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1.
Biomacromolecules ; 2021 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-34855378

RESUMO

We report a polypeptide-based thermogel as a new tool for hypothermic storage of stem cells at ambient temperature (25 °C). Stem cells were suspended in the sol state (10 °C) of an aqueous poly(ethylene glycol)-poly(l-alanine) (PEG-PA) solution (4.0 wt %) in phosphate-buffered saline (PBS), which turned into a stem cell-incorporated gel by a heat-induced sol-to-gel transition. The cell harvesting procedure from the thermogels was simply performed through a gel-to-sol transition by diluting and cooling the system. More than 99% of stem cells died in PBS and Pluronic F127 thermogel (control thermogel) when the cells were stored at 25 °C for 7 days. The cell recovery rate from the PEG-PA thermogel (64%) was significantly greater than that from the commercially available HypoThermosol FRS preservation solution (HTS) (26%). Additionally, the surviving stem cells from the PEG-PA thermogel were healthier than those from HTS in terms of (1) expression of stemness biomarkers (NANOG, OCT4, and SOX2), (2) proliferation rate, and (3) differentiation potentials into osteogenic, chondrogenic, and adipogenic lineages. Membrane stabilization was suggested as a cell protection mechanism in the cytocompatible PEG-PA thermogel. The PEG-PA thermogel provides a convenient cytocompatible way for the storage and recovery of cells and thus is a promising tool for the transportation and short-term banking of cells.

2.
ACS Appl Mater Interfaces ; 13(29): 33969-33980, 2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34275265

RESUMO

Precise control over the size and shape of ice crystals is a key factor to consider in designing antifreezing and cryoprotecting molecules for cryopreservation of cells. Here, we report that a poly(ethylene glycol)-poly(l-alanine) (PEG-PA) block copolymer exhibits excellent cryoprotecting properties for stem cells and antifreezing properties for water. As the molecular weight of PA increased from 500, 760, and 1750 Da (P1, P2, and P3) at the same PEG molecular weight of 5000 Da, the ß-sheet content decreased and α-helix content increased. Comparing P2 (PEG-PA; 5000-760) and P4 (PEG-PA: 1000-750), ß-sheets increased as the PEG block length decreased. The critical micelle concentration of the PEG-PA block copolymers was in a range of 0.5-3.0 mg/mL and was proportional to the hydrophobicity of the PEG-PA block copolymers. The P1, P2, and P3 self-assembled into spherical micelles, whereas P4 formed micelles with cylindrical morphology. The difference in the block copolymer structure affected ice recrystallization inhibition (IRI) activity and cryopreservation of cells. IRI activity was assayed via mean largest grain size (MLGS), and interactions between polymers and ice crystal surfaces were studied by dynamic ice-shaping studies. The MLGS decreased to 58 → 53 → 45 → 35 → 23% of that of PBS, as the polymer (PEG-PA 5000-500) concentration increased from 0.0 (PBS; control) → 1.0 → 5.0 → 10 → 30 → 50 mg/mL. The MLGS of PEG 5k solutions (negative control) decreased to 74 → 71 → 64 → 44 → 37% of that of PBS in the same concentration range. P3 and P4 with a longer hydrophobic PA block developed elongated ice crystals at above 30 mg/mL. The dynamic ice-shaping study exhibited that ice crystals became needle-shaped, as the hydrophobicity of the polymer increased as in P2-P4. The cell recovery in the P1 system after cryopreservation at -196 °C for 7 days was 87% of that of the dimethyl sulfoxide (DMSO) 10% system (positive control). The cell recovery was 48% for the P2 system and drastically decreased to less than 30% of that of the DMSO 10% system in the P3, P4, PEG 5k, PEG 1k, PVA 80H, and PVA 100H systems. Current studies suggest that IRI activity, round ice crystal shaping, and membrane stabilization activity of P1 cooperatively provide excellent cell recovery among the candidate systems. Recovered stem cells exhibited excellent proliferation and multilineage differentiation into osteocytes, chondrocytes, and adipocytes. To conclude, the PEG-PA (5000-500) block copolymer is suggested to be a promising antifreezing cryoprotectant for stem cells.


