Deficiency of the SMOC2 matricellular protein impairs bone healing and produces age-dependent bone loss.

Secreted extracellular matrix components which regulate craniofacial development could be reactivated and play roles in adult wound healing. We report a patient with a loss-of-function of the secreted matricellular protein SMOC2 (SPARC related modular calcium binding 2) presenting severe oligodontia, microdontia, tooth root deficiencies, alveolar bone hypoplasia, and a range of skeletal malformations. Turning to a mouse model, Smoc2-GFP reporter expression indicates SMOC2 dynamically marks a range of dental and bone progenitors. While germline Smoc2 homozygous mutants are viable, tooth number anomalies, reduced tooth size, altered enamel prism patterning, and spontaneous age-induced periodontal bone and root loss are observed in this mouse model. Whole-genome RNA-sequencing analysis of embryonic day (E) 14.5 cap stage molars revealed reductions in early expressed enamel matrix components (Odontogenic ameloblast-associated protein) and dentin dysplasia targets (Dentin matrix acidic phosphoprotein 1). We tested if like other matricellular proteins SMOC2 was required for regenerative repair. We found that the Smoc2-GFP reporter was reactivated in adjacent periodontal tissues 4 days after tooth avulsion injury. Following maxillary tooth injury, Smoc2-/- mutants had increased osteoclast activity and bone resorption surrounding the extracted molar. Interestingly, a 10-day treatment with the cyclooxygenase 2 (COX2) inhibitor ibuprofen (30 mg/kg body weight) blocked tooth injury-induced bone loss in Smoc2-/- mutants, reducing matrix metalloprotease (Mmp)9. Collectively, our results indicate that endogenous SMOC2 blocks injury-induced jaw bone osteonecrosis and offsets age-induced periodontal decay.

Functionalization of PTFE Materials Using a Combination of Polydopamine and Platelet-Rich Fibrin.

BACKGROUND: The diaphragm, which forms a physical barrier between the thoracic and the abdominal cavities, is also the major part of the respiratory system. Congenital diaphragmatic hernia (CDH) is a malformation of that partition muscle. Expanded polytetrafluoroethylene (e-PTFE), a synthetic nondegradable biomaterial, is currently used for the repair of diaphragm defects. Indeed, this hydrophobic biomaterial does not promote rapid and dense cell colonization. Surface modifications are needed to favor or even guide cellular responses. MATERIALS AND METHODS: In this context, we present here a practical and effective way of functionalization of the e-PTFE material. We investigated, by using electron microscopy, the coating with PRF (Platelet-Rich Fibrin) of PDA (Polydopamine) treated e-PTFE implant material. RESULTS: We demonstrate that this straightforward chemical functionalization with PDA increases the hydrophilicity of e-PTFE and thus improves tissue integration. Then, we demonstrated that whatever the contact time between PRF and e-PTFE and the centrifugation speed, the PDA coating on the e-PTFE biomaterial promotes further biological events like cell adhesion and spreading. CONCLUSIONS: Our findings clearly show that this composite coating (chemically by using PDA + biologically by using PRF) method of e-PTFE is a simple, interesting and promising way to favor tissular integration of such biomaterials.

Effect of etching the coronal dentin with the rubbing technique on the microtensile bond strength of a universal adhesive system.

BACKGROUND: The adhesion of composite resins to the dentin substrate is influenced by the treatment of the smear layer. While etch-and-rinse systems require dentin to be conditioned with phosphoric acid, self-etching systems preserve the smear layer by incorporating it into the adhesive layer. OBJECTIVES: The objective of this study was to evaluate the influence of etching with the rubbing technique on the microtensile bond strength (µTBS) of a universal adhesive to dentin. MATERIAL AND METHODS: Eighteen extracted teeth were selected. Two etch-and-rinse techniques (with and without rubbing) and a self-etching technique were used to bond the dentin surfaces with a universal adhesive system. After 24 h, the bonded samples were prepared for the µTBS testing. The specimens were loaded with a tensile force at a crosshead speed of 0.5 mm/min until failure. The scanning electron microscope (SEM) analyses were used to reveal the failure modes. The data were statistically analyzed with the one-way analysis of variance (ANOVA) and χ2 tests. RESULTS: The etch-and-rinse system with rubbing produced significantly lower bond strength (42.11 ±9.26 MPa,) than the etch-and-rinse system without rubbing (47.30 ±8.12 MPa) and significantly higher bond strength than the self-etching system (38.07 ±9.49 MPa). CONCLUSIONS: Under the conditions of this study, dentin etched with phosphoric acid for 3 s in the etchandrinse mode with the rubbing technique for a universal adhesive system decreases the µTBS of the composite to dentin.

