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1.
Mater Sci Eng C Mater Biol Appl ; 107: 110303, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31761191

RESUMO

Titanium dioxide (TiO2) nanoparticles (NPs), first developed in the 1990s, have been applied in numerous biomedical fields such as tissue engineering and therapeutic drug development. In recent years, TiO2-based drug delivery systems have demonstrated the ability to decrease the risk of tumorigenesis and improve cancer therapy. There is increasing research on the origin and effects of pristine and doped TiO2-based nanotherapeutic drugs. However, the detailed molecular mechanisms by which drug delivery to cancer cells alters sensing of gene mutations, protein degradation, and metabolite changes as well as its associated cumulative effects that determine the microenvironmental mechanosensitive metabolism have not yet been clearly elucidated. This review focuses on the microenvironmental influence of TiO2-NPs induced various mechanical stimuli on tumor cells. The differential expression of genome, proteome, and metabolome after treatment with TiO2-NPs is summarized and discussed. In the tumor microenvironment, mechanosensitive DNA mutations, gene delivery, protein degradation, inflammatory responses, and cell viability affected by the mechanical stimuli of TiO2-NPs are also examined.

2.
Biomacromolecules ; 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31714754

RESUMO

Biosourced and biodegradable polymers for additive manufacturing could enable the rapid fabrication of parts for a broad spectrum of applications ranging from healthcare to aerospace. However, a limited number of these materials are suitable for vat photopolymerization processes. Herein, we report a two-step additive manufacturing process to fabricate robust protein-based constructs using a commercially available laser-based stereolithography printer. Methacrylated bovine serum albumin (MA-BSA) was synthesized and formulated into aqueous resins that were used to print complex three-dimensional (3D) objects with a resolution comparable to a commercially available resin. The MA-BSA resins were characterized by rheometry to determine the viscosity and the cure rate, as both parameters can ultimately be used to predict the printability of the resin. In the first step of patterning these materials, the MA-BSA resin was 3D printed, and in the second step, the printed construct was thermally cured to denature the globular protein and increase the intermolecular noncovalent interactions. Thus, the final 3D printed part was comprised of both chemical and physical cross-links. Compression studies of hydrated and dehydrated constructs demonstrated a broad range of compressive strengths and Young's moduli that could be further modulated by adjusting the type and amount of co-monomer. The printed hydrogel constructs demonstrated good cell viability (>95%) after a 21 day culture period. These MA-BSA resins are expected to be compatible with other vat photopolymerization techniques including digital light projection and continuous liquid interface production.

3.
Nat Commun ; 10(1): 4671, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31604922

RESUMO

Mitochondrial trifunctional protein deficiency, due to mutations in hydratase subunit A (HADHA), results in sudden infant death syndrome with no cure. To reveal the disease etiology, we generated stem cell-derived cardiomyocytes from HADHA-deficient hiPSCs and accelerated their maturation via an engineered microRNA maturation cocktail that upregulated the epigenetic regulator, HOPX.  Here we report, matured HADHA mutant cardiomyocytes treated with an endogenous mixture of fatty acids manifest the disease phenotype: defective calcium dynamics and repolarization kinetics which results in a pro-arrhythmic state. Single cell RNA-seq reveals a cardiomyocyte developmental intermediate, based on metabolic gene expression. This intermediate gives rise to mature-like cardiomyocytes in control cells but, mutant cells transition to a pathological state with reduced fatty acid beta-oxidation, reduced mitochondrial proton gradient, disrupted cristae structure and defective cardiolipin remodeling. This study reveals that HADHA (tri-functional protein alpha), a monolysocardiolipin acyltransferase-like enzyme, is required for fatty acid beta-oxidation and cardiolipin remodeling, essential for functional mitochondria in human cardiomyocytes.