Assuntos
Crioprotetores/farmacologia , Gelo , Células-Tronco Mesenquimais/efeitos dos fármacos , Peptídeos/farmacologia , Polietilenoglicóis/farmacologia , Água/química , Proliferação de Células/efeitos dos fármacos , Criança , Crioprotetores/química , Humanos , Masculino , Tonsila Palatina/citologia , Tamanho da Partícula , Peptídeos/química , Polietilenoglicóis/química
3.
Polymers (Basel) ; 13(7)2021 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-33810521

RESUMO

Cancer progression and migration in the tumor microenvironment are related to cell types and three-dimensional (3D) matrices. Therefore, developing biomimetic tumor models, including co-culture systems and a tunable 3D matrix, could play an essential role in understanding the cancer environment. Here, multicellular spheroids using human adipose-derived mesenchymal stem cells (hADSCs) and breast cancer cells (MDA-MB-231) within the 3D matrix were used as a tumor microenvironment (TME) mimicking platform. The amphiphilic peptide block copolymer and hyaluronic acid (HA) formed a self-assembled structure, which provides a biocompatible 3D environment for the cells. Multicellular spheroids were formed on the optimized plate and were observed as cell migration from a spheroid within a 3D matrix, such as the invasive and metastatic cancer of TME. This study suggests a new 3D platform using polymer complexes and the importance of tumor complexities, including various cell types and microenvironments.

4.
Small ; 17(12): e2006110, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33721400

RESUMO

Chemical and functional anisotropy in Janus materials offer intriguing possibilities for constructing complex nanostructures and regulating chemical and biological reactions. Here, the authors report the fabrication of Janus nanosheets from molecular building blocks composed of two information-carrying biopolymers, DNA and peptides. Experimental and structural modeling studies reveal that DNA-peptide diblock conjugates assemble into Janus nanosheets with distinct DNA and peptide faces. The surprising level of structural control is attributed to the exclusive parallel ß-sheet formation of phenylalanine-rich peptides. This approach is extended to triblock DNA1-peptide-DNA2 conjugates, which assemble into nanosheets presenting two different DNA on opposite faces. The Janus nanosheets with independently addressable faces are utilized to organize an enzyme pair for concerted enzymatic reactions, where enhanced catalytic activities are observed. These results demonstrate that the predictable and designable peptide interaction is a promising tool for creating Janus nanostructures with regio-selective and sequence-specific molecular recognition properties.


Assuntos
DNA , Nanoestruturas , Peptídeos , Fenilalanina
5.
Biomater Res ; 24(1): 23, 2020 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-33334374

RESUMO

BACKGROUND: The aqueous solution behavior of thermosensitive PEG-PA block copolymers as well as secondary structure of PA is expected to significantly change through modification of the hydrophobic PA by long chain alkyl (C18) groups with different configurations. METHOD: Oleoyl and stearoyl (C18) groups were conjugated to poly(ethylene glycol)-poly(L-alanine) (PEG-PA; EG45A16) diblock copolymers to compare their conjugation effect on nano-assemblies and corresponding aqueous solution behavior of the polymers. RESULTS: Due to the nature of a hydrophilic PEG block and a hydrophobic PA or C18-modified PA, PEG-PA, oleoyl group-conjugated PEG-PA (PEG-PAO), and stearoyl group-conjugated PEG-PA (PEG-PAS) block copolymers form micelles in water. Compared with PEG-PA, the micelle size of PEG-PAO and PEG-PAS increased. Circular dichroism and FTIR spectra of aqueous polymer solutions showed that ß sheet content increased, whereas α helix content decreased by C18 modification of PEG-PA. PEG-PAS showed better performance in ice crystallization inhibition than PEG-PAO. The sol-to-gel transition temperatures of aqueous PEG-PAO solutions were 25-37 °C higher than those of aqueous PEG-PA solutions, whereas aqueous PEG-PAS solutions remained as gels in the temperature range of 0-80 °C. 1H-NMR spectra indicated that the oleoyl groups increased core mobility, whereas stearoyl groups decreased the core mobility of the micelles in water. The difference in micromobility between PAO and PAS interfered or promoted gelation of the aqueous polymer solutions, respectively. CONCLUSIONS: This study suggests that a hydrophobic C18-modification of polypeptide induces α helix-to-ß sheet transition of the polypeptide; however, aqueous solution behaviors including ice recrystallization inhibition and gelation are significantly affected by the nature of the hydrophobic molecule.

