Effect of RGD content in poly(ethylene glycol)-crosslinked poly(methyl vinyl ether-alt-maleic acid) hydrogels on the expansion of ovarian cancer stem-like cells.

The extracellular matrix (ECM) affects cell behaviors, such as survival, proliferation, motility, invasion, and differentiation. The arginine-glycine-aspartic acid (RGD) sequence is present in several ECM proteins, such as fibronectin, collagen type I, fibrinogen, laminin, vitronectin, and osteopontin. It is very critical to develop ECM-like substrates with well-controlled features for the investigation of influence of RGD on the behavior of tumor cells. In this study, poly(ethylene glycol) (PEG)-crosslinked poly(methyl vinyl ether-alt-maleic acid) (P(MVE-alt-MA)) hydrogels (PEMM) with different RGD contents were synthesized, fully characterized, and established as in vitro culture platforms to investigate the effects of RGD content on cancer stem cell (CSC) enrichment. The morphology, proliferation, and viability of SK-OV-3 ovarian cancer cells cultured on hydrogels with different RGD contents, the expression of CSC markers and malignant signaling pathway-related genes, and drug resistance were systematically evaluated. The cell aggregates formed on the hydrogel surface with a lower RGD content acquired certain CSC-like properties, thus drug resistance was enhanced. In contrast, the drug sensitivity of cells on the higher RGD content surface increased because of less CSC-like properties. However, the presence of RGD in the stiff hydrogels (PEMM2) had less effect on the stemness expression than did its presence in the soft hydrogels (PEMM1). The results suggest that RGD content and matrix stiffness can lead to synergetic effects on the expression of cancer cell stemness and the epithelial-mesenchymal transition (EMT), interleukin-6 (IL-6), and Wnt pathways.

Mussel-inspired hybrid network hydrogel for continuous adhesion in water.

The mussel-inspired catechol-based strategy has been widely used in the development of adhesives. However, the properties of the obtained adhesives were still severely limited in a humid environment, particularly in water. In this study, a facile and versatile approach was proposed to prepare an underwater adhesion hydrogel. First, dopamine (DA) was grafted on oxidized carboxymethylcellulose (OCMC) to obtain dopamine-grafted oxidized carboxymethylcellulose (OCMC-DA). Second, the acrylamide (AM) monomer was conjugated with OCMC-DA by a Schiff base reaction, and then polymerized to form an OCMC-DA/PAM hydrogel. The properties of the resulting hydrogel have been fully characterized. The underwater adhesion strength of the hydrogel can reach as high as 86.3 ± 7.2 kPa and reduced to 43 ± 3.4 kPa after being immersed in water for 9 days. More remarkably, we found that the maximal adhesion strength was shown when the G' and G'' of the hydrogel were very close. Moreover, we demonstrated the mechanical properties of our fabricated hydrogel by compressive tests and rheological analysis. The adhesive hydrogel also exhibits excellent biocompatibility.

Expansion of Ovarian Cancer Stem-like Cells in Poly(ethylene glycol)-Cross-Linked Poly(methyl vinyl ether-alt-maleic acid) and Alginate Double-Network Hydrogels.

