RESUMEN

The impact of electrical stimulation has been widely investigated on the wound healing process; however, its practicality is still challenging. This study explores the effect of electrical stimulation on fibroblasts in a culture medium containing different electrically-charged polysaccharide derivatives including alginate, hyaluronate, and chitosan derivatives. For this aim, an electrical stimulation, provided by a zigzag triboelectric nanogenerator (TENG), was exerted on fibroblasts in the presence of polysaccharides' solutions. The analyses showed a significant increase in cell proliferation and an improvement in wound closure (160 % and 90 %, respectively) for the hyaluronate-containing medium by a potential of 3 V after 48 h. In the next step, a photo-crosslinkable hydrogel was prepared based on hyaluronic acid methacrylate (HAMA). Then, the cells were cultured on HAMA hydrogel and treated by an electrical stimulation. Surprisingly, the results showed a remarkable increase in cell growth (280 %) and migration (82 %) after 24 h. Attributed to the electroosmosis phenomenon and an amplified transfer of soluble growth factors, a dramatic promotion was underscored in cell activities. These findings highlight the role of electroosmosis in wound healing, where TENG-based electrical stimulation is combined with bioactive polysaccharide-based hydrogels to promote wound healing.

Asunto(s)

Alginatos , Proliferación Celular , Fibroblastos , Ácido Hialurónico , Hidrogeles , Cicatrización de Heridas , Ácido Hialurónico/química , Ácido Hialurónico/farmacología , Alginatos/química , Proliferación Celular/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Hidrogeles/química , Hidrogeles/farmacología , Cicatrización de Heridas/efectos de los fármacos , Estimulación Eléctrica , Polielectrolitos/química , Animales , Ratones , Quitosano/química , Movimiento Celular/efectos de los fármacos , Humanos , Células 3T3 NIH

RESUMEN

Bats stand out among mammalian species for their exceptional traits, including the capacity to navigate through flight and echolocation, conserve energy through torpor/hibernation, harbor a multitude of viruses, exhibit resistance to disease, survive harsh environmental conditions, and demonstrate exceptional longevity compared to other mammals of similar size. In vivo studies of bats are challenging for several reasons, such as difficulty in locating and capturing them in their natural environments, limited accessibility, low sample size, environmental variation, long lifespans, slow reproductive rates, zoonotic disease risks, species protection, and ethical concerns. Thus, establishing alternative laboratory models is crucial for investigating the diverse physiological adaptations observed in bats. Obtaining quality cells from tissues is a critical first step for successful primary cell derivation. However, it is often impractical to collect fresh tissue and process the samples immediately for cell culture due to the resources required for isolating and expanding cells. As a result, frozen tissue is typically the starting resource for bat primary cell derivation, but cells in frozen tissue are usually damaged and have low integrity and viability. Isolating primary cells from frozen tissues thus poses a significant challenge. Herein, we present a successfully developed protocol for isolating primary dermal fibroblasts from frozen bat wing biopsies. This protocol marks a significant milestone, as this is the first protocol specifically focused on fibroblast isolation from bat frozen tissue. We also describe methods for primary cell characterization, genetic manipulation of primary cells through lentivirus transduction, and the development of stable cell lines. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Bat wing biopsy collection and preservation Support Protocol 1: Blood collection from bat venipuncture Basic Protocol 2: Isolation of primary fibroblasts from adult bat frozen wing biopsy Support Protocol 2: Primary fibroblast culture and subculture Support Protocol 3: Determination of growth curve and doubling time Support Protocol 4: Cell banking and thawing of primary fibroblasts Basic Protocol 3: Lentiviral transduction of bat primary fibroblasts Basic Protocol 4: Bat stable fibroblast cell line development Support Protocol 5: Bat fibroblast validation by immunofluorescence staining Basic Protocol 5: Chromosome counting.

