DSpace Repository :: Browsing by Author "Aguayo, Sebastian"

Browsing by Author "Aguayo, Sebastian"

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Aloe vera peel-derived nanovesicles display anti-inflammatory properties and prevent myofibroblast differentiation
(2024) Ramírez, Orlando; Pomareda, Florencia; Olivares, Belén; Huang, Ya-Lin; Zavala, Gabriela; Carrasco, Javiera; Álvarez, Simón; Leiva, Camila; Hidalgo, Valeria; Romo, Pablo; Sánchez, Matías; Vargas, Ayleen; Martínez, Jessica; Aguayo, Sebastian; Schuh, Christina
Background: Aloe vera (AV) is a medicinal plant, most known for its beneficial effects on a variety of skin conditions. Its known active compounds include carbohydrates and flavonoids such as quercetin and kaempferol, among others. In the past decade, plant nanovesicles (NVs) have gained considerable interest as interkingdom communicators, presenting an opportunity for clinical standardization of natural products. In this study, we aimed to assess the potential of AVpNVs for the treatment of burn wounds. Methods: AVpNVs were isolated and characterized regarding vesicle yield (nanoparticle tracking analysis) and structure (transmission electron microscopy and atomic force microscopy), as well as their protein content with proteomics. We assessed key characteristics for treating burn wounds in vitro, such as the anti-inflammatory potential in LPS-stimulated macrophages and keratinocytes, and the effect of AVpNVs on myofibroblast differentiation and contraction. Key findings: AVpNVs presented a homogenous NV population, vesicular shape, and NV-associated protein markers. AVpNVs significantly decreased the secretion of pro-inflammatory cytokines TNFα, IL-1β, and IL-6. Furthermore, AVpNVs inhibited myofibroblast differentiation and significantly decreased their contractile potential in collagen matrices. Observed effects were linked to proteins identified in the isolates through proteomics analysis. Conclusion: AVpNVs displayed characteristics as an inflammatory modulator, while simultaneously diminishing myofibroblast differentiation and contraction. Novel strategies for burn wound treatment seek to decrease scarring on a cellular and molecular level in the early stages of wound healing, which makes AVpNVs a promising candidate for future plant-vesicle-based treatments.
Antibacterial Effect of Honey-Derived Exosomes Containing Antimicrobial Peptides Against Oral Streptococci
(2021) Leiva-Sabadini, Camila; Alvarez, Simon; Barrera, Nelson P.; Schuh, Christina; Aguayo, Sebastian
Purpose: Recently, our group found exosome-like extracellular vesicles (EVs) in Apis mellifera honey displaying strong antibacterial effects; however, the underlying mechanism is still not understood. Thus, the aim of this investigation was to characterize the molecular and nanomechanical properties of A. mellifera honey-derived EVs in order to elucidate the mechanisms behind their antibacterial effect, as well as to determine differential antibiofilm properties against relevant oral streptococci. Methods: A. mellifera honey-derived EVs (HEc-EVs) isolated via ultracentrifugation were characterized with Western Blot and ELISA to determine the presence of specific exosomal markers and antibacterial cargo, and atomic force microscopy (AFM) was utilized to explore their ultrastructural and nanomechanical properties via non-destructive immobilization onto poly-L-lysine substrates. Furthermore, the effect of HEc-EVs on growth and biofilm inhibition of S. mutans was explored with microplate assays and compared to S. sanguinis. AFM was utilized to describe ultrastructural and nanomechanical alterations such as cell wall elasticity changes following HEc-EV exposure. Results: Molecular characterization of HEc-EVs identified for the first time important conserved exosome markers such as CD63 and syntenin, and the antibacterial molecules MRJP1, defensin-1 and jellein-3 were found as intravesicular cargo. Nanomechanical characterization revealed that honey-derived EVs were mostly <150nm, with elastic modulus values in the low MPa range, comparable to EVs from other biological sources. Furthermore, incubating oral streptococci with EVs confirmed their antibacterial and antibiofilm capacities, displaying an increased effect on S. mutans compared to S. sanguinis. AFM nanocharacterization showed topographical and nanomechanical alterations consistent with membrane damage on S. mutans. Conclusion: Honey is a promising new source of highly active EVs with exosomal origin, containing a number of antibacterial peptides as cargo molecules. Furthermore, the differential effect of HEC-EVs on S. mutans and S. sanguinis may serve as a novel biofilm-modulating strategy in dental caries.