A better understanding of cancer stem cells (CSCs) is essential for research on cancer therapy and drug resistance. Currently, increasingly more investigations are focused on obtaining CSCs to study the mechanism of their enhanced malignancy. In this work, three kinds of double-network hydrogels (PEMM/alginate), consisting of poly(ethylene glycol) (PEG) covalently cross-linked poly(methyl vinyl ether-alt-maleic acid) (P(MVE-alt-MA)) (network 1, denoted as PEMM) and Sr2+ (or Ca2+, Fe3+) ionically cross-linked alginates (network 2, denoted as SrAlg, CaAlg, or FeAlg), were prepared. The stiffness, morphology, and components of the PEMM/alginate hydrogels were systematically investigated to understand their effects on CSC enrichment. Only the PEMM/FeAlg hydrogels could support the long-term growth, proliferation, and spheroid formation of SK-OV-3 cells. The expression of CSC-related markers was evaluated with the levels of protein and gene at different stages. The cell spheroids cultured in the PEMM/FeAlg hydrogels acquired certain CSC-like properties, thus drug resistance was enhanced, especially in the PEMM-1/FeAlg hydrogel. In vivo tumorigenicity experiments also confirmed the presence of more CSCs in the PEMM-1/FeAlg hydrogel. The results suggest that matrix stiffness, morphology, and cations act synergistically on the regulation of the epithelial-mesenchymal transition (EMT), interleukin-6 (IL-6), and Wnt pathways, affecting the invasiveness of ovarian cancer and the conversion of the non-CSCs into CSCs. The PEMM-1/FeAlg hydrogel with lower elastic modulus, a more macroporous morphology, and higher swelling rate can significantly enhance the stemness, malignancy, and tumorigenicity of SK-OV-3 cells.

Self-Assembled Supramolecular Hybrid Hydrogels Based on Host-Guest Interaction: Formation and Application in 3D Cell Culture.

In recent decades, in vitro three-dimensional (3D) cell culture has been rapidly developed and widely used in many biomedical fields. Based on this background, a kind of self-assembled supramolecular hybrid hydrogel materials based on host-guest interaction of ß-cyclodextrin (ßCD) and adamantane (Ad) is designed for 3D cell culture. First, ßCD is grafted to poly(methyl vinyl ether-alt-maleic acid) (PMM) to obtain the host polymers of ßCD-grafted-PMM (PMM-ßCD). Second, the guest polymers of poly(acrylamide-co-N-adamantyl acrylamide) (PAAm-Ad) are synthesized through free-radical copolymerization of acrylamide and N-adamantyl acrylamide. Finally, the self-assembled supramolecular hybrid hydrogels of PMM-ßCD/PAAm-Ad are formed by simply mixing the aqueous solution of host and guest polymers with a total concentration of 3.3% (w/v) and a ßCD/Ad molar ratio of 1:1. The main cross-linking interactions come from the host-guest interaction of ßCD/Ad as well as hydrogen-bonding interaction of carboxyl/amide groups. The prepared hydrogels with good cytocompatibility have been successfully used as 3D cell culture scaffold for SKOV3, HUVEC, and L929 cells culture. Thus, this work provides a way and biomaterial for the preparation of a functionalized 3D cell culture scaffold, which lays an experimental and theoretical basis for cell follow-up research.

Thermoresponsive Poly(2-propyl-2-oxazoline) Surfaces of Glass for Nonenzymatic Cell Harvesting.

As one of the nonenzymatic cell-harvesting technologies, a thermal-responsive surface based on poly(2-oxazoline)s has achieved initial success in supporting the adhesion and thermal-induced detachment of animal cells. However, because of the laborious preparation procedure, this technique was only limited to research purposes. In this work, through using poly(glycidyl methacrylate) (PGMA) as the anchor layer, poly(2-propyl-2-oxazoline)s (PPOx) were grafted onto glass wafers through a facile two-step coating and annealing procedure for nonenzymatic cell harvesting. In the first step, the piranha solution-activated glass wafers were immersed into the chloroform solution of PGMA and then annealed for a given period of time to immobilize PGMA onto the glass wafers through the bonding between epoxy groups and hydroxyl groups. In the second step, the PGMA-coated glass wafers were further immersed into the chloroform solution of carboxyl-functionalized PPOx. After annealing, PPOx were immobilized onto the PGMA layer through the bonding between carboxyl groups and the residual epoxy groups. Atomic force microscopy, X-ray photoelectron spectroscopy, and ellipsometry were used to characterize the modified glass wafers. The results of cytocompatibility evaluation showed that the PPOx-coated glass wafers were almost nontoxic and were able to support the adhesion and proliferation of L929 cells well. By lowering the temperature to 8 °C, L929 and Vero cells were successfully detached from the PPOx-coated glass wafers without any enzymatic treatment. Further cultivation has demonstrated that the cooling procedure had little effect on cell viability, and the cells still retained good viability after harvesting.