Asunto(s)

Quirópteros , Criopreservación , Fibroblastos , Alas de Animales , Animales , Quirópteros/virología , Fibroblastos/citología , Criopreservación/métodos , Línea Celular , Biopsia/métodos , Técnicas de Cultivo de Célula/métodos , Congelación

RESUMEN

Skin identity is controlled by intrinsic features of the epidermis and dermis and their interactions. Modifying skin identity has clinical potential, such as the conversion of residual limb and stump (nonvolar) skin of amputees to pressure-responsive palmoplantar (volar) skin to enhance prosthesis use and minimize skin breakdown. Greater keratin 9 (KRT9) expression, higher epidermal thickness, keratinocyte cytoplasmic size, collagen length, and elastin are markers of volar skin and likely contribute to volar skin resiliency. Given fibroblasts' capacity to modify keratinocyte differentiation, we hypothesized that volar fibroblasts influence these features. Bioprinted skin constructs confirmed the capacity of volar fibroblasts to induce volar keratinocyte features. A clinical trial of healthy volunteers demonstrated that injecting volar fibroblasts into nonvolar skin increased volar features that lasted up to 5 months, highlighting a potential cellular therapy.

Asunto(s)

Refuerzo Biomédico , Bioimpresión , Dermis , Epidermis , Fibroblastos , Queratinocitos , Adulto , Femenino , Humanos , Masculino , Amputados , Diferenciación Celular , Colágeno/metabolismo , Dermis/citología , Dermis/metabolismo , Elastina/metabolismo , Epidermis/metabolismo , Fibroblastos/citología , Fibroblastos/trasplante , Mano , Queratina-9/metabolismo , Queratinocitos/citología , Queratinocitos/metabolismo , Refuerzo Biomédico/métodos

RESUMEN

Cell identities are defined by intrinsic transcriptional networks and spatio-temporal environmental factors. Here, we explored multiple factors that contribute to the identity of adipose stem cells, including anatomic location, microvascular neighborhood, and sex. Our data suggest that adipose stem cells serve a dual role as adipocyte precursors and fibroblast-like cells that shape the adipose tissue's extracellular matrix in an organotypic manner. We further find that adipose stem cells display sexual dimorphism regarding genes involved in estrogen signaling, homeobox transcription factor expression and the renin-angiotensin-aldosterone system. These differences could be attributed to sex hormone effects, developmental origin, or both. Finally, our data demonstrate that adipose stem cells are distinct from mural cells, and that the state of commitment to adipogenic differentiation is linked to their anatomic position in the microvascular niche. Our work supports the importance of sex and microvascular function in adipose tissue physiology.

Asunto(s)

Adipocitos , Tejido Adiposo , Fibroblastos , Caracteres Sexuales , Células Madre , Animales , Femenino , Adipocitos/citología , Adipocitos/metabolismo , Masculino , Tejido Adiposo/citología , Tejido Adiposo/metabolismo , Fibroblastos/metabolismo , Fibroblastos/citología , Células Madre/metabolismo , Células Madre/citología , Ratones , Diferenciación Celular , Adipogénesis/genética , Ratones Endogámicos C57BL , Matriz Extracelular/metabolismo , Humanos

RESUMEN

Absolute quantity imaging of biomolecules on a single cell level is critical for measurement assurance in biosciences and bioindustries. While infrared (IR) transmission microscopy is a powerful label-free imaging modality capable of chemical quantification, its applicability to hydrated biological samples remains challenging due to the strong IR absorption by water. Traditional IR imaging of hydrated cells relies on powerful light sources, such as synchrotrons, to mitigate the light absorption by water. However, we overcome this challenge by applying a solvent absorption compensation (SAC) technique to a home-built benchtop IR microscope based on an external-cavity quantum cascade laser. SAC-IR microscopy adjusts the incident light using a pair of polarizers to precompensate the IR absorption by water while retaining the full dynamic range. Integrating the IR absorbance over a cell yields the total mass of biomolecules per cell. We monitor the total mass of the biomolecules of live fibroblast cells over 12 h, demonstrating promise for advancing our understanding of the biomolecular processes occurring in live cells on the single-cell level.

Asunto(s)

Análisis de la Célula Individual , Animales , Ratones , Fibroblastos/citología , Fibroblastos/química , Espectrofotometría Infrarroja/métodos , Microscopía/métodos , Rayos Infrarrojos , Células 3T3 NIH

RESUMEN

Platelet-rich plasma (PRP) has become an important regenerative therapy. However, the preparation method of PRP has not been standardized, and the optimal platelet concentration for PRP used in skin wound repair is unclear, leading to inconsistent clinical efficacy of PRP. Therefore, the development of standardized preparation methods for PRP and the investigation of the dose-response relationship between PRP with different platelet concentrations and tissue regeneration plays an important role in the development and clinical application of PRP technology. This study has developed an integrated blood collection device from blood drawing to centrifugation. Response surface methodology was employed to optimize the preparation conditions, ultimately achieving a platelet recovery rate as high as 95.74% for PRP (with optimal parameters: centrifugation force 1730× g, centrifugation time 10 min, and serum separation gel dosage 1.4 g). Both in vitro and in vivo experimental results indicate that PRP with a (2×) enrichment ratio is the most effective in promoting fibroblast proliferation and skin wound healing, with a cell proliferation rate of over 150% and a wound healing rate of 78% on day 7.