Complex Interaction between Resident Microbiota and Misfolded Proteins: Role in Neuroinflammation and Neurodegeneration
(2020) González-Sanmiguel, Juliana; A P Schuh, Christina M; Muñoz-Montesino, Carola; Contreras-Kallens, Pamina; Aguayo, Luis G; Aguayo, Sebastian
Neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD) and Creutzfeldt-Jakob disease (CJD) are brain conditions affecting millions of people worldwide. These diseases are associated with the presence of amyloid-β (Aβ), alpha synuclein (α-Syn) and prion protein (PrP) depositions in the brain, respectively, which lead to synaptic disconnection and subsequent progressive neuronal death. Although considerable progress has been made in elucidating the pathogenesis of these diseases, the specific mechanisms of their origins remain largely unknown. A body of research suggests a potential association between host microbiota, neuroinflammation and dementia, either directly due to bacterial brain invasion because of barrier leakage and production of toxins and inflammation, or indirectly by modulating the immune response. In the present review, we focus on the emerging topics of neuroinflammation and the association between components of the human microbiota and the deposition of Aβ, α-Syn and PrP in the brain. Special focus is given to gut and oral bacteria and biofilms and to the potential mechanisms associating microbiome dysbiosis and toxin production with neurodegeneration. The roles of neuroinflammation, protein misfolding and cellular mediators in membrane damage and increased permeability are also discussed.
Exosome-like vesicles in Apis mellifera bee pollen, honey and royal jelly contribute to their antibacterial and pro-regenerative activity
(2019) Schuh, Christina; Aguayo, Sebastian; Zavala, Gabriela; Khoury, Maroun
Microvesicles are key players in cellular communication. As glandular secretions present a rich source of active exosomes, we hypothesized that exosome-like vesicles are present in Apis mellifera hypopharyngeal gland secretomal products (honey, royal jelly and bee pollen), and participate in their known antibacterial and pro-regenerative effects. We developed an isolation protocol based on serial centrifugation and ultracentrifugation steps and demonstrated the presence of protein-containing exosome-like vesicles in all three bee-derived products. Assessing their antibacterial properties, we found that exosome-like vesicles had bacteriostatic, bactericidal and biofilm-inhibiting effects on Staphylococcus aureus. Furthermore, we demonstrated that mesenchymal stem cells (MSCs) internalize bee-derived exosome- like vesicles and that these vesicles influence the migration potential of the MSCs. In an in vitro wound-healing assay, honey and royal jelly exosome-like vesicles increased migration of human MSCs, demonstrating their inter-kingdom activity. In summary, we have discovered exosome-like vesicles as a new, active compound in bee pollen, honey and royal jelly.
