Binary polymer brush patterns from facile initiator stickiness for cell culturing.

We report a new initiator stickiness method to fabricate micropatterned binary polymer brush surfaces, which are ideal platforms for studying cell adhesion behavior. The atom transfer radical polymerization (ATRP) initiator, ω-mercaptoundecyl bromoisobutyrate (MUDBr), is found to adsorb on several hosting polymer brushes, including poly[oligo(ethylene glycol)methyl ether methacrylate] (POEGMA), poly(2-hydroxyethyl methacrylate) (PHEMA), and poly(glycidyl methacrylate) (PGMA) brushes. Based on the initiator stickiness, micropatterned initiator molecules are printed onto a layer of homogenous hosting polymer brushes via microcontact printing (µCP), and then, vertically, a patterned second layer of polymer brushes is grown from the initiator areas. With this simple, fast, and additive method, we demonstrate the fabrication of various binary polymer brushes, and show their applications for patterning cell microarrays and controlling cell orientation. This new approach to generating binary polymer brushes shows great potential for the manipulation of interfacial phenomena, facilitating a range of applications from semiconductors and lubrication to fundamental cell biology studies.

Role of CPNE1 in Odontoblastic Differentiation of Rat Stem Cells from Apical Papilla.

CPNE1 is a calcium-dependent, phospholipid-binding protein that is ubiquitously expressed in various tissues and organs. This study investigates the expression and localization of CPNE1 in tooth germ development and the role of CPNE1 in odontoblastic differentiation. In rat tooth germs, CPNE1 is expressed in the odontoblasts and ameloblasts since the late bell stage. The depletion of CPNE1 in the stem cells from apical papilla (SCAPs) clearly inhibits the expression of odontoblastic-related genes and the formation of mineralized nodules during differentiation, while CPNE1 overexpression promotes this process. In addition, CPNE1 overexpression increases AKT phosphorylation during the odontoblastic differentiation of SCAPs. Furthermore, treatment with AKT inhibitor (MK2206) reduces the expression of odontoblastic-related genes in CPNE1 over-expressed SCAPs, and Alizarin Red staining shows reduced mineralization. These results suggest that CPNE1 plays a role in the tooth germ development as well as the odontblastic differentiation of SCAPs in vitro that is related to the AKT signaling pathway.

Interstitial Injection of Hydrogels with High-Mechanical Conductivity Relieves Muscle Atrophy Induced by Nerve Injury.

Injectable hydrogels have been extensively used in tissue engineering where high mechanical properties are key for their functionality at sites of high physiological stress. In this study, an injectable, conductive hydrogel is developed exhibiting remarkable mechanical strength that can withstand a pressure of 500 kPa (85% deformation rate) and display good fatigue resistance, electrical conductivity, and tissue adhesion. A stable covalent cross-linked network with a slip-ring structure by threading amino ß-cyclodextrin is formed onto the chain of a four-armed (polyethylene glycol) amino group, and then reacted with the four-armed (polyethylene glycol) maleimide under physiological conditions. The addition of silver nanowires enhances the hydrogel's electrical conductivity, enabling it to act as a good conductor in vivo. The hydrogel is injected into the fascial space, and the results show that the weight and muscle tone of the atrophied gastrocnemius muscle improve, subsequently alleviating muscle atrophy. Overall, this study provides a simple method for the preparation of a conductive hydrogel with high mechanical properties. In addition, the interstitial injection provides a strategy for the use of hydrogels in vivo.

Medical services for sports injuries and illnesses in the Beijing 2022 Olympic Winter Games.