Asunto(s)

Proliferación Celular , Plasma Rico en Plaquetas , Piel , Cicatrización de Heridas , Plasma Rico en Plaquetas/metabolismo , Plasma Rico en Plaquetas/química , Animales , Piel/lesiones , Piel/metabolismo , Humanos , Fibroblastos/citología , Ratones , Masculino , Plaquetas/metabolismo

RESUMEN

Yak is an excellent germplasm resource on the Tibetan Plateau and is able to live in high-altitude areas with hypoxic, cold, and harsh environments. Studies on induced pluripotent stem cells (iPSCs) in large ruminants commonly involve a combination strategy involving six transcription factors, Oct4, Sox2, Klf4, c-Myc, Nanog, and Lin28 (OSKMNL). This strategy tends to utilize genes from the same species to optimize pluripotency maintenance. In this study, we cloned the six pluripotency genes (OSKMNL) from yak and constructed a multi-cistronic lentiviral vector carrying these genes. This vector efficiently delivered the genes into yak fibroblasts, aiming to promote the reprogramming process. We verified that the treated cells had several pluripotency characteristics, marking the first successful construction of a lentiviral system carrying yak pluripotency genes. This achievement lays the foundation for subsequent establishment of yak iPSCs and holds significant implications for yak-breed improvement and germplasm-resource conservation.

Asunto(s)

Vectores Genéticos , Células Madre Pluripotentes Inducidas , Factor 4 Similar a Kruppel , Lentivirus , Lentivirus/genética , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/citología , Bovinos , Animales , Vectores Genéticos/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Reprogramación Celular/genética , Fibroblastos/metabolismo , Fibroblastos/citología

RESUMEN

BACKGROUND: Macrophage-based cell therapy is promising in solid tumors, but the efficient acquisition of macrophages remains a challenge. Induced pluripotent stem cell (iPSC)-induced macrophages are a valuable source, but time-consuming and costly. The application of reprogramming technologies allows for the generation of macrophages from somatic cells, thereby facilitating the advancement of cell-based therapies for numerous malignant diseases. METHODS: The composition of CD45+ myeloid-like cell complex (MCC) and induced macrophage (iMac) were analyzed by flow cytometry and single-cell RNA sequencing. The engraftment capacity of CD45+ MCC was evaluated by two transplantation assays. Regulation of c-Myc on MafB was evaluated by ChIP-qPCR and promoter reporter and dual luciferase assays. The phenotype and phagocytosis of iMac were explored by flow cytometry and immunofluorescence. Leukemia, breast cancer, and patient-derived tumor xenograft models were used to explore the anti-tumor function of iMac. RESULTS: Here we report on the establishment of a novel methodology allowing for reprogramming fibroblasts into functional macrophages with phagocytic activity by c-Myc overexpression. Fibroblasts with ectopic expression of c-Myc in iPSC medium rapidly generated CD45+ MCC intermediates with engraftment capacity as well as the repopulation of distinct hematopoietic compartments. MCC intermediates were stably maintained in iPSC medium and continuously generated functional and highly pure iMac just by M-CSF cytokine stimulation. Single-cell transcriptomic analysis of MCC intermediates revealed that c-Myc up-regulated the expression of MafB, a major regulator of macrophage differentiation, to promote macrophage differentiation. Characterization of the iMac activity showed NF-κB signaling activation and a pro-inflammatory phenotype. iMac cells displayed significantly increased in vivo persistence and inhibition of tumor progression in leukemia, breast cancer, and patient-derived tumor xenograft models. CONCLUSIONS: Our findings demonstrate that c-Myc alone is enough to reprogram fibroblasts into functional macrophages, supporting that c-Myc reprogramming strategy of fibroblasts can help circumvent long-standing obstacles to gaining "off-the-shelf" macrophages for anti-cancer immunotherapy.

