Yu Chi Wang

Decellularization and Recellularization Technologies in Tissue Engineering

Decellularization is the process by which cells are discharged from tissues/organs, but all of the essential cues for cell preservation and homeostasis are retained in a three-dimensional structure of the organ and its extracellular matrix components. During tissue decellularization, maintenance of the native ultrastructure and composition of the extracellular matrix (ECM) is extremely acceptable. For recellularization, the scaffold/matrix is seeded with cells, the final goal being to form a practical organ. In this review, we focus on the biological properties of the ECM that remains when a variety of decellularization methods are used, comparing recellularization technologies, including bioreactor expansion for perfusion-based bioartificial organs, and we discuss cell sources. In the future, decellularization–recellularization procedures may solve the problem of organ assembly on demand.

Differentiation of stem cells: strategies for modifying surface biomaterials.

Stem cells are a natural choice for cellular therapy because of their potential to differentiate into a variety of lineages, their capacity for self-renewal in the repair of damaged organs and tissues in vivo, and their ability to generate tissue constructs in vitro. Determining how to efficiently drive stem cell differentiation to a lineage of choice is critical for the success of cellular therapeutics. Many factors are involved in this process, the extracellular microenvironment playing a significant role in controlling cellular behavior. In recent years, researchers have focused on identifying a variety of biomaterials to provide a microenvironment that is conducive to stem cell growth and differentiation and that ultimately mimics the in vivo situation. Appropriate biomaterials support the cellular attachment, proliferation, and lineage-specific differentiation of stem cells. Tissue engineering approaches have been used to incorporate growth factors and morphogenetic factors—-factors known to induce lineage commitment of stem cells—into cultures with scaffolding materials, including synthetic and naturally derived biomaterials. This review focuses on various strategies that have been used in stem cell expansion and examines modifications of natural and synthetic materials, as well as various culture conditions, for the maintenance and lineage-specific differentiation of embryonic and adult stem cells.

n-butylidenephthalide protects against dopaminergic neuron degeneration and α-synuclein accumulation in Caenorhabditis elegans models of Parkinson's disease.

Background Parkinsons disease (PD) is the second most common degenerative disorder of the central nervous system that impairs motor skills and cognitive function. To date, the disease has no effective therapies. The identification of new drugs that provide benefit in arresting the decline seen in PD patients is the focus of much recent study. However, the lengthy time frame for the progression of neurodegeneration in PD increases both the time and cost of examining potential therapeutic compounds in mammalian models. An alternative is to first evaluate the efficacy of compounds in Caenorhabditis elegans models, which reduces examination time from months to days. n-Butylidenephthalide is the naturally-occurring component derived from the chloroform extract of Angelica sinensis. It has been shown to have anti-tumor and anti-inflammatory properties, but no reports have yet described the effects of n-butylidenephthalide on PD. The aim of this study was to assess the potential for n-butylidenephthalide to improve PD in C. elegans models. Methodology/Principal Findings In the current study, we employed a pharmacological strain that expresses green fluorescent protein specifically in dopaminergic neurons (BZ555) and a transgenic strain that expresses human α-synuclein in muscle cells (OW13) to investigate the antiparkinsonian activities of n-butylidenephthalide. Our results demonstrate that in PD animal models, n-butylidenephthalide significantly attenuates dopaminergic neuron degeneration induced by 6-hydroxydopamine; reduces α-synuclein accumulation; recovers lipid content, food-sensing behavior, and dopamine levels; and prolongs life-span of 6-hydroxydopamine treatment, thus revealing its potential as a possible antiparkinsonian drug. n-Butylidenephthalide may exert its effects by blocking egl-1 expression to inhibit apoptosis pathways and by raising rpn-6 expression to enhance the activity of proteasomes. Conclusions/Significance n-Butylidenephthalide may be one of the effective neuroprotective agents for PD.

Acetylcorynoline attenuates dopaminergic neuron degeneration and α-synuclein aggregation in animal models of Parkinson's disease