BACKGROUND: Beijing 2022 Olympic Winter Games was the second Games held amid the COVID-19 pandemic. To a certain extent, it has altered the way sporting activities operate. There is a lack of knowledge on injury risk and illness occurrence in elite winter sport athletes amid the COVID-19 pandemic. This study aimed to describe the incidence of injuries and illnesses sustained during the XXIV Olympic Winter Games in Beijing from February 4 to 20, 2022. METHODS: We recorded the daily number of injuries and illnesses among athletes reported by Beijing 2022 medical staff in the polyclinic, medical venues, and ambulance. We calculated injury and illness incidence as the number of injuries or illnesses occurring during competition or training, respectively, with incidence presented as injuries/illnesses per 100 athlete-days. RESULTS: In total, 2,897 athletes from 91 nations experienced injury or illness. Beijing 2022 medical staff reported 326 injuries and 80 illnesses, equaling 11.3 injuries and 2.8 illnesses per 100 athletes over the 17-day period. Altogether, 11% of the athletes incurred at least one injury and nearly 3% incurred at least one illness. The number of injured athletes was highest in the skating sports (n=104), followed by alpine skiing (n=53), ice track (n=37), freestyle skiing (n=36), and ice hockey (n=35), and was the lowest in the Nordic skiing disciplines (n=20). Of the 326 injuries, 14 (4.3%) led to an estimated absence from training or competition of more than 1 week. A total of 52 injured athletes were transferred to hospitals for further care. The number of athletes with illness (n=80) was the highest for skating (n=33) and Nordic skiing (n=22). A total of 50 illnesses (62.5%) were admitted to the department of dentistry/ophthalmology/otolaryngology, and the most common cause of illness was other causes, including preexisting illness and medicine (n=52, 65%). CONCLUSION: Overall, 11% of athletes incurred at least one injury during the Games, which is similar to the findings during the Olympic Winter Games in 2014 and 2018. Regarding illness, 2% of athletes were affected, which is approximately one-third of the number affected in the 2018 Olympic Winter Games.

Polymer-Assisted Metallization of Mammalian Cells.

Developing biotemplating techniques to translate microorganisms and cultured mammalian cells into metallic biocomposites is of great interest for biosensors, electronics, and energy. The metallization of viruses and microbial cells is successfully demonstrated via a genetic engineering strategy or electroless deposition. However, it is difficult to transform mammalian cells into metallic biocomposites because of the complicated genes and the delicate morphological features. Herein, "polymer-assisted cell metallization" (PACM) is reported as a general method for the transformation of mammalian cells into metallic biocomposites. PACM includes a first step of in situ polymerization of functional polymer on the surface and in the interior of the mammalian cells, and a subsequent electroless deposition of metal to convert the polymer-functionalized cells into metallic biocomposites, which retain the micro- and nanostructures of the mammalian cells. This new biotemplating method is compatible with different cell types and metals to yield a wide variety of metallic biocomposites with controlled structures and properties.

The involvement of genes related to bile secretion pathway in rat tooth germ development.

Tooth formation is accomplished under strict genetic control procedures. Therefore, exploring the gene network system of tooth development has a very positive practical significance for the study of tooth tissue regeneration and the prevention and treatment of tooth abnormalities. Early bell stage is the initial phase of odontoblast formation and dentin matrix deposition in the process of tooth development. Through RNA sequencing and differential gene analysis of the rat tooth germ samples at cap stage and early bell stage, we found that the bile secretion pathway was the most significant difference signal pathway during the development between cap stage and bell stage, which mainly included ABCC3, AQP4, SLC10A1, SLC2A1, SLC4A4, ADCY5, AQP9, CFTR, ATP1A2, ATP1B1 and ATP1A1, totally 11genes. Immunostaining revealed that SLC2A1, SLC4A4, ADCY5 and ATP1B1were mainly expressed in epithelium in bud stage and inner and outer enamel epithelium during the embryonic phase. In the postnatal 1 and postnatal 7, SLC2A1, SLC4A4 and ABCC3 were highly expressed in ameloblasts and odontoblasts while ADCY5, ATP1B1 and SLC10A1was expressed moderately only in odontoblasts. This finding illustrated that the bile secretion pathway related genes may participate in the development of tooth germ.

Advances of Wnt signalling pathway in dental development and potential clinical application.

Wnt signalling pathway is widely studied in many processes of biological development, like embryogenesis, tissue homeostasis and wound repair. It is universally known that Wnt signalling pathway plays an important role in tooth development. Here, we summarized the function of Wnt signalling pathway during tooth initiation, crown morphogenesis, root formation, and discussed the therapeutic potential of Wnt modulators.

Mechanical stabilization of proteolytically degradable polyethylene glycol dimethacrylate hydrogels through peptide interaction.