Asunto(s)

Reprogramación Celular , Fibroblastos , Macrófagos , Proteínas Proto-Oncogénicas c-myc , Macrófagos/metabolismo , Macrófagos/citología , Animales , Humanos , Ratones , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Fibroblastos/metabolismo , Fibroblastos/citología , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Femenino

RESUMEN

This study presents an innovative bi-layered three-dimensional skin-like nanopad (SLN) engineered for skin tissue regeneration. The SLN integrates a mechanically supportive polycaprolactone nanofibrous layer with a functional chitosan hydrogel film, mimicking natural skin. Our SLN exhibits superior flexibility, with a maximum elongation of 751.71 ± 125 % and exceptional porosity of 95 ± 4.5 %, ensuring effective exudate management due to its high water uptake capacity (4393 ± 72 %). FTIR analysis confirmed a distinctive fiber-hydrogel network within the SLN, which serves as a barrier against Staphylococcus aureus and Pseudomonas aeruginosa infiltration. In vitro cell viability assays with the human fibroblast have consistently demonstrated that 3D bi-layered SLN enhances fibroblast attachment, infiltration, and proliferation by 150 ± 20 %. In vivo studies in a rat model demonstrated significantly faster wound closure, with 60 % on day 7 and 87 % on day 10, compared to the 30 % and 60 % in controls, highlighting the efficacy of SLN. By mimicking the architecture of native skin, this biomimetic bi-layered SLN scaffold provides flexibility and support while accelerating in vivo wound closure by promoting fibroblast proliferation and infiltration. Customizable in size, depth, and shape, the engineered SLN has emerged as a promising platform for advanced wound care and tissue engineering.

Asunto(s)

Quitosano , Fibroblastos , Nanofibras , Ingeniería de Tejidos , Cicatrización de Heridas , Quitosano/química , Nanofibras/química , Cicatrización de Heridas/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Animales , Ingeniería de Tejidos/métodos , Ratas , Humanos , Andamios del Tejido/química , Piel , Proliferación Celular/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Hidrogeles/química , Hidrogeles/farmacología , Regeneración/efectos de los fármacos , Pseudomonas aeruginosa/efectos de los fármacos

RESUMEN

Pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic cancer, a leading cause of cancer-related deaths globally. Initial lesions of PDAC develop within the exocrine pancreas' functional units, with tumor progression driven by interactions between PDAC and stromal cells. Effective therapies require anatomically and functionally relevantin vitrohuman models of the pancreatic cancer microenvironment. We employed tomographic volumetric bioprinting, a novel biofabrication method, to create human fibroblast-laden constructs mimicking the tubuloacinar structures of the exocrine pancreas. Human pancreatic ductal epithelial (HPDE) cells overexpressing the KRAS oncogene (HPDE-KRAS) were seeded in the multiacinar cavity to replicate pathological tissue. HPDE cell growth and organization within the structure were assessed, demonstrating the formation of a thin epithelium covering the acini inner surfaces. Immunofluorescence assays showed significantly higher alpha smooth muscle actin (α-SMA) vs. F-actin expression in fibroblasts co-cultured with cancerous versus wild-type HPDE cells. Additionally,α-SMA expression increased over time and was higher in fibroblasts closer to HPDE cells. Elevated interleukin (IL)-6 levels were quantified in supernatants from co-cultures of stromal and HPDE-KRAS cells. These findings align with inflamed tumor-associated myofibroblast behavior, serving as relevant biomarkers to monitor early disease progression and target drug efficacy. To our knowledge, this is the first demonstration of a 3D bioprinted model of exocrine pancreas that recapitulates its true 3-dimensional microanatomy and shows tumor triggered inflammation.

Asunto(s)

Bioimpresión , Fibroblastos , Páncreas Exocrino , Humanos , Páncreas Exocrino/metabolismo , Fibroblastos/metabolismo , Fibroblastos/citología , Impresión Tridimensional , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/metabolismo , Carcinoma Ductal Pancreático/patología , Carcinoma Ductal Pancreático/metabolismo , Línea Celular Tumoral , Tomografía , Actinas/metabolismo , Interleucina-6/metabolismo , Ingeniería de Tejidos , Técnicas de Cocultivo , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética

