NIH 3T3 Mouse Fibroblast Cells Research Articles

Fluorescent half-sandwich iridium picolinamidate complexes for in-cell visualization

In this work, we report on the development of fluorescent half-sandwich iridium complexes using a fluorophore attachment strategy. These constructs consist of pentamethylcyclopentadienyl (Cp*) iridium units ligated by picolinamidate donors conjugated to green-emitting boron-dipyrromethene (bodipy) dyes. Reaction studies in H2O/THF mixtures showed that the fluorescent Ir complexes were active as catalysts for transfer hydrogenation, with activities similar to that of their non-fluorescent counterparts. The iridium complexes were taken up by NIH-3T3 mouse fibroblast cells, with 50% inhibition concentrations ranging from ~20–70 μM after exposure for 3 h. Visualization of the bodipy-functionalized Ir complexes in cells using fluorescence microscopy revealed that they were localized in the mitochondria and lysosome but not the nucleus. These results indicate that our fluorescent iridium complexes could be useful for future biological studies requiring intracellular catalyst tracking.

Effect of Lemongrass Essential Oil based Mouthwash against Microflora Associated with Dental Plaque

Mouthwash is one of the most convenient and effective method employed for dental plaque management. The aim of the undertaken study was to establish the antimicrobial and anti-biofilm properties of lemongrass essential oil (LGEO) based mouthwash on microbial flora from dental plaque and also check cytotoxicity of mouthwash formulation. Five main colonizers of dental plaque representing dental microflora and three different bacterial species mainly responsible for the formation of biofilm were selected in this study. LGEO based mouthwash was developed and its stability was also determined. The antimicrobial and anti-biofilm activity of LGEO based mouthwash has been evaluated against the representative dental microflora as per CLSI guidelines. Cytotoxicity of mouthwash was checked by globally used MTT assay employing NIH 3T3 mouse fibroblast cell line. The mouthwash has been found to exhibit the stability in its major component, citral and also found exhibit antimicrobial and anti-biofilm activity against dental microflora. No cytotoxic effect was observed on mouse fibroblast cell line. LGEO in formulated mouthwash being a natural, herbal material isolated from traditional medicinal plants appears as a good and effective substitute to control the microflora linked with dental plaque.

Systematic Studies of the Circadian Clock Genes Impact on Temperature Compensation and Cell Proliferation Using CRISPR Tools.

Simple SummaryOne of the major characteristics of the circadian clock is temperature compensation, and previous studies suggested a single clock gene may determine the temperature compensation. In this study, we report the first full collection of clock gene knockout cell lines using CRISPR/Cas9 tools. Our full collections indicate that the temperature compensation is a complex gene regulation system instead of being regulated by any single gene. Besides, we systematically compared the proliferation rates and circadian periods using our full collections, and we found that the cell growth rate is not dependent on the circadian period. Therefore, complex interaction between clock genes and their protein products may underlie the mechanism of temperature compensation, which needs further investigations.Mammalian circadian genes are capable of producing a self-sustained, autonomous oscillation whose period is around 24 h. One of the major characteristics of the circadian clock is temperature compensation. However, the mechanism underlying temperature compensation remains elusive. Previous studies indicate that a single clock gene may determine the temperature compensation in several model organisms. In order to understand the influence of each individual clock gene on the temperature compensation, twenty-three well-known mammalian clock genes plus Timeless and Myc genes were knocked out individually, using a powerful gene-editing tool, CRISPR/Cas9. First, Bmal1, Cry1, and Cry2 were knocked out as examples to verify that deleting genes by CRISPR is effective and precise. Cell lines targeting twenty-two genes were successfully edited in mouse fibroblast NIH3T3 cells, and off-target analysis indicated these genes were correctly knocked out. Through measuring the luciferase reporters, the circadian periods of each cell line were recorded under two different temperatures, 32.5 °C and 37 °C. The temperature compensation coefficient Q10 was subsequently calculated for each cell line. Estimations of the Q10 of these cell lines showed that none of the individual cell lines can adversely affect the temperature compensation. Cells with a longer period at lower temperature tend to have a shorter period at higher temperature, while cells with a shorter period at lower temperature tend to be longer at higher temperature. Thus, the temperature compensation is a fundamental property to keep cellular homeostasis. We further conclude that the temperature compensation is a complex gene regulation system instead of being regulated by any single gene. We also estimated the proliferation rates of these cell lines. After systematically comparing the proliferation rates and circadian periods, we found that the cell growth rate is not dependent on the circadian period.