Modulation of the biophysical and biochemical properties of collagen by glycation for tissue engineering applications
(2022) Vaez, Mina; Asgari, Meisam; Hirvonenc, Liisa; Bakir, Gorkem; Khattignavong, Emilie; Ezzo, Maya; Aguayo, Sebastian; Schuh, Christina; Gough, Kathleen; Bozec, Laurent
The structural and functional properties of collagen are modulated by the presence of intramolecular and intermolecular crosslinks. Advanced Glycation End-products (AGEs) can produce intermolecular crosslinks by bonding the free amino groups of neighbouring proteins. In this research, the following hypothesis is explored: The accumulation of AGEs in collagen decreases its proteolytic degradation rates while increasing its stiffness. Fluorescence Lifetime Imaging (FLIM) and Fourier-transform infrared spectroscopy (FTIR) detect biochemical changes in collagen scaffolds during the glycation process. The accumulation of AGEs increases exponentially in the collagen scaffolds as a function of Methylglyoxal (MGO) concentration by performing autofluorescence measurement and competitive ELISA. Glycated scaffolds absorb water at a much higher rate confirming the direct affinity between AGEs and interstitial water within collagen fibrils. In addition, the topology of collagen fibrils as observed by Atomic Force Microscopy (AFM) is a lot more defined following glycation. The elastic modulus of collagen fibrils decreases as a function of glycation, whereas the elastic modulus of collagen scaffolds increases. Finally, the enzymatic degradation of collagen by bacterial collagenase shows a sigmoidal pattern with a much slower degradation rate in the glycated scaffolds. This study identifies unique variations in the properties of collagen following the accumulation of AGEs. STATEMENT OF SIGNIFICANCE: In humans, Advanced Glycation End-products (AGEs) are naturally produced as a result of aging process. There is an evident lack of knowledge in the basic science literature explaining the biomechanical impact of AGE-mediated crosslinks on the functional and structural properties of collagen at both the nanoscale (single fibrils) and mesoscale (bundles of fibrils). This research, demonstrates how it is possible to harness this natural phenomenon in vitro to enhance the properties of engineered collagen fibrils and scaffolds. This study identifies unique variations in the properties of collagen at nanoscale and mesoscale following accumulation of AGEs. In their approach, they investigate the unique properties conferred to collagen, namely enhanced water sorption, differential elastic modulus, and finally sigmoidal proteolytic degradation behavior.
Nanomechanical and Molecular Characterization of Aging in Dentinal Collagen
(2022) Schuh, Christina; Leiva, Camila; Huang, Sui; Barrera, Nelson; Aguayo, Sebastian
Methylglyoxal (MGO) is an important molecule derived from glucose metabolism with the capacity of attaching to collagen and generating advanced glycation end products (AGEs), which accumulate in tissues over time and are associated with aging and diseases. However, the accumulation of MGO-derived AGEs in dentin and their effect on the nanomechanical properties of dentinal collagen remain unknown. Thus, the aim of the present study was to quantify MGO-based AGEs in the organic matrix of human dentin as a function of age and associate these changes with alterations in the nanomechanical and ultrastructural properties of dentinal collagen. For this, 12 healthy teeth from <26-y-old and >50-y-old patients were collected and prepared to obtain crown and root dentin discs. Following demineralization, MGO-derived AGEs were quantified with a competitive ELISA. In addition, atomic force microscopy nanoindentation was utilized to measure changes in elastic modulus in peritubular and intertubular collagen fibrils. Finally, principal component analysis was carried out to determine aging profiles for crown and root dentin. Results showed an increased presence of MGO AGEs in the organic matrix of dentin in the >50-y-old specimens as compared with the <26-y-old specimens in crown and root. Furthermore, an increase in peritubular and intertubular collagen elasticity was observed in the >50-y-old group associated with ultrastructural changes in the organic matrix as determined by atomic force microscopy analysis. Furthermore, principal component analysis loading plots suggested different "aging profiles" in crown and root dentin, which could have important therapeutic implications in restorative and adhesive dentistry approaches. Overall, these results demonstrate that the organic matrix of human dentin undergoes aging-related changes due to MGO-derived AGEs with important changes in the nanomechanical behavior of collagen that may affect diagnostic and restorative procedures in older people.