RESUMEN

In recent years, research on organ-on-a-chip technology has been flourishing, particularly for drug screening and disease model development. Fibroblasts and vascular endothelial cells engage in crosstalk through paracrine signaling and direct cell-cell contact, which is essential for the normal development and function of the heart. Therefore, to faithfully recapitulate cardiac function, it is imperative to incorporate fibroblasts and vascular endothelial cells into a heart-on-a-chip model. Here, we report the development of a human heart-on-a-chip composed of induced pluripotent stem cell (iPSC)-derived cardiomyocytes, fibroblasts, and vascular endothelial cells. Vascular endothelial cells cultured on microfluidic channels responded to the flow of culture medium mimicking blood flow by orienting themselves parallel to the flow direction, akin to in vivo vascular alignment in response to blood flow. Furthermore, the flow of culture medium promoted integrity among vascular endothelial cells, as evidenced by CD31 staining and lower apparent permeability. The tri-culture condition of iPSC-derived cardiomyocytes, fibroblasts, and vascular endothelial cells resulted in higher expression of the ventricular cardiomyocyte marker IRX4 and increased contractility compared to the bi-culture condition with iPSC-derived cardiomyocytes and fibroblasts alone. Such tri-culture-derived cardiac tissues exhibited cardiac responses similar to in vivo hearts, including an increase in heart rate upon noradrenaline administration. In summary, we have achieved the development of a heart-on-a-chip composed of cardiomyocytes, fibroblasts, and vascular endothelial cells that mimics in vivo cardiac behavior.

Asunto(s)

Células Endoteliales , Fibroblastos , Células Madre Pluripotentes Inducidas , Dispositivos Laboratorio en un Chip , Miocitos Cardíacos , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/fisiología , Fibroblastos/citología , Fibroblastos/metabolismo , Células Endoteliales/citología , Células Endoteliales/metabolismo , Células Endoteliales/fisiología , Diferenciación Celular , Células Cultivadas , Técnicas de Cocultivo/métodos , Sistemas Microfisiológicos

RESUMEN

OBJECTIVES: To establish a methodological system for reprogramming rat embryonic fibroblasts (REF) into chemically induced neurons (ciNCs) via small molecule compounds to provide safe and effective donor cells for treatment of neurodegenerative diseases. METHODS: Based on the method established by PEI Gang's research group to directly reprogram human fibroblasts into neurons, the induction medium and maturation medium was optimized by replacing the coating solution, mitigating oxidative stress injury, adding neurogenic protective factors, adjusting the concentration of trichothecenes, performing small-molecule removal experiments, and carrying out immunofluorescence and Western blotting on cells at different stages of induction to validate the effect of induction. RESULTS: When the original protocol was used for induction, the cell survival rate was (34.24±2.77)%. After replacing the coating solution gelatin with matrigel, the cell survival rate increased to (45.41±4.27)%; after adding melatonin, the cell survival rate increased to (67.95±5.61)% and (23.43±1.42)% were transformed into neural-like cells; after adding the small molecule P7C3-A20, the cell survival rate was further increased to (76.27±1.41)%, and (39.72±4.75)% of the cells were transformed into neural-like cells. When the concentration of trichothecene was increased to 30 µmol/L, the proportion of neural-like cells reached (55.79±1.90)%; after the removal of SP600125, (86.96±2.15)% of the cells survived, and the rate of neural-like cell production increased to (63.43±1.60)%. With the optimized protocol, REF could be successfully induced into ciNC through the neural precursor cell stage, in which the neural precursor cells were able to highly express the neural precursor cell markers SRY-related HMG-box gene 2 (Sox2) and paired box 6 (Pax6) as well as neuron-specific marker tubulin 1 (Tuj1), while the expression of fiber-associated protein vimentin was reduced. After two weeks of induction of neural precursor cells in a maturation medium, most cells displayed neuronal-like cell morphology. The induced ciNCs were able to highly express the mature neuronal surface markers Tuj1 and microtubule-associated protein 2 (MAP2), while the expression of vimentin was reduced. CONCLUSIONS: The small molecule combinations optimized in this study can reprogram REF to ciNCs under normoxic conditions.

