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Background: Neural cell-electrode coupling is crucial for effective neural and retinal prostheses. Enhancing this coupling can be achieved through surface modification and geometrical design to increase neuron-electrode proximity. In the current research, we focused on designing and studying various biomolecules as a method to elicit neural cell-electrode adhesion via cell-specific integrin mechanisms. Methods: We designed extracellular matrix biomimetic molecules with different head sequences (RGD or YIGSR), structures (linear or cyclic), and spacer lengths (short or long). These molecules, anchored by a thiol (SH) group, were deposited onto gold surfaces at various concentrations. We assessed the modifications using contact angle measurements, fluorescence imaging, and X-ray Photoelectron Spectroscopy (XPS). We then analyzed the adhesion of retinal cells and HEK293 cells to the modified surfaces by measuring cell density, surface area, and focal adhesion spots, and examined changes in adhesion-related gene and integrin expression. Results: Results showed that YIGSR biomolecules significantly enhanced retinal cell adhesion, regardless of spacer length. For HEK293 cells, RGD biomolecules were more effective, especially with cyclic RGD and long spacers. Both cell types showed increased expression of specific adhesion integrins and proteins like vinculin and PTK2; these results were in agreement with the adhesion studies, confirming the cell-specific interactions with modified surfaces. Conclusion: This study highlights the importance of tailored biomolecules for improving neural cell adhesion to electrodes. By customizing biomolecules to foster specific and effective interactions with adhesion integrins, our study provides valuable insights for enhancing the integration and functionality of retinal prostheses and other neural implants.
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Peripheral nerve injuries (PNI) currently have limited therapeutic efficacy, and functional scaffolds have been shown to be effective for treating PNI. Ovalbumin (OVA) is widely used as a natural biomaterial for repairing damaged tissues due to its excellent biocompatibility and the presence of various bioactive components. However, there are few reports on the repair of PNI by ovalbumin. In this study, a novel bionic functionalized topological scaffold based on ovalbumin and grafted with tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide was constructed by micro-molding method and surface-biomodification technology. The scaffolds were subjected to a series of evaluations in terms of morphology, mechanics, hydrophilicity, and biocompatibility, and the related molecular mechanisms were further penetrated. The results showed that the scaffolds prepared in this study had aligned ridge/groove structure, good mechanical properties and biocompatibility, and could be used as carriers to slowly release YIGSR, which effectively promoted the proliferation, migration and elongation of Schwann Cells (SCs), and significantly up-regulated the gene expression related to proliferation, apoptosis, migration and axon regeneration. Therefore, the bionic functional topological scaffold has significant application potential for promoting peripheral nerve regeneration and provides a new therapeutic option for repairing PNI.
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Axones , Traumatismos de los Nervios Periféricos , Humanos , Ovalbúmina/metabolismo , Regeneración Nerviosa/fisiología , Células de Schwann , Péptidos/química , Traumatismos de los Nervios Periféricos/terapia , Andamios del Tejido/químicaRESUMEN
Impaired microcirculation can cause lymphatic leakage which leads to a chronic swelling in the tissues of the body. However, no successful treatment gives any protection against lymphedema due to the lack of well-revealed pathophysiology of secondary lymphedema. Binary image of laminin immunohistochemical expression revealed that distribution of laminin expression localized during surgically induced lymphedema. 67 kDa laminin receptor (67LR) mRNA expression showed a peak at during lymphedema exacerbation. Since the response of 67LR molecules may affect the prevention of inflammation and edema, here we have hypothesized that 67LR ligand of YIGSR peptide could permit reconstructive environment for amelioration of lymphedema and evaluated the effect of YIGSR in a mouse tail model of lymphedema. Indeed, intra-abdominal injections of YIGSR for the first 3 days after inducing lymphedema in the mouse tail model reduced the tail lymphedema on day 14 by 27% (P = 0.035). Histology showed that YIGSR treatment protected lymphedema impairment in epidermis and dermis, and it also inhibited the expansion of intercellular spaces and enhanced especially cell adhesion in the basement membrane as revealed by transmission electron microscopy. Interestingly, the treatment also reduced the local expression of transforming growth factor (TGF)ß. Further elucidation of the mechanisms of 67LR-facilitated lymphangiogenesis contributes to find potential targets for the treatment of lymphedema.