Parkinsons disease (PD), the second most common neurodegenerative disease, impairs motor skills and cognitive function. To date, the drugs used for PD treatment provide only symptomatic relief. The identification of new drugs that show benefit in slowing the decline seen in PD patients is the focus of much current research. Acetylcorynoline is the major alkaloid component derived from Corydalis bungeana, a traditional Chinese medical herb. It has been shown to have anti-inflammatory properties, but no studies have yet described the effects of acetylcorynoline on PD. The aim of this study was to evaluate the potential for acetylcorynoline to improve PD in Caenorhabditis elegans models. In the present study, we used a pharmacological strain (BZ555) that expresses green fluorescent protein specifically in dopaminergic neurons, and a transgenic strain (OW13) that expresses human α-synuclein in muscle cells to study the antiparkinsonian effects of acetylcorynoline. Our experimental data showed that treatment with up to 10 mM acetylcorynoline does not cause toxicity in animals. Acetylcorynoline significantly decreases dopaminergic neuron degeneration induced by 6-hydroxydopamine in BZ555 strain; prevents α-synuclein aggregation; recovers lipid content in OW13 strain; restores food-sensing behavior, and dopamine levels; and prolongs life-span in 6-hydroxydopamine-treated N2 strain, thus showing its potential as a possible antiparkinsonian drug. Acetylcorynoline may exert its effects by decreasing egl-1 expression to suppress apoptosis pathways and by increasing rpn5 expression to enhance the activity of proteasomes.

Acetylcorynoline Impairs the Maturation of Mouse Bone Marrow-Derived Dendritic Cells via Suppression of IκB Kinase and Mitogen-Activated Protein Kinase Activities

Background Dendritic cells (DCs) are major modulators in the immune system. One active field of research is the manipulation of DCs as pharmacological targets to screen novel biological modifiers for the treatment of inflammatory and autoimmune disorders. Acetylcorynoline is the major alkaloid component derived from Corydalis bungeana herbs. We assessed the capability of acetylcorynoline to regulate lipopolysaccharide (LPS)-stimulated activation of mouse bone marrow-derived DCs. Methodology/Principal Findings Our experimental data showed that treatment with up to 20 µM acetylcorynoline does not cause cytotoxicity in cells. Acetylcorynoline significantly inhibited the secretion of tumor necrosis factor-α, interleukin-6, and interleukin-12p70 by LPS-stimulated DCs. The expression of LPS-induced major histocompatibility complex class II, CD40, and CD86 on DCs was also decreased by acetylcorynoline, and the endocytic capacity of LPS-stimulated DCs was restored by acetylcorynoline. In addition, LPS-stimulated DC-elicited allogeneic T-cell proliferation was blocked by acetylcorynoline, and the migratory ability of LPS-stimulated DCs was reduced by acetylcorynoline. Moreover, acetylcorynoline significantly inhibits LPS-induced activation of IκB kinase and mitogen-activated protein kinase. Importantly, administration of acetylcorynoline significantly attenuates 2,4-dinitro-1-fluorobenzene-induced delayed-type hypersensitivity. Conclusions/Significance Acetylcorynoline may be one of the potent immunosuppressive agents through the blockage of DC maturation and function.

Novel nanostructured biodegradable polymer matrices fabricated by phase separation techniques for tissue regeneration.

Biomimetic nanostructures have a wide range of applications. In particular, biodegradable polymer nanostructures that mimic the subtleties of extracellular matrix may provide favorable cell interactions. In this study, a co-solvent system was developed to configure a thermodynamically metastable biodegradable polymer solution, from which novel nanostructured matrices subsequently formed via wet phase separation (quaternary) or a combination with thermally induced phase separation. Three-dimensional (3D) nanostructured porous matrices were further fabricated by combination with particle-leaching (100-300μm glucose). The new co-solvent system may generate matrices with reproducible nanostructures from a variety of biodegradable polymers such as poly(d,l-lactide) (PLA), poly(ε-caprolactone) (PCL) and PCL-based polyurethane. In vitro cell culture experiments were performed with mouse pre-osteoblasts (MC3T3-E1) and human bone marrow-derived mesenchymal stem cells (hBM-MSC) to evaluate the osteoinductive potential of PLA nanostructures. The results showed that nanofibrous (<100nm) membranes promoted the bone-related marker gene expression and matrix mineralization of MC3T3-E1 at 14days. Nanofibrous 3D matrices seeded with hBM-MSC without osteogenic induction supplements demonstrated a 2.5-fold increase in bone matrix deposition vs. the conventional microporous matrices after 14 and 21days. Antimicrobial nanofibers were further obtained by plasma-assisted coating of chitosan on PLA nanofibers. This study reveals a platform for fabricating novel biodegradable nanofibrous architecture, with potential in tissue regeneration.