6.
J Am Chem Soc ; 142(45): 19142-19149, 2020 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-33074684

RESUMO

The efficient and selective light-driven conversion of carbon dioxide to formate is a scientific challenge for green chemistry and energy science, especially utilizing visible-light energy and earth-abundant catalytic materials. In this report, two mononuclear Ni(II) complexes of pyridylbenzimidazole (pbi) and pyridylbenzothiazole (pbt), such as Ni(pbt)(pyS)2 (1) and Ni(pbi)(pyS)2 (2) (pyS = pyridine-2-thiolate), were prepared and their reactivities studied. The two Ni complexes were examined for CO2 conversion using eosin Y as a photosensitizer upon visible-light irradiation in a H2O/ethanol solvent. The photoreaction of CO2 catalyzed by complexes 1 and 2 selectively affords formate with a high efficiency (14 000 turnover number) and a high catalytic selectivity of ∼99%. Undesirable proton reduction pathways were completely suppressed in the photocatalytic reactions with these sulfur-rich Ni catalysts under CO2. Hydrogen photoproduction was also studied under argon. Their kinetic isotope effects and influence of solution pH for formate and H2 production in the photocatalytic reactions are described in relation to the reaction mechanisms. These bioinspired Ni(II) catalysts with N/S ligation in relation to [NiFe]-hydrogenases are the first examples of early transition metal complexes affording such high selectivity and efficiencies, providing a future path to design solar-to-fuel processes for artificial photosynthesis.

7.
Biomacromolecules ; 21(8): 3176-3185, 2020 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-32640158

RESUMO

How to control osteochondral differentiation of mesenchymal stem cells at a proper stage is a key issue for articular cartilage regeneration. To solve this problem, injectable scaffolds with different chemical functional groups were designed by introducing one equivalent of α-cyclodextrin (α-CD) carboxylate and α-CD phosphate along poly(ethylene glycol)-poly(l-alanine) (PEG-L-PA) block copolymers. Dynamic light scattering, transmission electron microscopy images, and two-dimensional NMR spectra indicated that the PEG-L-PA block copolymers formed inclusion complexes with α-CD derivatives. Aqueous solutions of PEG-L-PA block copolymers (P), α-CD carboxylate/PEG-L-PA block copolymers (PCC), and α-CD phosphate/PEG-L-PA block copolymers (PCP) underwent sol-to-gel transition as the temperature increased. The storage moduli of P, PCC, and PCP gels ranged from 1000 to 1300 Pa at 37 °C. Tonsil-derived mesenchymal stem cells (TMSCs) were incorporated in situ in the gel during thermogelation of P, PCC, and PCP, which became the three-dimensional cell culture systems with different functional groups. After 21 days of incubation of TMSCs in the P, PCC, and PCP systems, the chondrogenic differentiation biomarker of type II collagen significantly increased in the P system, whereas the osteogenic biomarkers of osteocalcin and runt-related transcription factor 2 significantly increased in the PCP system. Both chondrogenic and osteogenic biomarkers were highly expressed in the PCC system. This study proved that thermogelling inclusion complex systems consisting of PEG-L-PA block copolymers and α-CD derivatives could be an excellent injectable matrix for fine-controlling osteochondral differentiation of mesenchymal stem cells.


Assuntos
Células-Tronco Mesenquimais , Diferenciação Celular , Condrogênese , Peptídeos , Polietilenoglicóis
8.
Biomacromolecules ; 21(1): 143-151, 2020 01 13.
Artigo em Inglês | MEDLINE | ID: mdl-31559819

