The effect of the shape of gold core-mesoporous silica shell nanoparticles on the cellular behavior and tumor spheroid penetration

Size, surface charge, and shape have a huge influence on the behavior, cellular uptake, and cytotoxic profile of nanoparticles. Herein, gold core and silica shell based nanoparticles (Au–MSSs) with spherical or rod-like shape were produced, in order to disclose the effect of the shape of nanomaterials on the cellular uptake, reactive oxygen species (ROS) production, 3D tumor spheroid penetration and cytotoxicity towards cancer cells. The Au–MSS spheres induced greater reduction of the ROS content of cancer cells and also displayed a more homogeneous distribution and penetration in 3D tumor spheroids. However, the Au–MSS rods show enhanced cellular migration and uptake in 2D cell cultures, which results in a higher drug delivering capacity. Furthermore, the Au–MSS rods displayed an enhanced cellular cytotoxicity upon laser irradiation (808 nm, 1.7 W cm−2, 10 min), where less than 10% of cells remained viable. In addition, both Au–MSSs have the potential to be used as imaging agents, which further expands their applicability as theranostic agents in the biomedical area. In summary, the obtained results show that the shape of Au–MSSs is crucial for their biological performance, which will have a great influence on their therapeutic outcome. Therefore, our findings may provide useful information for the development and design of new drug delivery systems towards a more effective therapy.info:eu-repo/semantics/publishedVersio

Stimuli-responsive mesoporous silica nanoparticles for cancer therapy: A review

The application of nanocarriers as selective drug delivery platforms, as imaging or as diagnostic agents has been evaluated in several studies in the area of biomedicine, namely for cancer therapy. Such systems have the potential to perform a controlled and site-specific delivery of therapeutic agents leading to a reduction of side effects and, ultimately, to an improved therapeutic outcome. Among the different nanocarriers developed so far, mesoporous silica nanoparticles have attracted the attention of the scientific community for being applied as drug delivery systems that are capable of controlling, both in space and time, the drug release. In this review, the modifications performed, so far, on mesoporous silica nanoparticles to imprint them a stimulus responsive behavior (namely, pH, redox potential, adenosine triphosphate, enzyme or temperature) in order to allow their application in cancer therapy are highlighted.info:eu-repo/semantics/publishedVersio

Thermo- and pH-responsive nano-in-micro particles for combinatorial drug delivery to cancer cells

Drug combinatorial therapy has been gaining the scientific community attention as a suitable approach to increase treatments efficacy and promote cancer eradication. In this study, a new pH- and thermo- responsive carrier was developed by combining doxorubicin-loaded gold-core silica shell nanorods with salicylic acid loaded poly (lactic-co-glycolic acid) based microparticles (NIMPS). The obtained results showed that the drugs and nanorods release could be triggered by the near-infrared (NIR) laser irradiation or by the exposition to an acidic environment. The in vitro 2D cell studies showed that the NIMPS are biocompatible and easily uptaken by HeLa cells. In addition, 3D cell culture models revealed that the NIMPS administration, combined with the NIR laser irradiation, was capable of reducing the size of the HeLa spheroids up to 48%. Overall, the attained data support the application of the nano-in-micro spheres as a dual stimuli responsive drug carrier system for the local administration of combined therapies to cervical cancer cells.info:eu-repo/semantics/publishedVersio

Optimization of liquid overlay technique to formulate heterogenic 3D co‐cultures models

Three‐dimensional (3D) cell culture models of solid tumors are currently having a tremendous impact in the in vitro screening of candidate anti‐tumoral therapies. These 3D models provide more reliable results than those provided by standard 2D in vitro cell cultures. However, 3D manufacturing techniques need to be further optimized in order to increase the robustness of these models and provide data that can be properly correlated with the in vivo situation. Therefore, in the present study the parameters used for producing multicellular tumor spheroids (MCTS) by liquid overlay technique (LOT) were optimized in order to produce heterogeneous cellular agglomerates comprised of cancer cells and stromal cells, during long periods. Spheroids were produced under highly controlled conditions, namely: (i) agarose coatings; (ii) horizontal stirring, and (iii) a known initial cell number. The simultaneous optimization of these parameters promoted the assembly of 3D characteristic cellular organization similar to that found in the in vivo solid tumors. Such improvements in the LOT technique promoted the assembly of highly reproducible, individual 3D spheroids, with a low cost of production and that can be used for future in vitro drug screening assays.info:eu-repo/semantics/publishedVersio

Production and characterization of polycaprolactone- hyaluronic acid/chitosan-zein electrospun bilayer nanofibrous membrane for tissue regeneration

A bilayered electrospun membrane was produced in this study, using the electrospinning technique, to be applied as a skin substitute. The upper layer of the membrane was comprised by hyaluronic acid and polycaprolactone in order to provide mechanical support and also to act as a physical barrier against external threats. Chitosan and zein were used to produce the bottom layer that was loaded with salicylic acid, in order to confer anti-inflammatory and antimicrobial activity to this layer. The physicochemical properties of the membranes were determined and the obtained results showed that the produced electrospun membrane display an ideal porosity, appropriate mechanical properties, controlled water loss and a suitable salicylic acid release profile. In addition, membranes did not exhibit any toxic effects for human fibroblast cells, since cells were able to adhere, spread and proliferate. Furthermore, no biofilm formation was noticed on membranes’ surface along the experiments. In conclusion, the gathered data reveal that this electrospun membrane has suitable properties to be used as a wound dressing.info:eu-repo/semantics/publishedVersio

