Jingchao Li

3D Culture of Chondrocytes in Gelatin Hydrogels with Different Stiffness

Gelatin hydrogels can mimic the microenvironments of natural tissues and encapsulate cells homogeneously, which makes them attractive for cartilage tissue engineering. Both the mechanical and biochemical properties of hydrogels can affect the phenotype of chondrocytes. However, the influence of each property on chondrocyte phenotype is unclear due to the difficulty in separating the roles of these properties. In this study, we aimed to study the influence of hydrogel stiffness on chondrocyte phenotype while excluding the role of biochemical factors, such as adhesion site density in the hydrogels. By altering the degree of methacryloyl functionalization, gelatin hydrogels with different stiffnesses of 3.8, 17.1, and 29.9 kPa Young’s modulus were prepared from the same concentration of gelatin methacryloyl (GelMA) macromers. Bovine articular chondrocytes were encapsulated in the hydrogels and cultured for 14 days. The influence of hydrogel stiffness on the cell behaviors including cell viability, cell morphology, and maintenance of chondrogenic phenotype was evaluated. GelMA hydrogels with high stiffness (29.9 kPa) showed the best results on maintaining chondrogenic phenotype. These results will be useful for the design and preparation of scaffolds for cartilage tissue engineering.

Facile preparation of albumin-stabilized gold nanostars for the targeted photothermal ablation of cancer cells

Gold nanostars (AuNSs) have been extensively studied as photothermal conversion agents for cancer therapy due to their high photothermal conversion efficiency. Surface modification is required to improve their colloidal stability and biocompatibility for biomedical applications. Moreover, the targeting delivery of AuNSs into the tumor sites is an effective approach to improve their therapeutic efficiency. In this study, we report the use of folic acid-bovine serum albumin conjugate (BSA-FA) stabilized AuNSs (BSA-FA-AuNSs) as agents for the targeted photothermal ablation of cervical cancer cells (HeLa). In our approach, BSA-FA conjugate was first synthesized via an amidation reaction, which was further used as a stabilizer to coat the AuNSs for surface modification. The BSA-FA-AuNSs showed good dispersibility and colloidal stability in different media. The strong absorption properties in the near-infrared (NIR) region enabled the increasing temperature of AuNSs under laser irradiation. The BSA-FA-AuNSs not only had a very low cytotoxicity in the studied concentrations, but they also showed targeting specificity to FA receptors-overexpressed cancer cells, which was confirmed by studying the cellular uptake. In addition, the BSA-FA-AuNSs displayed a much better therapeutic efficiency to HeLa cells under the NIR laser irradiation when compared with BSA-AuNSs without FA modification. The BSA-FA-AuNSs should have a great potential as photothermal conversion agents for the receptor-mediated treatment of cancer cells.

Composite scaffolds of gelatin and gold nanoparticles with tunable size and shape for photothermal cancer therapy

Photothermal therapy (PTT) has been extensively investigated as a promising strategy for cancer therapy. For successful application of this technique, various nanomaterials have been explored as photothermal conversion agents. Gold nanoparticles (AuNPs), especially Au nanorods and Au nanostars, have received much attention for photothermal therapy because of their facile preparation and high photothermal conversion efficiency. Due to the limited accumulation and easy diffusion of free nanoparticles, incorporation of nanoparticles into scaffolds for direct implantation has been demonstrated as an attractive way for cancer therapy applications. In this study, composite porous scaffolds of gelatin and AuNPs were prepared by introducing Au nanorods and Au nanostars with average sizes of around 35.0, 65.0 and 115.0 nm in gelatin scaffolds. The composite scaffolds were used for the localized PTT application of cancer cells. Gel/AuNP composite scaffolds supported cell adhesion and showed good biocompatibility. Temperature in the composite scaffolds increased quickly upon NIR laser irradiation. Photothermal efficiency and cancer cell killing efficiency were dependent on the shape, size and amount of AuNPs in the composite scaffolds. The composite scaffolds prepared with 65.0 nm Au nanorods showed the highest photothermal efficiency and cell killing efficiency. The results indicated the importance of the shape and size modulation of AuNPs for photothermal therapy applications.