4.
Circulation ; 140(20): 1647-1660, 2019 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-31587567

RESUMO

BACKGROUND: The giant sarcomere protein titin is important in both heart health and disease. Mutations in the gene encoding for titin (TTN) are the leading known cause of familial dilated cardiomyopathy. The uneven distribution of these mutations within TTN motivated us to seek a more complete understanding of this gene and the isoforms it encodes in cardiomyocyte (CM) sarcomere formation and function. METHODS: To investigate the function of titin in human CMs, we used CRISPR/Cas9 to generate homozygous truncations in the Z disk (TTN-Z-/-) and A-band (TTN-A-/-) regions of the TTN gene in human induced pluripotent stem cells. The resulting CMs were characterized with immunostaining, engineered heart tissue mechanical measurements, and single-cell force and calcium measurements. RESULTS: After differentiation, we were surprised to find that despite the more upstream mutation, TTN-Z-/--CMs had sarcomeres and visibly contracted, whereas TTN-A-/--CMs did not. We hypothesized that sarcomere formation was caused by the expression of a recently discovered isoform of titin, Cronos, which initiates downstream of the truncation in TTN-Z-/--CMs. Using a custom Cronos antibody, we demonstrate that this isoform is expressed and integrated into myofibrils in human CMs. TTN-Z-/--CMs exclusively express Cronos titin, but these cells produce lower contractile force and have perturbed myofibril bundling compared with controls expressing both full-length and Cronos titin. Cronos titin is highly expressed in human fetal cardiac tissue, and when knocked out in human induced pluripotent stem cell derived CMs, these cells exhibit reduced contractile force and myofibrillar disarray despite the presence of full-length titin. CONCLUSIONS: We demonstrate that Cronos titin is expressed in developing human CMs and is able to support partial sarcomere formation in the absence of full-length titin. Furthermore, Cronos titin is necessary for proper sarcomere function in human induced pluripotent stem cell derived CMs. Additional investigation is necessary to understand the molecular mechanisms of this novel isoform and how it contributes to human cardiac disease.

6.
Biol Pharm Bull ; 42(9): 1471-1481, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31474708

RESUMO

Sutaehwan (STH) has been used in Korean medicine for the treatment of abortus habitualis such as fetal restlessness in the uterus. Previously, we reported that a modified formulation of STH, Sutaehwan-Gami, has phytoestrogen-like properties in an ovariectomized menopausal rat model. However, the therapeutic effects of STH and the precise mechanisms by which STH affects various menopausal symptoms remain poorly understood. The current study was designed to explore the effects of a modified form of STH on menopausal anxiety, depression and heart hypertrophy and its mechanisms in 4-vinylcyclohexene diepoxide (VCD)-induced menopausal mouse models. VCD-induced menopausal model mice were fed a modified form of STH, which contained water extract of 3 herbs (called STH_KP17001) at a dose of 100 or 300 mg/kg/d or as a positive control, estradiol at a dose of 0.2 mg/kg/d with standard mouse pellets for 13 weeks. The results show that STH_KP17001 significantly restored the VCD-induced weight reduction of uterine and ovary through the phosphorylation of extracellular signal-regulated kinase (ERK) and protein kinase B (AKT) in the uterus and ovary. Moreover, STH_KP17001 showed slight proliferative effects and estrogen receptor α phosphorylation in MCF-7 cells. Treatment with STH_KP17001 reversed VCD-induced anxiety and depression through AMP-activated protein kinase (AMPK) activation and brain-derived neurotrophic factor (BDNF) expression in the cerebral cortex, while improving heart hypertrophy through inactivation of inhibitor of kappaB α (IκBα) in the heart. The results indicate that STH_KP17001 improves menopause-induced anxiety, depression and heart hypertrophy, implying its protective role for the management of menopausal symptoms.

