BACKGROUND: Mass production of exosomes is a prerequisite for their commercial utilization. This study investigated whether three-dimensional (3D) spheroid culture of mesenchymal stem cells (MSCs) could improve the production efficiency of exosomes and if so, what was the mechanism involved. METHODS: We adopted two models of 3D spheroid culture using the hanging-drop (3D-HD) and poly(2-hydroxyethyl methacrylate) (poly-HEMA) coating methods (3D-PH). The efficiency of exosome production from MSCs in the 3D spheroids was compared with that of monolayer culture in various conditions. We then investigated the mechanism of the 3D spheroid culture-induced increase in exosome production. RESULTS: The 3D-HD formed a single larger spheroid, while the 3D-PH formed multiple smaller ones. However, MSCs cultured on both types of spheroids produced significantly more exosomes than those cultured in conventional monolayer culture (2D). We then investigated the cause of the increased exosome production in terms of hypoxia within the 3D spheroids, high cell density, and non-adherent cell morphology. With increasing spheroid size, the efficiency of exosome production was the largest with the least amount of cells in both 3D-HD and 3D-PH. An increase in cell density in 2D culture (2D-H) was less efficient in exosome production than the conventional, lower cell density, 2D culture. Finally, when cells were plated at normal density on the poly-HEMA coated spheroids (3D-N-PH); they formed small aggregates of less than 10 cells and still produced more exosomes than those in the 2D culture when plated at the same density. We also found that the expression of F-actin was markedly reduced in the 3D-N-PH culture. CONCLUSION: These results suggested that 3D spheroid culture produces more exosomes than 2D culture and the non-adherent round cell morphology itself might be a causative factor. The result of the present study could provide useful information to develop an optimal process for the mass production of exosomes.
Actins ; Anoxia ; Cell Count ; Exosomes ; Mesenchymal Stromal Cells* ; Polyhydroxyethyl Methacrylate

OBJECTIVETo assess the effect of a liquid embolic agent 2-poly-hydroxyethyl -methacrylate (2-P-HEMA) for renal artery embolization in rabbits.

METHODSThe precipitation time of different concentrations (2%, 3.5%, 5%, 6.5%, 8% and 9.5%) of 2-P-HEMA dissolved in different solutions (ethanol, ethanol/iobitridol, and ethanol/Bi2O3) were determined in flowing water. The mixtures of 2-P-HEMA (2%, 5%, and 8%) with ethanol/ Bi2O3 were injected into the renal arteries of the rabbits, and the artery-embolizing effects were assessed using angiography at 2 and 12 weeks after the injection, with also macroscopic and microscopic examination of the embolized kidneys.

RESULTSThe mixtures of 2-P-HEMA and ethanol formed flocculent precipitation a few seconds after injection into flowing water, and the precipitation time showed no significant variations with the concentration of 2-P-HEMA in the mixture. Low and moderate concentrations of 2-P-HEMA could pass through the microcatheter smoothly with little injection resistance, and resulted in complete occlusion of the renal arteries without adhesion to the microcatheter. Angiography at 2 and 12 weeks detected no recanalization of the occluded renal arteries. Macroscopically, the lumen of the renal arteries was found to be occluded by the embolic agents, and deep penetration of the embolic agents into the glomerular arteries was observed microscopically. The mixture containing high-concentration 2-P-HEMA was difficult to deliver through the microcatheter due to high injection resistance.

CONCLUSION2-P-HEMA can be rapidly precipitated after injection into flowing water, and allows complete embolization of the renal arteries of rabbits at proper concentrations, suggesting its great potential as an endovascular liquid embolic agent.