Sub-10 nm gold nanoparticles promote adipogenesis and inhibit osteogenesis of mesenchymal stem cells

Sub-10 nm gold nanoparticles (Au NPs) have attracted extensive attention for different biomedical applications because of their small size. However, the influence of small NPs on the differentiation of human bone marrow-derived mesenchymal stem cells (hMSCs) is unclear. In this study, 4 nm small Au NPs (Au4-mPEG NPs) were synthesized to investigate their influence on the osteogenic and adipogenic differentiation of hMSCs and 40-nm counterparts (Au40-mPEG NPs) were used for comparison. Different from Au40-mPEG NPs, Au4-mPEG NPs reduced the ALP activity, calcium deposition and osteogenic marker gene expression, while increased the oil droplet formation and adipogenic marker gene expression in hMSCs. The mechanism study indicated that Au4-mPEG NP treatment did not affect the cellular mechanical property significantly but induced a high level of reactive oxygen species (ROS). The results provide some insights into the influence of sub-10 nm Au NPs on the stem cell functions and applications of these small particles for tissue engineering.

Preparation of gelatin/Fe3O4 composite scaffolds for enhanced and repeatable cancer cell ablation

Various nanomaterials have been extensively investigated for photothermal ablation of cancer cells because of their high photothermal conversion efficiency. However, the poor targeting specificity and low repeated heating efficiency of nanomaterials restrict their applications in the clinic. In this work, porous gelatin/iron oxide (Gel/Fe3O4) composite scaffolds were prepared by a facile ice particulate templating method for efficient and repeatable cancer cell ablation. Gel/Fe3O4 composite scaffolds showed controlled porous structure consisting of large pores and interconnecting small pores. The strong absorption in the near-infrared (NIR) region enabled the Gel/Fe3O4 composite scaffolds to elevate local temperature quickly under NIR laser irradiation. The composite scaffolds allowed cell adhesion and proliferation showing good biocompatibility. Cancer cells entrapped in the scaffolds could be efficiently killed during laser irradiation. Moreover, the therapeutic efficacy of Gel/Fe3O4 composite scaffolds could be enhanced by repeated laser irradiation treatment, which is important for clinical application because of the resistant and recurrent nature of cancer. The results indicated that the porous Gel/Fe3O4 composite scaffolds had good biocompatibility and excellent cancer cell ablation efficacy, which may provide an attractive way to use porous scaffolds for cancer therapy application.

Nanoencapsulation of individual mammalian cells with cytoprotective polymer shell

Nanoencapsulation of individual mammalian cells has great potential in biomedical, biotechnological and bioelectronic applications. However, existing techniques for cell nanoencapsulation generally yield short sustaining period and loose structure of encapsulation shell, which fails to meet the long-term cytoprotection and immunosuppression requirements. Here, we report a mild method to realize the nanoencapsulation of individual mammalian cells by layer-by-layer (LbL) assembly of gelatin inner layer and cross-linking of poly(ethylene glycol) (PEG) outer layer through thiol-click chemistry. With the present method, the encapsulated individual HeLa cells showed a high viability, long persistence period and effective resistance against macro external entities and high physical stress. Moreover, on-demand cell release could also be achieved by selective cleavage of succinimide thioether linkage in the outer PEG layer. The approach presented here may provide a new and versatile method for the cleavable nanoencapsulation of individual mammalian cells.

Single mammalian cell encapsulation by in situ polymerization

Encapsulation of single mammalian cells with biocompatible and protective materials has important applications in biomedical and single-cell biology research. To date, single mammalian cells have generally been encapsulated by physical approaches including hydrophobic and electrostatic interactions. However, the cytotoxicity of the used materials, short sustained period, and especially loose structure of the encapsulation layer have limited the application of the encapsulated cells. Here, we show a novel strategy based on in situ polymerization for encapsulation of single mammalian cells with a network structure of the polymeric shell under mild conditions. Using the present strategy, the encapsulated mammalian cells, HeLa cells, hMSCs, and BACs showed high viability. Furthermore, the encapsulation shell was able to prevent the penetration of macro-external entities, while maintaining the free exchange of smaller molecules. This work provides a new and versatile method for single mammalian cell encapsulation.

