and identifying circulating tumor cells.
View details for DOI 10.1063/1.4893348
View details for Web of Science ID 000344226200003
View details for PubMedID 25332725
View details for PubMedCentralID PMC4189394
DIFFERENTIATION OF NEURAL STEM/PROGENITOR CELLS USING LOW-INTENSITY ULTRASOUND
ULTRASOUND IN MEDICINE AND BIOLOGY
2014; 40 (9): 2195–2206
Herein, we report the evaluation of apoptosis, cell differentiation, neurite outgrowth and differentiation of neural stem/progenitor cells (NSPCs) in response to low-intensity ultrasound (LIUS) exposure. NSPCs were cultured under different conditions, with and without LIUS exposure, to evaluate the single and complex effects of LIUS. A lactic dehydrogenase assay revealed that the cell viability of NSPCs was maintained with LIUS exposure at an intensity range from 100 to 500 mW/cm(2). Additionally, in comparison with no LIUS exposure, the cell survival rate was improved with the combination of medium supplemented with nerve growth factor and LIUS exposure. Our results indicate that LIUS exposure promoted NSPC attachment and differentiation on a glass substrate. Neurite outgrowth assays revealed the generation of longer, thicker neurites after LIUS exposure. Furthermore, LIUS stimulation substantially increased the percentage of differentiating neural cells in NSPCs treated with nerve growth factor in comparison with the unstimulated group. The high percentage of differentiated neural cells indicated that LIUS induced neuronal networks denser than those observed in the unstimulated groups. Furthermore, the release of nitric oxide, an important small-molecule neurotransmitter, was significantly upregulated after LIUS exposure. It is therefore reasonable to suggest that LIUS promotes the differentiation of NSPCs into neural cells, induces neurite outgrowth and regulates nitric oxide production; thus, LIUS may be a potential candidate for NSPC induction and neural cell therapy.
View details for DOI 10.1016/j.ultrasmedbio.2014.05.001
View details for Web of Science ID 000341461100031
View details for PubMedID 25023110
Natural zwitterionic organosulfurs as surface ligands for antifouling and responsive properties
2014; 9 (2): 029010
Natural sulfur-containing zwitterionic compounds, l-cysteine (Cys), l-methionine, and glutathionine (GSH), have been employed as surface ligands to prevent protein nonspecific adsorption on planar substrates. These organosulfur compounds form self-assembled monolayers (SAMs) on gold substrates by gold-sulfur interaction. The chemical elements of SAMs were confirmed using x-ray photoelectron spectroscopy. The surface wetting tests for SAMs show that films prepared from Cys and GSH exhibited super-hydrophilicity (contact angles of θ = ~5°) due to their high coverage and strong hydration via ionic solvation and formation of hydrogen bonding. Quartz crystal microbalance with dissipation sensor was used to quantitatively and qualitatively monitor the adsorption of bovine serum albumin (BSA) from buffer onto these SAMs. It was found that the GSH film enables the resistance of BSA adsorption to the best extent at a physiological pH. Moreover, the surface charges of modified substrates were modulated by varying the pH value to control BSA adsorption. The effect of electrostatic repulsion on the antifouling behavior becomes prominent at a pH where the protein and the surface carry same charges. Consolidating the BSA adsorption measurements at different pH values, the antifouling properties of GSH-modified Au should be attributed to prevention of entropy gain and enthalpy loss, making BSA adsorption energetically unfavorable. It is believed that the surface modification with natural organosulfur ligands holds great potential in improving the biocompatibility of medical devices and in offering intelligent biointerfaces in response to environmental stimuli.
View details for DOI 10.1116/1.4869300
View details for Web of Science ID 000339817200010
View details for PubMedID 24985214
In Situ Surface Tailoring with Zwitterionic Carboxybetaine Moieties on Self-Assembled Thin Film for Antifouling Biointerfaces
2014; 7 (1): 130–42
A novel biointerface bearing zwitterionic carboxybetaine moieties was developed for effective resistance to nonspecific adsorption of proteins and blood cells. Self-assembled thin films (SAFs) of (N,N-dimethylaminopropyl) trimethoxysilane were formed as mattress layers by either vapor or solution deposition. Subsequently, the tertiary amine head groups on SAFs were reacted with β-propiolactone to give zwitterionic carboxybetaine moieties via in situ synthesis. The optimal reaction time of 8 h for both preparation methods was verified by static contact angle measurements. According to the X-ray photoelectron spectroscopy, 67.3% of amine groups on SAFs prepared from the vapor deposition was converted to the zwitterionic structures after reaction of β-propiolactone. The antifouling properties of the zwitterionic biointerfaces were quantitatively evaluated in the presence of protein solutions using a quartz crystal microbalance with dissipation, showing a great improvement by factors of 6.5 and 20.2 from tertiary amine SAFs and bare SiO₂ surfaces, respectively. More importantly, the zwitterionic SAFs were brought to contact with undiluted human blood in chaotic-mixer microfluidic systems; the results present their capability to effectively repel blood cell adhesion. Accordingly, in this work, development of carboxybetaine SAFs offers a facile yet effective strategy to fabricate biocompatible biointerfaces for a variety of potential applications in surface coatings for medical devices.