Effectiveness of Etching by Three Acids on the Morphological and Chemical Features of Dentin Tissue.

AIM: The purpose of this study was to evaluate the microscopic and chemical effects of phosphoric acid gel, phosphoric acid liquid, and polyacrylic acid application for 15 seconds (s) on coronal dentin. MATERIALS AND METHODS: Twelve extracted teeth were selected. Three etching acids were used to prepare the dentin surfaces. Scanning electron microscopy (SEM) and X-ray energy dispersive spectroscopy (EDX) were used to analyze the chemical and morphological changes of the dentinal surfaces, including the depth of demineralization. Collected data were statistically analyzed by the one-way analysis of variance test. RESULTS: Dentin etched with phosphoric acid gel or liquid showed greater peritubular dentin dissolution, including complete removal of the smear layer. In addition, there were many silica particles on the dentin etched by phosphoric acid gel 37%. The dentin that was etched with 25% polyacrylic acid for 15 seconds showed no smear layer removal. Chemical analysis (EDX) showed that dental surfaces etched with phosphoric acid liquid 37% for 15 seconds showed the strongest mineral dissolution at the calcium surface, with a calcium content of 5.25%. On the other hand, EDX analysis of the dental surface etched with 25% polyacrylic acid showed more surface enrichment in calcium (17.19%). CONCLUSION: Although phosphoric acid (gel or liquid) 37% cleans the dental surface, phosphoric acid gel precipitates silica particles on the etched dentin surface. These particles cannot be removed by rinsing off this acid. The application of polyacrylic acid for 15 seconds does not noticeably demineralized dentin, nor remove the smear layer. CLINICAL SIGNIFICANCE: The clinician should use phosphoric acid (gel or liquid) to clean dental surfaces to prepare them for the bonding process. The low demineralizing effects of the polyacrylic acid permits its use near the pulp.

Microscopic and Chemical Assessments of the Filling Ability in Oval-Shaped Root Canals Using Two Different Carrier-Based Filling Techniques.

OBJECTIVES: The aim of this study was to assess the filling ability in oval-shaped canals using two different carrier-based filling techniques. MATERIALS AND METHODS: Twenty-four human mandibular premolars with one oval canal were selected. Canals were shaped using WaveOne Gold Primary and ProGlider. Samples were divided into two groups and filled as follows: Thermafil and GuttaCore. The proportions of gutta-percha-filled areas (GPFAs), sealer-filled areas (SFAs), and void areas (VA), at 2 and 5 mm, were analyzed using optical numeric microscope, scanning electron microscope, and energy-dispersive X-ray. STATISTICAL ANALYSIS: Data were compared by Kruskal-Wallis one-way analysis of variance on ranks, with statistical significance set at α = 0.05. RESULTS: At 2 and 5 mm distances from the apex, this study discloses no statistically different filling ability for the two techniques. Concerning each sample treated using both filling systems, the presence of tags was visualized. At working length (WL)-5, and WL-2, the maximum tag penetration depth for the GuttaCore group into the dentinal tubules was, respectively, 96 µm and 48 µm, whereas the values in the thermafil group were 109 µm, and 55 µm, respectively. CONCLUSIONS: Our results clearly show that Thermafil and GuttaCore can fill oval-shaped canals in appropriate way. Furthermore, we can state that the GuttaCore obturator allows to preserve the same filling ability than Thermafil obturator, in view of the fact that there was no difference, in terms of GPFA, SFA, and VA between the two different carrier-based obturation techniques.

