The regional-dependent biaxial behavior of young and aged mouse skin: A detailed histomechanical characterization, residual strain analysis, and constitutive model Publication date: Available online 12 October 2019 Source: Acta Biomaterialia Author(s): William D. Meador, Gabriella P. Sugerman, Hannah M. Story, Ashley W. Seifert, Matthew R. Bersi, Adrian B. Tepole, Manuel K. Rausch Abstract Skin fulfills several vital functions, many of which are dependent on its mechanical properties. Therefore, as mice have become an invaluable model for skin research, determining murine skin's mechanical properties is important. Specifically, skin's mechanical properties are important for functional tests as well as for prognostic and diagnostic purposes. Additionally, computational simulations of skin behavior are becoming commonplace, rendering accurate models of murine skin's constitutive behavior necessary. To date, our knowledge of mouse skin mechanics shows significant gaps. For example, there are no comprehensive reports correlating skin's mechanical properties with region, age, and direction. Moreover, mouse skin's residual strain behavior has not been reported on. In our current work, we set out to fill these gaps. Based on histology, 2-photon microscopy, and planar biaxial testing, while accurately tracking various reference configurations, we report on differences in gross structure, microstructural organization, and constitutive response of skin, and cast those properties into a versatile Fung-type hyperelastic constitutive law for three reference configurations. Our data is the most comprehensive report contrasting the mechanical properties of young (12 weeks) and aged (52 weeks) mouse skin and will, thus, be valuable to basic science, as control data, and provide accurate constitutive laws for mouse skin modeling. STATEMENT OF SIGNIFICANCE: Our findings are significant as they fill several gaps in our understanding of mouse skin mechanics. This is particularly important as mouse skin is becoming a frequent and critical model of human skin for cosmetic and medical science. Specifically, we quantified how mechanical properties of mice skin vary with age, with location, and with direction. Additionally, we cast our findings into constitutive models that can be used by others for predictive computer simulations of skin behavior. Graphical abstract

Inflammatory response to magnesium-based biodegradable implant materials Publication date: Available online 12 October 2019 Source: Acta Biomaterialia Author(s): M.D. Costantino, A. Schuster, H. Helmholz, A. Meyer-Rachner, R. Willumeit-Römer, B.J.C. Luthringer-Feyerabend Abstract Biodegradability and mechanical properties of magnesium alloys are attractive for orthopaedic and cardiovascular applications. In order to study their cytotoxicity usually bone cells are used. However, after implantation, diverse and versatile cells are recruited and interact. Among the first ones coming into play are cells of the immune system, which are responsible for the inflammatory reaction. Macrophages play a central role in the inflammatory process due to the production of cytokines involved in the tissue healing but also in the possible failure of the implants. In order to evaluate the in vitro influence of the degradation products of magnesium-based alloys on cytokine release, the extracts of pure magnesium and two magnesium alloys (with gadolinium and silver as alloying elements) were examined in an inflammatory in vitro model. Human promonocytic cells (U937 cells) were differentiated into macrophages and further cultured with magnesium-based extracts for 1 and 3 days (simulating early and late inflammatory reaction phases), either at 37°C or at 39°C (mimicking normal and inflammatory conditions, respectively). All extracts exhibit very good cytocompatibility on differentiated macrophages. Results suggest that M1 and even more M2 profiles of macrophage were stimulated by the extracts of Mg. Furthermore, Mg-10Gd and Mg-2Ag extracts introduced a nuancing effect by rather inhibiting macrophage M1 profile. Magnesium-based biomaterials could thus induce a faster inflammation resolution while improving tissue repair. Statement of Significance Macrophage are the key-cells during inflammation and can influence the fate of tissue healing and implant performance. Magnesium-based implants are biodegradable and bioactive. Here we selected an in vitro system to model early and late inflammation and effect of pyrexia (37°C versus 39°C). We showed the beneficial and nuancing effects of magnesium (Mg) and the selected alloying elements (silver (Ag) and gadolinium (Gd)) on the macrophage polarisation. Mg extracts exacerbated simultaneously the macrophage M1 and M2 profiles while Mg-2Ag and Mg-10Gd rather inhibited the M1 differentiation. Furthermore, 39°C exhibited protective effect by either decreasing cytokine production or promoting anti-inflammatory ones, with or without extracts. Mg-based biomaterials could thus induce a faster inflammation resolution while improving tissue repair. Graphical abstract