View details for DOI 10.3390/ma7010130
View details for Web of Science ID 000336088500010
View details for PubMedID 28788445
View details for PubMedCentralID PMC5453132
Antibody conjugated supported lipid bilayer for capturing and purification of viable tumor cells in blood for subsequent cell culture
2013; 34 (21): 5191-5199
Interest in the identification and isolation of circulating tumor cells (CTCs) has been growing since the introduction of CTCs as an alternative to the tumor tissue biopsy, which can potentially be important indices for prognosis and cancer treatment. However, the contamination of non-specific binding of normal hematologic cells makes high purity CTCs detection problematic. Furthermore, preserving the viability of CTCs remains a challenge. In this study, we proposed to construct an anti-EpCAM functionalized supported lipid bilayer (SLB), a biomimetic and non-fouling membrane coating, for CTCs capturing, purification and maintaining the viability. Healthy human blood spiked with pre-stained colorectal cancer cell lines, HCT116 and colo205, were used to investigate interaction of cells with the anti-EpCAM functionalized SLB surfaces. Over 97% of HCT116, and 72% of colo205 were captured and adhered by the surface anti-EpCAM; conversely, the majority of blood cells were easily removed by gentle buffer exchange, with the overall purity of cancer cells exceeding 95%. The bound cancer cells were subsequently detached for cell culture. Both HCT116 and colo205 continued to proliferate over 2-week observation period, indicating that the anti-EpCAM functionalized SLB platform providing a simple strategy for capturing, purifying, and releasing viable targeted rare cells.
View details for DOI 10.1016/j.biomaterials.2013.03.096
View details for Web of Science ID 000319630000028
View details for PubMedID 23615560
Use of Surface Properties to Control the Growth and Differentiation of Mouse Fetal Liver Stem/Progenitor Cell Colonies
2012; 13 (11): 3483–93
Multilayers of poly-l-lysine/poly-l-glutamic acid (PLL/PLGA) were constructed by layer-by-layer deposition on an end-tethered cationic PLL brush film serving as an initial layer. Increasing the number of coupling layers increased the thickness and the hydration of the films, and decreased the films' shear modulus and serum adsorption. These films were used to culture primary mouse fetal liver cells. Fetal liver stem/progenitor cells (FLSPCs) were isolated and maintained on the PLGA-terminal PLL/PLGA surfaces, forming colonies with clear boundaries that were partially attached to the surface, with cross-sectional areas of ~500 to ~2500 μm(2) after 2 days culture. Long-term studies showed that the cluster size of colonies slowly expanded and was correlated with the surface properties. For example, on the thicker films with shear modulus, G, less than 5 kPa, FLSPCs cluster size was constrained within a small distribution with less than 4000 μm(2) of projected area, whereas on the thinner films with G > 30 kPa, clusters were expanded and widely distributed, with projected areas over 4000 um(2). Immunostaining studies suggested that clusters with a small size maintained the self-renewal characteristics of stem cells, while the expanded clusters were clearly the results of spontaneous differentiation, exhibiting hepatocyte-like properties. On PLL-terminal t-(PLL/PLGA) films, which are less favorable for stem cell cultures than PLGA-terminal t-(PLL/PLGA) films, the cluster size distribution was also correlated with the film thickness, with more clusters of small size preserved on the thicker films. We observed that a soft, hydrated, serum-free surface could restrict the FLSPC expansion, resulting in self-maintenance of FLSPC colonies.