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Dual-modal magnetic resonance/fluorescent imaging (MRI/FI) attracts moreandmoreattentions in diagnosis of tumors. A corresponding dual-modal imaging agent with sufficient tumor sensitivity and specificity should be matched to improve imaging quality. Tripeptide (RGD) and pentapeptide (YIGSR) were selected as the tumor-targeting groups and attached to gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) and rhodamine B (RhB), and then make two novel polypeptide-based derivatives (RGD-Gd-DTPA-RhB and YIGSR-Gd-DTPA-RhB), respectively. These derivatives were further characterized and their properties, such as cell uptake, cell cytotoxicity, MRI and FI assay, were measured. YIGSR-Gd-DTPA-RhB and RGD-Gd-DTPA-RhB had high relaxivity, good tumor-targeting property, low cell cytotoxicity and good red FI in B16F10 melanoma cells. Moreover, YIGSR-Gd-DTPA-RhB and RGD-Gd-DTPA-RhB possessed high uptake to B16F10 melanoma, and then achieve highly enhanced FI and MRI of tumors in mice for a prolonged time. Therefore, YIGSR-Gd-DTPA-RhB and RGD-Gd-DTPA-RhB can be applied as the potential agents for tumor targeted MRI/FI in vivo.
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Medios de Contraste , Melanoma , Ratones , Animales , Medios de Contraste/química , Gadolinio DTPA/farmacología , Gadolinio DTPA/química , Gadolinio/química , Imagen por Resonancia Magnética/métodos , Oligopéptidos/farmacología , Imagen Óptica/métodos , Espectroscopía de Resonancia MagnéticaRESUMEN
This study describes a method for the modification of polyurethane small-diameter (5 mm) vascular prostheses obtained with the phase inversion method. The modification process involves two steps: the introduction of a linker (acrylic acid) and a peptide (REDV and YIGSR). FTIR and XPS analysis confirmed the process of chemical modification. The obtained prostheses had a porosity of approx. 60%, Young's Modulus in the range of 9-11 MPa, and a water contact angle around 40°. Endothelial (EC) and smooth muscle (SMC) cell co-culture showed that the surfaces modified with peptides increase the adhesion of ECs. At the same time, SMCs adhesion was low both on unmodified and peptide-modified surfaces. Analysis of blood-materials interaction showed high hemocompatibility of obtained materials. The whole blood clotting time assay showed differences in the amount of free hemoglobin present in blood contacted with different materials. It can be concluded that the peptide coating increased the hemocompatibility of the surface by increasing ECs adhesion and, at the same time, decreasing platelet adhesion. When comparing both types of peptide coatings, more promising results were obtained for the surfaces coated with the YISGR than REDV-coated prostheses.
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Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Prótesis Vascular , Poliuretanos/química , Poliuretanos/farmacología , Animales , Materiales Biocompatibles/síntesis química , Coagulación Sanguínea/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Técnicas de Cocultivo , Células Endoteliales de la Vena Umbilical Humana/citología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Humanos , Ensayo de Materiales , Fenómenos Mecánicos , Ratones , Microscopía Electrónica de Rastreo , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/efectos de los fármacos , Adhesividad Plaquetaria/efectos de los fármacos , Poliuretanos/síntesis química , Porosidad , Propiedades de SuperficieRESUMEN
In recent years, there has been an increasing interest toward the covalent binding of bioactive peptides from extracellular matrix proteins on scaffolds as a promising functionalization strategy in the development of biomimetic matrices for tissue engineering. A totally new approach for scaffold functionalization with peptides is based on Molecular Imprinting technology. In this work, imprinted particles with recognition properties toward laminin and fibronectin bioactive moieties were synthetized and used for the functionalization of biomimetic sponges, which were based on a blend of alginate, gelatin, and elastin. Functionalized sponges underwent a complete morphological, physicochemical, mechanical, functional, and biological characterization. Micrographs of functionalized sponges showed a highly porous structure and a quite homogeneous distribution of imprinted particles on their surface. Infrared and thermal analyses pointed out the presence of interactions between blend components. Biodegradation and mechanical properties appeared adequate for the aimed application. The results of recognition tests showed that the deposition on sponges did not alter the specific recognition and binding behavior of imprinted particles. In vitro biological characterization with cardiac progenitor cells showed that early cell adherence was promoted. In vivo analysis showed that developed scaffolds improved cardiac progenitor cell adhesion and differentiation toward myocardial phenotypes.