Dryocrassin Suppresses Immunostimulatory Function of Dendritic Cells and Prolongs Skin Allograft Survival

Dendritic cells (DCs) are the major specialized antigen-presenting cells for the development of optimal T-cell immunity. DCs can be used as pharmacological targets to monitor novel biological modifiers for the cure of harmful immune responses, such as transplantation rejection. Dryopteris crassirhizoma Nakai (Aspiadaceae) is used for traditional herbal medicine in the region of East Asia. The root of this fern plant has been listed for treating inflammatory diseases. Dryocrassin is the tetrameric phlorophenone component derived from Dryopteris. Here we tested the immunomodulatory potential of dryocrassin on lipopolysaccharide (LPS)-stimulated activation of mouse bone marrow-derived DCs in vitro and in skin allograft transplantation in vivo. Results demonstrated that dryocrassin reduced the emission of tumor necrosis factor-α, interleukin-6, and interleukin-12p70 by LPS-stimulated DCs. The expression of LPS-induced major histocompatibility complex class II, CD40, and CD86 on DCs was also blocked by dryocrassin. Moreover, LPS-stimulated DC-elicited allogeneic T-cell proliferation was alleviated by dryocrassin. In addition, dryocrassin inhibited LPS-induced activation of IkB kinase, JNK/p38 mitogen-activated protein kinase, and the translocation of NF-κB. Treatment with dryocrassin noticeably diminished 2,4-dinitro-1-fluorobenzene-reduced delayed-type hypersensitivity and extended skin allograft survival. Dryocrassin may be one of the potent immunosuppressive agents for transplant rejection via the destruction of DC maturation and function.

Spatial control of cells, peptide delivery and dynamic monitoring of cellular physiology with chitosan-assisted dual color quantum dot FRET peptides

Cell-based assays have become important tools in the pharmaceutical and biotechnology industries. However, observing and monitoring molecules in cells that mimic the physiological environment is often difficult. Dynamic processes not only increase the accuracy of simulations, but also improve our understanding of the function and regulation of molecules within cells. In this study we used chitosan as a multifunctional biomaterial for selective micropatterning of cells, peptide delivery and covalent bonding with quantum dots (QD) to decrease the cytotoxicity of QD. Our results demonstrate the efficacy of chitosan-QD-peptide-Alexa Fluor 488 in controlling the spread and spatial organization of cells. Cationic chitosan also provided an efficient delivery mechanism to live cells. We used the shift from green to red fluorescence of the chitosan dual color QD peptide to detect biological activity. This methodology has potential applications in high throughput screening of inhibitors and activators of biological mechanisms and pathways and for use in the pharmaceutical industry.

Irisflorentin modifies properties of mouse bone marrow-derived dendritic cells and reduces the allergic contact hypersensitivity responses

Irisflorentin is an isoflavone component derived from the roots of Belamcanda chinensis (L.) DC. In traditional Chinese medicine, this herb has pharmacological properties to treat inflammatory disorders. Dendritic cells (DCs) are crucial modulators for the development of optimal T-cell immunity and maintenance of tolerance. Aberrant activation of DCs can induce harmful immune responses, and so agents that effectively improve DC properties have great clinical value. We herein investigated the effects of irisflorentin on lipopolysaccharide (LPS)-stimulated maturation of mouse bone marrow-derived DCs in vitro and in the contact hypersensitivity response (CHSR) in vivo. Our results demonstrated that treatment with up to 40 μM irisflorentin does not cause cellular toxicity. Irisflorentin significantly lessened the proinflammatory cytokine production (tumor necrosis factor-α, interleukin-6, and interleukin-12p70) by LPS-stimulated DCs. Irisflorentin also inhibited the expression of LPS-induced major histocompatibility complex class II and costimulatory molecules (CD40 and CD86) on LPS-stimulated DCs. In addition, irisflorentin diminished LPS-stimulated DC-elicited allogeneic T-cell proliferation. Furthermore, irisflorentin significantly interfered with LPS-induced activation of IκB kinase, c-Jun N-terminal kinase, and p38, as well as the nuclear translocation of NF-κB p65. Subsequently, treatment with irisflorentin obviously weakened 2,4-dinitro-1-fluorobenzene-induced delayed-type hypersensitivity. These findings suggest new insights into the role of irisflorentin as an immunotherapeutic adjuvant through its capability to modulate the properties of DCs.

Nonlinear analysis of micro piezoelectric energy harvesters

This article proposes a framework for determining the types of nonlinearity observed in the frequency response of microscale energy harvesters made of a piezoelectric film deposited on a stainless-steel substrate. The model accounts for inertial, geometrical and material nonlinearities due to amplified excitation and induced hysteresis. The simulations based on the multiple scale analysis reveals the softening type of nonlinearity for the case of a 15 μm PZT thick film deposited on a 60 μm stainless-steel substrate. They agree quite well with the experimental observations. In addition, the further investigation shows the existence of the critical film thickness such that the hardening (softening) nonlinearity is observed if the film thickness is below (above) this critical value. It is also found that such a key parameter is mainly affected by the ratio of the bending stiffness due to material nonlinearity to that based on linear moduli. Finally, the hardening type of nonlinearity was also observed in different samples with very small film thickness, as predicted by the proposed framework.