RESUMO

A poly(ethylene glycol)-based thermogel can capture an iron ion (Fe3+) through a crown ether-like coordination bond between the oxygen atom and metal ions, thus, providing a sustained Fe3+-releasing system. Poly(ethylene glycol)-l-poly(alanine) thermogel was used in this study. The polypeptide forms a rather robust gel, and the degradation products are a neutral amino acid, which provides cyto-compatible neutral pH environments during the cell culture. During the heat-induced sol-to-gel transition at 37 °C, tonsil-derived mesenchymal stem cells (TMSCs) and iron ions were incorporated, leading to the formation of a three-dimensional matrix toward neuronal differentiation of the incorporated TMSCs. The initial concentration of the iron ions was varied between 0, 15, 30, and 60 mM. About 10% of the loaded iron ions was released over 21 days, which continuously supplied iron ions to the cells. The incorporation of iron ions not only increased the gel modulus at 37 °C from 107 to 680 Pa, but also promoted cell aggregation with a significant secretion of the cell adhesion signal of FAK. Expression of biomarkers related to the neuronal differentiation of TMSCs, including NFM, MAP2, GFAP, NURR1, NSE, and TUBB3, increased 4-35-fold at the mRNA level in the Fe3+-containing system compared to that of the system without Fe3+. Immunofluorescence studies also confirmed pronounced cell aggregation and a significant increase in neuronal biomarkers at the protein level. This study suggests that an iron ion-releasing thermogelling system can be a promising injectable scaffold toward neuronal differentiation of stem cells.


Assuntos
Géis/química , Géis/farmacocinética , Ferro/farmacocinética , Células-Tronco Mesenquimais/efeitos dos fármacos , Neurônios/citologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Criança , Feminino , Quinase 1 de Adesão Focal/genética , Marcadores Genéticos/genética , Temperatura Alta , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Neurônios/fisiologia , Tonsila Palatina/citologia , Peptídeos/química , Peptídeos/farmacocinética , Polietilenoglicóis/química , Temperatura de Transição
9.
Small ; 16(12): e1903045, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31523921

RESUMO

A reactive oxygen species (ROS)-sensitive degradable polymer would be a promising material in designing a disease-responsive system or accelerating degradation of polymers with slow hydrolysis kinetics. Here, a thermogelling poly(ethylene glycol)-polycaprolactone-poly(ethylene glycol) (PEG-PCL-PEG or EG12 -CL20 -EG12 ) triblock copolymer with an oxalate group at the middle of the polymer is reported. The polymers form micelles with an average size of 100 nm in water. Thermogelation is observed in a concentration range of 8.0-37.0 wt%. In particular, the aqueous PEG-PCL-PEG triblock copolymer solutions are in a gel state at 37 °C in a concentration range of 25.0-37.0 wt%, whereas the aqueous PEG-PCL diblock copolymer solutions are in a sol state in the same concentration range at 37 °C. Thus, the gel depot could dissolve out once degradation of the triblock copolymers occurs at the oxalate group as confirmed by the in vitro experiment. In vivo gel formation is confirmed by injecting an aqueous PEG-PCL-PEG solution (36.0 wt%) into the subcutaneous layer of rats. The gel completely disappears in 21 d. A model polypeptide drug (cyclosporine A) is released over 21 d from the in situ formed gel. The micelle-based thermogel of PEG-PCL-PEG with ROS-triggering degradability is a promising injectable material for biomedical applications.


Assuntos
Micelas , Poliésteres , Animais , Polietilenoglicóis , Polímeros , Ratos , Espécies Reativas de Oxigênio
10.
World J Stem Cells ; 11(8): 506-518, 2019 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-31523370

RESUMO

Located near the oropharynx, the tonsils are the primary mucosal immune organ. Tonsil tissue is a promising alternative source for the high-yield isolation of adult stem cells, and recent studies have reported the identification and isolation of tonsil-derived stem cells (T-SCs) from waste surgical tissue following tonsillectomies in relatively young donors (i.e., under 10 years old). As such, T-SCs offer several advantages, including superior proliferation and a shorter doubling time compared to bone marrow-derived mesenchymal stem cells (MSCs). T-SCs also exhibit multi-lineage differentiation, including mesodermal, endodermal (e.g., hepatocytes and parathyroid-like cells), and even ectodermal cells (e.g., Schwann cells). To this end, numbers of researchers have evaluated the practical use of T-SCs as an alternative source of autologous or allogenic MSCs. In this review, we summarize the details of T-SC isolation and identification and provide an overview of their application in cell therapy and regenerative medicine.

