The macroporous synthetic poly(2-hydroxyethyl methacrylate) (pHEMA) hydrogels as 3D cellular scaffolds with specific internal morphology, so called dual pore size, were designed and studied. The morphological microstructure of hydrogels was characterized in the gel swollen state and the susceptibility of gels for stem cells was evaluated. The effect of specific chemical groups covalently bound in the hydrogel network by copolymerization on cell adhesion and growth, followed by effect of laminin coating were investigated. The evaluated gels contained either carboxyl groups of the methacrylic acid or quaternary ammonium groups brought by polymerizable ammonium salt or their combinations. The morphology of swollen gel was visualized using the laser scanning confocal microscopy. All hydrogels had very similar porous structures – their matrices contained large pores (up to 102 μm) surrounded with gel walls with small pores (100 μm). The total pore volume in hydrogels swollen in buffer solution ranged between 69 and 86 vol%. Prior to the seeding of the mouse embryonal stem cells, the gels were coated with laminin. The hydrogel with quaternary ammonium groups (with or without laminin) stimulated the cell growth the most. The laminin coating lead to a significant and quaternary ammonium groups. The gel chemical modification influenced also the topology of cell coverage that ranged from individual cell clusters to well dispersed multi cellular structures. Findings in this study point out the laser scanning confocal microscopy as an irreplaceable method for a precise and quick assessment of the hydrogel morphology. In addition, these findings help to optimize the chemical composition of the hydrogel scaffold through the combination of chemical and biological factors leading to intensive cell attachment and proliferation.

Spinal cord injury (SCI), is a devastating condition leading to the loss of locomotor and sensory function below the injured segment. Despite some progress in acute SCI treatment using stem cells and biomaterials, chronic SCI remains to be addressed. We have assessed the use of laminin-coated hydrogel with dual porosity, seeded with induced pluripotent stem cell-derived neural progenitors (iPSC-NPs), in a rat model of chronic SCI. iPSC-NPs cultured for 3 weeks in hydrogel in vitro were positive for nestin, glial fibrillary acidic protein (GFAP) and microtubule-associated protein 2 (MAP2). These cell-polymer constructs were implanted into a balloon compression lesion, 5 weeks after lesion induction. Animals were behaviorally tested, and spinal cord tissue was immunohistochemically analyzed 28 weeks after SCI. The implanted iPSC-NPs survived in the scaffold for the entire experimental period. Host axons, astrocytes and blood vessels grew into the implant and an increased sprouting of host TH+ fibers was observed in the lesion vicinity. The implantation of iPSC-NP-LHM cell-polymer construct into the chronic SCI led to the integration of material into the injured spinal cord, reduced cavitation and supported the iPSC-NPs survival, but did not result in a statistically significant improvement of locomotor recovery.

Since the discovery of the hydrophilic swollen gels prepared by the radical polymerization of 2-hydroxyethyl methacrylate by prof. O. Wichterle and his group there has been a great effort to investigate this polymer. It was due not only to the use of poly(2-hydroxyethyl methacrylate) (PHEMA) for hydrogel contact lenses but also to its interesting physical and chemical properties. It is characterized by its ability to swell in both hydrophilic and hydrophobic media and a very good compatibility with living tissues. Even after being swollen in aqueous environment, PHEMA retains a sufficient mechanical strength, flexibility and stability over time. The material has therefore been used in various applications. In addition to ophthalmology, it was investigated for implants or polymer systems for controlled drug delivery. Less known is its use in sorbents or monoliths for chromatographic column packing. However, it remains an object of great scientific interest, as evidenced annually by the number of publications with this key word. At the same time, it represents an important model polymer in both the scientific research of synthetic hydrogels and biomedical applications including tissue engineering. It is the aim of this article to give a clear overview of the history, structure and properties of this unique monomer, its polymers and applications, based on the findings of literature and the results of our own experiments.