Assembled anti-adhesion polypropylene mesh with self-fixable and degradable in situ mussel-inspired hydrogel coating for abdominal wall defect repair.

Abdominal adhesion to polypropylene (PP) mesh remains one of the major complications in hernia repair. Thus, a challenge exists to endow PP mesh with powerful anti-adhesion properties in hernia repair. To investigate potential options, the assembled PP mesh was developed with effective anti-adhesion properties through an in situ coating of the mesh surface with self-fixable and biodegradable mussel-inspired hydrogels. Through mixing oxidized-carboxymethylcellulose functionalized with dopamine (OCMC-DA) with carboxymethylchitosan (CMCS), a layer of hydrogel (OCMC-DA/CMCS) can be formed in situ on the PP mesh without the addition of crosslinking agents; the dopamine then acts as an immobilization group to fix these hydrogels to the PP mesh and the tissue surface. In this way, the assembled PP mesh (OCMC-DA/CMCS/PP) was obtained. The properties of the OCMC-DA/CMCS hydrogels were optimized, and the OCMC-DA4/CMCS hydrogel was selected to construct the assembled PP mesh. The lap-shear test revealed that OCMC-DA4/CMCS has tissue-adhesive properties. In vitro cell tests proved the excellent biocompatibility of the hydrogel. An optimized bioabsorption time and significant anti-adhesion properties were demonstrated through an in vivo test with a rat model. The adhesion area and tenacity of the OCMC-DA4/CMCS/PP group were more than 80% lower than those of the native PP mesh group and created a slightly inflammatory reaction.

H727 Multicellular Spheroids and Its Resistance to Antitumor Drugs Sunitinib and Axitinib.

The three-dimensional (3D) culture model of neuroendocrine tumor H727 cells was established by using the agarose gel as culture matrix, which provided a new method for drug screening of neuroendocrine tumors. As VEGFR inhibitor, sunitinib and axitinib were applied to inhibit human neuroendocrine H727 cell line in two-dimensional (2D) and 3D culture models. The inhibitory rate of H727 cells with different drug concentration were assessed by CCK-8 assay method and combined with using the FDA/PI double staining and the digital microscope analysis system. When the concentration of sunitinib ≥4.0 µmol/L, the H727 spheroids began to split, and the apoptosis of H727 cells occurred, the sizes of multicellular spheroids was significantly reduced in the groups of high-dose axitinib. These results illustrated that sunitinib and axitinib can effectively inhibit the growth and proliferation of neuroendocrine tumor H727 cells. Sunitinib and axitinib can also promote apoptosis of H727 cells.

Thermo-Sensitive PLGA-PEG-PLGA Tri-Block Copolymer Hydrogel as Three-Dimensional Cell Culture Matrix for Ovarian Cancer Cells.

Thermo-sensitive hydrogels which could encapsulate cells and provide a three dimensional (3D) microenvironment have great potential in building new cell culture models in vitro. In this study, a thermal responsive hydrogel based on PLGA-PEG-PLGA tri-block copolymers was developed as matrix for 3D ovarian cancer cell culturing. The gelation of PLGA-PEG-PLGA tri-block copolymer was concentration-dependent. SEM images showed the pores were suitable for the formation of 3D cell structures. Cell morphological results showed that large aggregates of ovarian cancer cells (HO8910) were formed after cultured for 10 days. Therefore, hydrogel based on PLGA-PEG-PLGA tri-block copolymers hold potential as in vitro cell culture matrix for ovarian cancer cells.

Shape-controlled fabrication of magnetite silver hybrid nanoparticles with high performance magnetic hyperthermia.