Single-Dose Combination Nanovaccine Induces Both Rapid and Long-Lived Protection Against Pneumonic Plague Publication date: Available online 11 October 2019 Source: Acta Biomaterialia Author(s): Danielle A Wagner-Muñiz, Sean M Kelly, Andrew C Petersen, Nathan Peroutka-Bigus, Ross J Darling, Brian H Bellaire, Michael J Wannemuehler, Balaji Narasimhan Abstract Yersinia pestis, the causative agent of pneumonic plague, induces a highly lethal infection if left untreated. Currently, there is no FDA-approved vaccine against this pathogen; however, USAMRIID has developed a recombinant fusion protein, F1-V, that has been shown to induce protection against pneumonic plague. Many F1-V-based vaccine formulations require prime-boost immunization to achieve protective immunity, and there are limited reports of rapid induction of protective immunity (≤ 14 days post-immunization (DPI)). The STimulator of INterferon Genes agonists cyclic dinucleotides (CDNs) have been shown to be promising vaccine adjuvants. Polyanhydride nanoparticle-based vaccines (i.e., nanovaccines) have also shown to enhance immune responses due to their dual functionality as adjuvants and delivery vehicles. In this work, a combination nanovaccine was designed that comprised F1-V-loaded nanoparticles combined with the CDN, dithio-RP,RP-cyclic di-guanosine monophosphate, to induce rapid and long-lived protective immunity against pneumonic plague. All mice immunized with a single dose combination nanovaccine were protected from Y. pestis lethal challenge within 14 DPI and demonstrated enhanced protection over F1-V adjuvanted with CDNs alone at challenge doses ≥7,000 CFU Y. pestis CO92. In addition, 75 percent of mice receiving the single dose of the combination nanovaccine were protected from challenge at 182 DPI, while maintaining high levels of antigen-specific serum IgG. ELISPOT analysis of vaccinated animals at 218 DPI revealed F1-V-specific long-lived plasma cells in bone marrow in mice vaccinated with CDN adjuvanted F1-V or the combination nanovaccine. Microarray analysis of serum from these vaccinated mice revealed the presence of serum antibody that bound to a broad range of F1 and V linear epitopes. These results demonstrate that combining the adjuvanticity of CDNs with a nanovaccine delivery system enables induction of both rapid and long-lived protective immunity against Y. pestis. Graphical abstract

In vitro Evaluation of Pt-coated Electrospun Nanofibers for Endovascular Coil Embolization Publication date: Available online 11 October 2019 Source: Acta Biomaterialia Author(s): Min-Woo Kim, Seongpil An, Hyunjun Seok, Hyunchul Jung, Dong-Hyuk Park, Alexander L. Yarin, Sam S. Yoon Abstract Recently, endovascular coil embolization has been introduced to treat intracranial aneurysms because it has lower morbidity and mortality than surgical clipping. The endovascular coils prevent the extravasation of blood by decreasing the permeability of an aneurysm flow governed by Darcy's law. Here, we developed and explored Pt-coated micro-ropes for potential use as endovascular coils. Electrospinning with subsequent electroplating were employed to fabricate Pt-coated nanofibers, which were tightly twisted to form micro-ropes. The compatibility of Pt micro-ropes with commercial delivery catheters was verified and their performance was experimentally explored in an in vitro experimental model. The developed Pt-coated micro-ropes demonstrated feasibility as efficient and low-cost endovascular coils. Statement of Significance The use of Platinum(Pt)-coated polymer nanofibers to prevent blood extravasation has been demonstrated. These Pt nanofibers were installed within a microfluidic channel, and the resulting reduced permeability was evaluated using a fluid similar to blood. Based on the obtained results, these newly developed nanofibers are expected to decrease the operation cost for aneurysmal subarachnoid hemorrhage (SAH), owing their reduced size and low material cost. Overall, the use of this new material should reduce the operational risk associated with the multiple steps required to place the Pt coils at the SAH site. And the compatibility of Pt micro-ropes with commercial delivery catheters was verified and their performance was experimentally explored in an in vitro experimental model. The developed Pt-coated micro-ropes demonstrated feasibility as efficient and low-cost endovascular coils. Graphical Abstract