To determine whether PHEMA [poly(2-hydroxyethylmethacrylate)] is suitable for portal vein embolization in patients scheduled to right hepatectomy and whether it is as effective as the currently used agent (a histoacryl/lipiodol mixture). Two groups of nine patients each scheduled for extended right hepatectomy for primary or secondary hepatic tumor, had right portal vein embolization in an effort to induce future liver remnant (FLR) hypertrophy. One group had embolization with PHEMA, the other one with the histoacryl/lipiodol mixture. In all patients, embolization was performed using the right retrograde transhepatic access. Embolization was technically successful in all 18 patients, with no complication related to the embolization agent. Eight patients of either group developed FLR hypertrophy allowing extended right hepatectomy. Likewise, one patient in each group had recanalization of a portal vein branch. Histology showed that both embolization agents reach the periphery of portal vein branches, with PHEMA penetrating somewhat deeper into the periphery. PHEMA has been shown to be an agent suitable for embolization in the portal venous system comparable with existing embolization agent (histoacryl/lipiodol mixture).

An extensive study of various properties of gel electrolyte systems based on 2-ethoxyethyl methacrylate (EOEMA) was conducted with several experimental techniques in order to clarify the influence of varying composition of the samples. The addition of 3-(trimethoxysilyl) propyl methacrylate (SPMA), used here with the aim of decreasing flammability of the ether-oxygen rich system, was investigated in terms of resulting mechanical, thermal and electrochemical properties. Furthermore, changes in these properties based on the use of different cross-linker monomers, salt and its solvent were studied.

Cíl: Primárním cílem je zjistit, zda nesíťovaný poly(2-hydroxyethylmethakrylát) (PHEMA) je vhodný pro embolizaci pravostranného portálního řečiště u pacientů před rozšířenou pravostrannou hepatektomií. Sekundárním cílem je srovnání dvou způsobů určování objemu jater: CT volumetrie a určení objemu jater podle povrchu těla. Metodika: Na třech pracovištích byla prováděna embolizace pravostranné porty látkou PHEMA u celkem 22 pacientů. Výkon byl prováděn retrográdní katetrizací pravostranné porty z transhepatického vpichu. Za průměrně 39,7 (10–120) dní byla zhodnocena hypertrofie levého laloku. Hypertrofie levého laloku byla posuzována vzhledem k objemu celých jater vypočteného CT volumetrií a k objemu jater určeného pomocí povrchu těla. Výsledky: U všech pacientů byl výkon technicky úspěšný a podařilo se uzavřít pravostranné portální řečiště. U 14 pacientů stoupla velikost levého laloku o nejméně pět procent celkového objemu jater. V žádném případě nedošlo ke komplikaci, která by souvisela s typem embolizačního materiálu nebo jeho aplikací. Objemy jaterní tkáně určené dvěma různými metodami se od sebe dosti podstatně liší, zejména u extrémně velkých či extrémně malých jater. Jan H. Peregrin1 Jozef Kováč1 Dana Kautznerová1 Eva Honsová3 Martin Oliverius2 Martin Köcher4 Marie Černá4 Vlastimil Válek5 Tomáš Andrašina5 Martin Přádný6 Jiří Michálek6

This chapter contains sections titled: Introduction Methods of Preparation of Porous Hydrogels Hydrogels Crosslinked With Degradable Crosslinkers Hydrogels Degradable in the Main Chain Conclusions Acknowledgments

Macroporous hydrogels of dual porosity based on cross-linked poly(2-hydroxyethyl methacrylate) (pHEMA) were prepared, and their swelling, mechanical responses, and hydraulic permeability, as well as cell adhesion and proliferation, were examined on gel-based scaffolds. Large pores on the order of tens and hundreds of microns were generated by adding a solid porogen (fractionated particles of sodium chloride) to the polymerization mixture, which was subsequently washed out of the gel. Small pores on the order of a few microns were formed by the reaction-induced phase separation mechanism caused by addition of a diluent (1-dodecanol), which is a poor solvent for pHEMA. Morphological studies using light and electron microscopy techniques revealed that the large pores were embedded in the pHEMA matrix containing small fused spherical pores. Mechanical behavior of the dual-porosity hydrogels characterized by oscillatory shear measurements revealed that despite very high pore volume, the hydrogels were sufficiently stiff and self-supportive. The cell cultivation pilot experiment showed that the morphology (dual porosity) demonstrated marked effects on the promotion of cell adhesion, growth, and proliferation, not only on the gel surface but in the large pores within the gel bulk as well.