Superparamagnetic Fe3O4 nanoparticles (NPs)-based hyperthermia is a promising non-invasive approach for cancer therapy. However, the heat transfer efficiency of Fe3O4 NPs is relative low, which hinders their practical clinical applications. Therefore, it is promising to improve the magnetic hyperthermia efficiency by exploring the higher performance magnetic NPs-based hybrid nanostructures. In the current study, it presents a straightforward in situ reduction method for the shape-controlled preparation of magnetite (Fe3O4) silver (Ag) hybrid NPs designed as magnetic hyperthermia heat mediators. The magnetite silver hybrid NPs with core-shell (Fe3O4@Ag) or heteromer (Fe3O4-Ag) structures exhibited a higher biocompatibility with SMMC-7721 cells and L02 cells than the individual Ag NPs. Importantly, in the magnetic hyperthermia, with the exposure to alternating current magnetic field, the Fe3O4@Ag and Fe3O4-Ag hybrid NPs indicated much better tumor suppression effect against SMMC-7721 cells than the individual Fe3O4 NPs in vitro and in vivo. These results demonstrate that the hybridisation of Fe3O4 and Ag NPs could greatly enhance the magnetic hyperthermia efficiency of Fe3O4 NPs. Therefore, the Fe3O4@Ag and Fe3O4-Ag hybrid NPs can be used to be as high performance magnetic hyperthermia mediators based on a simple and effective preparation approach.

Self-healing pH-sensitive poly[(methyl vinyl ether)-alt-(maleic acid)]-based supramolecular hydrogels formed by inclusion complexation between cyclodextrin and adamantane.

Self-healing materials are of interest for drug delivery, cell and gene therapy, tissue engineering, and other biomedical applications. In this work, on the base of biocompatible polymer poly(methyl vinyl ether-alt-maleic acid) (P(MVE-alt-MA)), host polymer ß-cyclodextrin-grafted P(MVE-alt-MA) (P(MVE-alt-MA)-g-ß-CD) and guest polymer adamantane-grafted P(MVE-alt-MA) (P(MVE-alt-MA)-g-Ad) were first prepared. Then through taking advantage of the traditional host-guest interaction of ß-cyclodextrin and adamantane, a novel self-healing pH-sensitive physical P(MVE-alt-MA)-g-ß-CD/P(MVE-alt-MA)-g-Ad supramolecular hydrogels were obtained after simply mixing the aqueous solution of host polymer and guest polymer. This kind of supramolecular hydrogels not only possess pH-sensitivity, but also possess the ability to repair themselves after being damaged.

Poly(dopamine)-inspired surface functionalization of polypropylene tissue mesh for prevention of intra-peritoneal adhesion formation.

Polypropylene (PP), as one of the most common prosthetic materials, has been widely used in intra-peritoneal repair. However, its adhesion to viscera has severely limited its application. Therefore it is critical to improve the PP surface with an anti-adhesion property. In this work, based on dopamine-inspired chemistry, virgin PP (V-PP) mesh was first pretreated with O2 plasma, subsequently dipped in dopamine aqueous solution for 24 h, and then chitosan (CS) was grafted onto it. Finally the anti-adhesion mesh (O-PP/PDA/CS) was obtained. The formation procedure of a PDA/CS ad-layer was characterized by water contact angle measurements, ATR-FTIR, SEM, and XPS. The results show that a PDA/CS ad-layer could be coated on the PP surface efficiently. NIH/3T3 cells were first cultured on O-PP/PDA/CS meshes to evaluate the availability of anti-adhesion and biocompatibility in vitro, and then the efficacy of the PDA/CS-coating as a barrier for reducing postsurgical adhesions was evaluated using a rat abdominal wall defect model. Compared with the V-PP group, NIH/3T3 cells exhibited higher viability in the O-PP/PDA/CS groups as evaluated by the CCK-8 method. In addition, NIH/3T3 cells grow into round-shapes on the O-PP/PDA/CS surface. This indicates that the modification strategy can facilely lead to excellent properties of anti-adhesion. In vivo tests further indicate that O-PP/PDA/CS meshes were effective in reducing adhesion formation.

Sliced Magnetic Polyacrylamide Hydrogel with Cell-Adhesive Microarray Interface: A Novel Multicellular Spheroid Culturing Platform.