A highly transparent tri-polymer complex in situ hydrogel of HA, collagen and four-arm-PEG as potential vitreous substitute

There is a requirement of removal and replacement of vitreous for various ophthalmic diseases, e.g. retinopathy and retinal detachment. Clinical tamponades, e.g. silicone oil and fluorinated gases are used but limited due to their toxicity and some complications. A lot of polymer-based materials have been tested and proposed as vitreous substitute, but till date, there is no ideal vitreous substitute available. Thus, it requires to develop an improved vitreous substitute which will be highly suitable for vitreous replacement. We have developed tri-polymer complex in situ hydrogels by crosslinking among hyaluronic acid (HA), collagen (Coll) and four-arm-polyethylene glycol (PEG). All the developed hydrogels are biocompatible with NIH 3T3 mouse fibroblast cells, having pH in the range 7–7.44 and refractive index in the range 1.333–1.345. The developed hydrogels are highly transparent, showing transmittance >97%. FTIR study shows that the hydrogel was crosslinked by amide bond formation between HA and PEG, and between Coll and PEG. The rheological study shows that all the developed hydrogels exhibit viscoelastic behavior and all the hydrogels have storage modulus values (>100 pa) which is greater than loss modulus values—indicating sufficient elasticity for vitreous application. The elastic nature of the hydrogel increases with the increase in PEG concentration. The gel is formed in between 2 and 3 min—indicating its application in situ. The viscosity of the developed hydrogels shows shear thinning behavior. The pre-gel solution of the hydrogel is injectable through a 22 G needle—indicating its application in situ through vitrectomy surgery. All the hydrogels are hydrophilic and have water content of 96% approximately. Thus, the results show the positive properties for its application as a potential vitreous substitute.

Comparative analysis of cytocompatible properties of the root surface exposed to diode laser (970 nm) as adjunct to scaling and root planing: An in vitro study.

Aims:The aim of the current study was to examine the effects of diode laser (970 nm) when used as adjunct to mechanical debridement on reattachment of fibroblast cell cultures to the root surface.Materials and Methods:Eighteen extracted single-rooted human teeth were used in the study. Briefly, the anatomical crowns were removed; roots were sectioned and preserved in antiseptic solution. The sections were divided into three groups: (i) scaling and root planing (SRP), (ii) SRP followed by diode laser exposure and (iii) untreated control group. All the sections were inoculated with NIH 3T3 mouse fibroblast cell lines and cell attachment was confirmed by scanning electron microscopy analysis and growth was quantitatively assessed by MTT assay after days 1, 3 and 5.Results:On intragroup analysis, there was significant cell proliferation seen on day 1 and day 5. However, no significant differences were observed between the treatment groups at the various time points.Conclusion:970-nm diode laser exposure had a positive effect in enhancing the fibroblast reattachment to root surfaces. However, to ascertain the role of laser treatment in enhancing the rapid cell growth compared to SRP, further quantitative studies with more sample numbers are required.

DNA with zwitterionic and negatively charged phosphate modifications: Formation of DNA triplexes, duplexes and cell uptake studies.

