An in vitro assessment of the response of THP-1 macrophages to varying stiffness of a glycol-chitosan hydrogel for vocal fold tissue engineering applications.
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ABSTRACT:
SUBMITTER: Coburn PT
PROVIDER: S-EPMC8079542 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The human vocal folds (VFs) undergo complex biomechanical stimulation during phonation. The aim of the present study was to develop and validate a phono-mimetic VF flow perfusion bioreactor, which mimics the mechanical microenvironment of the human VFs in vitro. The bioreactor uses airflow-induced self-oscillations, which have been shown to produce mechanical loading and contact forces that are representative of human phonation. The bioreactor consisted of two synthetic VF replicas within a silicone body. A cell-scaffold mixture (CSM) consisting of human VF fibroblasts, hyaluronic acid, gelatin, and a polyethylene glycol cross-linker was injected into cavities within the replicas. Cell culture medium (CCM) was perfused through the scaffold by using a customized secondary flow loop. After the injection, the bioreactor was operated with no stimulation over a 3-day period to allow for cell adaptation. Phonation was subsequently induced by using a variable speed centrifugal blower for 2 h each day over a period of 4 days. A similar bioreactor without biomechanical stimulation was used as the nonphonatory control. The CSM was harvested from both VF replicas 7 days after the injection. The results confirmed that the phono-mimetic bioreactor supports cell viability and extracellular matrix proteins synthesis, as expected. Many scaffold materials were found to degrade because of challenges from phonation-induced biomechanical stimulation as well as due to biochemical reactions with the CCM. The bioreactor concept enables future investigations of the effects of different phonatory characteristics, that is, voice regimes, on the behavior of the human VF cells. It will also help study the long-term functional outcomes of the VF-specific biomaterials before animal and clinical studies.
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Project description:Three-dimensional porous scaffolds are widely employed in tissue engineering and regenerative medicine for their ability to carry bioactives and cells; and for their platform properties to allow for bridging-the-gap within an injured tissue. This study describes the effect of various methoxypolyethylene glycol (mPEG) derivatives (mPEG (-OCH3 functionality), mPEG-aldehyde (mPEG-CHO) and mPEG-acetic acid (mPEG-COOH)) on the morphology and physical properties of chemically crosslinked, semi-interpenetrating polymer network (IPN), chitosan (CHT)/mPEG blend cryosponges. Physicochemical and molecular characterization revealed that the -CHO and -COOH functional groups in mPEG derivatives interacted with the -NH2 functionality of the chitosan chain. The distinguishing feature of the cryosponges was their unique morphological features such as fringe thread-, pebble-, curved quartz crystal-, crystal flower-; and canyon-like structures. The morphological data was well corroborated by the image processing data and physisorption curves corresponding to Type II isotherm with open hysteresis loops. Functionalization of mPEG had no evident influence on the macro-mechanical properties of the cryosponges but increased the matrix strength as determined by the rheomechanical analyses. The cryosponges were able to deliver bioactives (dexamethasone and curcumin) over 10 days, showed varied matrix degradation profiles, and supported neuronal cells on the matrix surface. In addition, in silico simulations confirmed the compatibility and molecular stability of the CHT/mPEG blend compositions. In conclusion, the study confirmed that significant morphological variations may be induced by minimal functionalization and crosslinking of biomaterials.
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Project description:The biomechanical function of the vocal folds (VFs) depends on their viscoelastic properties. Many conditions can lead to VF scarring that compromises voice function and quality. To identify candidate replacement materials, the structure, composition, and mechanical properties of native tissues need to be understood at phonation frequencies. Previously, the authors developed the torsional wave experiment (TWE), a stress-wave-based experiment to determine the linear viscoelastic shear properties of small, soft samples. Here, the viscoelastic properties of porcine and human VFs were measured over a frequency range of 10-200 Hz. The TWE utilizes resonance phenomena to determine viscoelastic properties; therefore, the specimen test frequency is determined by the sample size and material properties. Viscoelastic moduli are reported at resonance frequencies. Structure and composition of the tissues were determined by histology and immunochemistry. Porcine data from the TWE are separated into two groups: a young group, consisting of fetal and newborn pigs, and an adult group, consisting of 6-9-month olds and 2+-year olds. Adult tissues had an average storage modulus of 2309±1394 Pa and a loss tangent of 0.38±0.10 at frequencies of 36-200 Hz. The VFs of young pigs were significantly more compliant, with a storage modulus of 394±142 Pa and a loss tangent of 0.40±0.14 between 14 and 30 Hz. No gender dependence was observed. Histological staining showed that adult porcine tissues had a more organized, layered structure than the fetal tissues, with a thicker epithelium and a more structured lamina propria. Elastin fibers in fetal VF tissues were immature compared to those in adult tissues. Together, these structural changes in the tissues most likely contributed to the change in viscoelastic properties. Adult human VF tissues, recovered postmortem from adult patients with a history of smoking or disease, had an average storage modulus of 756±439 Pa and a loss tangent of 0.42±0.10. Contrary to the results of some other investigators, no significant frequency dependence was observed. This lack of observable frequency dependence may be due to the modest frequency range of the experiments and the wide range of stiffnesses observed within nominally similar sample types.