Background/Aims: The development of hepatocyte-based Bioartificial Liver Assist Devices, intended for the therapy of chronic and fulminant liver failure, is one of the important tasks in the area of tissue engineering. New advances in the development of semipermeable non-woven nanofiber biomaterials and the co-cultivation of bone marrow mesenchymal stromal cells (BMSC) and hepatocytes could be utilized in order to maintain hepatocyte cultures in these devices. Methodology: We have compared rat hepatocyte growth on nanofiber biomaterials from different polymers, 2-hydroxyethylmethacrylate (HEMA) and ethoxyethylmethacrylate (EOEMA) copolymers, polyurethane (PUR), chitosan and polycapronolactone (PCL) spun from different solvent mixtures. Results: In all cases the adhesion of hepatocytes to nanofibers was significantly better/stronger than to unstructured polymer surfaces; coating the nanofibers with collagen did not increase cell adhesion. We found the best hepatocyte adhesion on HEMA/EOEMA copolymer nanofibers and PCL nanofibers spun from a mixture of ethylacetate and dimethyl sulphoxide. Using a migration assay, we observed the migration of BMSC towards hepatocytes; hepatocytes cocultivated with BMSC excreted lower amounts of stress enzymes. Conclusions: The results demonstrate that nonwoven nanofiber layers, particularly those containing BMSC, are a suitable biocompatible support for functional hepatocyte cultures and that they can be used in a laboratory bioreactor or potentially in clinical setting.

We describe a new biopolymer-based nanofibrous material possibly suitable for tissue engineering prepared by an environment-friendly organic solvent-free method. Glycogen, a biodegradable hyperbranched D-glucose polymer, comes from renewable resources and is normally present in man. It forms nanofibres by simple freeze-drying from aqueous solutions with concentration less than 0.5%. However, the architecture of the freeze-dried material depends on the starting biopolymer concentration within the tested range 0.1-5 wt%; in particular higher concentrations produce porous sponge-like structures with communicating pores. Because of the solubility of glycogen in water, nanofibres were modified by solvent-free grafting biodegradable poly(ethyl cyanoacrylate) from vapor phase. Exposing glycogen nanofibres to vapors of ethyl cyanoacrylate only slightly changed the material architecture while producing a water-insoluble biodegradable material with glycogen-to-poly(ethyl cyanoacrylate) ratio depending on the polymerization time. The material was proven to be hydrolytically degradable over the course of several months.

A comparative evaluation of the morphological and swelling properties corresponding to the porous poly(2-hydroxyethyl methacrylate) (PHEMA) hydrogels and to the chitosan (CS) hydrogels was developed in this paper. The porous structure of hydrogels based on PHEMA or CS was tailored by ice-templating process and porogen leaching. Poly(methylmethacrylate) (PMMA), as fractionated particles, was used as polymer porogen. The influence of the average size of the fractionated PMMA particles on the internal morphology and swelling properties of the hydrogels was followed. The average pore diameter of PHEMA cryogels increased from 10 ± 2 μm up to 22 ± 5 μm with the increase of the size of the fractionated PMMA particles from below 32 μm up to 50–90 μm. On the other hand, in the case of CS cryogels prepared in the presence of the PMMA particles with different sizes, pores with an average size of 74 ± 6 μm, irrespective of the size of PMMA particles were formed, strong changes being observed in the morphology of the pore walls, these being less compact and, therefore, more accessible for the diffusion of low molecular weight species. Hydrogels based on PHEMA or CS with microchanneled structures arranged along the freezing direction, were generated by unidirectional freezing. The swelling measurements showed that the cryogels prepared in the presence of PMMA particles attained the equilibrium swelling much faster than those prepared without PMMA.