Two phosphate modifications were introduced into the DNA backbone using the Staudinger reaction between the 3’,5’-dinucleoside β-cyanoethyl phosphite triester formed during DNA synthesis and sulfonyl azides, 4-(azidosulfonyl)-N,N,N-trimethylbutan-1-aminium iodide (N+ azide) or p-toluenesulfonyl (tosyl or Ts) azide, to provide either a zwitterionic phosphoramidate with N+ modification or a negatively charged phosphoramidate for Ts modification in the DNA sequence. The incorporation of these N+ and Ts modifications led to the formation of thermally stable parallel DNA triplexes, regardless of the number of modifications incorporated into the oligodeoxynucleotides (ONs). For both N+ and Ts-modified ONs, the antiparallel duplexes formed with complementary RNA were more stable than those formed with complementary DNA (except for ONs with modification in the middle of the sequence). Additionally, the incorporation of N+ modifications led to the formation of duplexes with a thermal stability that was less dependent on the ionic strength than native DNA duplexes. The thermodynamic analysis of the melting curves revealed that it is the reduction in unfavourable entropy, despite the decrease in favourable enthalpy, which is responsible for the stabilisation of duplexes with N+ modification. N+ONs also demonstrated greater resistance to nuclease digestion by snake venom phosphodiesterase I than the corresponding Ts-ONs. Cell uptake studies showed that Ts-ONs can enter the nucleus of mouse fibroblast NIH3T3 cells without any transfection reagent, whereas, N+ONs remain concentrated in vesicles within the cytoplasm. These results indicate that both N+ and Ts-modified ONs are promising for various in vivo applications.

Glucuronoxylan-based quince seed hydrogel: A promising scaffold for tissue engineering applications

Natural gums and mucilages from plant-derived polysaccharides are potential candidates for a tissue-engineering scaffold by their ability of gelation and biocompatibility. Herein, we utilized Glucuronoxylan-based quince seed hydrogel (QSH) as a scaffold for tissue engineering applications. Optimization of QSH gelation was conducted by varying QSH and crosslinker glutaraldehyde (GTA) concentrations. Structural characterization of QSH was done by Fourier Transform Infrared Spectroscopy (FTIR). Furthermore, morphological and mechanical investigation of QSH was performed by Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). The protein adsorption test revealed the suitability of QSH for cell attachment. Biocompatibility of QSH was confirmed by culturing NIH-3T3 mouse fibroblast cells on it. Cell viability and proliferation results revealed that optimum parameters for cell viability were 2 mg mL−1 of QSH and 0.03 M GTA. SEM and DAPI staining results indicated the formation of spheroids with a diameter of approximately 300 μm. Furthermore, formation of extracellular matrix (ECM) microenvironment was confirmed with the Collagen Type-I staining. Here, it was demonstrated that the fabricated QSH is a promising scaffold for 3D cell culture and tissue engineering applications provided by its highly porous structure, remarkable swelling capacity and high biocompatibility.

One-step vapor deposition of fluorinated polycationic coating to fabricate antifouling and anti-infective textile against drug-resistant bacteria and viruses.

The ongoing pandemic caused by the novel coronavirus has turned out to be one of the biggest threats to the world, and the increase of drug-resistant bacterial strains also threatens the human health. Hence, there is an urgent need to develop novel anti-infective materials with broad-spectrum anti-pathogenic activity. In the present study, a fluorinated polycationic coating was synthesized on a hydrophilic and negatively charged polyester textile via one-step initiated chemical vapor deposition of poly(dimethyl amino methyl styrene-co-1H,1H,2H,2H-perfluorodecyl acrylate) (P(DMAMS-co-PFDA), PDP). The surface characterization results of SEM, FTIR, and EDX demonstrated the successful synthesis of PDP coating. Contact angle analysis revealed that PDP coating endowed the polyester textile with the hydrophobicity against the attachment of different aqueous foulants such as blood, coffee, and milk, as well as the oleophobicity against paraffin oil. Zeta potential analysis demonstrated that the PDP coating enabled a transformation of negative charge to positive charge on the surface of polyester textile. The PDP coating exhibited excellent contact-killing activity against both gram-negative Escherichia coli and gram-positive methicillin-resistant Staphylococcus aureus, with the killing efficiency of approximate 99.9%. In addition, the antiviral capacity of PDP was determined by a green fluorescence protein (GFP) expression-based method using lentivirus-EGFP as a virus model. The PDP coating inactivated the negatively charged lentivirus-EGFP effectively. Moreover, the coating showed good biocompatibility toward mouse NIH 3T3 fibroblast cells. All the above properties demonstrated that PDP would be a promising anti-pathogenic polymeric coating with wide applications in medicine, hygiene, hospital, etc., to control the bacterial and viral transmission and infection.