7.
J Cell Biol ; 218(9): 2919-2944, 2019 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-31395619

RESUMO

Mutations in A-type nuclear lamins cause dilated cardiomyopathy, which is postulated to result from dysregulated gene expression due to changes in chromatin organization into active and inactive compartments. To test this, we performed genome-wide chromosome conformation analyses in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with a haploinsufficient mutation for lamin A/C. Compared with gene-corrected cells, mutant hiPSC-CMs have marked electrophysiological and contractile alterations, with modest gene expression changes. While large-scale changes in chromosomal topology are evident, differences in chromatin compartmentalization are limited to a few hotspots that escape segregation to the nuclear lamina and inactivation during cardiogenesis. These regions exhibit up-regulation of multiple noncardiac genes including CACNA1A, encoding for neuronal P/Q-type calcium channels. Pharmacological inhibition of the resulting current partially mitigates the electrical alterations. However, chromatin compartment changes do not explain most gene expression alterations in mutant hiPSC-CMs. Thus, global errors in chromosomal compartmentation are not the primary pathogenic mechanism in heart failure due to lamin A/C haploinsufficiency.

8.
Nat Commun ; 10(1): 2797, 2019 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-31243273

RESUMO

Collective cell migration occurs in many patho-physiological states, including wound healing and invasive cancer growth. The integrity of the expanding epithelial sheets depends on extracellular cues, including cell-cell and cell-matrix interactions. We show that the nano-scale topography of the extracellular matrix underlying epithelial cell layers can strongly affect the speed and morphology of the fronts of the expanding sheet, triggering partial and complete epithelial-mesenchymal transitions (EMTs). We further demonstrate that this behavior depends on the mechano-sensitivity of the transcription regulator YAP and two new YAP-mediated cross-regulating feedback mechanisms: Wilms Tumor-1-YAP-mediated downregulation of E-cadherin, loosening cell-cell contacts, and YAP-TRIO-Merlin mediated regulation of Rho GTPase family proteins, enhancing cell migration. These YAP-dependent feedback loops result in a switch-like change in the signaling and the expression of EMT-related markers, leading to a robust enhancement in invasive cell spread, which may lead to a worsened clinical outcome in renal and other cancers.


Assuntos
Células Epiteliais/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Nanoestruturas , Proteínas WT1/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Cães , Técnicas de Silenciamento de Genes , Proteínas de Fluorescência Verde/metabolismo , Células Madin Darby de Rim Canino , Propriedades de Superfície , Proteínas WT1/genética , Proteínas rho de Ligação ao GTP/genética
9.
Cardiovasc Res ; 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31049579

RESUMO

AIMS: Heart failure invariably affects patients with various forms of Muscular Dystrophy (MD), but the onset and molecular sequelae of altered structure and function resulting from full-length dystrophin (Dp427) deficiency in MD heart tissue are poorly understood.To better understand the role of dystrophin in cardiomyocyte development and the earliest phase of DMD cardiomyopathy, we studied human cardiomyocytes differentiated from induced pluripotent stem cells (hiPSC-CMs) obtained from the urine of a Deuchenne Muscular Dystrophy (DMD) patient. METHODS AND RESULTS: The contractile properties of patient-specific hiPSC-CMs, with no detectable dystrophin (DMD-CMs with a deletion of exon 50), were compared to CMs containing a CRISPR-Cas9 mediated deletion of a single G base at position 263 of the dystrophin gene (c.263delG -CMs) isogenic to the parental line of hiPSC-CMs from a healthy individual. We hypothesized that the absence of a dystrophin-actin linkage would adversely affect myofibril and cardiomyocyte structure and function. Cardiomyocyte maturation was driven by culturing long-term (80-100 days) on a nanopatterned surface, which resulted in hiPSC-CMs with adult-like dimensions and aligned myofibrils. CONCLUSIONS: Our data demonstrate that lack of Dp427 results in reduced myofibril contractile tension, slower relaxation kinetics and to Ca2+ handling abnormalities, similar to DMD cells, suggesting either retarded or altered maturation of cardiomyocyte structures associated with these functions. This study offers new insights into the functional consequences of Dp427 deficiency at an early stage of cardiomyocyte development in both patient-derived and CRISPR-generated models of dystrophin deficiency.