Balancing enhancement of neurite extension against loss of matrix support in synthetic hydrogels containing proteolytically degradable and bioactive signaling peptides to optimize tissue formation is difficult. Using a systematic approach, polyethylene glycol hydrogels containing concurrent continuous concentration gradients of the laminin derived bioactive signaling peptide, Ile-Lys-Val-Ala-Val (IKVAV), and collagen derived matrix metalloprotease degradable peptide, GPQGIWGQ, were fabricated and characterized. During proteolytic degradation of the concentration gradient hydrogels, the IKVAV and IWGQ cleavage fragment from GPQGIWGQ were found to interact and stabilize the bulk Young's Modulus of the hydrogel. Further testing of discrete samples containing GPQGIWGQ or its cleavage fragments, GPQG and IWGQ, indicates hydrophobic interactions between the peptides are not necessary for mechanical stabilization of the hydrogel, but changes in the concentration ratio between the peptides tethered in the hydrogel and salts and ions in the swelling solution can affect the stabilization. Encapsulation of human induced pluripotent stem cell derived neural stem cells did not reduce the mechanical properties of the hydrogel over a 14 day neural differentiation culture period, and IKVAV was found to maintain concentration dependent effects on neurite extension and mRNA gene expression of neural cytoskeletal markers, similar to previous studies. As a result, this work has significant implications for the analysis of biological studies in matrices, as the material and mechanical properties of the hydrogel may be unexpectedly temporally changing during culture due to interactions between peptide signaling elements, underscoring the need for greater matrix characterization during the degradation and cell culture. STATEMENT OF SIGNIFICANCE: Greater emulation of the native extracellular matrix is necessary for tissue formation. To achieve this, matrices are becoming more complex, often including multiple bioactive signaling elements. However, peptide signaling in polyethylene glycol matrices and amino acids interactions between peptides can affect hydrogel material and mechanical properties, but are rarely studied. The current study identifies such an interaction between laminin derived peptide, IKVAV, and collagen derived matrix metalloprotease degradable peptide, GPQGIWGQ. Previous studies using these peptides did not identify their interactions' ability to mechanically stabilize the hydrogel during degradation. This work underscores the need for greater matrix characterization and consideration of bioactive signaling element effects temporally on the matrix's material and mechanical properties, as they can contribute to cellular response.

[Effect of GLUD1 on proliferation, osteogenic differentiation and mineralization of human dental pulp stem cells].

PURPOSE: To investigate the effect of glutamate dehydrogenase 1 (GLUD1) on proliferation, osteogenic differentiation and mineralization of human dental pulp stem cells (hDPSCs). METHODS: hDPSCs were isolated by tissue-explant method in vitro, and shGLUD1 lentivirus was transfected to knock down the expression of GLUD1. RT-PCR and Western blot were performed to detect the expression of GLUD1. CCK8 assay was used to evaluate cell proliferation. After culture with osteogenic inducing medium for 14 days, alizarin red staining was used to detect the formation of mineralization nodules, and RT-PCR and immunofluorescence staining were performed to detect the expression of Runx2 and OCN, respectively. The data were analyzed with SPSS 20.0 software package. RESULTS: The expression of GLUD1 was significantly increased in hDPSCs after osteogenic induction compared with the control. After transfection with shGLUD1 lentivirus, GLUD1 expression was significantly decreased (P<0.05). Compared with the control group, mineralization nodule formation was significantly decreased in shGLUD1 group after osteogenic induction. The expression of OCN (late-staged markers for osteogenic differentiation) were significantly decreased both in mRNA and protein levels, while the expression of Runx2 (early-staged markers for osteoblast differentiation) was up-regulated. CONCLUSIONS: shGLUD1 inhibits the proliferation, mineralization and the late stage of osteogenic differentiation of hDPSCs in vitro. GLUD1 may play an important role in osteogenic differentiation of hDPSCs.

RNA-Sequencing Analyses Demonstrate the Involvement of Canonical Transient Receptor Potential Channels in Rat Tooth Germ Development.