Asunto(s)

Reprogramación Celular , Fibroblastos , Neuronas , Animales , Ratas , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Neuronas/citología , Neuronas/efectos de los fármacos , Reprogramación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Diferenciación Celular/efectos de los fármacos

RESUMEN

Cellular senescence is a cellular state characterized by irreversible growth arrest, resistance to apoptosis and secretion of inflammatory molecules, which is causally linked to the pathogenesis of many age-related diseases. Besides, there is accumulating evidence that selective removal of senescent cells can benefit therapies for cancer and fibrosis by modulating the inflammatory microenvironment. While the field of so-called senolytics has spawned promising small molecules and peptides for the selective removal of senescent cells, there is still no effective means to detect senescent cells in vivo, a prerequisite for understanding the role of senescence in pathophysiology and to assess the effectiveness of treatments aimed at removing senescent cells. Here, we present a strategy based on an mRNA logic circuit, that yields mRNA-dependent protein expression only when a senescence-specific miRNA signature is present. Following a validation of radiation-induced senescence induction in primary human fibroblasts, we identify miRNAs up- and downregulated in association with cellular senescence using RT-qPCR. Incorporating binding sites to these miRNAs into the 3' untranslated regions of the mRNA logic circuit, we demonstrate the senescence-specific expression of EGFP for detection of senescent cells and of a constitutively active caspase-3 for selective removal. Altogether, our results pave the way for a novel approach to execute an mRNA-based programme specifically in senescent cells aimed at their detection or selective removal.

Asunto(s)

Senescencia Celular , MicroARNs , ARN Mensajero , Humanos , Senescencia Celular/genética , MicroARNs/genética , MicroARNs/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Fibroblastos/metabolismo , Fibroblastos/citología , Caspasa 3/metabolismo , Caspasa 3/genética , Regiones no Traducidas 3'/genética , Regulación de la Expresión Génica

RESUMEN

Additive manufacturing can develop regenerative scaffolds for wound healing. 3D printing offers meticulous porosity, mechanical integrity, cell adhesion and cost-effectiveness. Herein, we prepared ink composed of carboxymethyl cellulose (CMC), polyvinylpyrrolidone (PVP), collagen, and oregano extract for the fabrication of tissue constructs. The blend was optimized to form a homogeneous ink and rheological characterization demonstrated shear thinning behavior. The scaffolds were printed using Direct Ink Write (DIW) at a flow speed of 4 mm3/s and a layer height of 0.18 mm. The fabricated scaffolds demonstrated an ultimate tensile strength (UTS) and toughness of 730 KPa and 2.72 MJ/m3, respectively. Scanning Electron Microscopy (SEM) revealed an average pore size of 300 ± 30 µm. Fourier transform infrared spectroscopy (FTIR) analysis confirmed that all materials were present. The contact angle of the composite scaffold was 68° ± 1°. Moreover, the scaffolds presented 82 % mass loss (degradation) in phosphate buffer saline (PBS) over 14 days. The composite scaffold exhibited inhibition zones of 9 mm and 12 mm against Staphylococcus aureus and Escherichia coli, respectively. The PVP/CMC/collagen/oregano 3D printed scaffolds exhibited excellent biocompatibility with the mesenchymal stem cells and humman dermal fibroblast cells, confirmed by water-soluble tetrazolium - 8 (WST-8) assay (test conducted for 7 days). The enhanced angiogenic potential of said scaffold was assesed by release of vascular endothelial growth factor followed by further validation through in-vivo CAM assay. Thus, confirming suitability for the potential wound healing application.

Asunto(s)

Carboximetilcelulosa de Sodio , Colágeno , Origanum , Povidona , Andamios del Tejido , Cicatrización de Heridas , Povidona/química , Cicatrización de Heridas/efectos de los fármacos , Carboximetilcelulosa de Sodio/química , Carboximetilcelulosa de Sodio/farmacología , Andamios del Tejido/química , Colágeno/química , Colágeno/farmacología , Humanos , Origanum/química , Staphylococcus aureus/efectos de los fármacos , Impresión Tridimensional , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Escherichia coli/efectos de los fármacos , Tinta , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/citología , Antibacterianos/farmacología , Antibacterianos/química , Porosidad , Resistencia a la Tracción , Animales