Three-dimensional differentiation of human pluripotent stem cell-derived neural precursor cells using tailored porous polymer scaffolds Publication date: Available online 11 October 2019 Source: Acta Biomaterialia Author(s): Ashley R. Murphy, John M. Haynes, Andrew L. Laslett, Neil R. Cameron, Carmel M. O'Brien Abstract This study investigates the utility of a tailored poly(ethylene glycol) diacrylate-crosslinked porous polymeric tissue engineering scaffold, with mechanical properties specifically optimised to be comparable to that of mammalian brain tissue for 3D human neural cell culture. Results obtained here demonstrate the attachment, proliferation and terminal differentiation of both human induced pluripotent stem cell- and embryonic stem cell-derived neural precursor cells (hPSC-NPCs) throughout the interconnected porous network within laminin-coated scaffolds. Phenotypic data and functional analyses are presented demonstrating that this material supports terminal in vitro neural differentiation of hPSC-NPCs to a mixed population of viable neuronal and glial cells for periods of up to 49 days. This is evidenced by the upregulation of TUBB3, MAP2, SYP and GFAP gene expression, as well as the presence of the proteins βIII-TUBULIN, NEUN, MAP2 and GFAP. Functional maturity of neural cells following 49 days 3D differentiation culture was tested via measurement of intracellular calcium. These analyses revealed spontaneously active, synchronous and rhythmic calcium flux, as well as response to the neurotransmitter glutamate. This tailored construct has potential application as an improved in vitro human neurogenesis model with utility in platform drug discovery programs. Graphical abstract

2020 Acta Award Recipients Publication date: Available online 11 October 2019 Source: Acta Biomaterialia Author(s):

Biomimetic Hydrogel for Rapid and Scar-free Healing of Skin Wounds inspired by the healing process of oral mucosa Publication date: Available online 10 October 2019 Source: Acta Biomaterialia Author(s): Xiaoying Kong, Jun Fu, Kai Shao, Lili Wang, Xuefang Lan, Jinsheng Shi Abstract Inspired by the wound healing characteristics of the oral mucosa, a biomimetic hydrogel was prepared to realize the rapid and scar-free healing of skin wounds. Through monitoring the healing process of injured oral mucosa, we find out that the combination of high, rapid and sequential expression of some growth factors and the sterile-moist microenvironment are crucial for re-epithelialization and precise control of the inflammation process. On the base of our findings, a hydrogel loaded with several functional compounds was prepared to achieve a comprehensive simulation of the oral mucosal trauma microenvironment for skin wound healing. After 7 days treatment, the skin wound area of the treated group was only about 20% of that of the untreated group, and the proportion of collagen type III and type I in the treated group was much higher than that of the untreated group, suggesting lighter scar hyperplasia. The comprehensive treatment strategy of sequential expression of growth factors in combination with maintaining of a sterile and humid environment is expected to have great application prospect in the field of chronic trauma repair and cosmetic surgery. Statement of significance Long healing time and scar hyperplasia during wound healing have been a serious problem in the past decades of wound healing research. Oral cavity wound healing occurs in an environment that sustains ongoing physical trauma and is rich in bacteria. Despite this, injuries to the mucosal surface often heal faster than cutaneous wounds and leave less noticeable scars. Therefore, in recent years, many scholars have begun to study the healing mechanism of oral mucosa, which supports a new inspiration for the study of skin wound repair: whether the injured skin can achieve a rapid scar-free healing effect similar to oral mucosa? Imitating the biological process of oral mucosa wound healing would be a promising therapeutic strategy in wound healing. Therefore, inspired by the wound healing characteristics of the oral mucosa, a biomimetic gel was prepared to realize the rapid and scar-free healing of skin wounds. Through monitoring the healing process of injured oral mucosa, the combination of high, rapid and sequential expression of some growth factors and sterile-moist microenvironment was crucial for re-epithelialization and precise control of the inflammation process. The comprehensive treatment strategy of sequential expression of growth factors in combination with maintance of a sterile and humid environment implies its potential use in the field of chronic trauma repair and cosmetic surgery. Graphical abstract