View details for DOI 10.1021/bm301074j
View details for Web of Science ID 000310931900008
View details for PubMedID 23004572
Tethered Fibronectin Liposomes on Supported Lipid Bilayers as a Prepackaged Controlled-Release Platform for Cell-Based Assays
2012; 13 (8): 2254–62
A biomimetic construct containing an extracellular matrix protein-liposome composite tethered on supported lipid bilayers (SLBs) was formed with fibronectin (FN), and polyethylene glycol (PEG) and cholesterol-containing liposomes. The construct can serve as a multifunctional platform for cell attachment and drug release. The successful fabrication of the FN-liposome-SLB model platform was analyzed in situ with a quartz crystal microbalance with dissipation. The long-term stability of the surface tethered liposomes was measured via an encapsulated fluorescent probe. Less than 20% of the fluorescent probe content was released in 8 days, which compared favorably to the release of 90% of the probe content in one day from a similar construct made without PEG and cholesterol. HeLa cells were used to study the cellular interactions with the model platform. The extracellular matrix composition, FN, was found to be essential to promote HeLa cell adhesion on the liposome-SLB surfaces. Upon cell adhesion, the liposomes were spatially reorganized and absorbed by the cells. The rate of HeLa cell apoptosis was correlated with the surface density of doxorubicin-loaded liposomes, confirming the effective drug delivery through liposomes. The multifunctional model platform could be useful as preadministered, controlled-release platforms for cell- and tissue-based assays.
View details for DOI 10.1021/bm300426u
View details for Web of Science ID 000307422300006
View details for PubMedID 22721275
The influence of collagen film nanostructure on pulmonary stem cells and collagen-stromal cell interactions
2010; 31 (32): 8271–80
We have recently identified a rare subpopulation of lung colony cells with the characteristics of pulmonary stem cells, and discovered that stem cell colonies grew preferentially on type I collagen films in a serum-free medium. In order to further optimize culture conditions and determine stem cell growth in relation to microenvironments (including the stroma, medium and nanostructures of type I collagen films), both primary and pre-sorted stem cells were cultured on the type I collagen films with controllable degree of polymerization and film thickness, as confirmed by an atomic force microscope and surface profiler. We found that in a primary culture, the spreading of stromal cells is greatly restrained and both the size and number of colonies are significantly reduced on highly polymerized collagen films. In contrast, in a pre-sorted stem cell culture without stromal cells, the intrinsic stem cell properties and cell number are independent of the degree of collagen polymerization. Our results indicate that the nanostructures of type I collagen films primarily affect stem colony formation through the collagen-stroma interactions. In those cases, collagen film thickness shows no effect on colony formation.
View details for DOI 10.1016/j.biomaterials.2010.07.038
View details for Web of Science ID 000283112700018
View details for PubMedID 20673998
Effects of extracellular matrix protein functionalized fluid membrane on cell adhesion and matrix remodeling
2010; 31 (27): 7183–95
In order to study cellular responses and extracellular matrix protein remodeling mediated by biomaterials coating, we proposed a biomimetic construct containing protein-conjugated supported lipid bilayers (SLBs) as a cell culture platform. Single or multi-component proteins-bound SLBs were fabricated by conjugating type I collagen and/or fibronectin on the N-hydroxysulfosuccinimide-functionalized SLBs. The proposed protein-conjugated systems were quantitatively characterized by the quartz crystal microbalance with dissipation. NIH 3T3 fibroblasts were cultured on the model constructs and on oxygen plasma pretreated polystyrene (PSo) for parallel comparison. The retards of mobility of SLB after protein conjugation and cell culture were estimated by fluorescence recovery after photobleaching. The resulting cell morphology, adsorption kinetics and somatic dynamics were examined microscopically. We found that, on the SLB based cultures, the largest spreading size and cell number counts of 3T3 fibroblasts were found on the fibronectin containing surfaces. However, on the protein-coated PSo surfaces, no such distinguishable differences can be observed on all protein contents. Immunofluorescent staining results revealed that adsorption of endogenously produced fibronectin by 3T3 cells on PSo based surfaces is significantly more than that on SLB based surfaces. This suggests that the anti-fouling nature of underneath SLBs have played an important role in preventing 3T3 cells from effectively remodeling their microenvironment, whereas cells can easily remodel the nonspecific adsorption prone surfaces such as PSo based platforms. In summary, the protein conjugated SLB surfaces can serve as a platform for determining and regulating cell specific binding and subsequent signaling events with extracellular environments.