11.
Biomacromolecules ; 19(6): 2302-2307, 2018 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-29742350

RESUMO

Polycaprolactone (PCL) was reported a long time ago; however, its biomedical applications has not been extensively investigated in comparison with poly(lactide- co-glycolide) (PLGA) due to its too slow degradation profile. Here, we are reporting an oxalate-connected oligocaprolactone multiblock copolymer (PCL-OX) as a fast degradable PCL while maintaining its crystalline properties and low melting point of PCL. The in vivo application of the paclitaxel-loaded PCL-OX microspheres provided a steady plasma drug concentration of 6-9 µg/mL over 28 days, similar to that of the PLGA microspheres. Both PCL and PLGA microspheres were completely cleared two months after in vivo implantation. The PCL-OX microspheres showed a similar tissue compatibility to that of PLGA microspheres in the subcutaneous layer of rats. These findings suggest that PCL-OX is a useful biomaterial that solves the slow degradation problems of PCL and, thus, may find uses in other biomedical applications as an alternative to PLGA.


Assuntos
Plásticos Biodegradáveis , Sistemas de Liberação de Medicamentos , Microesferas , Paclitaxel , Poliésteres , Animais , Plásticos Biodegradáveis/síntese química , Plásticos Biodegradáveis/química , Plásticos Biodegradáveis/farmacocinética , Plásticos Biodegradáveis/farmacologia , Implantes de Medicamento , Masculino , Paclitaxel/química , Paclitaxel/farmacocinética , Paclitaxel/farmacologia , Poliésteres/química , Poliésteres/farmacocinética , Poliésteres/farmacologia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacocinética , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacologia , Ratos , Ratos Sprague-Dawley
12.
Biomaterials ; 159: 91-107, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29316455

RESUMO

Thermogel is an aqueous polymer solution that undergoes sol-to-gel transition as the temperature increases. Cells, growth factors, and signaling molecules can be incorporated simultaneously during the sol-to-gel transition. The cytocompatible procedure makes the thermogel an excellent platform for 3D culture of stem cells. This review focuses on the crucial questions that need to be addressed to achieve effective differentiation of stem cells into target cells, comprising low modulus, cell adhesion, and controlled supply of the growth factors. Recent progress in the use of thermogel as a 3D culture system of stem cells is summarized, and our perspectives on designing a new thermogel for 3D culture and its eventual application to injectable tissue engineering of stem cells are presented.


Assuntos
Polímeros/química , Células-Tronco/citologia , Engenharia Tecidual/métodos , Adesão Celular/fisiologia , Diferenciação Celular/fisiologia , Temperatura
13.
Tissue Eng Regen Med ; 15(5): 521-530, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30603576

RESUMO

Background: Thermogel is an aqueous solution that exhibits a sol-to-gel transition as the temperature increases. Stem cells, growth factors, and differentiating factors can be incorporated in situ in the matrix during the sol-to-gel transition, leading to the formation of a three-dimensional (3D) cell-culture scaffold. Methods: The uses of thermogelling polypeptides, such as collagen, Matrigel™, elastin-like polypeptides, and synthetic polypeptides, as 3D scaffolds of cells, are summarized in this paper. Results: The timely supply of growth factors to the cells, cell survival, and metabolite removal is to be insured in the cell culture matrix. Various growth factors were incorporated in the matrix during the sol-to-gel transition of the thermogelling polypeptide aqueous solutions, and preferential differentiation of the incorporated stem cells into specific target cells were investigated. In addition, modulus of the matrix was controlled by post-crosslinking reactions of thermogels or employing composite systems. Chemical functional groups as well as biological factors were selected appropriately for targeted differentiation of the incorporated stem cells. Conclusion: In addition to all the advantages of thermogels including mild conditions for cell-incorporation and controlled supplies of the growth factors, polypeptide thermogels provide neutral pH environments to the cells during the degradation of the gel. Polypeptide thermogels as an injectable scaffold can be a promising system for their eventual in vivo applications in stem cell therapy.