RESUMEN

The multibiocomponent hybrid alginate hydrogels based on brown and sea algae, containing 100 % ingredients of natural origin were prepared by ionic crosslinking reaction of a polymeric matrix with lipid nanodispersion. To the best of the Authors' knowledge such multicomponent biobased hydrogel of promising medical and cosmetical applications for the first time was obtained in the environment of flower water, received earlier as a waste by-product from various chemical processes. An innovative hybrid alginate hydrogel that is completely biodegradable and eco-friendly was obtained following waterless and upcycling trends that are in line with the principles of sustainable development. The optimal composition of the lipid nanodispersion and the polymeric matrix was selected using the statistical method of design of the experiment. Based on obtained results, multibiocomponent hybrid alginate hydrogels with various ratios of lipid nanodispersion were obtained. Subsequently, the porous structure and elasticity of the hybrid hydrogels were analyzed. Moreover, to confirm the safety of the multibiocomponent alginate hybrid hydrogels the cytotoxic tests were carried out using human fibroblasts and keratinocytes cell lines. As the final product hybrid of hydrolate-swollen alginate hydrogel and lipid nanodispersion containing several active ingredients (silymarin, bakuchiol, spirulina) was obtained.

Asunto(s)

Alginatos , Materiales Biocompatibles , Hidrogeles , Lípidos , Hidrogeles/química , Alginatos/química , Humanos , Lípidos/química , Materiales Biocompatibles/química , Cosméticos/química , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Queratinocitos/efectos de los fármacos , Queratinocitos/citología , Línea Celular

RESUMEN

Androgenic alopecia (AGA) typically manifests post-puberty, resulting in decreases in hair density, disruptions in the hair growth cycle, and alterations in hair follicle micro structure. Dihydrotestosterone (DHT) is a key hormone implicated in hair loss, especially on male. In this study, we found that each of arginine (Arg), arterial extract (AE) or biotin tripeptide-1 (BT-1), when combined with low level light therapy (LLLT, at 630 nm, 2 J/cm2), showed the efficacy in enhancing mitochondrial functions, cell proliferation and collagen synthesis in fibroblasts. Additionally, CARRIPOWER (the complexes of AE, BT-1, Arg, and Diaminopyrimidine derivatives), in conjunction with LLLT (630 nm, 2 J/cm2), showed promising results in dermal papilla cells (DPCs). The promising results contained not also inflammatory cytokines (IL-1ß and IL-6) and cell pro apoptotic factor (TGF-ß2) reduction, but also Wnt pathway inhibition by decreasing DKK1 level, and pro-hair growth factors (vascular endothelial growth factor (VEGF) and ß-catenin) increase. This innovative combination therapy offers a potential solution for the treatment of AGA, addressing both hormonal and cellular factors involved in hair loss.

Asunto(s)

Proliferación Celular , Fibroblastos , Cabello , beta Catenina , Humanos , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Fibroblastos/metabolismo , Fibroblastos/citología , Fibroblastos/efectos de la radiación , Fibroblastos/efectos de los fármacos , beta Catenina/metabolismo , Cabello/efectos de la radiación , Cabello/crecimiento & desarrollo , Cabello/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/metabolismo , Arginina/química , Arginina/farmacología , Alopecia/terapia , Folículo Piloso/efectos de la radiación , Folículo Piloso/metabolismo , Folículo Piloso/efectos de los fármacos , Terapia por Luz de Baja Intensidad , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Oligopéptidos/química , Oligopéptidos/farmacología , Masculino , Colágeno/metabolismo , Colágeno/química , Vía de Señalización Wnt/efectos de los fármacos , Vía de Señalización Wnt/efectos de la radiación , Línea Celular , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacos , Mitocondrias/efectos de la radiación

RESUMEN

Scaffolds with multiphasic structures are considered to be superior for guided tissue regeneration. Two types of tilapia skin collagen gradient membranes (stepped gradient and linear gradient) with multiphasic structures were prepared by controlling the collagen concentrations and the freezing rates. The results revealed that collagen gradient membranes were more capable of guiding tissue regeneration compared to homogeneous membranes. These two gradient membranes featured a dense outer layer and a loose inner layer, with good mechanical properties as indicated by tensile strengths of more than 250 Kpa and porosities exceeding 85 %. Additionally, these membranes also showed good hydrophilicity and water absorption, with an inner layer contact angle of less than 91° and a water absorption ratio greater than 40 times. Furthermore, the multiphasic scaffolds were proved to be biocompatible by the acute toxicity assay, the intradermal irritation test and so on. Gradient membranes could effectively promote the adhesion and proliferation of fibroblasts and osteoblasts, through elevating the TGF-ß/Smad signaling pathway by TGF-ß and Smads, and activating the Wnt/ß-catenin signaling pathway by LRP5 and ß-catenin, similar to homogenous membranes. Therefore, collagen gradient membranes from tilapia skin show important application value in guiding tissue regeneration.