Polydopamine Functionalization: A Smart and Efficient Way to Improve Host Responses to e-PTFE Implants.

Among the different materials used as protheses for the treatment of Congenital Diaphragmatic Hernia, expanded polytetrafluoroethylene (e-PTFE) plays a leading role owing to its mechanical properties as explained in the first part of this review. However, this material is poorly cell adhesive, which is expected for its contact on the abdominal face, but should display specific tissue adhesion on its thoracic exposed faced. A side specific functionalization method is hence required. The deposition of a nanosized polydopamine film on PTFE is known to be possible but immersion of the e-PTFE membrane in an aerated dopamine solution leads to a functionalization not only on both faces of the membrane but also in its porous volume. The fact that polydopamine also forms at the water/air interface has allowed to transfer a polydopamine film on only one face of the e-PTFE membrane. The deposition method and applications of such Janus like membranes are reviewed in the second part of the review.

A New SLC10A7 Homozygous Missense Mutation Responsible for a Milder Phenotype of Skeletal Dysplasia With Amelogenesis Imperfecta.

Amelogenesis imperfecta (AI) is a heterogeneous group of rare inherited diseases presenting with enamel defects. More than 30 genes have been reported to be involved in syndromic or non-syndromic AI and new genes are continuously discovered (Smith et al., 2017). Whole-exome sequencing was performed in a consanguineous family. The affected daughter presented with intra-uterine and postnatal growth retardation, skeletal dysplasia, macrocephaly, blue sclerae, and hypoplastic AI. We identified a homozygous missense mutation in exon 11 of SLC10A7 (NM_001300842.2: c.908C>T; p.Pro303Leu) segregating with the disease phenotype. We found that Slc10a7 transcripts were expressed in the epithelium of the developing mouse tooth, bones undergoing ossification, and in vertebrae. Our results revealed that SLC10A7 is overexpressed in patient fibroblasts. Patient cells display altered intracellular calcium localization suggesting that SLC10A7 regulates calcium trafficking. Mutations in this gene were previously reported to cause a similar syndromic phenotype, but with more severe skeletal defects (Ashikov et al., 2018;Dubail et al., 2018). Therefore, phenotypes resulting from a mutation in SLC10A7 can vary in severity. However, AI is the key feature indicative of SLC10A7 mutations in patients with skeletal dysplasia. Identifying this important phenotype will improve clinical diagnosis and patient management.

Technical Quality of Root Canal Filling in Preclinical Training at Strasbourg University Using Two Teaching Protocols.

OBJECTIVES: The aim of this study was to compare two teaching protocols according to the technical quality of root canal therapy (RCT) and the procedural errors occurred in preclinical training. MATERIALS AND METHODS: Two different groups of students were concerned. The first one (G1) performed a crown-down technique to shape the root canal systems and cold lateral condensation technique to fill them. The second one (G2) performed a step-down technique without initial manual scouting to shape the root canal systems, and cold hydraulic condensation technique, to fill them. G2 used clinical operative microscope to check the access cavity preparation. STATISTICAL ANALYSIS: The quality of RCTs and procedural errors were recorded and analyzed using chi-squared test and t-test. RESULTS: Four hundred sixty-eight root canals from 152 maxillary molars were treated by the G1 students: 46.6% canals were judged as acceptable. Four hundred sixty-nine root canals from 152 mandibular molars were treated by G1: 58.8% canals were judged as acceptable. Five hundred fifteen root canals from 156 maxillary molars were treated by G2 students: 84.1% canals were judged as acceptable. Four hundred ninety-three root canals from 156 mandibular molars were treated by G2: 90.9% canals were judged as acceptable. Among the errors, the incidence of "ledges" and "fractured instruments" was statistically significant in G1 compared with G2, both on maxillary and on mandibular molars. CONCLUSIONS: The molar RCTs performed by G2, who got benefit from the new teaching protocol, resulted in a better quality of root filling and in fewer procedural errors compared with the molar RCTs performed by G1.