14.
ACS Appl Mater Interfaces ; 9(49): 42668-42675, 2017 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-29165981

RESUMO

Stem cell therapy for damaged cartilage suffers from low rates of retention, survival, and differentiation into chondrocytes at the target site. To solve these problems, here we propose a two-dimensional/three-dimensional (2D/3D) nanocomposite system. As a new two-dimensional (2D) material, hexagonal layered double hydroxides (LDHs) with a uniform lateral length of 2-3 µm were prepared by a hydrothermal process. Then, tonsil-derived mesenchymal stem cells (TMSCs), arginylglycylaspartic acid-coated LDHs, and kartogenin (KGN) were incorporated into the gel through the thermal-energy-driven gelation of the system. The cells exhibited a tendency to aggregate in the nanocomposite system. In particular, chondrogenic biomarkers of type II collagen and transcription factor SOX 9 significantly increased at both the mRNA and protein levels in the nanocomposite system, compared to the pure thermogel systems. The inorganic 2D materials increased the rigidity of the matrix, slowed down the release of a soluble factor (KGN), and improved cell-material interactions in the gel. The current 2D/3D nanocomposite system of bioactive LDH/thermogel can be a new platform material overcoming drawbacks of hydrogel-based 3D cell culture systems and is eventually expected to be applied as an injectable stem cell therapy.


Assuntos
Nanocompostos , Diferenciação Celular , Células Cultivadas , Condrócitos , Condrogênese , Hidróxidos , Células-Tronco Mesenquimais , Peptídeos , Células-Tronco , Engenharia Tecidual
15.
Biomacromolecules ; 18(7): 2214-2219, 2017 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-28605182

RESUMO

Pluronics F127 (P, PEG-PPG-PEG triblock copolymer) was coupled with diphenylalanine (FF) to prepare FF-end-capped Pluronics (FFPFF). With increasing temperature from 10 to 60 °C, the FFPFF self-assembled to vesicles in water. The unimer-to-vesicle transition accompanies endothermic enthalpy of 53.9 kcal/mol. Aqueous P and FFPFF solutions exhibited thermogelation in 15.0-24.0 wt %. The gel phase of FFPFF was stable up to 90 °C, whereas that of P turned into a sol again at 55-86 °C, indicating that end-capping with FF improved the gel stability against heat. In addition, the carboxylic acids of the FF end-groups can form coordination bonds with metal ions, and the gel modulus at 37 °C increased from 15-21 KPa (P) to 20-25 KPa (FFPFF) to 24-28 KPa (FFPFF-Zn), and the duration of gel against water-erosion increased from 24 h (P) to 60 h (FFPFF-Zn), leading to a useful biomaterial for sustained drug delivery. The FFPFF-Zn gels implanted in the rats' subcutaneous layer induced a mild inflammatory responses. Contrary to the previous end-capping of Pluronics by poly(lactic acid), polycarprolactone, carboxylic acid, and so on that weakened the gel stability, the diphenylalanine end-capping strengthened the stability of Pluronics gel against heat and water-erosion. This paper suggests that the control of polymer nanoassemblies directed by FF end-groups improves the mechanical properties and stability of the resulting thermogel and, thus, provides a useful drug delivery carrier with prolonged durability.


Assuntos
Micelas , Poloxâmero , Animais , Portadores de Fármacos/síntese química , Portadores de Fármacos/química , Portadores de Fármacos/farmacologia , Géis , Poloxâmero/química , Poloxâmero/farmacologia , Ratos , Água
16.
ACS Chem Neurosci ; 8(7): 1455-1458, 2017 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-28452458

RESUMO

Nuclear receptor related 1 (NURR1) is an essential protein for maintenance of dopaminergic neurons in adult midbrain of which deficiency leads to Parkinson's disease. To enhance the NURR1 production of neural cells, various approaches are under investigation. Here we report that NURR1 is highly expressed in stem cells by exposure to an L-polarized blue light emitting diode (LED). Compared to stem cells cultured in the absence of a LED, under polarized green and red LEDs, the stem cells exposed to a polarized blue LED significantly enhanced neuronal biomarkers such as neurofilament M (NFM) and neuron specific enolase (NSE) at both mRNA and protein levels. In particular, NURR1 was selectively enhanced by the stem cells exposed to the L-polarized blue LED. Stem cells exposed to the L-polarized blue LED increased mitochondrial ATP and intracellular calcium ions, which support neuronal differentiation of the stem cells. This study suggests that chiro-optical treatments by using polarized light with a specific wavelength can be used for engineering of stem cells with enhanced specific biochemicals, which may open a new method for a specific disease.