Cell-adhesive properties are of great significance to materials serving as extracellular matrix mimics. Appropriate cell-adhesive property of material interface can balance the cell-matrix interaction and cell-cell interaction and can promote cells to form 3D structures. Herein, a novel magnetic polyacrylamide (PAM) hydrogel fabricated via combining magnetostatic field induced magnetic nanoparticles assembly and hydrogel gelation was applied as a multicellular spheroids culturing platform. When cultured on the cell-adhesive microarray interface of sliced magnetic hydrogel, normal and tumor cells from different cell lines could rapidly form multicellular spheroids spontaneously. Furthermore, cells which could only form loose cell aggregates in a classic 3D cell culture model (such as hanging drop system) were able to be promoted to form multicellular spheroids on this platform. In the light of its simplicity in fabricating as well as its effectiveness in promoting formation of multicellular spheroids which was considered as a prevailing tool in the study of the microenvironmental regulation of tumor cell physiology and therapeutic problems, this composite material holds promise in anticancer drugs or hyperthermia therapy evaluation in vitro in the future.

Identification and localization of gastrointestinal hormones in the skin of the bullfrog Rana catesbeiana during periods of activity and hibernation.

Amphibian skin and its secretions contain a wide variety of biogenic amines and biologically active peptides, some of which are either identical or highly homologous to gastrointestinal hormones (GHs) of higher vertebrates. This study investigated the distribution density and immunoreactive (IR) intensity of 5-hydroxytryptamine (5-HT), gastrin (GAS), somatostatin (SS), pancreatic polypeptide (PP), neuropeptide Y (NPY) and glucagon (GLU) IR cells in the skin of the bullfrog Rana catesbeiana during periods of activity and hibernation. The results indicated that the six types of GHs were all present in the bullfrog skin and were most predominant in the epidermis and mucous glands. In dorsal skin, the density of the GHs-IR cells in mucous glands was higher than that in epidermis except for GAS-IR cells. In ventral skin, the density of 5-HT, PP and NPY-IR cells in mucous glands was also higher than that in the epidermis. During hibernation, the density of the six types of GHs-IR cells and the IR intensity of GAS, SS, NPY and GLU-IR cells in the epidermis of dorsal skin increased significantly. The IR intensity of SS, PP and NPY-IR cells in granular glands of ventral skin also increased significantly during hibernation. These results suggested that multiple types of GHs-IR cells present in the skin of R. catesbeiana, may play important roles in the regulation of the physiological functions of skin. Also, adaptive changes in the density and IR intensity of GHs-IR cells occurred during hibernation.

Distribution of endocrine cells in the digestive tract of Alligator sinensis during the active and hibernating period.

The digestive tract is the largest endocrine organ in the body; the distribution pattern of endocrine cells varies with different pathological and physiological states. The aim of the present study was to investigate the distributed density of 5-hydroxytryptamine (5-HT), gastrin (GAS), somatostatin (SS) and vasoactive intestinal peptide (VIP) immunoreactive (IR) cells in the digestive tract of Alligator sinensis during the active and hibernating period by immunohistochemical (IHC) method. The results indicated that 5-HT-IR cells were distributed throughout the entire digestive tract, which were most predominant in duodenum and jejunum. The density increased significantly in stomach and duodenum during hibernation. GAS-IR cells were limited in small stomach and small intestine. The density decreased significantly in small stomach during hibernation, while increased in duodenum. What's more, most of the endocrine cells in duodenum were generally spindle shaped with long cytoplasmic processes ending in the lumen during hibernation. SS-IR cells were limited in stomach and small stomach. The density increased in stomach while decreased in small stomach during hibernation, meanwhile, fewer IR cells occurred in small intestine. VIP-IR cells occurred in stomach and small stomach. The density decreased in small stomach, while increased in stomach during hibernation. These results indicated that the endocrine cells in different parts of digestive tract varied differently during hibernation, their changes were adaptive response to the hibernation.