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Project description:In this paper, polyvinyl alcohol/Ag-Metal-organic framework (PVA/Ag@MOF) and polyvinyl alcohol/chitosan (PVA/CS) were used as the inner and outer layers to successfully prepare a bilayer composite hydrogel for tissue engineering scaffold. The performance of bilayer hydrogels was evaluated. The outer layer (PVA/CS) has a uniform pore size distribution, good water retention, biocompatibility and cell adhesion ability. The inner layer (PVA/Ag@MOF) has good antibacterial activity and poor biocompatibility. PVA, PVA/0.1%Ag@MOF, PVA/0.5%Ag@MOF, and PVA/1.0%Ag@MOF show anti-microbial activity in ascending order. However, its use as an inner layer avoids direct contact with cells and prevents infection. The cell viability of all samples was above 90%, indicating that the bilayer hydrogel was non-toxic to A549 cells. The bilayer hydrogel scaffold combines the advantages of the inner and outer layers. In summary, this new bilayer composite is an ideal lung scaffold for tissue engineering.
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Project description:Vocal cord healing is a dynamic process, and many genes and proteins are involved, which play varying roles at different regeneration stages after injury. Previous studies have shown that inflammatory responses occur at the early stage of vocal cord injury, where the fibroblasts proliferate exuberantly with intensive secretion and deposition of ECM. These activities reach the peak at 3-7 days and their intensity begins to decline 15 days later. A study based on the dermal system has shown that ECM remodeling during the repair of injury can last for several months. However, few studies have been conducted as to the dynamic changes of gene expressions and signaling pathway during the healing process of vocal cord injury. Plotting these changes will facilitate the understanding about the physiological changes during healing and the identification of key time points and target genes in fibrosis formation.
2019-10-26 | GSE139383 | GEO
Project description:Vocal cord healing is a dynamic process, and many genes and proteins are involved, which play varying roles at different regeneration stages after injury. Previous studies have shown that inflammatory responses occur at the early stage of vocal cord injury, where the fibroblasts proliferate exuberantly with intensive secretion and deposition of ECM. These activities reach the peak at 3-7 days and their intensity begins to decline 15 days later. A study based on the dermal system has shown that ECM remodeling during the repair of injury can last for several months. However, few studies have been conducted as to the dynamic changes of gene and microRNA expressions during the healing process of vocal cord injury. Plotting these changes will facilitate the understanding about the physiological changes during healing and the identification of key time points and target genes and microRNAs in fibrosis formation.
2020-03-25 | GSE147473 | GEO
Project description:We used microarrays to characterize transcriptome profiles of rat vocal fold tissue following surgical injury (vs. naive tissue); rat vocal fold fibroblasts harvested from scar tissue at the 60 d time point (vs. naive fibroblasts); rat vocal fold scar fibroblasts treated with siRNA against the collagen chaperone protein rat gp46 (vs. scramble siRNA). Adult Fischer 344 rat vocal fold tissue was harvested at 3, 14, and 60 days following surgical injury (control = age-matched naive tissue); rat vocal fold scar fibroblasts were obtained via explant culture of tissue obtained 60 days following surgical injury and harvested at 80% confluence during passage 1 (control = age-matched naive rat vocal fold fibroblasts); rat vocal fold scar fibroblasts were treated for 1 h with 50 nM liposome-delivered siRNA against rat gp46 when 80% confluent at passage 1, cultured for an additional 24 h in fresh media, then harvested (control = rat vocal fold scar fibroblasts treated with 50 nM liposome-delivered scramble siRNA).
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Project description:Glycol chitosan (GC) is a hydrophilic chitosan derivative, known for its aqueous solubility. Previously, we have demonstrated the feasibility of preparing injectable, enzymatically crosslinked hydrogels from HPP [3-(4-Hydroxyphenyl)-propionic acid (98%)]-modified GC. However, HPP-GC gels showed very slow degradation, which presents challenges as an in vivo protein delivery vehicle. This study reports the potential of acetylated HPP-GC hydrogels as a biodegradable hydrogel platform for sustained protein delivery. Enzymatic crosslinking was used to prepare injectable, biodegradable hydrogels from HPP-GC with various degrees of acetylation (DA). The acetylated polymers were characterized using Fourier transform infrared and nuclear magnetic resonance spectroscopy. Rheological methods were used to characterize the mechanical behavior of the hydrogels. In vitro degradation and protein release were performed in the presence and absence of lysozyme. In vivo degradation was studied using a mouse subcutaneous implantation model. Finally, two hydrogel formulations with distinct in vitro/in vivo degradation and in vitro protein release were evaluated in 477-SKH1-Elite mice using live animal imaging to understand in vivo protein release profiles. The lysozyme-mediated degradation of the gels was demonstrated in vitro and the degradation rate was found to be dependent on the DA of the polymers. In vivo degradation study further confirmed that gels formed from polymers with higher DA degraded faster. In vitro protein release demonstrated the feasibility to achieve lysozyme-mediated protein release from the gels and that the rate of protein release can be modulated by varying the DA. In vivo protein release study further confirmed the feasibility to achieve differential protein release by varying the DA. The feasibility to develop degradable enzymatically crosslinked GC hydrogels is demonstrated. Gels with a wide spectrum of degradation time ranging from less than a week and more than 6 weeks can be developed using this approach. The study also showed the feasibility to fine tune in vivo protein release by modulating HPP-GC acetylation. The hydrogel platform therefore holds significant promise as a protein delivery vehicle for various biomedical and regenerative engineering applications. Impact statement The study describes the feasibility to develop a novel enzyme sensitive biodegradable and injectable hydrogel, where in the in vivo degradation rate and protein release profile can be modulated over a wide range. The described hydrogel platform has the potential to develop into a clinically relevant injectable and tunable protein delivery vehicle for a wide range of biomedical applications.
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