Assuntos
Luz , Células-Tronco Mesenquimais/metabolismo , Neurogênese , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/biossíntese , Trifosfato de Adenosina/metabolismo , Cálcio/metabolismo , Técnicas de Cultura de Células/instrumentação , Sobrevivência Celular , Criança , Feminino , Imunofluorescência , Expressão Gênica/efeitos da radiação , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos da radiação , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Proteínas de Neurofilamentos/biossíntese , Proteínas de Neurofilamentos/genética , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Tonsila Palatina , Fosfopiruvato Hidratase/biossíntese , Fosfopiruvato Hidratase/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
17.
ACS Appl Mater Interfaces ; 9(13): 11568-11576, 2017 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-28290667

RESUMO

A poly(ethylene glycol)-b-poly(l-alanine) (PEG-l-PA) hydrogel incorporating tonsil-derived mesenchymal stem cells (TMSCs), tauroursodeoxycholic acid (TUDCA), hepatocyte growth factor (HGF), and fibroblast growth factor 4 (FGF4) was prepared through thermal gelation of an aqueous polymer solution for an injectable tissue engineering application. The thermal gelation accompanied conformational changes of both PA and PEG blocks. The gel modulus at 37 °C was controlled to be 1000 Pa by using a 14.0 wt % aqueous polymer solution. The gel preserved its physical integrity during the 3D culture of the cells. TUDCA, HGF, and FGF4 were released from the PEG-l-PA hydrogel over 21 days of the 3D cell culture period. TMSCs initially exhibited a spherical shape, whereas some fibers protruded from the cells on days 14-21 of 3D culture. The injectable system exhibited pronounced expressions of the hepatic biomarkers at both mRNA and protein levels, which are significantly better than the commercially available hyaluronic acid gel. In particular, the hepatogenically differentiated cells from the TMSCs in the injectable system demonstrated hepatic biofunctions comparable to HepG2 cells for the uptakes of low density lipoproteins (52%) and indocyanine green (76%), and the production of albumin (40%) and urea (52%), which are also significantly better than the 3D-cultured cells in the commercially available hyaluronic acid gel. Our studies suggest that the PEG-l-PA thermogel incorporating TMSCs, TUDCA, and growth factors is highly promising as an in situ forming tissue engineering system.


Assuntos
Engenharia Tecidual , Diferenciação Celular , Células-Tronco Mesenquimais , Tonsila Palatina , Peptídeos , Polietilenoglicóis
18.
J Biomed Nanotechnol ; 12(5): 1023-34, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27305823

RESUMO

The development of efficient and safe gene delivery carriers has been a major challenge in the clinical application of non-viral gene therapy. Herein, we report novel bioreducible poly(amido amine)s for the efficient delivery of genetic material such as plasmid DNA. A library of 34 different bioreducible polymer compounds was synthesized and screened to find lead materials for in vitro gene transfection. Our lead material (CBA-106) allows effortless polyplex formation with genetic materials by electrostatic interactions at the weight ratio of 1:5 (DNA/polymer). Polyplexes were further characterized by DLS and AFM analysis. Enhanced serum stability and bioreducibility under physiological conditions were confirmed, in addition to low cellular cytotoxicity. When compared with a commercially available gene delivery carrier (Lipofectamine 2000), CBA-1 06 shows comparable or even surpassing gene transfection efficiency. Furthermore, BMP-2 plasmids were efficiently delivered to tonsil-derived mesenchymal stem cells (TMSCs) for osteogenic commitment in vitro and in vivo. Taken together, our results clearly demonstrate the potential of novel bioreducible polymeric systems for gene delivery applications. We suggest that our system can provide a valuable platform for the broad application of gene regulation in cell therapy and regenerative medicine.


Assuntos
Materiais Biocompatíveis/química , Diferenciação Celular , Técnicas de Transferência de Genes , Células-Tronco Mesenquimais/citologia , Osteogênese , Tonsila Palatina/citologia , Poliaminas/química , Animais , Proteína Morfogenética Óssea 2/metabolismo , Cátions , DNA/metabolismo , Difusão Dinâmica da Luz , Proteínas de Fluorescência Verde/metabolismo , Camundongos , Microscopia de Força Atômica , Plasmídeos/metabolismo , Poliaminas/síntese química , Transfecção
19.
Eur Arch Otorhinolaryngol ; 273(11): 3827-3834, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26994900