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Abnormal migration and proliferation of endothelial cells (EC) drive neovascular retinopathies. While anti-VEGF treatment slows progression, pathology is often supported by decrease in intraocular pigment epithelium-derived factor (PEDF), an endogenous inhibitor of angiogenesis. A surface helical 34-mer peptide of PEDF, comprising this activity, is efficacious in animal models of neovascular retina disease but remains impractically large for therapeutic use. We sought smaller fragments within this sequence that mitigate choroidal neovascularization (CNV). Expecting rapid intravitreal (IVT) clearance, we also developed a method to reversibly attach peptides to nano-carriers for extended delivery. Synthetic fragments of 34-mer yielded smaller anti-angiogenic peptides, and N-terminal capping with dicarboxylic acids did not diminish activity. Charge restoration via substitution of an internal aspartate by asparagine improved potency, achieving low nM apoptotic response in VEGF-activated EC. Two optimized peptides (PEDF 335, 8-mer and PEDF 336, 9-mer) were tested in a mouse model of laser-induced CNV. IVT injection of either peptide, 2-5 days before laser treatment, gave significant CNV decrease at day +14 post laser treatment. The 8-mer also decreased CNV, when administered as eye drops. Also examined was a nanoparticle-conjugate (NPC) prodrug of the 9-mer, having positive zeta potential, expected to display longer intraocular residence. This NPC showed extended efficacy, even when injected 14 days before laser treatment. Neither inflammatory cells nor other histopathologic abnormalities were seen in rabbit eyes harvested 14 days following IVT injection of PEDF 336 (>200 µg). No rabbit or mouse eye irritation was observed over 12-17 days of PEDF 335 eye drops (10 mM). Viability was unaffected in 3 retinal and 2 choroidal cell types by PEDF 335 up to 100 µM, PEDF 336 (100 µM) gave slight growth inhibition only in choroidal EC. A small anti-angiogenic PEDF epitope (G-Y-D-L-Y-R-V) was identified, variants (adipic-Sar-Y-N-L-Y-R-V) mitigate CNV, with clinical potential in treating neovascular retinopathy. Their shared active motif, Y - - - R, is found in laminin (Ln) peptide YIGSR, which binds Ln receptor 67LR, a known high-affinity ligand of PEDF 34-mer.
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Inhibidores de la Angiogénesis/uso terapéutico , Neovascularización Coroidal/prevención & control , Proteínas del Ojo/uso terapéutico , Factores de Crecimiento Nervioso/uso terapéutico , Oligopéptidos/uso terapéutico , Serpinas/uso terapéutico , Administración Oftálmica , Inhibidores de la Angiogénesis/química , Animales , Apoptosis , Neovascularización Coroidal/metabolismo , Neovascularización Coroidal/patología , Modelos Animales de Enfermedad , Portadores de Fármacos , Electrorretinografía , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Proteínas del Ojo/química , Ratones , Ratones Endogámicos C57BL , Factores de Crecimiento Nervioso/química , Oligopéptidos/química , Soluciones Oftálmicas , Profármacos , Conejos , Ratas , Serpinas/químicaRESUMEN
The basement membrane is a specialized extracellular matrix substrate responsible for support and maintenance of epithelial and endothelial structures. Engineered basement membrane-like hydrogel systems have the potential to advance understanding of cell-cell and cell-matrix interactions by allowing precise tuning of the substrate or matrix biochemical and biophysical properties. In this investigation, we developed tunable hydrogel substrates with conjugated bioactive peptides to modulate cell binding and growth factor signaling by endothelial cells. Hydrogels were formed by employing a poly(ethylene glycol) crosslinker to covalently crosslink gelatin polymers and simultaneously conjugate laminin-derived YIGSR peptides or vascular endothelial growth factor (VEGF)-mimetic QK peptides to the gelatin. Rheological characterization revealed rapid formation of hydrogels with similar stiffnesses across tested formulations, and swelling analysis demonstrated dependency on peptide and crosslinker concentrations in hydrogels. Levels of phosphorylated VEGF Receptor 2 in cells cultured on hydrogel substrates revealed that while human umbilical vein endothelial cells (HUVECs) responded to both soluble and conjugated forms of the QK peptide, conditionally-immortalized human glomerular endothelial cells (GEnCs) only responded to the conjugated presentation of the peptide. Furthermore, whereas HUVECs exhibited greatest upregulation in gene expression when cultured on YIGSR- and QK-conjugated hydrogel substrates after 5 days, GEnCs exhibited greatest upregulation when cultured on Matrigel control substrates at the same time point. These results indicate that conjugation of bioactive peptides to these hydrogel substrates significantly influenced endothelial cell behavior in cultures but with differential responses between HUVECs and GEnCs.