10.
11.
Adv Healthc Mater ; 8(4): e1900098, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30789678
12.
Adv Healthc Mater ; 8(4): e1801332, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30614636

RESUMO

Cells respond and actively adapt to environmental cues in the form of mechanical stimuli. This extends to immune cells and their critical role in the maintenance of tissue homeostasis. Multiple recent studies have begun illuminating underlying mechanisms of mechanosensation in modulating immune cell phenotypes. Since the extracellular microenvironment is critical to modify cellular physiology that ultimately determines the functionality of the cell, understanding the interactions between immune cells and their microenvironment is necessary. This review focuses on mechanoregulation of immune responses mediated by macrophages, dendritic cells, and T cells, in the context of modern mechanobiology.

13.
Adv Healthc Mater ; 8(4): e1801395, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30657652

RESUMO

Self-assembled RNA particles have been exploited widely to maximize the therapeutic potential of RNA. However, the immune response via RNA particles is not fully understood. In addition, the investigation of the immunogenicity from RNA-based particles is required owing to inherent immunostimulatory effects of RNA for clinical translation. To examine the immune stimulating potency, rationally designed microsized RNA particles, called RNA microspheres (RMSs), are generated with single or double strands via rolling circle transcription. The RMSs show an exceptional stability in the presence of serum, while they are selectively degraded under endolysosomal conditions. With precisely controlled size, both RMSs are successfully taken up by macrophages. Unlike the nature of RNA fragments, RMSs induce only basal-level expression of inflammatory cytokines as well as type I interferon from macrophages, suggesting that RMSs are immunocompatible in the therapeutic dose range. Taken together, this study could help accelerate clinical translation and broaden the applicability of the self-assembled RNA-based particles without being limited by their potential immunotoxicity, while a systematic controllability study observing the release of RNA fragments from RMSs would provide self-assembled RNA-based structures with a great potential for immunomodulation.

14.
Acta Biomater ; 95: 337-347, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-30529081

RESUMO

Bioengineering approaches to regulate stem cell fates aim to recapitulate the in vivo microenvironment. In recent years, manipulating the micro- and nano-scale topography of the stem cell niche has gained considerable interest for the purposes of controlling extrinsic mechanical cues to regulate stem cell fate and behavior in vitro. Here, we established an optimal nanotopographical system to improve 3-dimensional (3D) differentiation of pancreatic cells from human pluripotent stem cells (hPSCs) by testing gradient-pattern chips of nano-scale polystyrene surface structures with varying sizes and shapes. The optimal conditions for 3D differentiation of pancreatic cells were identified by assessing the expression of developmental regulators that are required for pancreatic islet development and maturation. Our results showed that the gradient chip of pore-part 2 (Po-2, 200-300 nm diameter) pattern was the most efficient setting to generate clusters of pancreatic endocrine progenitors (PDX1+ and NGN3+) compared to those of other pore diameters (Po-1, 100-200 or Po-3, 300-400 nm) tested across a range of pillar patterns and flat surfaces. Furthermore, the Po-2 gradient pattern-derived clusters generated islet-like 3D spheroids and tested positive for the zinc-chelating dye dithizone. The spheroids consisted of more than 30% CD200 + endocrine cells and also expressed NKX6.1 and NKX2.2. In addition, pancreatic ß- cells expressing insulin and polyhormonal cells expressing both insulin and glucagon were obtained at the final stage of pancreatic differentiation. In conclusion, our data suggest that an optimal topographical structure for differentiation to specific cell types from hPSCs can be tested efficiently by using gradient-pattern chips designed with varying sizes and surfaces. STATEMENT OF SIGNIFICANCE: Our study provides demonstrates of using gradient nanopatterned chips for differentiation of pancreatic islet-like clusters. Gradient nanopatterned chips are consisted of two different shapes (nanopillar and nanopore) in three different ranges of nano sizes (100-200, 200-300, 300-400 nm). We found that optimal nanostructures for differentiation of pancreatic islet-like clusters were 200-300 nm nano pores. Cell transplantation is one of the major therapeutic option for type 1 diabetes mellitus (DM) using stem cell-derived ß-like cells. We generated 50 um pancreatic islet-like clusters in size, which would be an optimal size for cell transplantation. Futuremore, the small clusters provide a powerful source for cell therapy. Our findings suggest gradient nanopatterned chip provides a powerful tool to generate specific functional cell types of a high purity for potential uses in cell therapy development.