Electrochemistry on Stretchable Nanocomposite Electrodes: Dependence on Strain.

Stretchable nanocomposite conductors are essential for engineering of bio-inspired deformable electronics, human-machine interfaces, and energy storage devices. While the effect of strain on conductivity for stretchable conductors has been thoroughly investigated, the strain dependence of multiple other electrical-transport processes and parameters that determine the functionalities and biocompatibility of deformable electrodes has received virtually no attention. The constancy of electrochemical parameters at electrode-fluid interfaces such as redox potentials, impedances, and charge-transfer rate constants on strain is often tacitly assumed. However, it remains unknown whether these foundational assumptions actually hold true for deformable electrodes. Furthermore, it is also unknown whether the previously used charge-transport circuits describing electrochemical processes on rigid electrodes are applicable to deformable electrodes. Here, we investigate the validity of the strain invariability assumptions for an elastic composite electrode based on gold nanoparticles (AuNPs). A comprehensive model of electrode reactions that accurately describes electrochemical processes taking place on nanocomposite electrodes for ferro-/ferricyanide electrochemicals pair at different strains is developed. Unlike rigid gold electrodes, the model circuit for stretchable electrodes is comprised of two parallel impedance segments describing (a) diffusion and redox processes taking place on the open surface of the composite electrode and (b) redox processes that occur in nanopores. AuNPs forming the open-surface circuit support the redox process, whereas those forming the nanopores only increase the double-layer capacitance. The redox potential was found to be strain-independent for tensile deformations as high as 40%. Other parameters, however, display strong strain dependence, exemplified by the 2-2.5 and 27 times increases of active area of the open and nanopore surface area, respectively, after application of 40% strain. Gaining better understanding of the strain-dependent and -independent electrochemical parameters enables both fundamental and practical advances in technologies based on deformable electrodes.

Boric Acid as an Efficient Agent for the Control of Polydopamine Self-Assembly and Surface Properties.

The deposition of polydopamine (PDA) films on surfaces, a versatile deposition method with respect to the nature of the used substrate, is unfortunately accompanied by deposition of insoluble precipitates in solution after a prolonged oxidation time of dopamine solutions. Therefore, there is evident interest to find methods able to stop the deposition of PDA on surfaces and to simultaneously control the self-assembly of PDA in solution to get stable colloidal aggregates. In addition to proposed methods relying on the use of polymers like poly(vinyl alcohol) and proteins like human serum albumin, we show herein that boric acid is an efficient adjuvant that is simultaneously able to stop the self-assembly of PDA in solution as well as on surfaces and to change the adhesive properties of the resulting PDA coatings.

Control of Surface-Localized, Enzyme-Assisted Self-Assembly of Peptides through Catalyzed Oligomerization.

Localized self-assembly allowing both spatial and temporal control over the assembly process is essential in many biological systems. This can be achieved through localized enzyme-assisted self-assembly (LEASA), also called enzyme-instructed self-assembly, where enzymes present on a substrate catalyze a reaction that transforms noninteracting species into self-assembling ones. Very few LEASA systems have been reported so far, and the control of the self-assembly process through the surface properties represents one essential step toward their use, for example, in artificial cell mimicry. Here, we describe a new type of LEASA system based on α-chymotrypsin adsorbed on a surface, which catalyzes the production of (KL)nOEt oligopeptides from a KLOEt (K: lysine; L: leucine; OEt ethyl ester) solution. When a critical concentration of the formed oligopeptides is reached near the surface, they self-assemble into ß-sheets resulting in a fibrillar network localized at the interface that can extend over several micrometers. One significant feature of this process is the existence of a lag time before the self-assembly process starts. We investigate, in particular, the effect of the α-chymotrypsin surface density and KLOEt concentration on the self-assembly kinetics. We find that the lag time can be finely tuned through the surface density in α-chymotrypsin and KLOEt concentration. For a given surface enzyme concentration, a critical KLOEt concentration exists below which no self-assembly takes place. This concentration increases when the surface density in enzyme decreases.