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Materiales Biocompatibles/química , Células Endoteliales/efectos de los fármacos , Gelatina/química , Hidrogeles/química , Péptidos/química , Polietilenglicoles/química , Polímeros/química , Aminas/metabolismo , Membrana Basal , Células Endoteliales de la Vena Umbilical Humana , Humanos , Concentración de Iones de Hidrógeno , Microscopía Electrónica de Rastreo , Péptidos/farmacología , Polímeros/farmacologíaRESUMEN
Substrates for embryonic stem cell culture are typified by poorly defined xenogenic, whole proteins or cellular components that are difficult and expensive to generate, characterize, and recapitulate. Herein, the generation of well-defined scaffolds of Gly-Tyr-Ile-Gly-Ser-Arg (GYIGSR) peptide-functionalized poly(ε-caprolactone) (PCL) aligned nanofibers are used to accelerate the neural lineage commitment and differentiation of D3 mouse embryonic stem cells (mESCs). Gene expression trends and immunocytochemistry analysis were similar to laminin-coated glass, and indicated an earlier differentiation progression than D3 mESCs on laminin. Further, GYIGSR-functionalized nanofiber substrates yielded an increased gene expression of Sox1, a neural progenitor cell marker, and Tubb3, Cdh2, Syp, neuronal cell markers, at early time points. In addition, guidance of neurites was found to parallel the fiber direction. We demonstrate the fabrication of a well-defined, xeno-free functional nanofiber scaffold and demonstrates its use as a surrogate for xenogenic and complex matrixes currently used for the neural differentiation of stem cells ex vivo. STATEMENT OF SIGNIFICANCE: In this paper, we report the use of GYIGSR-functionalized poly(ε-caprolactone) aligned nanofibers as a tool to accelerate the neural lineage commitment and differentiation of D3 mouse embryonic stem cells. The results indicate that functional nanofiber substrates promote faster differentiation than laminin coated substrates. The data suggest that aligned nanofibers and post-electrospinning surface modification with bioactive species can be combined to produce translationally relevant xeno-free substrates for stem cell therapy. Future development efforts are focused on additional bioactive species that are able to function as surrogates for other xenogenic factors found in differentiation media.
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Diferenciación Celular , Células Madre Embrionarias de Ratones/metabolismo , Nanofibras/química , Neuronas/metabolismo , Péptidos/química , Andamios del Tejido/química , Animales , Antígenos de Diferenciación/biosíntesis , Línea Celular , Regulación de la Expresión Génica , Humanos , Ratones , Células Madre Embrionarias de Ratones/citología , Neuronas/citologíaRESUMEN
As an effective clinic treatment for cardiovascular disease, vascular transplantation gains much acceptance recently. However, due to the acute thrombosis and intimal hyperplasia, long-term failure of synthetic grafts after implanted in small diameter blood vessel decelerates its commercial use. The continued acute inflammation and delayed endothelialization have been considered as fundamental reasons. To enhance the adhesion and organization of endothelial cells (ECs) and improve the vascular remodeling process, we have constructed a vascular graft based on electrospun polycaprolactone (PCL) matrix, on which organoselenium-immobilized polyethyleneimine (SePEI) for in situ nitric oxide (NO) generation and hyaluronic acid (HA) grafted with poly (ethylene glycol) (PEG) modified Tyr-Ile-Gly-Ser-Arg (YIGSR) for antifouling and EC adhesion were deposited through electrostatic layer-by-layer assembly. The in vitro results showed that SePEI deposited on the grafts could catalyze stable generation of NO. After in situ implantation in rats for 4 and 8weeks, the graft promoted the transformation of macrophages into an anti-inflammatory phenotype (M2), which helped endothelium remodeling. YIGSR on the outmost layer facilitated more rapid and organized EC adhesion compared to PCL and non-modified grafts. PEG polymer chain on the outmost layer mitigated nonspecific adsorption of undesirable blood components. In our study, we first demonstrated the regulation of macrophage polarization by an NO-generating vascular graft. The results indicated that the approach of anti-inflammatory macrophage polarization and enhanced endothelialization through NO generation and PEG-modified YIGSR in our study may provide a new perspective for the clinic application of cell-free small-diameter vascular grafts.