View details for DOI 10.1016/j.biomaterials.2010.05.076
View details for Web of Science ID 000280616300031
View details for PubMedID 20580428
Type I Collagen-Functionalized Supported Lipid Bilayer as a Cell Culture Platform
2010; 11 (5): 1231-1240
The supported phospholipid bilayer serves as an important biomimetic model for the cell membrane in both basic and applied scientific research. We have constructed a biomimetic platform based on a supported phospholipid bilayer that is functionalized with type I collagen to serve as a substrate for cell culture. To create the type I collagen-functionalized lipid bilayer assembly, a simple chemical approach was employed: lipid vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-(glutaryl) (DP-NGPE), a carboxylic acid-functionalized phospholipid, were prepared and then fused onto an SiO(2) substrate to form a supported lipid bilayer. Subsequently, type I collagen molecules were introduced to form stable collagen-lipid conjugates via amide linkages with activated DP-NGPE lipids. The binding kinetics of the conjugation process and the resultant changes in film thickness and viscoelasticity were followed using the quartz crystal microbalance with dissipation (QCM-D) monitoring. The morphology of the conjugated collagen adlayer was investigated with atomic force microscopy (AFM). We observed that the adsorbed collagen molecules tended to self-assemble into fibrillar structures. Fluorescence recovery after photobleaching (FRAP) was utilized to estimate lateral lipid mobility, which was reduced by up to 20% after the coupling of type I collagen to the underlying lipid bilayer. As a cell culture platform, the collagen-conjugated supported lipid bilayer showed promising results. Smooth muscle cells (A10) retained normal growth behavior on the collagen-functionalized platform, unlike the bare POPC lipid bilayer and the POPC/DG-NGPE bilayer without collagen. The biomimetic functionalized lipid system presented here is a simple, yet effective approach for constructing a cell culture platform to explore the interactions between extracellular matrix components and cells.
View details for DOI 10.1021/bm901445r
View details for Web of Science ID 000277355800013
View details for PubMedID 20361729
Effect of Solvents and Temperature on the Conformation of Poly(beta-benzyl-L-aspartate) Brushes
2010; 11 (5): 1308–13
We report on the synthesis and characterization of end-tethering polypeptide monolayers based on poly(beta-benzyl-L-aspartate) (PBLA) homopolymer and PBLA-b-poly(gamma-benzyl-L-glutamate) block copolymer. The homopolypeptide and copolypeptide brushes were fabricated by the sequential, surface-initiated vapor deposition polymerization of the N-carboxyanhydride of beta-benzyl-L-aspartate or gamma-benzyl-L-glutamate, yielding 80-nm-thick, chemically grafted films after 30 min of reaction time. Both Fourier transform infrared spectrometry and circular dichroism showed that the polypeptide brushes could be reversibly and repeatedly switched between left-handed and right-handed alpha-helical structures in response to solvent vapor exposure or permanently converted to a beta-sheet structure when heated to 160 degrees C in air. The facile, in vacuo manufacturability and the robustness of the films of PBLA-based brushes could allow them to be incorporated as active components for biosensing and nanofabricated devices.
View details for DOI 10.1021/bm1000907
View details for Web of Science ID 000277355800023
View details for PubMedID 20415412
Selection, Enrichment, and Maintenance of Self-Renewal Liver Stem/Progenitor Cells Utilizing Polypeptide Polyelectrolyte Multilayer Films
2010; 11 (4): 994–1001
Recent progress has led to the identification of liver stem/progenitor cells as suitable sources for generating transplantable liver cells. However, the great variability in methods utilized to isolate liver stem/progenitor cells is a considerable challenge for clinical applications. The polyelectrolyte-multilayer technique can constitute a useful method for selective cell adhesion. Whether enrichment of liver stem/progenitor cells can be achieved utilizing polypeptide polyelectrolyte-multilayer films was investigated in current work. Fetal liver cells isolated from E13.5 mouse embryos were seeded on the poly-l-glutamic acid/poly-l-lysine alternating films, and we revealed that fetal liver stem/progenitor cells were selected and formed colonies. These undifferentiated colonies were maintained on the films composed of four alternating layers, with the topmost poly-l-glutamic acid layer judged by the constitutive expression of stem-cell markers such as Dlk-1, CD49f, and CD133 and self-renew marker-beta-catenin. Our work has demonstrated that highly tunable polyelectrolyte-multilayer films were suitable for selective enrichment of liver stem/progenitor cells in vitro.