Enamel and dental anomalies in latent-transforming growth factor beta-binding protein 3 mutant mice.

Latent-transforming growth factor beta-binding protein 3 (LTBP-3) is important for craniofacial morphogenesis and hard tissue mineralization, as it is essential for activation of transforming growth factor-ß (TGF-ß). To investigate the role of LTBP-3 in tooth formation we performed micro-computed tomography (micro-CT), histology, and scanning electron microscopy analyses of adult Ltbp3-/- mice. The Ltbp3-/- mutants presented with unique craniofacial malformations and reductions in enamel formation that began at the matrix formation stage. Organization of maturation-stage ameloblasts was severely disrupted. The lateral side of the incisor was affected most. Reduced enamel mineralization, modification of the enamel prism pattern, and enamel nodules were observed throughout the incisors, as revealed by scanning electron microscopy. Molar roots had internal irregular bulbous-like formations. The cementum thickness was reduced, and microscopic dentinal tubules showed minor nanostructural changes. Thus, LTBP-3 is required for ameloblast differentiation and for the formation of decussating enamel prisms, to prevent enamel nodule formation, and for proper root morphogenesis. Also, and consistent with the role of TGF-ß signaling during mineralization, almost all craniofacial bone components were affected in Ltbp3-/- mice, especially those involving the upper jaw and snout. This mouse model demonstrates phenotypic overlap with Verloes Bourguignon syndrome, also caused by mutation of LTBP3, which is hallmarked by craniofacial anomalies and amelogenesis imperfecta phenotypes.

Computed Tomographic Study of the Pediatric Diaphragmatic Growth: Application to the Treatment of Congenital Diaphragmatic Hernia.

Background The prosthesis commonly used for the treatment of congenital diaphragmatic hernia (CDH) lacks elasticity to replace the diaphragm's mechanical properties and does not follow the natural growth of the child treated. Objective To determine the appropriate properties required for the prostheses, a CT study on healthy patients was conducted. Methods Two methods of diaphragmatic surface analysis are assessed: the diaphragmatic surface is either estimated using surface 2D estimations (method 1), or calculated using length measures on thoracoabdominal CT scans from children (method 2). Patients are divided into two groups depending on their age: group 1: n = 9; median age: 2.0 months (0.1-9.5); group 2: n = 9; median age: 182.6 months (158.5-235.5). Growth factor between the two groups is calculated and the two methods are statistically compared. Results The ratio group 2/group 1 of the diaphragmatic surfaces was 4.3 ± 0.2 on the left side and 4.0 ± 0.2 on the right side for method 1, and 5.1 ± 0.2 on the left side and 5.1 ± 0.3 on the right side for method 2. The difference in the median values between both methods is statistically significant for both the left and right sides (p = 0.022 and p = 0.002, respectively). Hence, the two methods cannot be used exchangeably. Conclusion The treatment of CDH with large defect remains a challenge because of the high incidence of hernia recurrence probably due to prosthesis defect; thus it is important to estimate the diaphragmatic surface precisely. We aim to develop a prosthesis material that can be commonly used and found a mean diaphragmatic growth factor of approximately 4 to 5 from early childhood to adolescence.

New insight in the biological integration of polytetrafluoroethylene from an explant used for diaphragm repair.

Congenital diaphragmatic hernia is a severe disease requiring diaphragm replacement mostly with expanded polytetrafluoroethylene. Unfortunately, the recurrence rate is high due to prosthesis failure with significant morbidity for the child. To provide a better understanding of the integration and possible failure processes of the biomaterial implanted in humans, we conducted electron microscopical and mechanical assessments on a prosthesis explant from a child with congenital diaphragmatic hernia presenting a recurrence. Our findings show a major penetration of connective tissue into the expanded polytetrafluoroethylene on the rough side, whereas the smooth side presents few tissue colonization. This penetration is more important in the central area (area A) with large collagen bundles and layers, in comparison to the peripheral areas without prosthesis failure (area B), where few extracellular matrix is produced. The connective tissue penetrates the prosthesis in depth. In contrast, the peripheral areas with prosthesis failure (area C) show very few cells and extracellular matrix, with an oriented organization in comparison to areas A and B. Obviously, the forces applied on the implanted material modulate the cellular behavior of the newly developed tissues. Atomic force microscopic measurements of the biomaterials' surfaces may explain some cellular behaviors according to areas with or without failure.