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Óxido Nítrico/metabolismo , Oligopéptidos/química , Polietilenglicoles/química , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/metabolismo , Adhesión Celular , Polaridad Celular , Proliferación Celular , Endotelio Vascular/citología , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Macrófagos/citología , Macrófagos/metabolismo , Masculino , Ratones , Óxido Nítrico/química , Poliésteres/química , Células RAW 264.7 , Ratas , Ratas Sprague-Dawley , Electricidad Estática , Injerto VascularRESUMEN
Since the use of animal experimentation is restricted with regard to cosmetic materials, alternative in vitro models such as skin equivalents (SEs) are needed. Laminin is one of the major non-collagenous glycoproteins. The pentapeptide YIGSR (Tyr-Ile-Gly-Ser-Arg) is a functional motif of laminin that binds to the laminin receptor. In the present study, we examined whether YIGSR could improve the reconstruction of SEs. YIGSR has no effects on monolayer cell proliferation of CCD25-Sk fibroblasts or HaCaT keratinocytes. Interestingly, YIGSR decreased TGF-ß1 levels, although it promoted type Ι collagen synthesis in CCD25-Sk cells. In HaCaT cells, YIGSR decreased the expression of involucrin and loricrin, which are differentiation markers. Furthermore, YIGSR increased levels of proliferating cell nuclear antigen (PCNA), p63, and integrin α6, and decreased involucrin in SE models. In addition, two models containing YIGSR (mixed with dermal equivalents or added into media) did not show any differences in expression levels of PCNA, p63, integrin α6, and involucrin. Therefore, YIGSR is a useful agent for reconstruction of SEs, independent of its method of application. These results indicate that YIGSR stimulates epidermal proliferation and basement membrane formation while inhibiting keratinocyte differentiation of SEs. Taken together, these results indicate that YIGSR promotes the reconstruction of SEs, potentially via decreased TGF-ß1 levels and consequent inhibition of epidermal differentiation.
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Biomimética/normas , Laminina/biosíntesis , Oligopéptidos/biosíntesis , Piel/patología , Fibroblastos/patología , Humanos , República de CoreaRESUMEN
Controlling the cell-matrix interaction is a critical factor in the design and fabrication of tissue engineering scaffolds. A particular peptide sequence, Arg-Gly-Asp (RGD peptide), is often used as an adhesion ligand in the engineering of different types of tissues. While in some cases this has been adequate, the use of multiple ligands may be required for the successful engineering of some tissue types. We hypothesized that hydrogels presenting both the RGD peptide and the YIGSR peptide (Tyr-Ile-Gly-Ser-Arg) could successfully regulate the phenotype of PC12 cells, thereby providing a new platform for effective tissue engineering applications. We prepared alginate hydrogels modified with both RGD and YIGSR peptides at several different bulk ligand densities and determined the ways in which PC12 cells can respond to them in vitro. We demonstrate that alginate hydrogels presenting both RGD and YIGSR peptides successfully regulate the proliferation, morphological change, and neuronal differentiation of PC12 cells in vitro. Successful adhesion and proliferation of PC12 cells were dependent on the bulk density of RGD peptides, while neuronal differentiation was significantly enhanced by increasing the YIGSR density. These results suggest that hydrogels presenting multiple adhesion ligands offer many useful applications in tissue engineering approaches.