Sauropus androgynus (L.) leaf phytochemical activated biocompatible zinc oxide nanoparticles: An antineoplastic agent against human triple negative breast cancer and a potent nanocatalyst for dye degradation

The present study delineates the biosynthesis of ZnOSA NPs by availing S. androgynous (L.) aqueous leaf extract. The formation of ZnOSA NPs was substantiated by FT-IR and UV–visible spectroscopy which was further corroborated by the elemental composition study through EDS spectroscopy. The FE-SEM and HRTEM micrographs have revealed the formation of NPs with different shapes and sizes ranging from 12 to 23 nm. The BET isotherms obtained have manifested the mesoporous nature of ZnOSA NPs. The in-vitro cytotoxicity studies of ZnOSA NPs carried out on MDAMB468 human triple-negative breast cancer (TNBC) and NIH3T3 mouse fibroblast cells have evinced their potential cytotoxic effect (75.98 ± 1.07) on cancer cells sparing the normal cells. Further, the results of the AO-EB dual staining assay indicated early apoptosis in TNBC cells by displaying greenish yellow-fluorescence in the nuclei at a IC50 concentration of 53.79 µg/mL of NPs. Additionally; the human erythrocyte membrane stabilization assay has corroborated the haemocompatibility of ZnOSA NPs by exhibiting a % HRBCMS up to 89.15 ± 0.02%. Furthermore, the photocatalytic experiments performed on Methylene Blue dye have revealed the excellent degradation efficiencies of ZnOSA NPs up to 88.01% with high stability and reusability tested during 4 catalytic cycles. Thus, the obtained results have shown greater potential in the anticancer and photocatalytic dye degradation activities of bio-fabricated ZnOSA NPs.

Preparation of protein nanoparticle-coated poly(hydroxybutyrate) electrospun nanofiber based scaffold for biomedical applications

In this study, soybean protein nanoparticles (SPN) modified poly(hydroxybutyrate) (PHB) electrospun nanofiber scaffold is fabricated by a facile two-step method combination of PHB electrospinning and SPN evaporation-induced self-assembly. The surface characteristics, structure and properties are investigated by SEM, EDS, XPS, FTIR, XRD, DMA, WCA, tensile test and water uptake capacity measurements. To characterize biodegradability and cytocompatibility, biodegradation tests are performed in simulated body fluid with and without lysozyme and cell cultures are carried out using NIH3T3 mouse fibroblast cells by in vitro studies. The as-obtained SPN-modified PHB nanofiber scaffold presents better cytocompatibility and is more suitable for biomedical applications.

Ginger facilitates cell migration and heat tolerance in mouse fibroblast cells.

The components of ginger root (Zingiber officinale Roscoe) are widely used for various medicinal purposes. Several bioactive compounds have been identified in ginger, including 6-, 8- and 10-gingerols, and 6-shogaol, which are agonists of the thermo-sensors transient receptor potential (TRP) cation channel subfamily V member 1 and TRP ankyrin 1. Our previous study demonstrated that ginger powder may affect human metabolism in vivo. However, the effects of the bioactive compounds of ginger on cells have not been completely elucidated. The present study investigated whether ginger powder extracts could modify cell functions in mouse fibroblast cells. The active components of ginger powder extracts were characterized using high-performance liquid chromatography. The activation of protein kinases, actin assembly, cell migration, expression levels of heat shock proteins (HSPs) and cell viability after heat shock were analyzed in NIH3T3 mouse fibroblast cells. Subsequently, 6-, 8-, 10- and 12-gingerols, as well as 6-, 8- and 10-shogaols, were detected in ginger powder extracts. The levels of phosphorylated Akt, mTOR, ERK and p38 MAPK increased after a 10-min stimulation with ginger powder extracts. In addition, HSP expression levels, lamellipodia formation occurring at cell edges, cell migration and tolerance against heat shock were facilitated following ginger powder extract stimulation. These results suggest that ginger modified cell functions, including actin assembly and heat tolerance, in vitro.