Aberration-Corrected Transmission Electron Microscopic Study of the Central Dark Line Defect in Human Tooth Enamel Crystals.

Angstrom resolution images of human tooth enamel (HTE) crystallites were obtained using aberration-corrected high-resolution transmission electron microscopy and atomic-resolution scanning transmission electron microscopy in the modes of bright field, annular dark field, and high-angle annular dark-field. Images show that the central dark line (CDL) defect observed around the center of the HTE crystals is a site for caries formation in the HTE and has a thickness of ~0.2 nm. Results also suggest that the CDL goes through one of the OH- planes.

Retinoic Acid Excess Impairs Amelogenesis Inducing Enamel Defects.

Abnormalities of enamel matrix proteins deposition, mineralization, or degradation during tooth development are responsible for a spectrum of either genetic diseases termed Amelogenesis imperfecta or acquired enamel defects. To assess if environmental/nutritional factors can exacerbate enamel defects, we investigated the role of the active form of vitamin A, retinoic acid (RA). Robust expression of RA-degrading enzymes Cyp26b1 and Cyp26c1 in developing murine teeth suggested RA excess would reduce tooth hard tissue mineralization, adversely affecting enamel. We employed a protocol where RA was supplied to pregnant mice as a food supplement, at a concentration estimated to result in moderate elevations in serum RA levels. This supplementation led to severe enamel defects in adult mice born from pregnant dams, with most severe alterations observed for treatments from embryonic day (E)12.5 to E16.5. We identified the enamel matrix proteins enamelin (Enam), ameloblastin (Ambn), and odontogenic ameloblast-associated protein (Odam) as target genes affected by excess RA, exhibiting mRNA reductions of over 20-fold in lower incisors at E16.5. RA treatments also affected bone formation, reducing mineralization. Accordingly, craniofacial ossification was drastically reduced after 2 days of treatment (E14.5). Massive RNA-sequencing (RNA-seq) was performed on E14.5 and E16.5 lower incisors. Reductions in Runx2 (a key transcriptional regulator of bone and enamel differentiation) and its targets were observed at E14.5 in RA-exposed embryos. RNA-seq analysis further indicated that bone growth factors, extracellular matrix, and calcium homeostasis were perturbed. Genes mutated in human AI (ENAM, AMBN, AMELX, AMTN, KLK4) were reduced in expression at E16.5. Our observations support a model in which elevated RA signaling at fetal stages affects dental cell lineages. Thereafter enamel protein production is impaired, leading to permanent enamel alterations.

Generating and characterizing the mechanical properties of cell-derived matrices using atomic force microscopy.

Mechanical interaction between cells and their surrounding extracellular matrix (ECM) controls key processes such as proliferation, differentiation and motility. For many years, two-dimensional (2D) models were used to better understand the interactions between cells and their surrounding ECM. More recently, variation of the mechanical properties of tissues has been reported to play a major role in physiological and pathological scenarios such as cancer progression. The 3D architecture of the ECM finely tunes cellular behavior to perform physiologically relevant tasks. Technical limitations prevented scientists from obtaining accurate assessment of the mechanical properties of physiologically realistic matrices. There is therefore a need for combining the production of high-quality cell-derived 3D matrices (CDMs) and the characterization of their topographical and mechanical properties. Here, we describe methods that allow to accurately measure the young modulus of matrices produced by various cellular types. In the first part, we will describe and review several protocols for generating CDMs matrices from endothelial, epithelial, fibroblastic, muscle and mesenchymal stem cells. We will discuss tools allowing the characterization of the topographical details as well as of the protein content of such CDMs. In a second part, we will report the methodologies that can be used, based on atomic force microscopy, to accurately evaluate the stiffness properties of the CDMs through the quantification of their young modulus. Altogether, such methodologies allow characterizing the stiffness and topography of matrices deposited by the cells, which is key for the understanding of cellular behavior in physiological conditions.