Matrix stiffening induces endothelial dysfunction via the TRPV4/microRNA-6740/ET-1 mechanotransduction pathway Publication date: Available online 10 October 2019 Source: Acta Biomaterialia Author(s): Xiang Song, Zhenwei Sun, Gan Chen, Pan Shang, Guoxing You, Jingxiang Zhao, Sisi Liu, Dong Han, Hong Zhou Abstract Vascular stiffening is associated with the prognosis of cardiovascular disease (CVD). Endothelial dysfunction, as shown by reduced vasodilation and increased vasoconstriction, not only affects vascular tone, but also accelerates the progression of CVD. However, the precise effect of vascular stiffening on endothelial function and its mechanism is unclear and a possible underlying has not been determined. In this study, we found that increasing substrate stiffness promoted endothelin-1 (ET-1) expression and inhibited endothelial nitric oxide synthase expression in human umbilical vein endothelial cells. Additionally, miR-6740-5p was identified as a stiffness-sensitive microRNA, which was downregulated by a stiff substrate, resulting in increased ET-1 expression. Furthermore, we found that substrate stiffening reduced the expression and activity of the calcium channel TRPV4, which subsequently enhanced ET-1 expression by inhibiting miR-6740-5p. Finally, analysis of clinical plasma samples showed that plasma miR-6740-5p levels in patients with carotid atherosclerosis were significantly lower than those in healthy people. Taken together, our findings show a novel mechanically regulated TRPV4/miR-6740/ET-1 signaling axis by which substrate stiffness affects endothelial function. Our findings indicate that vascular stiffening induces endothelial dysfunction, thereby accelerating progression of CVD. Furthermore, this study indicates that endothelial dysfunction induced by improper biophysical cues from cardiovascular implants may be an important reason for complications arising from the use of cardiovascular implants. Graphical abstract

Bioinspired Multifunctional Biomaterials with Hierarchical Microstructure for Wound Dressing Publication date: Available online 10 October 2019 Source: Acta Biomaterialia Author(s): Jianmin Xue, Xiaocheng Wang, Endian Wang, Tian Li, Jiang Chang, Chengtie Wu Abstract Developing multifunctional wound dressing with desired mechanical strength is of great significance for the treatment of different types of skin wounds. Inspired by the close relationship between strength and hierarchical structure of nacre, hierarchical and porous graphene oxide-chitosan-calcium silicate (GO-CTS-CS) film biomaterials are fabricated in combination of vacuum filtration-assisted assembly and freeze-drying methods. The bioinspired hierarchical materials emulate orderly porous lamellar micron-scale structure and the “brick-and-mortar” layered nanostructure. The hierarchical microstructure endows the GO-CTS-CS biomaterials with good tensile strength, compatible breathability and water absorption. Furthermore, the hierarchical GO-CTS-CS biomaterials exhibit ideal photothermal performance, leading to significant photothermal antibacterial and antitumor efficiency. Meanwhile, the hierarchical GO-CTS-CS biomaterials show stimulatory effect on in vivo chronic wound healing. Therefore, such a high performance and multifunctional biomaterial is believed to offer a promising alternative to traditional wound dressing in future. Graphical Abstract

A unique methionine-rich protein – aragonite crystal complex: structure and mechanical functions of the Pinctada fucata bivalve hinge ligament Publication date: Available online 8 October 2019 Source: Acta Biomaterialia Author(s): Michio Suzuki, Kazuki Kubota, Ryo Nishimura, Lumi Negishi, Kazuki Komatsu, Hioryuki Kagi, Katya Rehav, Sidney Cohen, Steve Weiner Abstract The bivalve hinge ligament holds the two shells together. The ligament functions as a spring to open the shells after they were closed by the adductor muscle. The ligament is a mineralized tissue that bears no resemblance to any other known tissue. About half the ligament is composed of a protein-rich matrix, and half of long and extremely thin segmented aragonite crystals. Here we study the hinge ligament of the pearl oyster Pinctada fucata. FIB SEM shows that the 3D organization is remarkably ordered. The full sequence of the major protein component contains a continuous segment of 30 repeats of MMMLPD. There is no known homologous protein. Knockdown of this protein prevents crystal formation, demonstrating that the integrity of the matrix is necessary for crystals to form. X-ray diffraction shows that the aragonite crystals are more aligned in the compressed ligament, indicating that the crystals may be actively contributing to the elastic properties. The fusion interphase that joins the ligament to the shell nacre is composed of a prismatic mineralized tissue with a thin organic-rich layer at its center. Nanoindentation of the dry interphase shows that the elastic modulus of the nacre adjacent to the interphase gradually decreases until it approximates that of the interphase. The interphase modulus slightly increases until it matches the ligament. All these observations demonstrate that the ligament shell complex is a remarkable biological tissue that has evolved unique properties that enable bivalves to open their shell effectively innumerable times during the lifetime of the animal. Graphical abstract