RESUMO

Surgical transplantation of parathyroid tissue into the forearm muscle is one of the most commonly used surgical techniques. While simple, the procedure suffers from drawbacks. This study evaluated the feasibility of thermoreversible gel as an injectable carrier for parathyroid autotransplantation. Polyethyleneglycol-polyalanine-co-phenylalanine (PEG-PAF) thermoreversible gel (sol form at 4 °C, gel form at 37 °C) were manufactured. Thirty-eight Sprague-Dawley rats were divided into two groups (19 control, C group; 19 experimental, P group). The parathyroid glands of rats were excised. Parathyroid tissues were transplanted into the muscle pocket in sternocleidomastoid muscle in the C group. In the P group, the tissues were injected into the same muscle mixed with 0.3 ml thermoreversible gel. The serum levels of parathyroid hormone (PTH), ionized calcium, and phosphorous were measured before surgical procedure, on 7, 21, 56, and 70 days after surgery. Histology and immunohistochemistry were performed. Preoperative median PTH level of the C and the P group were 60.80 and 43.85 pg/ml, respectively (p = 0.641). Seventy days after surgery, median PTH level was 32.8 and 25.61 pg/ml, respectively. On day 70, the PTH level was restored by 54 % in the C group and 56 % in the P group compared to the preoperative value (p = 0.620). There were no significant intergroup differences in the ionized calcium/phosphorous level. Histology and immunohistochemistry revealed the successful transplantation of parathyroid tissues into the muscles in both groups. In conclusion, the PEG-PAF-based thermoreversible gel is a good candidate carrier material for intramuscular parathyroid autotransplantation.


Assuntos
Glândulas Paratireoides/transplante , Tecidos Suporte , Animais , Estudos de Viabilidade , Feminino , Géis , Humanos , Injeções Intramusculares , Masculino , Pessoa de Meia-Idade , Hormônio Paratireóideo/sangue , Fenilalanina/química , Polietilenoglicóis/química , Ratos , Ratos Sprague-Dawley , Transplante Autólogo
20.
Biomacromolecules ; 17(3): 1075-82, 2016 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-26909543

RESUMO

There are four EF-hand polypeptides in calmodulin, a natural ubiquitous calcium binding protein that activates the enzymes involved in Ca(2+)-mediated signal transduction. An EF-hand polypeptide has six carboxylate functional groups in the middle loop region between two rigid polypeptides. In this study, a calcium binding polymer (CBP) with a structure of poly(L-alanine)-poly(L-alanine-co-L-glutamic acid)-poly(ethylene glycol)-poly(L-alanine-co-L-glutamic acid)-poly(L-alanine) (PA-PAE-PEG-PAE-PA; A11.1-A3.4E3.2-EG40.1-A3.4E3.2-A11.1) was synthesized by mimicking the EF-hand polypeptide. The 6-7 carboxylate functional groups from PAE are expected to form a binding site for Ca(2+). As the Ca(2+) bound to CBP, small changes in the circular dichroism spectra and (13)C NMR spectra were observed, indicating that Ca(2+) binding to CBP induced changes in the conformation of CBP. The binding constant of CBP to Ca(2+) was investigated by using the competitive binding of 2,2',2″,2‴-{ethane-1,2-diylbis[oxy(4-bromo-2,1-phenylene)nitrilo]} tetraacetic acid (5,5-Br2-BAPTA). The binding constant obtained with a CaLigator program by least-squares fitting of the absorbance profile as a function of Ca(2+) concentration was 5.1 × 10(5) M(-1), which was similar to that of calmodulin. The selectivity of CBP for metal ion binding was compared among Ca(2+), Cu(2+), and Zn(2+). The binding constant was obtained through a similar competitive binding study with murexide. The binding constants for Ca(2+), Cu(2+), and Zn(2+) were 7.0 × 10(5), 4.2 × 10(5), and 1.7 × 10(5) M(-1), respectively, indicating 2-4-fold higher selectivity of CBP for Ca(2+) compared to Cu(2+) and Zn(2+). The CBP has selectivity for Ca(2+), and binding affinity for Ca(2+) was similar to the biological Ca(2+) binding motif of calmodulin.


Assuntos
Cálcio/metabolismo , Motivos EF Hand , Peptídeos/química , Sítios de Ligação , Calmodulina/química , Calmodulina/metabolismo , Cobre/metabolismo , Peptídeos/síntese química , Peptídeos/metabolismo , Ligação Proteica , Zinco/metabolismo
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