Tooth development depends on multiple molecular interactions between the dental epithelium and mesenchyme, which are derived from ectodermal and ectomesenchymal cells, respectively. We report on a systematic RNA sequencing analysis of transcriptional expression levels from the bud to hard tissue formation stages of rat tooth germ development. We found that GNAO1, ENO1, EFNB1, CALM1, SIAH2, ATP6V0A1, KDELR2, GTPBP1, POLR2C, SORT1, and members of the canonical transient receptor potential (TRPC) channel family are involved in tooth germ development. Furthermore, Cell Counting Kit 8 (CCK8) and Transwell migration assays were performed to explore the effects of these differentially expressed genes (DEGs) on the proliferation and migration of dental pulp stem cells. Immunostaining revealed that TRPC channels are expressed at varying levels during odontogenesis. The identified genes represent novel candidates that are likely to be vital for rat tooth germ development. Together, the results provide a valuable resource to elucidate the gene regulatory mechanisms underlying mammalian tooth germ development.

The epidemiology of supernumerary teeth and the associated molecular mechanism.

Supernumerary teeth are common clinical dental anomalies. Although various studies have provided abundant information regarding genes and signaling pathways involved in tooth morphogenesis, which include Wnt, FGF, BMP, and Shh, the molecular mechanism of tooth formation, especially for supernumerary teeth, is still unclear. In the population, some cases of supernumerary teeth are sporadic, while others are syndrome-related with familial hereditary. The prompt and accurate diagnosis of syndrome related supernumerary teeth is quite important for some distinctive disorders. Mice are the most commonly used model system for investigating supernumerary teeth. The upregulation of Wnt and Shh signaling in the dental epithelium results in the formation of multiple supernumerary teeth in mice. Understanding the molecular mechanism of supernumerary teeth is also a component of understanding tooth formation in general and provides clinical guidance for early diagnosis and treatment in the future.

Effects of free radical initiators on polyethylene glycol dimethacrylate hydrogel properties and biocompatibility.

Many studies have utilized Irgacure 2959 photopolymerized poly(ethylene glycol) (PEG) hydrogels for tissue engineering application development. Due to the limited penetration of ultraviolet light through tissue, Irgacure 2959 polymerized hydrogels are not suitable for use in tissues where material injection is desirable, such as the spinal cord. To address this, several free radical initiators (thermal initiator VA044, ammonium persulfate (APS)/TEMED reduction-oxidation reaction, and Fenton chemistry) are evaluated for their effects on the material and mechanical properties of PEG hydrogels compared with Irgacure 2959. To emulate the effects of endogenous thiols on in vivo polymerization, the effects of chain transfer agent (CTA) dithiothreitol on gelation rates, material properties, Young's and shear modulus, are examined. Mouse embryonic stem cells and human induced pluripotent stem cell derived neural stem cells were used to investigate the cytocompatibility of each polymerization. VA044 and Fenton chemistry polymerization of PEG hydrogels both had gelation rates and mechanical properties that were highly susceptible to changes in CTA concentration and showed poor cytocompatibility. APS/TEMED polymerized hydrogels maintained consistent gelation rates and mechanical properties at high CTA concentration and had a similar cytocompatibility as Irgacure 2959 when cells were encapsulated within the PEG hydrogels. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3059-3068, 2017.

Indirect Solid Freeform Fabrication of an Initiator-Free Photocrosslinkable Hydrogel Precursor for the Creation of Porous Scaffolds.

In the present work, a photopolymerized urethane-based poly(ethylene glycol) hydrogel is applied as a porous scaffold material using indirect solid freeform fabrication (SFF). This approach combines the benefits of SFF with a large freedom in material selection and applicable concentration ranges. A sacrificial 3D poly(ε-caprolactone) structure is generated using fused deposition modeling and used as template to produce hydrogel scaffolds. By changing the template plotting parameters, the scaffold channel sizes vary from 280 to 360 µm, and the strut diameters from 340 to 400 µm. This enables the production of scaffolds with tunable mechanical properties, characterized by an average hardness ranging from 9 to 43 N and from 1 to 6 N for dry and hydrated scaffolds, respectively. Experiments using mouse calvaria preosteoblasts indicate that a gelatin methacrylamide coating of the scaffolds results in an increased cell adhesion and proliferation with improved cell morphology.

The growth of filaments under macromolecular confinement using scaling theory.