15.
Int J Nanomedicine ; 13: 7217-7227, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30510414

RESUMO

Background: Tendon-derived stem cells (TDSCs) are key factors associated with regeneration and healing in tendinopathy. The aim of this study was to investigate the effects of mechanical stiffness and topographic signals on the differentiation of TDSCs depending on age and pathological conditions. Materials and methods: We compared TDSCs extracted from normal tendon tissues with TDSCs from tendinopathic Achilles tendon tissues of Sprague Dawley rats in vitro and TDSCs cultured on nanotopographic cues and substrate stiffness to determine how to control the TDSCs. The tendinopathy model was created using a chemical induction method, and the tendon injury model was created via an injury-and-overuse method. Norland Optical Adhesive 86 (NOA86) substrate with 2.48 GPa stiffness with and without 800 nm-wide nanogrooves and a polyurethane substrate with 800 nm-wide nanogrooves were used. Results: TDSCs from 5-week-old normal tendon showed high expression of type III collagen on the flat NOA86 substrate. In the 15-week normal tendon model, expression of type III collagen was high in TDSCs cultured on the 800 nm NOA86 substrates. However, in the 15-week tendon injury model, expression of type III collagen was similar irrespective of nanotopographic cues or substrate stiffness. The expression of type I collagen was also independent of nanotopographic cues and substrate stiffness in the 15-week normal and tendon injury models. Gene expression of scleraxis was increased in TDSCs cultured on the flat NOA86 substrate in the 5-week normal tendon model (P=0.001). In the 15-week normal tendon model, scleraxis was highly expressed in TDSCs cultured on the 800 nm and flat NOA86 substrate (P=0.043). However, this gene expression was not significantly different between the substrates in the 5-week tendinopathy and 15-week tendon injury models. Conclusion: Development and maturation of tendon are enhanced when TDSCs from normal tendons were cultured on stiff surface, but not when the TDSCs came from pathologic models. Therapeutic applications of TDSCs need to be flexible based on tendon age and tendinopathy.

16.
Nano Converg ; 5(1): 27, 2018 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-30467757

RESUMO

Advancing as a biosensing nanotechnology, nanohybrids present a new class of functional materials with high selectivity and sensitivity, enabling integration of nanoscale chemical/biological interactions with biomedical devices. The unique properties of ZnO combined with metal oxide nanostructures were recently demonstrated to be an efficient approach for sensor device fabrication with accurate, real-time and high-throughput biosensing, creating new avenues for diagnosis, disease management and therapeutics. This review article collates recent advances in the modified ZnO nanostructured metal oxide nanohybrids for efficient enzymatic and non-enzymatic biosensor applications. Furthermore, we also discussed future prospects for nanohybrid materials to yield high-performance biosensor devices.

