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Background: Modulation of macrophage polarization is required for effective tissue repair and regenerative therapies. Therapeutic modulation of macrophages from an inflammatory M1 to a fibrotic M2 phenotype could help in diseases, such as chronic wounds, which are stalled in a prolonged and heightened inflammatory stage within the wound healing process., Objective: This study evaluates the efficiency of a pullulan/gelatin nanofiber scaffold loaded with retinoic acid (RA) and adipose-derived mesenchymal stem cells (ASCs) to modulate M1 to M2 anti-inflammatory transition., Methods: Scaffolds were fabricated by electrospinning, and crosslinked using ethylene glycol diglycidyl ether (EGDE). Exposure of RA and/or ASCs to cultured macrophages have been shown to promote M1 to M2 transition. Pullulan was chosen as a scaffold material due to its ability to quench reactive oxygen species, key signaling molecules that play an important role in the progression of inflammation, as well as for its excellent mechanical properties. Gelatin was chosen as an additional scaffold component due to the presence of cell-binding motifs and its biocompatibility. Scaffold compositions examined were 75:25 and 50:50, pullulan:gelatin. The scaffolds were crosslinked in 1:70 and 1:50 EGDE:EtOH. The scaffold composition was determined via FTIR. For the present study, the 75:25 pullulan:gelatin crosslinked with 1:70 EGDE:EtOH, forming nanofibers 328 ± 47.9 nm (mean ± SD) in diameter, was chosen as the scaffold composition due to its lower degradation and release rate, which allows a sustained delivery of RA., Results: The scaffold composition degraded to approximately 80% after 14 days, with approximately 38% of the drug released after 7 days. THP-1 monocytic cells were induced into a M1 macrophage phenotype through stimulation with lipopolysaccharide (LPS) and gamma interferon (IFN-γ). These M1 macrophages were the exposed to scaffolds loaded with RA and ASCs, to induce differentiation to an M2 phenotype., Conclusion: Gene expression quantitation by qPCR showed a reduction of M1 biomarkers, tumor necrosis factor alpha (TNFα) and interleukin 1β (IL1β), and an increase of M2 biomarker CCL22 after 2 days of exposure, suggesting successful M1 to M2 transition.

An important challenge in the fabrication of tissue engineered constructs for regenerative medical applications is the development of processes capable of delivering cells and biomaterials to specific locations in a consistent manner. Electrospraying live cells has been introduced in recent years as a cell seeding method, but its effect on phenotype nor genotype has not been explored. A promising candidate for the cellular component of these constructs are human adipose-derived stem cells (hASCs), which are multipotent stem cells that can be differentiated into fat, bone, and cartilage cells. They can be easily and safely obtained from adipose tissue, regardless of the age and sex of the donor. Moreover, these cells can be maintained and expanded in culture for long periods of time without losing their differentiation capacity. In this study, hASCs directly incorporated into a polymer solution were electrosprayed, inducing differentiation into chondrocytes, without the addition of any exogenous factors. Multiple studies have demonstrated the effects of exposing hASCs to biomolecules-such as soluble growth factors, chemokines, and morphogens-to induce chondrogenesis. Transforming growth factors (e.g., TGF-β) and bone morphogenetic proteins are particularly known to play essential roles in the induction of chondrogenesis. Although growth factors have great therapeutic potential for cell-based cartilage regeneration, these growth factor-based therapies have presented several clinical complications, including high dose requirements, low half-life, protein instability, higher costs, and adverse effects in vivo. The present data suggests that electrospraying has great potential as hASCs-based therapy for cartilage regeneration., (© 2021. The Author(s).)

Alternative bone regeneration strategies that do not rely on harvested tissue or exogenous growth factors are needed. One of the major challenges in tissue reconstruction is recreating the bone tissue microenvironment using the appropriate combination of cells, scaffold, and stimulation to direct differentiation. This study presents a bone regeneration formulation that involves the use of human adipose-derived mesenchymal stem cells (hASCs) and a three-dimensional (3D) hydrogel scaffold based on self-assembled RADA16 peptides containing superparamagnetic iron oxide nanoparticles (NPs). Although superparamagnetic NPs could be used as stimulus to manipulate the cell proliferation and differentiation, in this paper their use is explored for assisting osteogenic differentiation of hASCs in conjunction with direct stimulation by extremely low-frequency pulsed electromagnetic fields (pEMFs). Cellular morphology, proliferation, and viability, as well as alkaline phosphatase activity, calcium deposition, and osteogenic capacity were monitored for cells cultured up to 21 days in the 3D construct. The results show that the pEMFs and NPs do not have any negative effect on cell viability, but instead distinctly induced early differentiation of hASCs to an osteoblastic phenotype, when compared with cells without biophysical stimulation. This effect is attributed to synergy between the pEMFs and NPs, which may have stimulated mechanotransduction pathways, which, in turn activated biochemical signals between cells to differentiate or proliferate. This approach may offer a safe and effective option for the treatment of non-union bone fractures. Bioelectromagnetics. © 2020 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc., (© 2020 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc.)