Airway administration of a highly versatile peptide-based liposomal construct for local and distant antitumoral vaccination.

With the discovery of tumor-associated antigens such as ErbB2, vaccination is considered as a promising strategy to prevent the development of cancer or treat the existing disease. Among routes of immunization, the respiratory route provides the opportunity to develop non-invasive approach for vaccine delivery. In the current study, this administration route was used in order to investigate the potency of a highly versatile di-epitopic liposomal construct to exhibit local or distant antitumoral efficiency after prophylactic or therapeutic vaccination in mice. Well-characterized liposomes, containing the ErbB2 (p63-71) TCD8(+) and HA (p307-319) TCD4(+) peptide epitopes and the Pam2CAG adjuvant, were formulated and administered into the airway of naïve BALB/c mice. The nanoparticle vaccine candidate induced local and specific systemic immune response, as measured by immune cell infiltration and chemokine and cytokine production in BALF or lung tissue, and by spleen T-cell activation ex vivo, respectively. This potent immune response resulted in an efficient antitumor activity against both lung and solid s.c. tumors. Interestingly, the antitumor efficacy was observed after both prophylactic and therapeutic vaccinations, which are the most judicious ones to fight cancer. Our data showed an undeniable interest of liposomal peptide-based vaccines in antitumor vaccination by the respiratory route, opening new perspectives for cancer treatment.

Priming cells for their final destination: microenvironment controlled cell culture by a modular ECM-mimicking feeder film.

Mammalian cell culture is the starting point in many research studies focusing on biomedical applications. However, researchers have little control over the standardized cell microenvironment parameters. Here a modular ECM-mimicking surface coating for cell culture environment is designed. This substrate is a new and versatile thin film obtained by spin-coating of concentrated gelatin crosslinked by transglutaminase. It can be modified with respect to the biochemical and biophysical needs of the final cell destination, i.e. it delivers loaded multi-growth factors and serum components and allows for cell culture in a serum-free culture medium. Also, a well-known cell behavior modulator, the substrate stiffness, is controlled exogenously by addition of nanoparticles. In addition to growth factors, antimicrobial agents such as natural peptides are added to the substrate for limiting the repeated addition of antimicrobial agents to the culture medium and to prevent the increase of resistant bacterial strains in the culture environment. Finally, this substrate contains simultaneously ECM components, growth factors, stiffening elements and antimicrobial agents. It provides a favorable microenvironment and sterile conditions. It is a free-of-maintenance system, as cells will grow without addition of serum or antimicrobial cocktails. This low cost and easy-to-use substrate could emerge as a new standard for cell culture.

Bioactive Seed Layer for Surface-Confined Self-Assembly of Peptides.

The design and control of molecular systems that self-assemble spontaneously and exclusively at or near an interface represents a real scientific challenge. We present here a new concept, an active seed layer that allows to overcome this challenge. It is based on enzyme-assisted self-assembly. An enzyme, alkaline phosphatase, which transforms an original peptide, Fmoc-FFY(PO4 (2-) ), into an efficient gelation agent by dephosphorylation, is embedded in a polyelectrolyte multilayer and constitutes the "reaction motor". A seed layer composed of a polyelectrolyte covalently modified by anchoring hydrogelator peptides constitutes the top of the multilayer. This layer is the nucleation site for the Fmoc-FFY peptide self-assembly. When such a film is brought in contact with a Fmoc-FFY(PO4 (2-) ) solution, a nanofiber network starts to form almost instantaneously which extents up to several micrometers into the solution after several hours. We demonstrate that the active seed layer allows convenient control over the self-assembly kinetics and the geometric features of the fiber network simply by changing its peptide density.