Ligand density-dependent influence of arginine–glycine–aspartate functionalized gold nanoparticles on osteogenic and adipogenic differentiation of mesenchymal stem cells

Extracellular matrix (ECM) plays a very important role in regulating cell function and fate. It is highly desirable to fabricate biomimetic models to investigate the role of ECM in stem cell differentiation. In this study, arginine–glycine–aspartate (RGD)-modified gold nanoparticles (Au NPs) with tunable surface ligand density were prepared to mimic the ECM microenvironment. Their effect on osteogenic and adipogenic differentiation of human mesenchymal stem cells (MSCs) was investigated. The biomimetic Au NPs were taken up by MSCs in a ligand density-dependent manner. The biomimetic NPs with a high RGD density had an inhibitive effect on the alkaline phosphatase (ALP) activity, calcium deposition, and osteogenic marker gene expression of MSCs. Their effect on oil droplet formation and adipogenic marker gene expression was negative when RGD density was low, while their effect was promotive when RGD density was high. The biomimetic Au NPs regulated the osteogenic and adipogenic differentiation of MSCs mainly through affecting the focal adhesion and cytoskeleton. This study highlights the roles of biomimetic NPs on stem cell differentiation that could provide a meaningful strategy in fabricating functional biomaterials for tissue engineering and biomedical applications.

Preparation of dexamethasone-loaded calcium phosphate nanoparticles for the osteogenic differentiation of human mesenchymal stem cells

As an earliest known and readily available osteogenic inducer for stem cells, dexamethasone (DEX) plays a key role in affecting cell functions and cellular processes, especially for cell proliferation and differentiation. However, the clinical application of DEX has been limited because of its uncontrolled release. An ideal carrier is desired to control the DEX release for the osteogenic differentiation of stem cells and bone tissue engineering. Biphasic calcium phosphate nanoparticles (BCP-NPs) should be potential carriers for DEX due to their osteoconductive properties and good biocompatibility as a bone graft biomaterial. In this study, DEX-loaded BCP-NPs were prepared by two methods: (1) immersion of BCP-NPs in a DEX solution (denoted DEX/BCP-NPs), (2) DEX incorporation during BCP-NP formation in a calcifying solution (denoted DEX@BCP-NPs). The DEX@BCP-NPs showed a higher DEX loading amount and more sustainable DEX release than did the DEX/BCP-NPs. The DEX@BCP-NPs were used for the culture of human bone marrow-derived mesenchymal stem cells (hMSCs) and showed a promotive effect on the proliferation of hMSCs. Furthermore, the DEX@BCP-NPs significantly increased the alkaline phosphatase (ALP) activity, calcium deposition and gene expressions of the osteogenic markers of hMSCs when compared to BCP-NPs without DEX loading. The results demonstrated BCP-NPs were good carriers for DEX loading and the DEX@BCP-NPs should be useful for bone tissue engineering.

Induction of Chondrogenic Differentiation of Human Mesenchymal Stem Cells by Biomimetic Gold Nanoparticles with Tunable RGD Density

&NA; Nanostructured materials have drawn a broad attention for their applications in biomedical fields. Ligand‐modified nanomaterials can well mimic the dynamic extracellular matrix (ECM) microenvironments to regulate cell functions and fates. Herein, ECM mimetic gold nanoparticles (Au NPs) with tunable surface arginine‐glycine‐aspartate (RGD) density are designed and synthesized to induce the chondrogenic differentiation of human mesenchymal stem cells (hMSCs). The biomimetic Au NPs with an average size of 40 nm shows good biocompatibility without affecting the cell proliferation in the studied concentration range. The RGD motifs on Au NPs surface facilitate cellular uptake of NPs into monolayer hMSCs through integrin‐mediated endocytosis. The biomimetic NPs have a promotive effect on cartilaginous matrix production and marker gene expression in cell pellet culture, especially for the biomimetic Au NPs with high surface RGD density. This study provides a novel strategy for fabricating biomimetic NPs to regulate cell differentiation, which holds great potentials in tissue engineering and biomedical applications.