Animals ; Embolization, Therapeutic ; methods ; Female ; Male ; Polyhydroxyethyl Methacrylate ; Rabbits ; Radiography ; Random Allocation ; Renal Artery ; diagnostic imaging ; pathology

Poly (hydroxyethyl methacrylate) (PHEMA) hydrogel for intraocular lens (IOL) materials was synthesized by solution polymerization using 2-hydroxyethyl methacrylate (HEMA) as raw material, ammonium persulfate and sodium pyrosulfite (KPS/SMBS) as catalyst, and trietyleneglycole dimethacrylate (TEGDMA) as cross-linking additive. Effects of reaction time, temperature, dosage of catalyst and cross-linking additive on mechanical strength and the equilibrium water content (EWC) of the PHEMA hydrogel were systematically investigated and their structure and optical property were also characterized. The experimental results showed that the optimum conditions for preparing PHEMA hydrogel are: catalyst 0.5 wt%, cross-linking additive 1.0 wt%, reaction temperature 40 degrees C, reaction time 36 h. Under the optimum conditions, the tensile strength of PHEMA hydrogel prepared is as high as 0.57 MPa, hardness of Shore A is 23.0, EWC is over 40%, and light transmittance is over 97%.
Biocompatible Materials ; chemistry ; Humans ; Hydrogel, Polyethylene Glycol Dimethacrylate ; chemistry ; Lenses, Intraocular ; Polyhydroxyethyl Methacrylate ; chemical synthesis ; chemistry ; Tensile Strength

Artificial corneas have been developed as an alternative to natural donor tissue to replace damaged or diseased corneas. This study was conducted to evaluate the stability and biocompatibility of PHEMA-PMMA [poly (2-hydroxyl methacrylate)-poly (methyl methacrylate)] keratoprostheses in rabbits following penetrating keratoplasty. Sixteen male New Zealand White rabbits aged 16 weeks were divided into three groups. Group I and group II contained six rabbits each, while the control group had four rabbits. Experimental surgery was conducted under general anesthesia. The cornea was penetrated using an 8 mm diameter biopsy punch. In group I (core 5 mm & skirt 3 mm) and group II (core 6 mm & skirt 2 mm), the keratoprosthesis was placed into the recipient full thickness bed and sutured into position with double-layer continuous. In the control group, corneal transplantation using normal allogenic corneal tissue was performed with the same suture method. After four and eight weeks, keratoprosthesis devices were evaluated by histopathological analysis of gross lesions. Post-operative complications were observed, such as extrusion and infection in experimental groups. Most corneas were maintained in the defect site by double-layer continuous suture materials for 4 weeks and kept good light transmission. However, most artificial cornea were extruded before 8 weeks. Overall, combined PHEMA and PMMA appears to have sufficient advantages for production of artificial corneas because of its optical transparency, flexibility and other mechanical features. However, the stability and biocompatibility were not sufficient to enable application in humans and animals at the present time using penetrating keratoplasty. Further studies are essential to improve the stability and biocompatibility with or without other types of keratoplasty.
Anesthesia, General ; Animals ; Biopsy ; Cornea ; Corneal Transplantation ; Humans ; Keratoplasty, Penetrating* ; Male ; Methods ; Pliability ; Polyhydroxyethyl Methacrylate ; Polymethyl Methacrylate ; Rabbits* ; Sutures ; Tissue Donors

A 28-year-old female presented with a palpable mass lesion on the superonasal aspect of her right globe and she had a progressive diplopia. She had a scleral encircling surgery with a Miragel explant (MIRA, Waltham, Mass, USA) for the tractional retinal detachment associated with pars planitis 9 years previously. On examination, she revealed restricted eye movements of her right eye. The magnetic resonance imaging documented a swelling of the Miragel explant that mimicked a periorbital mass lesion. The Miragel explant was removed and fragmentation of the explant was found intraoperatively. The removed Miragel explant was examined by a scanning electron microscopy, and this demonstrated a disintergrated and swollen structural composition of the Miragel explant. Postoperatively, her extraocular movement was almost restored and the retina remained well attached. Alterations in the structural composition of the Miragel explant results in an excessive swelling that causes a restriction of the extraocular movement, and this can mimick a periorbital mass lesion.
Adult ; Diplopia/*etiology ; Female ; Granuloma, Plasma Cell, Orbital/*etiology ; Humans ; Magnetic Resonance Imaging ; Polyhydroxyethyl Methacrylate/*adverse effects/*analogs & derivatives ; Retinal Detachment/surgery ; Scleral Buckling/*adverse effects ; Visual Fields