Currently, there is no effective strategy for the treatment of spinal cord injury (SCI). A suitable combination of modern hydrogel materials, modified to effectively bridge the lesion cavity, combined with appropriate stem cell therapy seems to be a promising approach to repair spinal cord damage. We demonstrate the synergic effect of porosity and surface modification of hydrogels on mesenchymal stem cell (MSC) adhesiveness in vitro and their in vivo survival in an experimental model of SCI. MSCs were seeded on four different hydrogels: hydroxypropylmethacrylate-RGD prepared by heterophase separation (HPMA-HS-RGD) and three other hydrogels polymerized in the presence of a solid porogen: HPMA-SP, HPMA-SP-RGD, and hydroxy ethyl methacrylate [2-(methacryloyloxy)ethyl] trimethylammonium chloride (HEMA-MOETACl). Their adhesion capability and cell survival were evaluated at 1, 7, and 14 days after the seeding of MSCs on the hydrogel scaffolds. The cell-polymer scaffolds were then implanted into hemisected rat spinal cord, and MSC survival in vivo and the ingrowth of endogenous tissue elements were evaluated 1 month after implantation. In vitro data demonstrated that HEMA-MOETACl and HPMA-SP-RGD hydrogels were superior in the number of cells attached. In vivo, the highest cell survival was found in the HEMA-MOETACl hydrogels; however, only a small ingrowth of blood vessels and axons was observed. Both HPMA-SP and HPMA-SP-RGD hydrogels showed better survival of MSCs compared with the HPMA-HS-RGD hydrogel. The RGD sequence attached to both types of HPMA hydrogels significantly influenced the number of blood vessels inside the implanted hydrogels. Further, the porous structure of HPMA-SP hydrogels promoted a statistically significant greater ingrowth of axons and less connective tissue elements into the implant. Our results demonstrate that the physical and chemical properties of the HPMA-SP-RGD hydrogel show the best combination for bridging a spinal cord lesion, while the HEMA-MOETACl hydrogel serves as the best carrier of MSCs.

Currently, there is no effective strategy for the treatment of spinal cord injury (SCI). A combination of biomaterials and stem cell therapy seems to be a promising approach to increase regenerative potential after SCI. We evaluated the use of a cell-polymer construct based on a combination of the conditionally immortalized spinal progenitor cell line SPC-01_GFP3, derived from human fetal spinal cord tissue, with a serotonin-modified poly(2-hydroxyethyl methacrylate) hydrogel (pHEMA-5HT). We compared the effect of treatment with a pHEMA-5HT hydrogel seeded with SPC-01 _GFP3 cells, treatment with a pHEMA-5HT only and no treatment on functional outcome and tissue reconstruction in hemisected rats. Prior to transplantation the cell-polymer construct displayed a high potential to support the growth, proliferation and differentiation of SPC-01 cells in vitro. One month after surgery, combined hydrogel-cell treatment reduced astrogliosis and tissue atrophy and increased axonal and blood vessel ingrowth into the implant; however, two months later only the ingrowth of blood vessels remained increased. SPC-01_GFP3 cells survived well in vivo and expressed advanced markers of neuronal differentiation. However, a majority of the transplanted cells migrated out of the lesion and only rarely remained in the hydrogel. No differences among the groups in motor or sensory recovery were observed. Despite the support of the hydrogel as a cell carrier in vitro, and good results in vivo one month postsurgery, there was only a small effect on long term recovery, mainly due to the limited ability of the hydrogels to support the in vivo growth and differentiation of cells within the implant. Further modifications will be necessary to achieve stable long term improvement in functional outcome.

Hydrogel nanoparticles, referred to also as nanogels, are of special interest for medical and pharmaceutical applications. Due to small size in the range below the diameter of the capillaries, they are proposed as drug delivery carriers. The aim of the study was to estimate the influence of composition and reaction conditions during synthesis of poly-N-isopropyl acrylamide cross-linked by polyethylene glycol diacrylate on the purification rates of the polymer. Six types of thermosensitive nanogels were prepared by surfactant-free dispersion polymerization and assessed in terms of process yield, composition, and size at temperatures below and over volume phase temperature. During the diffusion of impurities, in the course of dialysis, assessed by the conductometric method, the remarkable influence of temperature and initiator concentration on the process was revealed. The release rates varied in the range between 9.63 · 10−2 and 1.39 · 10−1 h−1 in the first stage of the process, whereas in the second stage they were between 2.09 · 10−2 and 6.28 · 10−2 h−1. The evaluated time to obtain acceptable purity of the preparation was estimated to be in the range of 18 days. More detailed research should be directed towards the influence of the structure of obtained material on the purification process.

