Complex vascular anomalies arise from perturbed development of blood or lymphatic vessels, often caused by germline or post-zygotic variants. Complex vascular anomalies give rise to complications including pain, coagulopathies, respiratory failure, and abnormal fluid accumulation in different body compartments, which are often intractable and can be life-threatening despite interventions such as sclerotherapy, embolization, or surgery. Recent work has demonstrated the importance of phenotype-genotype correlation to guide biologically based medical treatments.
The ability to regulate cell-material interactions is important in various applications such as regenerative medicine and cell separation. This study successfully demonstrates that the binding states of cells on a hydrogel surface can be programmed by using hybridized aptamers and triggering complementary sequences (CSs). In the absence of the triggering CSs, the aptamers exhibit a stable, hybridized state in the hydrogel for cell-type-specific catch. In the presence of the triggering CSs, the aptamers are transformed into a new hybridized state that leads to the rapid dissociation of the aptamers from the hydrogel. As a result, the cells are released from the hydrogel. The entire procedure of cell catch and release during the transformation of the aptamers is biocompatible and does not involve any factor destructive to either the cells or the hydrogel. Thus, the programmable hydrogel is regenerable and can be applied to a new round of cell catch and release when needed.
Central conducting lymphatic anomaly (CCLA) due to congenital maldevelopment of the lymphatics can result in debilitating and life-threatening disease with limited treatment options. We identified 4 individuals with CCLA, lymphedema, and microcystic lymphatic malformation due to pathogenic, mosaic variants in KRAS. To determine the functional impact of these variants and identify a targeted therapy for these individuals, we used primary human dermal lymphatic endothelial cells (HDLECs) and zebrafish larvae to model the lymphatic dysplasia. Expression of the p.Gly12Asp and p.Gly13Asp variants in HDLECs in a 2‑dimensional (2D) model and 3D organoid model led to increased ERK phosphorylation, demonstrating these variants activate the RAS/MAPK pathway. Expression of activating KRAS variants in the venous and lymphatic endothelium in zebrafish resulted in lymphatic dysplasia and edema similar to the individuals in the study. Treatment with MEK inhibition significantly reduced the phenotypes in both the organoid and the zebrafish model systems. In conclusion, we present the molecular characterization of the observed lymphatic anomalies due to pathogenic, somatic, activating KRAS variants in humans. Our preclinical studies suggest that MEK inhibition should be studied in future clinical trials for CCLA due to activating KRAS pathogenic variants.
Growth factors are potent signaling molecules that regulate numerous physiological processes. However, the safe and efficient delivery of growth factors remains an unmet goal when growth factors are applied to treat various pathologies. To control the spatiotemporal delivery of growth factors, material carriers such as hydrogels have been investigated. Hydrogels are promising growth factor delivery systems for their high water content and similarities to native tissue. However, the high permeability of hydrogel releases the loaded growth factors rapidly. In addition, the release rate of multiple growth factors cannot be controlled individually when multiple growth factors are incorporated. In order to better control the release rates of growth factors from hydrogels, functionalization strategies using affinity ligands should be explored.
Nucleic acid aptamers are synthetic oligonucleotides that bind to target molecules with high specificity and high affinity. In addition, these aptamers can be chemically modified with various functional groups and conjugated to a variety of biomaterials. For these reasons, hydrogels functionalized with aptamers could be a valuable tool for controlling the release of growth factors. Two main objectives were pursued in this work: 1) characterizing the molecular interactions of aptamers with cognate biomolecules, and 2) develop aptamer-functionalized delivery systems for the controlled release of growth factors. Modulation of the binding affinity can occur through the introduction or removal of steric hindrance or the use base substitutions. In addition, the use of an oligonucleotide complementary to the aptamer can be used to inactivate the aptamer. The aptamers were then used to develop an aptamer-functionalized system for the high retention and regulated release of growth factors. The work described herein presents a promising method to control growth factor delivery for the treatment of many human diseases.
Surfaces functionalized with affinity ligands have been widely studied for applications such as biological separations and cell regulation. While individual ligands can be directly conjugated onto a surface, it is often important to conjugate polyvalent ligands onto the surface to enhance ligand display. This study was aimed at exploring a method for surface functionalization via polymerization of affinity ligands, which was achieved through ligand hybridization with DNA polymers protruding from the surface. The surface with polyvalent ligands was evaluated via aptamer-mediated cell binding. The results show that this surface bound target cells more effectively than a surface directly functionalized with individual ligands in situations with either equal amounts of ligand display or equal amounts of surface reaction sites. Therefore, this study has demonstrated a new strategy for surface functionalization to enhance ligand display and cell binding. This strategy may find broad applications in settings where surface area is limited or the surface of a material does not possess sufficient reaction sites.
DNA growth on nanoparticles: Nanoparticle-mediated molecular recognition and DNA polymerization are integrated for extracellular matrix (ECM) imaging via programmable signal amplification. By using DNA monomers labeled with identical or different imaging reagents, it is promising to achieve single- or multiple-modality imaging of the ECM.
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