Poly (acrylic ester) hydrogel materials were widely used in intraocular lens and contact lens because of their excellent optical performance and biocompatibility. In this paper, the bulk copolymerization behavior of hydrophilic hydroxyethyl methacrylat with hydrophobic methyl metharylate was studied; and the optical performance, calcium deposits, equilibrium water content of polymers and its hydrogels obtained by different ratios of monomers were systematically investigated. The experimental results showed that the average light transmittance and the equilibrium water content of the obtained hydrogels increased with the increasing of the hydrophilic monomer content from 0 to 100%; however, the hardness decreased. The highest light transmittance reached 97% and the hardness of Shore A fell from 92 to 25, the equilibrium water content of hydrogel increased from 16% to 64%. The absorbent capacity of copolymers reduced with the adding of cross-linking monomer. When m(hydrophilic monomer): m(hydrophobic monomer) = 90 : 10, the combination property of the polymer and its hydrogel obtained is optimum.
Acrylic Resins ; chemical synthesis ; chemistry ; Biocompatible Materials ; chemical synthesis ; chemistry ; Humans ; Hydrogels ; chemical synthesis ; chemistry ; Lenses, Intraocular ; Polyhydroxyethyl Methacrylate ; chemical synthesis ; chemistry ; Tensile Strength

OBJECTIVETo develop a composite material containing human hair keratin (HHK), collagen sponge (inner layer) and poly 2-hydroxyethyl methacrylate (PHEMA) film that allows sustained release of polydatin and test its effect as a biological dressing in promoting burn wound healing in SD rats.

METHODSThree HHK materials with fast, moderate, and low degradation rates were mixed at the ratio of 4:3:3 to prepare a reticular structure, which was processed into a composite material with bovine tendon-derived collagen sponge, and further complexed with HEMA film containing PD prepared by polymerization. Degree II burn wound was induced in SD rats by scalding and within postburn day 2-5, the wounds were cleansed and covered with the composite material or with glutaraldehyde-treated porcine skin (positive control). At week 1, 2, 4, 6 and 8 following wound dressing, 6 full-thickness skin samples were harvested from the wounds for histological observation and immunohistochemical detection of collagen and elastic fibers, and the wound healing time and healing rate were recorded.

RESULTSThe prepared collagen sponge film was transparent and porous (50-300 microm in diameter) and allowed sustained PD release into normal saline within 48 h. Compared with the porcine skin, the composite material reduced exudation and maintained ideal moisture of the wound, and significantly shortened the wound healing time (P=0.000). On day 7, 14, and 21 following dressing, the composite material and porcine skin significantly increased the wound healing rate as compared with the negative control group (P=0.000), and on day 14, the composite achieved significantly greater healing rate than the porcine skin (P<0.05).

CONCLUSIONHHK-collagen sponge-PHEMA/PD composite as a dressing material promotes burn wound healing in rats by allowing in vivo construction of tissue engineered epidermis. PHEMA is feasible for sustained drug delivery in this composite.

Animals ; Biological Dressings ; Burns ; drug therapy ; Cattle ; Collagen ; therapeutic use ; Drug Delivery Systems ; Drugs, Chinese Herbal ; pharmacology ; Glucosides ; pharmacology ; Humans ; Keratins ; therapeutic use ; Polyhydroxyethyl Methacrylate ; therapeutic use ; Rats ; Rats, Sprague-Dawley ; Stilbenes ; pharmacology ; Swine ; Tissue Engineering ; Wound Healing