17.
Sci Rep ; 8(1): 14210, 2018 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-30242256

RESUMO

The root cause of cancer mortality and morbidity is the metastatic spread of the primary tumor, but underlying mechanisms remain elusive. Here we investigate biomechanical interactions that may accompany invasive spread of melanoma cells. We find that metastatic cells can exert considerable traction forces and modify local collagen organization within a 3D matrix. When this re-organization is mimicked using a nano-fabricated model of aligned extracellular matrix fibers, metastatic cells, including less invasive melanoma cells, were in turn induced to align, elongate and migrate, guided by the local ridge orientations. Strikingly, we found that this aligned migration of melanoma cells was accompanied by long-range regulation of cytoskeletal remodeling that show anisotropic stiffening in the direction of fiber orientation suggestive of a positive feedback between ECM fiber alignment and forces exerted by cancer cells. Taken together, our findings suggest that the invasive spread of cancer cells can be defined by complex interplay with the surrounding matrix, during which they both modify the matrix and use the matrix alignment as a persistent migration cue, leading to more extensive and rapid invasive spread.

18.
J Pharmacopuncture ; 21(2): 120-125, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30151313

RESUMO

Objective: The purpose of this study was to investigate the clinical effects of BU pharmacopuncture therapy consisting of bear's gall(fel ursi) and ox bezoar(bovis calculus) on acute lumbar sprain. Methods: 12 patients diagnosed as acute lumbar sprain in 6 designated local Korean medicine clinics from October 2017 to February 2018 were treated by BU pharmacopuncture. Several acupoints in abdomen and lumbar region were selected by clinicians at their own discretion. The effectiveness of the therapy was evaluated using VAS and ODI. After that we reviewed the medical records of all these patients to evaluate the effectiveness and safety of the therapy. Results: The VAS and ODI scales were significantly decreased after BU pharmacopuncture therapy. And no major complications and adverse effects were reported. Conclusion: BU pharmacopuncture showed rapid pain relief in patients with acute lumbar sprain. It is possible to shorten the treatment period of acute lumbar sprain and prevent progressing to chronic back pain in advance. To establish the effects of BU pharmacopuncture therapy, more succeeding clinical and laboratory studies are needed.

19.
Biophys J ; 114(6): 1257-1263, 2018 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-29590582

RESUMO

Living cells orient the cytoskeleton polarity and directional migration in response to spatial gradients of multiple types of cues. The resulting tactic behaviors are critical for the proper cell localization in the context of complex single-cell and tissue behaviors. In this perspective, we highlight the recent discovery of, to our knowledge, a new -taxis phenomenon, the topotaxis, which mediates directional cell migration in response to the gradients of such topographic features as the density of extracellular matrix fibers. The direction of topotactic migration critically depends on the effective stiffness of the cortical cytoskeleton, which is controlled by the balance between two parallel signaling pathways activated by the extracellular matrix input. Topotaxis can account for such striking cell behaviors as the opposite directionality of migration of benign and metastatic cancer cells and certain aspects of the wound-healing process. We anticipate that, in conjunction with other tactic phenomena, topotaxis can provide critical information for understanding and design of tissue structure and function.

20.
J Biomed Mater Res A ; 106(6): 1543-1551, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29368451

RESUMO

Skeletal muscle has a well-organized tissue structure comprised of aligned myofibers and an encasing extracellular matrix (ECM) sheath or lamina, within which reside satellite cells. We hypothesize that the organization of skeletal muscle tissues in culture can affect both the structure of the deposited ECM and the differentiation potential of developing myotubes. Furthermore, we posit that cellular and ECM cues can be a strong determinant of myoblast fusion and morphology in 3D tissue culture environments. To test these, we utilized a thermoresponsive nanofabricated substratum to engineer anisotropic sheets of myoblasts which could then be transferred and stacked into multilayered tissues. Within such engineered tissues, we found that myoblasts rapidly sense topography and deposit structurally organized ECM proteins. Furthermore, the initial tissue structure was found to exert significant control over myoblast fusion and eventual myotube organization. These results highlight the importance of ECM structure on myoblast fusion and organization, and provide insights into substrate-mediated control of myotube formation in the development of novel, more effective, engineered skeletal muscle tissues. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1543-1551, 2018.

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