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Hidrogeles/química , Alginatos/química , Animales , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Neuronas/citología , Neuronas/efectos de los fármacos , Oligopéptidos/química , Oligopéptidos/farmacología , Células PC12 , RatasRESUMEN
This article presents data related to the research article "Systematic optimization of an engineered hydrogel allows for selective control of human neural stem cell survival and differentiation after transplantation in the stroke brain" (P. Moshayedi, L.R. Nih, I.L. Llorente, A.R. Berg, J. Cinkornpumin, W.E. Lowry et al., 2016) [1] and focuses on the biocompatibility aspects of the hydrogel, including its stiffness and the inflammatory response of the transplanted organ. We have developed an injectable hyaluronic acid (HA)-based hydrogel for stem cell culture and transplantation, to promote brain tissue repair after stroke. This 3D biomaterial was engineered to bind bioactive signals such as adhesive motifs, as well as releasing growth factors while supporting cell growth and tissue infiltration. We used a Design of Experiment approach to create a complex matrix environment in vitro by keeping the hydrogel platform and cell type constant across conditions while systematically varying peptide motifs and growth factors. The optimized HA hydrogel promoted survival of encapsulated human induced pluripotent stem cell derived-neural progenitor cells (iPS-NPCs) after transplantation into the stroke cavity and differentially tuned transplanted cell fate through the promotion of glial, neuronal or immature/progenitor states. The highlights of this article include: (1) Data of cell and bioactive signals addition on the hydrogel mechanical properties and growth factor diffusion, (2) the use of a design of Experiment (DOE) approach (M.W. 2 Weible and T. Chan-Ling, 2007) [2] to select multi-factorial experimental conditions, and (3) Inflammatory response and cell survival after transplantation.
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The 37/67-kDa laminin receptor (LAMR/RPSA) was originally identified as a 67-kDa binding protein for laminin, an extracellular matrix glycoprotein that provides cellular adhesion to the basement membrane. LAMR has evolutionary origins, however, as a 37-kDa RPS2 family ribosomal component. Expressed in all domains of life, RPS2 proteins have been shown to have remarkably diverse physiological roles that vary across species. Contributing to laminin binding, ribosome biogenesis, cytoskeletal organization, and nuclear functions, this protein governs critical cellular processes including growth, survival, migration, protein synthesis, development, and differentiation. Unsurprisingly given its purview, LAMR has been associated with metastatic cancer, neurodegenerative disease and developmental abnormalities. Functioning in a receptor capacity, this protein also confers susceptibility to bacterial and viral infection. LAMR is clearly a molecule of consequence in human disease, directly mediating pathological events that make it a prime target for therapeutic interventions. Despite decades of research, there are still a large number of open questions regarding the cellular biology of LAMR, the nature of its ability to bind laminin, the function of its intrinsically disordered C-terminal region and its conversion from 37 to 67 kDa. This review attempts to convey an in-depth description of the complexity surrounding this multifaceted protein across functional, structural and pathological aspects.
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Regulación de la Expresión Génica/fisiología , Laminina/metabolismo , Receptores de Laminina/metabolismo , Proteínas Ribosómicas/metabolismo , Humanos , Laminina/genética , Receptores de Laminina/genética , Proteínas Ribosómicas/genéticaRESUMEN
Understanding ß cell-extracellular matrix (ECM) interactions can advance our knowledge of the mechanisms that control glucose homeostasis and improve culture methods used in islet transplantation for the treatment of diabetes. Laminin is the main constituent of the basement membrane and is involved in pancreatic ß cell survival and function, even enhancing glucose-stimulated insulin secretion. Most of the studies on cell responses towards laminin have focused on integrin-mediated interactions, while much less attention has been paid on non-integrin receptors, such as the 67 kDa laminin receptor (67LR). The specificity of the receptor-ligand interaction through the adhesion of INS-1 cells (a rat insulinoma cell line) to CDPGYIGSR-, GRGDSPC- or CDPGYIGSR + GRGDSPC-covered surfaces was evaluated. Also, the effects of the 67LR knocking down over glucose-stimulated insulin secretion were investigated. Culture of the INS-1 cells on the bioactive surfaces was improved compared to the low-fouling carboxymethyl dextran (CMD) surfaces, while downregulation of the 67LR resulted in reduced cell adhesion to surfaces bearing the CDPGYIGSR peptide. Glucose-stimulated insulin secretion was hindered by downregulation of the 67LR, regardless of the biological motif available on the biomimetic surfaces on which the cells were cultured. This finding illustrates the importance of the 67LR in glucose-stimulated insulin secretion and points to a possible role of the 67LR in the mechanisms of insulin secretion.