Quantitatively describing macromolecular confinement is still a challenge. Using the assembly of DNA tiles in a polyacrylamide network as a model, we studied the effect of macromolecular confinement on the growth of the filament by scaling theory. The results show that the confinement regulates the morphology, the initial growth rate v, and the eventual length of the filament Nm. The initial growth rate is dependent on the medium viscosity η as ν∝η(-0.94), and the filament adjusts its length in the given confined space as Nm∝ (ξ/Rg)(1.8), with ξ being the mesh size of the polyacrylamide solution and Rg being the radius of gyration of polyacrylamide.

Characterization and optimization of glycerol/sebacate ratio in poly(glycerol-sebacate) elastomer for cell culture application.

Poly(glycerol-sebacate) (PGS) is an elastomeric biodegradable polyester. Our previous series of studies have showed that PGS has good biocompatibility. In view of the potential use of PGS in bioengineering, we attempt to characterize the PGS polymer with different ratio of glycerol and sebacic acid, and the cell adhesion and growth on these polymers. PGSs with different proportion of glycerol and sebacic acid were synthesized by polycondensation reaction. The microstructure of the series PGSs were characterized by infrared spectroscopy and X-ray diffraction analysis (XRD). Results showed that, with the increase of the ratio of sebacic acid in PGS from 1:0.8, 1:1, to 1:1.2 (ratio of glycerol to sebacic acid), the main diffraction peak in XRD, the sol content and gel swelling increased but then decreased, suggesting that the degree of crosslinking and the inherent degree of order of the series PGS increased and then decreased. With the increase of sebacic acid proportion, water absorption increased and then decreased, and the water absorption ranged from 9.62% to 10.66%. The mass loss of the series of samples in degradation experiments ranged from 24.63% to 40.06% on the 32nd day of degradation. Cell culture data suggested that the polymer with the ratio of 1:0.8 for glycerol and sebacate was suitable for cell adhesion and growth. In conclusion, PGS can be used as the cell culture matrix by modifying the composition ratio of glycerol and sebacic acid to improve the properties of cell adhesion and growth.

Effect of hydrion evolution by polylactic-co-glycolic acid coating on degradation rate of pure iron.

For biodegradable iron coronary stents, the major problem is the low degradation rate in body environment. In this study, a new strategy was proposed to increase the degradation rate of iron in vitro. The hydrion evolution was intended to be introduced into the degradation system to increase the degradation rate. To realize this strategy, polylactic-co-glycolic acid (PLGA) was coated onto the surface of pure iron. The degradation process and mechanism of pure iron coated with PLGA were investigated. The results showed that iron coated with PLGA exhibited higher degradation rate in the static immersion test all along. With the degradation of PLGA, the oligomers of PLGA could release abundant H(+) which could dissolve the ferrous oxide to make the electrolyte and oxygen to reach the surface of iron again and simultaneity trigger the hydrion evolution at the middle stage of the degradation. The study also revealed that the solution ions failed to permeate the PLGA coating and the deposition of calcium and phosphorus in the degradation layer was inhibited which further enhanced the degradation.

A poly(glycerol-sebacate-curcumin) polymer with potential use for brain gliomas.

Curcumin has multiple biological and pharmacological activities, including antioxidant, anti-inflammatory, antiviral, antibacterial, antifungal, and antitumor activities. However, the clinical use of curcumin is limited because of its poor oral absorption and extremely poor bioavailability. In order to overcome these limitations, we conjugate curcumin chemically into the known biocompatible and biodegradable polymer, poly(glycerol-sebacate), and prepare the unitary poly(glycerol-sebacate-curcumin) polymer. The structure, the in vitro degradation, the drug release, and antitumor activity as well as the in vivo degradation and tissue biocompatibility of poly(glycerol-sebacate-curcumin) polymer are investigated. The in vitro degradation and drug release profile of poly(glycerol-sebacate-curcumin) are in a linear manner. The in vitro antitumor assay shows that poly(glycerol-sebacate-curcumin) polymer significantly inhibits human malignant glioma cells, U87 and T98 cells. In view of the cytotoxicity against brain gliomas, local use of this polymer would be a potential method for brain tumors.

