Bioactive Scaffolds as a Promising Alternative for Enhancing Critical-Size Bone Defect Regeneration in the Craniomaxillofacial Region.

Reconstruction of critical-size bone defects (CSDs) in the craniomaxillofacial (CMF) region remains challenging. Scaffold-based bone-engineered constructs have been proposed as an alternative to the classical treatments made with autografts and allografts. Scaffolds, a key component of engineered constructs, have been traditionally viewed as biologically passive temporary replacements of deficient bone lacking intrinsic cues to promote osteogenesis. Nowadays, scaffolds are functionalized, giving rise to bioactive scaffolds promoting bone regeneration more effectively than conventional counterparts. This review focuses on the three approaches most used to bioactivate scaffolds: (1) conferring microarchitectural designs or surface nanotopography; (2) loading bioactive molecules; and (3) seeding stem cells on scaffolds, providing relevant examples of in vivo (preclinical and clinical) studies where these methods are employed to enhance CSDs healing in the CMF region. From these, adding bioactive molecules (specifically bone morphogenetic proteins or BMPs) to scaffolds has been the most explored to bioactivate scaffolds. Nevertheless, the downsides of grafting BMP-loaded scaffolds in patients have limited its successful translation into clinics. Despite these drawbacks, scaffolds containing safer, cheaper, and more effective bioactive molecules, combined with stem cells and topographical cues, remain a promising alternative for clinical use to treat CSDs in the CMF complex replacing autografts and allografts.

Multifaceted plant responses to circumvent Phe hyperaccumulation by downregulation of flux through the shikimate pathway and by vacuolar Phe sequestration.

Detrimental effects of hyperaccumulation of the aromatic amino acid phenylalanine (Phe) in animals, known as phenylketonuria, are mitigated by excretion of Phe derivatives; however, how plants endure Phe accumulating conditions in the absence of an excretion system is currently unknown. To achieve Phe hyperaccumulation in a plant system, we simultaneously decreased in petunia flowers expression of all three Phe ammonia lyase (PAL) isoforms that catalyze the non-oxidative deamination of Phe to trans-cinnamic acid, the committed step for the major pathway of Phe metabolism. A total decrease in PAL activity by 81-94% led to an 18-fold expansion of the internal Phe pool. Phe accumulation had multifaceted intercompartmental effects on aromatic amino acid metabolism. It resulted in a decrease in the overall flux through the shikimate pathway, and a redirection of carbon flux toward the shikimate-derived aromatic amino acids tyrosine and tryptophan. Accumulation of Phe did not lead to an increase in flux toward phenylacetaldehyde, for which Phe is a direct precursor. Metabolic flux analysis revealed this to be due to the presence of a distinct metabolically inactive pool of Phe, likely localized in the vacuole. We have identified a vacuolar cationic amino acid transporter (PhCAT2) that contributes to sequestering excess of Phe in the vacuole. In vitro assays confirmed PhCAT2 can transport Phe, and decreased PhCAT2 expression in PAL-RNAi transgenic plants resulted in 1.6-fold increase in phenylacetaldehyde emission. These results demonstrate mechanisms by which plants maintain intercompartmental aromatic amino acid homeostasis, and provide critical insight for future phenylpropanoid metabolic engineering strategies.