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Regulación hacia Abajo/efectos de los fármacos , Glucosa/farmacología , Insulina/metabolismo , Péptidos/farmacología , Receptores de Laminina/agonistas , Animales , Línea Celular Tumoral , Secreción de Insulina , Ratas , Receptores de Laminina/biosíntesisRESUMEN
Defects in the corneal stroma caused by trauma or diseases such as macular corneal dystrophy and keratoconus can be detrimental for vision. Development of therapeutic methods to enhance corneal regeneration is essential for treatment of these defects. This paper describes a bioactive peptide nanofiber scaffold system for corneal tissue regeneration. These nanofibers are formed by self-assembling peptide amphiphile molecules containing laminin and fibronectin inspired sequences. Human corneal keratocyte cells cultured on laminin-mimetic peptide nanofibers retained their characteristic morphology, and their proliferation was enhanced compared with cells cultured on fibronectin-mimetic nanofibers. When these nanofibers were used for damaged rabbit corneas, laminin-mimetic peptide nanofibers increased keratocyte migration and supported stroma regeneration. These results suggest that laminin-mimetic peptide nanofibers provide a promising injectable, synthetic scaffold system for cornea stroma regeneration.
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Materiales Biocompatibles/farmacología , Sustancia Propia/fisiología , Nanofibras/química , Péptidos/farmacología , Regeneración/efectos de los fármacos , Secuencia de Aminoácidos , Animales , Proliferación Celular/efectos de los fármacos , Forma de la Célula/efectos de los fármacos , Células Cultivadas , Sustancia Propia/citología , Sustancia Propia/efectos de los fármacos , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/ultraestructura , Humanos , Inmunohistoquímica , Datos de Secuencia Molecular , Nanofibras/ultraestructura , Péptidos/química , Conejos , Tensoactivos/químicaRESUMEN
End-functional PLLA nanofibers were fabricated into mats of random or aligned fibers and functionalized post-spinning using metal-free "click chemistry" with the peptide Tyr-Ile-Gly-Ser-Arg (YIGSR). Fibers that were both aligned and functionalized with YIGSR were found to significantly increase the fraction of mouse embryonic stem cells (mESC) expressing neuron-specific class III beta-tubulin (TUJ1), the level of neurite extension and gene expression for neural markers compared to mESC cultured on random fiber mats and unfunctionalized matrices. Precise functionalization of degradable polymers with bioactive peptides created translationally-relevant materials that capitalize on the advantages of both synthetic and natural systems, while mitigating the classic limitations of each.
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Diferenciación Celular/efectos de los fármacos , Células Madre Embrionarias/citología , Nanofibras/química , Neuritas/metabolismo , Oligopéptidos/farmacología , Poliésteres/farmacología , Animales , Cromatografía en Gel , Células Madre Embrionarias/efectos de los fármacos , Células Madre Embrionarias/metabolismo , Humanos , Ratones , Nanofibras/ultraestructura , Neuritas/efectos de los fármacos , Poliésteres/químicaRESUMEN
The validity of 99mTc-YIGSR, a novel receptor radio-tracer, in imaging the Ehrlich ascites tumor was evaluated. YIGSR, a pentapeptide of laminin, was labeled with 99mTc by using a bifunctional chelator S-Acetly-NH3-MAG3. The MIBI was labeled with 99mTc by following the kit instruction. The mice of tumor group were intravenously injected 1-2 mCi of 99mTc-YIGSR or 99mTc-MIBI via caudal vein, immobilized and imaged under a Gamma camera. The same procedure was performed in mice of blockade group, in which the unlabeled YIGSR was previously injected to block the receptor-recognition sites, and inflammation group serving as control. The reverse-phase Sep-Pak C18 chromatogram was found to have an essentially complete conjugation between YIGSR and S-Acetly-NH3-MAG3. The conjugated YIGSR could be radio-labeled successfully with 99mTc at room temperature and neutral pH, with a radio-labeling yield of 62%. Without the chelator S-Acetly-NH3-MAG3, the YIGSR was labeled with 99mTc at an efficiency of 4%. The imagological study revealed obvious tumor accumulation of 99mTc-YIGSR 15 min after the injection, and the uptake peaked after 3 h with a tumor-to-muscle ratio (T/M) of 11.36. The radio-tracer was slowly cleared up and resulted in a T/M of 3.01 at the 8th h after the injection. As for blocked group, the tumor uptake of radiotracer was significantly lower, with the highest T/M being 4.61 after 3 h and 0.89 after 8 h. The T/M was 3.72 at the 3rd h and 1.29 at the 8th h after the 99mTc-YIGSR injection in the inflammatory group. The T/M was significantly higher in tumor group than in inflammatory group or control group (P＜0.001). In the 99mTc-MIBI group, the T/M was 1.40 at the 3rd h and 0.55 at the 8th h after the injection, which showed a significant difference as compared with 99mTc-YIGSR (P＜0.001).It is concluded that YIGSR can be successfully radiolabelled by using S-Acetly-NH3-MAG3.99mTc-YIGSR has many advantages in tumor imaging, such as quick and clear visualization, high sensitivity and specificity, and satisfactory target/non-target ratio (N/NT). It promises to be tumor radio-tracer.