Much of the current understanding on molecular and cellular events of adipose developmental biology comes from monolayer cell culture models using preadipocyte cell lines, although in vivo adipose tissue consists of a much more complex three-dimensional microenvironment of diverse cell types, extracellular network, and tissue-specific morphological and functional features. Added to this fact, the preadipocytes, on which the adipogenesis mechanisms are mostly explored, possess some serious limitations (e.g., time of initial subculture and adipogenic differentiation time), which, perhaps, can efficiently be replaced with progenitor cells such as adipose tissue-derived stem cells (ASCs). With the objective of developing a better in vitro model for adipose developmental biology, this project involves gene expression profiling of human ASCs (hASCs) during their differentiation to adipocytes in a 2D versus 3D culture model. This transcriptional-level analysis revealed that gene expression patterns of adipogenesis-induced hASCs in a 3D self-assembled polypeptide hydrogel are relatively different from the 2D monolayered cells on plastic hard substrate. Moreover, analysis of adipogenic lineage progression 9 days after adipogenic induction shows earlier differentiation of hASCs in 2D over their 3D counterparts. However, differentiation in 2D shows some unexpected behavior in terms of gene expression, which does not seem to be related to adipogenic lineage specification. Since hASCs are already being used in clinical trials due to their therapeutic potential, it is important to have a clear understanding of the molecular mechanisms in an in vivo model microenvironment like the one presented here., (© 2020 S. Karger AG, Basel.)

Aim: Chloroquine (Chl) has shown its potential in cancer therapy and graphene oxide (GO) exhibited excellent tumor-targeting ability, biocompatibility and low toxicity. We have endeavored to conjugate Chl to GO sheets and investigated the nonproliferation action on A549 cell lines along with cell signaling pathways., Materials & Methods: Cellular toxicity, autophagic flux modulation and cell death mechanism induced by GO-Chl have been investigated on A549 cell lines., Results & Conclusion: GO-Chl induces accumulation of autophagosomes (monodansylcadaverine staining, green fluorescence protein-tagged LC3 plasmid and transmission electron microscopy observations) in A549 cells through the blockade of autophagic flux that serves as scaffold for necrosome assembling and activates necroptotic cell death. GO-Chl nanoconjugate could be used as an effective cancer therapeutic agent, by targeting the autophagy necroptosis axis.

Heart disease remains one of the leading causes of death in industrialized nations with myocardial infarction (MI) contributing to at least one fifth of the reported deaths. The hypoxic environment eventually leads to cellular death and scar tissue formation. The scar tissue that forms is not mechanically functional and often leads to myocardial remodeling and eventual heart failure. Tissue engineering and regenerative medicine principles provide an alternative approach to restoring myocardial function by designing constructs that will restore the mechanical function of the heart. In this review, we will describe the cellular events that take place after an MI and describe current treatments. We will also describe how biomaterials, alone or in combination with a cellular component, have been used to engineer suitable myocardium replacement constructs and how new advanced culture systems will be required to achieve clinical success., Competing Interests: Statement No competing financial interests exist.

Beta protein 1 (BP 1 ) is a homeobox protein expressed in 80% of breast cancer cells in either estrogen receptor (ER) positive or ER negative breast cancer. However, it is barely detectable in normal breast tissues. In this project we present an electrochemical DNA nanostructured gold biosensor for detection of BP 1 . The gold sensor is first electrochemically nanostructured in 0.5 M sulfuric acid to reach superior conductivity, larger surface area, and higher stability. Nanostructured gold surface was characterized by atomic force microscopy (AFM) and scanning electron microscopy (SEM). The nanostructured gold sensor is then modified with double-stranded (ds) DNA mapping the genomic sequence that contains the binding site for BP 1 . A redox-active probe (methylene blue) was intercalated in dsDNA to monitor the binding event of BP 1 . A linear correlation of the electrochemical response by concentration of BP 1 was obtained (R 2 = 0.998) with a limit of detection of 1.2 nM. This nanostructured gold dsDNA sensor is shown to be sensitive, selective, stable, and reusable allowing for its potential clinical use.

The use of bone grafts is the standard to treat skeletal fractures, or to replace and regenerate lost bone, as demonstrated by the large number of bone graft procedures performed worldwide. The most common of these is the autograft, however, its use can lead to complications such as pain, infection, scarring, blood loss, and donor-site morbidity. The alternative is allografts, but they lack the osteoactive capacity of autografts and carry the risk of carrying infectious agents or immune rejection. Other approaches, such as the bone graft substitutes, have focused on improving the efficacy of bone grafts or other scaffolds by incorporating bone progenitor cells and growth factors to stimulate cells. An ideal bone graft or scaffold should be made of biomaterials that imitate the structure and properties of natural bone ECM, include osteoprogenitor cells and provide all the necessary environmental cues found in natural bone. However, creating living tissue constructs that are structurally, functionally and mechanically comparable to the natural bone has been a challenge so far. This focus of this review is on the evolution of these scaffolds as bone graft substitutes in the process of recreating the bone tissue microenvironment, including biochemical and biophysical cues.

