Tree diversity in agroforestry systems of native fine-aroma cacao, Amazonas, Peru.

Cocoa cultivation is of considerable economic and social importance to the Amazonas region and is commonly associated with forest species in the region. However, the diversification level and composition of cacao agroforestry systems in Peru are poorly understood. The objective of this study is, therefore, to describe the diversity of tree species in cocoa AFS by plantation age. Accordingly, the number of species of 15 plots covering a total of 1.5 hectares was recorded. Moderately low levels of tree species diversity were reported (H´ ranged 0.89-1.45). In total 17 species were reported throughout the study area. The most abundant botanical family was represented by a single Musa sp. species. The dissimilarity indices show a moderate similarity between the age ranges evaluated (over 62%). Additionally, the IVI indicates that the most important species are used for food and timber apart from providing shade, additionally major of this species are introduced intentionally for the farmers. Based on the observations, it may be concluded that the farmer's interest in obtaining further benefits from the plot, mostly economic benefits affect the diversification of cocoa agroforestry systems.

Mimicking the Organic and Inorganic Composition of Anabolic Bone Enhances Human Mesenchymal Stem Cell Osteoinduction and Scaffold Mechanical Properties.

Engineered bone graft designs have been largely inspired by adult bone despite functionally significant differences from the composition of anabolic bone in both the mineralized and non-mineralized fractions. Specifically, anabolic bone contains hydroxyapatite with ionic substitutions that facilitate bone turnover and relatively rare collagens type VI and XII that are important for normal bone development. In this work, human mesenchymal stem cells (hMSCs) were cultured in lyophilized collagen type I scaffolds mineralized with hydroxyapatite containing Mg2+ substitutions, then induced to deposit an extracellular matrix (ECM) containing collagens VI and XII by exposure to GW9662, a PPARγ inhibitor. Delivery of GW9662 was accomplished through either Supplemented Media or via composite microspheres embedded in the scaffolds for localized delivery. Furthermore, hMSCs and scaffolds were cultured in both static and perfuse conditions to investigate the interaction between GW9662 treatment and perfusion and their effects on ECM deposition trends. Perfusion culture enhanced cell infiltration into the scaffold, deposition of collagen VI and XII, as well as osteogenic differentiation, as determined by gene expression of osteopontin, BMP2, and ALP. Furthermore, scaffold mineral density and compressive modulus were increased in response to both GW9662 treatment and perfusion after 3 weeks of culture. Local delivery of GW9662 with drug-eluting microspheres had comparable effects to systemic delivery in the perfusate. Together, these results demonstrate a strategy to create a scaffold mimicking both organic and inorganic characteristics of anabolic bone and its potential as a bone graft.

Conditioning of 3D Printed Nanoengineered Ionic-Covalent Entanglement Scaffolds with iP-hMSCs Derived Matrix.

Additive manufacturing is a promising method for producing customized 3D bioactive constructs for regenerative medicine. Here, 3D printed highly osteogenic scaffolds using nanoengineered ionic-covalent entanglement ink (NICE) for bone tissue engineering are reported. This NICE ink consists of ionic-covalent entanglement reinforced with Laponite, a 2D nanosilicate (nSi) clay, allowing for the printing of anatomic-sized constructs with high accuracy. The 3D printed structure is able to maintain high structural stability in physiological conditions without any significant swelling or deswelling. The presence of nSi imparts osteoinductive characteristics to the NICE scaffolds, which is further augmented by depositing pluripotent stem cell-derived extracellular matrix (ECM) on the scaffolds. This is achieved by stimulating human induced pluripotent stem cell-derived mesenchymal stem cells (iP-hMSCs) with 2-chloro-5-nitrobenzanilide, a PPARγ inhibitor that enhances Wnt pathway, resulting in the deposition of an ECM characterized by high levels of collagens VI and XII found in anabolic bone. The osteoinductive characteristics of these bioconditioned NICE (bNICE) scaffolds is demonstrated through osteogenic differentiation of bone marrow derived human mesenchymal stem cells. A significant increase in the expression of osteogenic gene markers as well as mineralized ECM are observed on bioconditioned NICE (bNICE) scaffolds compared to bare scaffolds (NICE). The bioconditioned 3D printed scaffolds provide a unique strategy to design personalized bone grafts for in situ bone regeneration.

Nanoengineered Ionic-Covalent Entanglement (NICE) Bioinks for 3D Bioprinting.

We introduce an enhanced nanoengineered ionic-covalent entanglement (NICE) bioink for the fabrication of mechanically stiff and elastomeric 3D biostructures. NICE bioink formulations combine nanocomposite and ionic-covalent entanglement (ICE) strengthening mechanisms to print customizable cell-laden constructs for tissue engineering with high structural fidelity and mechanical stiffness. Nanocomposite and ICE strengthening mechanisms complement each other through synergistic interactions, improving mechanical strength, elasticity, toughness, and flow properties beyond the sum of the effects of either reinforcement technique alone. Herschel-Bulkley flow behavior shields encapsulated cells from excessive shear stresses during extrusion. The encapsulated cells readily proliferate and maintain high cell viability over 120 days within the 3D-printed structure, which is vital for long-term tissue regeneration. A unique aspect of the NICE bioink is its ability to print much taller structures, with higher aspect ratios, than can be achieved with conventional bioinks without requiring secondary supports. We envision that NICE bioinks can be used to bioprint complex, large-scale, cell-laden constructs for tissue engineering with high structural fidelity and mechanical stiffness for applications in custom bioprinted scaffolds and tissue engineered implants.

Hybrid nonlinear photoacoustic and reflectance confocal microscopy for label-free subcellular imaging with a single light source.

Nonlinear photoacoustic microscopy is capable of achieving subcellular optically resolved absorption contrast in three dimensions but cannot provide structural context for the acquired images. We have developed a dual-modality imaging system that combines the optical absorption contrast of a nonlinear photoacoustic microscope with the optical scattering contrast of a reflectance confocal microscope. By integrating the confocal detection optics into the optical setup of the nonlinear photoacoustic microscope, the two systems were co-registered and may be acquired at the same time and with the same light source. Simultaneous images of fixed erythrocytes and fibroblasts were measured to demonstrate the complementary information that is provided by the two modalities.

Photocrosslinkable and elastomeric hydrogels for bone regeneration.

Nanocomposite biomaterials are extensively investigated for cell and tissue engineering applications due their unique physical, chemical and biological characteristics. Here, we investigated the mechanical, rheological, and degradation properties of photocrosslinkable and elastomeric nanocomposite hydrogels from nanohydroxyapatite (nHAp) and gelatin methacryloyl (GelMA). The addition of nHAp resulted in a significant increase in mechanical stiffness and physiological stability. Cells readily adhere and proliferate on the nanocomposite surfaces. Cyclic stretching of cells on the elastomeric nanocomposites revealed that nHAp elicited a stronger alignment response in the direction of strain. In vitro studies highlight enhanced bioactivity of nanocomposites as determined by alkaline phosphate (ALP) activity. Overall, the elastomeric and photocrosslinkable nanocomposite hydrogels can be used for minimally invasive therapy for bone regeneration.