RESUMEN
The validity of 99mTc-YIGSR, a novel receptor radio-tracer, in imaging the Ehrlich ascites tumor was evaluated. YIGSR, a pentapeptide of laminin, was labeled with 99mTc by using a bifunctional chelator S-Acetly-NH3-MAG3. The MIBI was labeled with 99mTc by following the kit instruction. The mice of tumor group were intravenously injected 1-2 mCi of 99mTc-YIGSR or 99mTc-MIBI via caudal vein, immobilized and imaged under a Gamma camera. The same procedure was performed in mice of blockade group, in which the unlabeled YIGSR was previously injected to block the receptor-recognition sites, and inflammation group serving as control. The reverse-phase Sep-Pak C18 chromatogram was found to have an essentially complete conjugation between YIGSR and S-Acetly-NH3-MAG3. The conjugated YIGSR could be radio-labeled successfully with 99mTc at room temperature and neutral pH, with a radio-labeling yield of 62%. Without the chelator S-Acetly-NH3-MAG3, the YIGSR was labeled with 99mTc at an efficiency of 4%. The imagological study revealed obvious tumor accumulation of 99mTc-YIGSR 15 min after the injection, and the uptake peaked after 3 h with a tumor-to-muscle ratio (T/M) of 11.36. The radio-tracer was slowly cleared up and resulted in a T/M of 3.01 at the 8th h after the injection. As for blocked group, the tumor uptake of radiotracer was significantly lower, with the highest T/M being 4.61 after 3 h and 0.89 after 8 h. The T/M was 3.72 at the 3rd h and 1.29 at the 8th h after the 99mTc-YIGSR injection in the inflammatory group. The T/M was significantly higher in tumor group than in inflammatory group or control group (P＜0.001). In the 99mTc-MIBI group, the T/M was 1.40 at the 3rd h and 0.55 at the 8th h after the injection, which showed a significant difference as compared with 99mTc-YIGSR (P＜0.001).It is concluded that YIGSR can be successfully radiolabelled by using S-Acetly-NH3-MAG3.99mTc-YIGSR has many advantages in tumor imaging, such as quick and clear visualization, high sensitivity and specificity, and satisfactory target/non-target ratio (N/NT). It promises to be tumor radio-tracer.
RESUMEN
To investigate a new kind of tumor tracer 99mTc-YIGSR developed from a five amino structure (YIGSR) of the Laminin -chain,which can bind to the laminin receptors of tumor specifically, and radiolabeled with MAG3. (1) Preparation of the 99m Tc-YIGSR probe: with S-Acetly-NH3-MAG3as the chelator and with proper reductants YIGSR was labeled with 99mTc; (2) Cell culture and viability measurement: EAC was maintained in RPMI 1640 supplemented with calf serum; the trypan blue exclusion was applied to calculate the cell viability; (3) Study of the cell dynamic: The EACs uptake of 99mTc-YIGSR and99mTc-MIBI was observed at 37 ℃ and 22 ℃, respectively. (1)The labeling efficiencies of 99mTc-YIGSR and99mTc-MIBI were (62±3) % and (96±2) %, respectively; (2) The cell viability was declined with time of incubation; (3) At 37 ℃, the EACS uptake of 99mTc-YIGSR and99mTc-MIBI reached the peak of (43. 16±2.4) % and (24.4±1.8) % at 60min, respectively; and at 22 ℃, the highest uptake was (26.5±2.1) % and (9. 47±1.9) % at 60min, respectively. The in vitro study suggests that 99mTc-YIGSR is superior to 99mTc-MIBI in cell uptake and has potential value in tumor imaging.