Improvements in osteoconduction of implant biomaterials require focusing on the bone-implant interface, which is a complex multifactorial system. Surface topography of implants plays a crucial role at this interface. Nanostructured surfaces have been shown to promote serum protein adsorption and osteoblast adhesion when compared to micro-structured surfaces for bone-implant materials. We studied the influence of the serum proteins fibronectin and vitronectin on the attachment and proliferation of osteoblasts onto nanostructured titania surfaces. Human fetal osteoblastic cells hFOB 1.19 were used as model osteoblasts and were grown on nanoporous TiO2 templates, using Ti6AI4V and commercially pure Ti substrates as controls. Results show a significant increase in cell proliferation'on nanoporous TiO2 over flat substrates. Initial cell attachment data exhibited a significant effect by either fibronectin or vitronectin on cell adhesion at the surface of any of the tested materials. In addition, the extent of cell adhesion was significantly different between the nanoporous TiO2 and both Ti6AI4V and commercially pure Ti substrates, with the first showing the highest surface coverage. There was no significant difference on osteoblast attachment or proliferation between the presence of fibronectin or vitronectin using any of the material substrates. Taken together, these results suggest that the increase in osteoblast attachment and proliferation shown on the nanoporous TiO2 is due to an increase in the adsorption of fibronectin and vitronectin because of the higher surface area and to an enhanced protein unfolding, which allows access to osteoblast binding motifs within these proteins.

Biosensors have shown great potential for health care and environmental monitoring. The performance of biosensors depends on their components, among which the matrix material, i.e., the layer between the recognition layer of biomolecule and transducer, plays a crucial role in defining the stability, sensitivity and shelf-life of a biosensor. Recently, zinc oxide (ZnO) nanostructures and thin films have attracted much interest as materials for biosensors due to their biocompatibility, chemical stability, high isoelectric point, electrochemical activity, high electron mobility, ease of synthesis by diverse methods and high surface-to-volume ratio. ZnO nanostructures have shown the binding of biomolecules in desired orientations with improved conformation and high biological activity, resulting in enhanced sensing characteristics. Furthermore, compatibility with complementary metal oxide semiconductor technology for constructing integrated circuits makes ZnO nanostructures suitable candidate for future small integrated biosensor devices. This review highlights recent advances in various approaches towards synthesis of ZnO nanostructures and thin films and their applications in biosensor technology., (Copyright © 2012 Elsevier B.V. All rights reserved.)

In the present study, 11-mercaptoundecanoic acid-modified gold nanoparticles (∼7 nm) were conjugated with chloroquine to explore their potential application in cancer therapeutics. The anticancer activity of chloroquine-gold nanoparticle conjugates (GNP-Chl) was demonstrated in MCF-7 breast cancer cells. The MCF-7 cells were treated with different concentrations of GNP-Chl conjugates, and the cell viability was assayed using trypan blue, resulting in an IC(50) value of 30 ± 5 μg/mL. Flow cytometry analysis revealed that the major pathway of cell death was necrosis, which was mediated by autophagy. The drug release kinetics of GNP-Chl conjugates revealed the release of chloroquine at an acidic pH, which was quantitatively estimated using optical absorbance spectroscopy. The nature of stimuli-responsive drug release and the inhibition of cancer cell growth by GNP-Chl conjugates could pave the way for the design of combinatorial therapeutic agents, particularly nanomedicine, for the treatment of cancer., (Copyright © 2012 Elsevier B.V. All rights reserved.)

We report a novel method for high-throughput investigations on cell-material interactions based on metal oxide nanoscaffolds. These scaffolds possess a continuous gradient of various titanium alloys allowing the compositional and morphological variation that could substantially improve the formation of an osseointegrative interface with bone. The model nanoscaffold has been fabricated on commercially pure titanium (cp-Ti) substrate with a compositional gradients of tin (Sn), chromium (Cr), and niobium (Nb) deposited using a combinatorial approach followed by annealing to create native oxide surface. As an invitro test system, the human fetal osteoblastic cell line (hFOB 1.19) has been used. Cell-adhesion of hFOB 1.19 cells and the suitability of these alloys have been evaluated for cell-morphology, cell-number, and protein adsorption. Although, cell-morphology was not affected by surface composition, cell-proliferation rates varied significantly with surface metal oxide composition; with the Sn- and Nb-rich regions showing the highest proliferation rate and the Cr-rich regions presenting the lowest. The results suggest that Sn and Nb rich regions on surface seems to promote hFOB 1.19 cell proliferation and may therefore be considered as implant material candidates that deserve further analysis.