The polycondensing temperature rather than time determines the degradation and drug release of poly(glycerol-sebacate) doped with 5-fluorouracil.

Poly(glycerol-sebacate) (PGS) is an elastomeric biodegradable polyester that could be used as biodegradable drug carrier. We have previously prepared PGS implants doped with 5-fluorouracil (5-FU-PGSs) and found that 5-FU-PGSs exhibited an initial burst of 5-FU release during in vitro degradation. The synthesis temperature and time are two of the most important reaction conditions for polymer synthesis. Therefore, in order to establish a controllable drug-release manner, we prepared a series of 5-FU-PGS with 2% weight of 5-FU under synthesis conditions with different polycondensing temperature and time and characterized the infrared spectrum properties, in vitro degradation and drug release. Results showed that the polycondensing temperature determined the mechanical properties, degradation and drug release of 5-FU-PGSs. With the polycondensing temperature increasing, the elastic modulus and hardness of 5-FU-PGSs increased, and the mass loss and 5-FU release rate decreased. The polycondensing time had no significant influence on the mechanical property, degradation and drug release of 5-FU-PGSs. We suggest that the polycondensing temperature is the factor to control the drug-release manner.

Glycolic acid modulates the mechanical property and degradation of poly(glycerol, sebacate, glycolic acid).

The development of biodegradable materials with controllable degradation properties is beneficial for a variety of applications. Poly(glycerol-sebacate) (PGS) is a promising candidate of biomaterials; so we synthesize a series of poly(glycerol, sebacate, glycolic acid) (PGSG) with 1:2:0, 1:2:0.2, 1:2:0.4, 1:2:0.6, 1:2:1 mole ratio of glycerol, sebacate, and glycolic acid to elucidate the relation of doped glycolic acid to the degradation rate and mechanical properties. The microstructures of the polymers with different doping of glycolic acid were dissimilar. PGSG with glycolic acid in the ratio of 0.2 displayed an integral degree of ordering, different to those with glycolic acid in the ratio of 0, 0.4, 0.6, and 1, which showed mild phase separation structure. The number, DeltaH(m), and temperature of the PGSG melting peaks tended to decrease with the increasing ratio of doped glycolic acid. In vitro and in vivo degradation tests showed that the degradation rate of PGSG with glycolic acid in the ratio of 0.2 was slowest, but in the ratio range of 0, 0.4, and 0.6, the degradation rate increased with the increase of glycolic acid. All PGSG samples displayed good tissue response and anticoagulant effects. Our data suggest that doping glycolic acid can modulate the microstructure and degree of crosslinking of PGS, thereby control the degradation rate of PGS.

Thin-film enhanced nerve guidance channels for peripheral nerve repair.

It has been demonstrated that nerve guidance channels containing stacked thin-films of aligned poly(acrylonitrile-co-methylacrylate) fibers support peripheral nerve regeneration across critical sized nerve gaps, without the aid of exogenous cells or proteins. Here, we explore the ability of tubular channels minimally supplemented with aligned nanofiber-based thin-films to promote endogenous nerve repair. We describe a technique for fabricating guidance channels in which individual thin-films are fixed into place within the lumen of a polysulfone tube. Because each thin-film is <10 microm thick, this technique allows fine control over the positioning of aligned scaffolding substrate. We evaluated nerve regeneration through a 1-film guidance channel--containing a single continuous thin-film of aligned fibers--in comparison to a 3-film channel that provided two additional thin-film tracks. Thirty rats were implanted with one of the two channel types, and regeneration across a 14 mm tibial nerve gap was evaluated after 6 weeks and 13 weeks, using a range of morphological and functional measures. Both the 1-film and the 3-film channels supported regeneration across the nerve gap resulting in functional muscular reinnervation. Each channel type characteristically influenced the morphology of the regeneration cable. Interestingly, the 1-film channels supported enhanced regeneration compared to the 3-film channels in terms of regenerated axon profile counts and measures of nerve conduction velocity. These results suggest that minimal levels of appropriately positioned topographical cues significantly enhance guidance channel function by modulating endogenous repair mechanisms, resulting in effective bridging of critically sized peripheral nerve gaps.