Potential novel strategies for the treatment of dental pulp-derived pain: pharmacological approaches and beyond
(2019) Schuh, Christina; Benso, Bruna; Aguayo, Sebastian
The diagnosis and management of pain is an everyday occurrence in dentistry, and its effective control is essential to ensure the wellbeing of patients. Most tooth-associated pain originates from the dental pulp, a highly vascularized and innervated tissue, which is encased within mineralized dentin. It plays a crucial role in the sensing of stimuli from the local environment, such as infections (i.e. dental caries) and traumatic injury, leading to a local inflammatory response and subsequently to an increase in intra-pulp pressure, activating nerve endings. However, thermal, chemical, and mechanical stimuli also have the ability to generate dental pulp pain, which presents mechanisms highly specific to this tissue and which have to be considered in pain management. Traditionally, the management of dental pulp pain has mostly been pharmacological, using non-steroidal anti-inflammatory drugs (NSAIDs) and opioids, or restorative (i.e. removal of dental caries), or a combination of both. Both research areas continuously present novel and creative approaches. This includes the modulation of thermo-sensitive transient receptor potential cation channels (TRP) by newly designed drugs in pharmacological research, as well as the use of novel biomaterials, stem cells, exosomes and physical stimulation to obtain pulp regeneration in regenerative medicine. Therefore, the aim of this review is to present an up-to-date account of causes underlying dental pain, novel treatments involving the control of pain and inflammation and the induction of pulp regeneration, as well as insights in pain in dentistry from the physiological, pharmacological, regenerative and clinical perspectives
Royal jelly extracellular vesicles promote wound healing by modulating underlying cellular responses
(2023) Álvarez, Simón; Contreras, Pamina; Aguayo, Sebastian; Ramírez, Orlando; Vallejos, Catalina; Ruiz, Jorge; Carrasco, Eva; Troncoso, Stefanie; Morales, Bernardo; Schuh, Christina
Apis mellifera royal jelly (RJ) is a well-known remedy in traditional medicine around the world and its versatile effects range from antibacterial to anti-inflammatory properties and pro-regenerative properties. As a glandular product, RJ has been shown to contain a substantial number of extracellular vesicles (EVs), and, in this study, we aimed to investigate the extent of involvement of RJEVs in wound healing-associated effects. Molecular analysis of RJEVs verified the presence of exosomal markers such as CD63 and syntenin, and cargo molecules MRJP1, defensin-1, and jellein-3. Furthermore, RJEVs were demonstrated to modulate mesenchymal stem cell (MSC) differentiation and secretome, as well as decrease LPS-induced inflammation in macrophages by blocking the mitogen-activated protein kinase (MAPK) pathway. In vivo studies confirmed antibacterial effects of RJEVs and demonstrated an acceleration of wound healing in a splinted mouse model. This study suggests that RJEVs play a crucial role in the known effects of RJ by modulating the inflammatory phase and cellular response in wound healing. Transfer of RJ into the clinics has been impeded by the high complexity of the raw material. Isolating EVs from the raw RJ decreases the complexity while allowing standardization and quality control, bringing a natural nano-therapy one step closer to the clinics.
Type I collagen hydrogels as a delivery matrix for royal jelly derived extracellular vesicles
(2020) Ramírez, Orlando J.; Alvarez, Simón; Contreras-Kallens, Pamina; Barrera, Nelson P.; Aguayo, Sebastian; Schuh, Christina
Throughout the last decade, extracellular vesicles (EVs) have become increasingly popular in several areas of regenerative medicine. Recently, Apis mellifera royal jelly EVs (RJ EVs) were shown to display favorable wound healing properties such as stimulation of mesenchymal stem cell migration and inhibition of staphylococcal biofilms. However, the sustained and effective local delivery of EVs in nonsystemic approaches – such as patches for chronic cutaneous wounds – remains an important challenge for the development of novel EV-based wound healing therapies. Therefore, the present study aimed to assess the suitability of type I collagen -a well-established biomaterial for wound healing – as a continuous delivery matrix. RJ EVs were integrated into collagen gels at different concentrations, where gels containing 2mg/ml collagen were found to display the most stable release kinetics. Functionality of released RJ EVs was confirmed by assessing fibroblast EV uptake and migration in a wound healing assay. We could demonstrate reliable EV uptake into fibroblasts with a sustained promigratory effect for up to 7 d. Integrating fibroblasts into the RJ EV-containing collagen gel increased the contractile capacity of these cells, confirming availability of RJ EVs to fibroblasts within the collagen gel. Furthermore, EVs released from collagen gels were found to inhibit Staphylococcus aureus ATCC 29213 biofilm formation. Overall, our results suggest that type I collagen could be utilized as a reliable, reproducible release system to deliver functional RJ EVs for wound healing therapies.

Browsing by Author "Aguayo, Sebastian"

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