View details for DOI 10.1021/bm901461e
View details for Web of Science ID 000276557300022
View details for PubMedID 20337459
Controlled Molecular Organization of Surface Macromolecular Assemblies Based on Stimuli-Responsive Polypeptide Brushes
2009; 10 (1): 58–65
End-tethered cationic polypeptide brushes of poly(L-lysine) (t-PLL) were combined with three anionic polymers, poly(acrylic acid) (PAA), poly(L-glutamic acid) (PLGA), and poly(L-aspartic acid) (PLAA), to form reversible polyelectrolyte complex films at surfaces at neutral pH. The polyelectrolyte complex formation was confirmed by an in situ zeta-potential study and by positive fluorescent images after adding prelabeled anionic polymers. The secondary conformations of the t-PLL complex films depend upon the specific polyelectrolyte with which t-PLL was coupled as studied by circular dichroism and FTIR. Specifically, the random coil chain configuration of the t-PLL film was converted to an alpha-helical, beta-sheet, or random coil structure after forming complexes with PAA, PLGA, or PLAA, respectively. Each of these complexes could be returned to the original random coil t-PLL structure by a dilute acid rinse. Additional thickness and morphological studies from ellipsometry and atomic force microscopy have further shown that the corresponding film thicknesses of the individual solvated films were affected more by the secondary structures in films than by the adsorbed mass or surface net charges. The solvated thickness was reduced significantly after the random coil t-PLL film was coupled with polyanions in forming compact regulated structures in films. This biomimetic approach provides a new opportunity for controlling the molecular organization in surface macromolecular assemblies and may provide a model for structural study of protein complexes on a chip.
View details for DOI 10.1021/bm8007956
View details for Web of Science ID 000262399600008
View details for PubMedID 19053808
- Biomimetic synthesis of silica films directed by polypeptide brushes CHEMISTRY OF MATERIALS 2008; 20 (19): 6148–56
Preparation of end-grafted polymer brushes by nitroxide-mediated free radical polymerization of vaporized vinyl monomers
2005; 21 (21): 9562–67
In this work, we report a gas-phase polymerization approach to create end-grafted vinyl based polymer films on silicon oxide based substrates. The "surface-initiated vapor deposition polymerization" (SI-VDP) of vaporized vinyl monomers, via the nitroxide-mediated free radical polymerization mechanism, was developed to fabricate various homo- and block copolymer brushes from surface-bound initiators, 1-(4'-oxa-2'-phenyl-12'-trimethoxysilyldodecyloxy)-2,2,6,6-tetra-methylpiperidine ("TEMPO"). The resulting polymer thin films were characterized by the Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, ellipsometry, and contact angle goniometry, respectively, to identify the surface composition, film thickness, surface coverage, and water contact angles. Through the SI-VDP, end-grafted polymer films of polystyrene (PSt), poly(acrylic acid) (PAAc), poly(N-(2-hydroxypropyl) methacrylamide) (PHPMA), and poly(N-isopropylacrylamide) (PNIPAAm) with 10-200 nm thicknesses were fabricated. Furthermore, the block copolymer films of PAAc (1st block)-b-PSt (2nd block), PSt (1st block)-b-PAAc (2nd block), and a triblock copolymer film of PAAc (1st)-b-PSt (2nd)-b-PHPMA (3rd), were also fabricated, suggesting the "renewability" of the TEMPO-initiated polymerization in the SI-VDP scheme. It is also noticed that the SI-VDP is more efficient than the conventional solution phase polymerization in producing functional polymer brushes such as PNIPAAm, PAAc, or PAAc-b-PSt end-grafted films. In summary, our studies have shown clear advantages of the SI-VDP setup for the nitroxide-mediated polymerization scheme in controlling synthesis of end-grafted homo- and copolymer thin films.
View details for DOI 10.1021/la051109t
View details for Web of Science ID 000232453300030
View details for PubMedID 16207036
Preparation of unidirectional end-grafted alpha-helical polypeptides by solvent quenching
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
2003; 125 (21): 6376–77
A simple "solvent quenching" approach to align an alpha-helical end-grafted poly(gamma-benzyl l-glutamate) monolayer is presented. By sequentially treating poly(gamma-benzyl l-glutamate) with good solvent (chloroform) and bad solvent (acetone), we obtained a highly aligned monolayer with an average tilt angle as small as 3 degrees .
View details for DOI 10.1021/ja034428k
View details for Web of Science ID 000183031800017
View details for PubMedID 12785771