Asunto(s)

Materiales Biocompatibles , Colágeno , Animales , Colágeno/química , Colágeno/metabolismo , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Andamios del Tejido/química , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/citología , Proliferación Celular/efectos de los fármacos , Membranas Artificiales , Tilapia/metabolismo , Osteoblastos/efectos de los fármacos , Osteoblastos/citología , Osteoblastos/metabolismo , Ratones , Ensayo de Materiales , Piel/metabolismo , Adhesión Celular/efectos de los fármacos , Resistencia a la Tracción

Asunto(s)

Hematopoyesis Clonal , Fibroblastos , Humanos , Hematopoyesis Clonal/genética , Fibroblastos/metabolismo , Fibroblastos/citología , Metaanálisis como Asunto , Mutación , Proliferación Celular

RESUMEN

Bioprinted hydrogels are extensively studied to provide an artificial matrix for 3D cell culture. The success of bioprinting hydrogels relies on fine-tuning their rheology and composition to achieve shear-thinning behavior. However, a challenge arises from the limited viscoelastic and stiffness range accessible from a single hydrogel formulation. Nevertheless, hydrogel mechanical properties are recognized as essential cues influencing cell phenotype, migration, and differentiation. Thus, it is crucial to develop a system to easily modulate bioprinted hydrogels' mechanical behaviors. In this work, we modulated the viscoelastic properties and stiffness of bioprinted hydrogels composed of fibrinogen, alginate, and gelatin by tuning the crosslinking bath solution. Various concentrations of calcium ionically crosslinked alginate, while transglutaminase crosslinked gelatin. Subsequently, we characterized the mechanical behavior of our bioprinted hydrogels from the nanoscale to the macroscale. This approach enabled the production of diverse bioprinted constructs, either with similar elastic behavior but different elastic moduli or with similar elastic moduli but different viscoelastic behavior from the same hydrogel formulation. Culturing fibroblasts in the hydrogels for 33 days revealed a preference for cell growth and matrix secretion in the viscoelastic hydrogels. This work demonstrates the suitability of the method to decouple the effects of material mechanical from biochemical composition cues on 3D cultured cells.

Asunto(s)

Bioimpresión , Elasticidad , Hidrogeles , Hidrogeles/química , Viscosidad , Animales , Alginatos/química , Ratones , Técnicas de Cultivo Tridimensional de Células , Técnicas de Cultivo de Célula , Fibroblastos/citología , Fibroblastos/efectos de los fármacos

RESUMEN

Injectable cell therapies offer several advantages compared with traditional open surgery, including less trauma to the patient, shorter recovery time, and lower risk of infection. However, a significant problem is the difficulty in developing effective cell delivery carriers that are cyto-compatible and maintain cell viability both during and after injection. In the presented study, it was aimed to develop poly(butylene adipate-co-terephthalate) (PBAT) microcarriers using the emulsion preparation-solvent evaporation technique. The optimized diameter of the PBAT microcarriers was determined as 104 ± 15 µm at 700 rpm and there would be no blockage after injection due to the nonswelling feature of microcarriers. Furthermore, the cellular activities of PBAT microcarriers were evaluated in static culture for 7 days using L929 mouse fibroblasts, MC3T3-E1 mouse pre-osteoblasts, and rat adipose-derived mesenchymal cells (AdMSCs). 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide results and Sscanning electron microscope images showed that PBAT microcarriers increased the adhesion and proliferation properties of pre-osteoblasts and stem cells, while L929 fibroblasts formed aggregates by adhering to certain regions of the microcarrier surface and did not spread on the surface. These results emphasize that PBAT microcarriers can be used as injectable carriers, especially in stem cell therapies, but their surface properties need to be modified for some cells.

Asunto(s)

Poliésteres , Animales , Ratones , Poliésteres/química , Ratas , Fibroblastos/metabolismo , Fibroblastos/citología , Línea Celular , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Osteoblastos/metabolismo , Osteoblastos/citología , Propiedades de Superficie , Proliferación Celular/efectos de los fármacos , Células 3T3 , Técnicas de Cultivo de Célula , Adhesión Celular/efectos de los fármacos