Preparation and morphological characterization of some novel hydrogels based on chitosan (CS) with porous structure tailored by ice-templating and porogen leaching are presented in the paper. Poly(methylmethacrylate) (PMMA), as fractionated particles, has been used as polymer porogen. The influence of the mesh of the fractionated PMMA particles, the weight ratio between CS and fractionated PMMA particles, and the speed of the crystallization, on the internal morphology of the hydrogels have been deeply investigated. The morphology of the obtained hydrogels was observed by scanning electron microscopy (SEM). As a function of the synthesis conditions, hydrogels with a heterogeneous morphology consisting of randomly and evenly distributed polyhedral pores, or with an oriented structure, which has microchanneled structures arranged along the freezing direction, were generated.

The structural properties of microfiber meshes made from poly(2-hydroxyethyl methacrylate) (PHEMA) were found to significantly depend on the chemical composition and subsequent cross-linking and nebulization processes. PHEMA microfibres showed promise as scaffolds for chondrocyte seeding and proliferation. Moreover, the peak liposome adhesion to PHEMA microfiber scaffolds observed in our study resulted in the development of a simple drug anchoring system. Attached foetal bovine serum-loaded liposomes significantly improved both chondrocyte adhesion and proliferation. In conclusion, fibrous scaffolds from PHEMA are promising materials for tissue engineering and, in combination with liposomes, can serve as a simple drug delivery tool.

Reductively biodegradable hydrogels based on poly[N-(2-hydroxypropyl)methacrylamide] crosslinked with N-[3-(methacryloylamino)propyl] -6-{[5-({[3-(methacryloylamino)propyl]amino}carbonyl)-2-pyridyl]disulfanyl}nicotinamide intended for tissue engineering were synthesized and characterized. The rate of irreversible reductive degradation with thiol l-cysteine (a model of human body where several thiols are present in extracellular space) was studied using several l-cysteine concentrations. The mechanism of the irreversible reductive bond cleavage in the crosslinker structure was confirmed on a low-molecular-weight model. The hydrogel is stable during storage in phosphate buffered saline and is degraded relatively quickly in a concentration-dependent manner after addition of l-cysteine to the surrounding medium.

We describe the synthesis of new crosslinker N′-{3-[2-(4-{2-[3-(methacryloylamino)propanoyl]hydrazono}cyclohexyliden)hydrazino]-3-oxopropyl}-2-methylacrylamide, which is stable at physiological pH, but is hydrolytically cleaved in acidic pH. Study of acid hydrolysis showed that the crosslinker is hydrolyzed into two N′-(3-hydrazino-3-oxopropyl)-2-methylacrylamide molecules and one 1,4-cyclohexanedione molecule. Hydrogels based on poly[N-(2-hydroxypropyl)methacrylamide] crosslinked with this crosslinker were prepared and their degradation rate was studied as a function of pH (in the range of pH 1.5–7.4). The hydrogel is cleaved within 8–27 days at pH between 1.5 and 6 and is stable at pH 7.4 making it promising for the construction of oesophageal stents.

Synthetic hydrogels are often used in biomedical applications as many of them are compatible with living tissue and moreover they can meet most criteria for artificial tissue properties. For applications in tissue engineering modification of polymer surface using some bioactive compounds (e. g. saccharides, proteins) for promoting the process of cell adhesion and proliferation is widely used. In this work, a series of modified hydrogels was prepared by three different methods: by copolymerization of 2-hydroxyethyl methacrylate with methacrylic acid, by hydrolysis and oxidation of poly(2-hydroxyethyl methacrylate) to obtain carboxyl-rich supports. The influence of the reaction conditions of the hydrogel surface treatment on the total carboxylic group content and the swelling degree was studied. The modified hydrogels were characterized by attenuated total reflectance FT-IR spectroscopy and refractive index measurements. Obtained carboxylic groups on the hydrogel surface have allowed the immobilization of avidin in two ways: 1) electrostatically through dissociated carboxylic groups and 2) covalently bonded through activated carboxylic groups by N-hydroxysuccinimide. The capacity of hydrogels for avidin immobilization was determined by Bradford spectrophotometric method. The results so far obtained from the preliminary biological tests showed that immobilized avidin on the hydrogel surface provides better adhesion and proliferation of keratinocytes compared to supports without avidin.