Unravelling the human triple negative breast cancer suppressive activity of biocompatible zinc oxide nanostructures influenced by Vateria indica (L.) fruit phytochemicals

The present study delineates the biosynthesis of ZnOVI nanostructures by using aqueous fruit extract of V. indica. The study has disclosed the role of V. indica fruit extract as both reducing and capping agents, ushering the formation of ZnOVI nanostructures with distinct morphologies. The formation of ZnOVI nanostructures was corroborated by FT-IR and UV–visible spectroscopy which was further substantiated by the elemental composition study through EDS spectroscopy. The nanostructures were also investigated by Rietveld refinement of PXRD data, FE-SEM, and BET analysis. The morphology, size, and surface area were found to be precursor stoichiometry dependent. The in-vitro cytotoxicity study of ZnOVI nanostructures carried out on MDA-MB468 human triple-negative breast cancer (TNBC) cells has revealed their potential cytotoxicity (91.18 ± 1.98). MTT assay performed on the NIH3T3 mouse fibroblast cells has unfolded the non-toxic nature of ZnOVI nanostructures. Additionally, the results of the AO-EB dual staining assay indicated early apoptosis in TNBC cells by displaying greenish yellow-fluorescence in the nuclei. Reactive oxygen species (ROS) measurement study has confirmed the elevated intracellular levels of ROS, supporting the oxidative-stress induced cytotoxicity in ZnOVI nanostructures treated TNBC cells. Furthermore, the haemocompatibility of ZnOVI nanostructures was evaluated using human erythrocytes. Thus, the obtained results have shown greater potential in the anticancer activity of bio-fabricated ZnOVI nanostructures.

In VivoComparison of Three Human Acellular Dermal Matrices for Breast Reconstruction

Acellular dermal matrices (ADMs) have become popular in implant-based breast reconstruction. The aim of this study was to compare three commonly used ADM products in vivo in an animal model. The nucleic acid content (residual double-stranded DNA) and the levels of the remaining growth factors after decellularization were measured for each ADM. Cytocompatibility with ADMs was documented using NIH 3T3 mouse fibroblast cells. In vivo, the implanted ADMs were histologically evaluated at 1, 2, 3, and 6 months (n=5) using male 8-week-old Sprague-Dawley rats. Fibroblasts grew in the SureDerm HD and DermACELL with no cytotoxicity. In a rat model, SureDerm HD and DermACELL incorporated more readily into the surrounding host tissue, as measured by rapid cell influx and collagen deposition, and showed more delayed tissue remodeling with decreased matrix metalloproteinases levels compared to AlloDerm. SureDerm HD and DermACELL can be used as biological materials for breast reconstruction.

Silencing PCBP2 normalizes desmoplastic stroma and improves the antitumor activity of chemotherapy in pancreatic cancer.

Rationale: Dense desmoplastic stroma is a fundamental characteristic of pancreatic ductal adenocarcinoma (PDAC) and comprises up to 80% of the tumor mass. Type I collagen is the major component of the extracellular matrix (ECM), which acts as a barrier to impede the delivery of drugs into the tumor microenvironment. While the strategy to deplete PDAC stroma has failed in clinical trials, normalization of the stroma to allow chemotherapy to kill the tumor cells in the “nest” could be a promising strategy for PDAC therapy. We hypothesize that silencing the poly(rC)-binding protein 2 (αCP2, encoded by the PCBP2 gene) leads to the destabilization and normalization of type I collagen in the PDAC stroma.Methods: We develop a micro-flow mixing method to fabricate a peptide-based core-stabilized PCBP2 siRNA nanocomplex to reverse the accumulation of type I collagen in PDAC tumor stroma. Various in vitro studies were performed to evaluate the silencing activity, cellular uptake, serum stability, and tumor penetration of the PCBP2 siRNA nanocomplex. We also investigated the penetration of small molecules in stroma-rich pancreatic cancer spheroids after the treatment with the PCBP2 siRNA nanocomplex. The anti-tumor activity of the PCBP2 siRNA nanocomplex and its combination with gemcitabine was evaluated in an orthotopic stroma-rich pancreatic cancer mouse model.Results: Silencing the PCBP2 gene using siRNA reverses the accumulation of type I collagen in human pancreatic stellate cells (PSCs) and mouse NIH 3T3 fibroblast cells. The siRNA nanocomplex significantly reduces ECM production and enhances drug penetration through desmoplastic tumor stroma. The combination of gemcitabine with the PCBP2 siRNA nanocomplex markedly suppresses the tumor progression in a desmoplastic PDAC orthotopic mouse model.Conclusion: This approach provides a new therapeutic avenue to improve the antitumor efficacy of PDAC therapies by normalizing tumor stroma using the PCBP2 siRNA nanocomplex.