IR780-loaded TPGS-TOS micelles for breast cancer photodynamic therapy

IR780 iodide is a near-infrared (NIR) dye with a huge potential for cancer imaging and phototherapy. However, its biomedical application is strongly impaired by its lipophilic character. Herein, amphiphilic micelles based on d-α-tocopheryl polyethylene glycol succinate (TPGS) and d-α-tocopheryl succinate (TOS), two vitamin E derivatives with intrinsic anticancer activity, are explored to load IR780. IR780-loaded micelles with suitable sizes are obtained by using specific TPGS and TOS weight feed ratios during micelles formulation and these are able to encapsulate IR780 with high efficiency. In in vitro assays, the IR780-loaded micelles induce a cytotoxic effect in cancer cells upon exposure to NIR irradiation through the generation of reactive oxygen species (photodynamic therapy). This effective ablation of cancer cells is achieved using an ultra-low IR780 concentration. Moreover, IR780-loaded micelles also have the ability to act as photothermal and imaging agents, which widens their therapeutic and diagnostic potential. Overall, TPGS-TOS micelles are promising nanoplatforms for IR780-mediated cancer phototherapy and imaging.info:eu-repo/semantics/publishedVersio

Polyethylene glycol (PEG) molecular weight influences the ClearT2 optical clearing method for spheroids imaging by confocal laser scanning microscopy

Some fluorescence microscopies, like confocal laser scanning microscopy (CLSM), have limited penetration depth. Consequently, the visualization and imaging of 3D cell cultures, such as spheroids, can be a significant challenge. Therefore, to improve the imaging of 3D tissues, clearing methods have been optimized to render transparency to the opaque spheroids. In this work, the influence of the polyethylene glycol (PEG) molecular weight (MW) used in the ClearT2 method for the imaging of PI stained spheroids was investigated. The results demonstrated that ClearT2 clearing method contribution for spheroids transparency and preservation of the PI fluorescence intensity is independent on the PEG MW. However, the ClearT2 method performed using PEG 4000 Da allowed a better PI signal penetration depth and cross-section depth. Overall, the optimization of PEG MW can improve the imaging of intact spheroids by CLSM. Further, this work may also contribute to increase the application of 3D cell culture models by pharmaceutical industry in therapeutics high-throughput screening.info:eu-repo/semantics/acceptedVersio

Isolation and culture of human umbilical artery smooth muscle cells expressing functional calcium channels

The human umbilical cord is a biological sample that can be easily obtained just after birth. A methodology was developed to perform cultures of human umbilical artery smooth muscle cells (HUASMC) expressing contractile proteins and functional ionic channels. To avoid fibroblast and endothelial cell contamination, we mechanically separated the tunica media, which only contains HUASMC and matrix proteins. To isolate the cells, collagenase V and elastase were used as hydrolyzing enzymes. The isolated cells were plated in collagen-coated dishes to obtain cultures of HUASMC. The cells obtained after different passages (1 to 6) exhibit the characteristic vascular smooth cell morphology and express smooth muscle alpha-2 actin, myosin heavy chain SM1, and alpha subunits of L- and T-type calcium channels (Cav 1.2, Cav 1.2, and Cav 3.2). Electrophysiology recordings for L- and T-type calcium channels were made, indicating that these channels are functional in the cultured cells. In conclusion, the procedure developed allows obtaining cultures of HUASMC expressing contractile proteins and also functional ionic channels. These cells could be used to study cellular and molecular aspects about the regulation of the vascular function.info:eu-repo/semantics/publishedVersio

Chitosan/arginine–chitosan polymer blends for assembly of nanofibrous membranes for wound regeneration

Frequently, skin is subjected to damaging events, such as deep cuts, burns or ulcers, which may compromise the integrity of this organ. To overcome such lesions, different strategies have been employed. Among them, wound dressings aimed to re-establish skin native properties and decreased patient pain have been pursued for a long time. Herein, an electrospun membrane comprised by deacetylated/arginine modified chitosan (CH-A) was produced to be used as a wound dressing. The obtained results showed that the membrane has a highly hydrophilic and porous three-dimensional nanofibrous network similar to that found in human native extracellular matrix. In vitro data indicate that human fibroblasts adhere and proliferate in contact with membranes, thus corroborating their biocompatibility. This nanofiber-based biomaterial also demonstrated bactericidal activity for two bacterial strains. In vivo application of CH-A nanofibers in full thickness wounds resulted in an improved tissue regeneration and faster wound closure, when compared to non-modified membranes. Such findings support the suitability of using this membrane as a wound dressing in a near future.info:eu-repo/semantics/publishedVersio

Electrodynamic tailoring of self-assembled three-dimensional electrospun constructs

The rational design of three-dimensional electrospun constructs (3DECs) can lead to striking topographies and tailored shapes of electrospun materials. This new generation of materials is suppressing some of the current limitations of the usual 2D non-woven electrospun fiber mats, such as small pore sizes or only flat shaped constructs. Herein, we pursued an explanation for the self-assembly of 3DECs based on electrodynamic simulations and experimental validation. We concluded that the self-assembly process is driven by the establishment of attractive electrostatic forces between the positively charged aerial fibers and the already collected ones, which tend to acquire a negatively charged network oriented towards the nozzle. The in situ polarization degree is strengthened by higher amounts of clustered fibers, and therefore the initial high density fibrous regions are the preliminary motifs for the self-assembly mechanism. As such regions increase their in situ polarization electrostatic repulsive forces will appear, favoring a competitive growth of these self-assembled fibrous clusters. Highly polarized regions will evidence higher distances between consecutive micro-assembled fibers (MAFs). Different processing parameters – deposition time, electric field intensity, concentration of polymer solution, environmental temperature and relative humidity – were evaluated in an attempt to control material's design.info:eu-repo/semantics/publishedVersio