Dielectrophoretic manipulation of particles on a microfluidics platform with planar tilted electrodes

Cell/particle separation enabled by lab-on-a-chip (LOC) devices has been a promising solution for separating target cells from the body fluids. Understanding the effects of fabrication and performance parameters on the separation efficiency can further advance applications of such devices in life sciences. This study presents a parametric study on the displacement of the cell-representative polystyrene particles in dielectrophoresis (DEP) based microseparators. The proposed device includes slanted planar interdigitated electrodes to apply a lateral DEP force on the particles. The effect of fabrication parameters (i.e., the channel length and width, and the deflection angle of the electrodes), as well as operational factors (i.e., the volumetric throughput and particle size) on the DEP-induced displacement, are studied. The results show that the volumetric throughput has the most contribution to the position of the particles. Among the geometrical parameters, the channel length is found to be the most effective factor in the particles position, while the effect of the channel width is shown to be negligible. The device was also tested with NIH 3T3 mouse fibroblast cells, which exhibited the same behavior. The results of this research render the desired separation performance to be attained via the design of low-angle slanted electrodes alongside with a balance between the volumetric throughput and the length of the DEP region. These findings lead to a successful size-selective sorting or focusing of 5 μm–15 μm particles, which can provide valuable insights into the design of microfluidic platforms to selectively separate different cell types in biofluids.

Production of Porous Films Based on Biodegradable Polyesters by the Casting Solution Technique Using a Co-Soluble Porogen (Camphor).

Porous films have been prepared from degradable polymers—poly-3-hydroxybutyrate (PHB), poly-ε-caprolactone (PCL) and a blend of these polymers (1:3)—by adding porogen (camphor) to the polymer solution at 10%, 30% or 50% of the total mass of the polymer and porogen, and leaching it out afterwards. After the rinse, camphor content in films decreased to about 0.025%. The structure, physical/mechanical and biological properties of the films were investigated as dependent on their composition and porosity, which varied depending on the amount of camphor added. The surface of PHB films was porous, the PCL films were relatively smooth, and the PHB/PCL films had an intermediate structure. The addition of camphor increased the thickness (from 35 to 45 µm, from 40 to 80 µm and from 20 to 65 µm for PHB, PCL and PHB/PCL, respectively) and porosity (from 4.2(±3.6)% to 50.0(±12.8)%, from 6.4(±5.5)% to 54.5(±6.0)% and from 4.9(±4.8)% to 51.5(±5.8)%, respectively) of the films. The introduction (and removal) of 10% camphor into the PHB and PHB/PCL films led to an approximately twofold increase in the polar component of the free surface energy (from 5.4 ± 0.38 to 11.8 ± 1.33 and from 2.7 ± 0.13 to 5.2 ± 0.09 mN/m, respectively) but in other cases, on the contrary, a decrease in this indicator was registered. The increase of camphor addition from 0% to 50% gradually impaired mechanical properties of the films: so, Young’s modulus decreased from 3.6 to 1.8 GPa, from 0.30 to 0.12 GPa and from 0.50 to 0.20 GPa for PHB, PCL and PHB/PCL, respectively. At the same time, the water vapor transmission rate considerably increased from 197.37 ± 23.62 to 934.03 ± 114.34 g/m2/d for PHB films; from 1027.99 ± 154.10 to 7014.62 ± 280.81 g/m2/d for PCL films; and from 715.47 ± 50.08 to 4239.09 ± 275.54 g/m2/d for PHB/PCL films. Results of biocompatibility testing in the culture of NIH 3T3 mouse fibroblast cells showed that for the most of experimental samples cell adhesion and proliferation were comparable or superior to the corresponding parameters on the initial nonporous films. The best results were obtained for PHB films where at Day 3 of the experiment the registered cell density for experimental samples arrived at 2.66(±0.26) × 105 cells/cm2 versus 1.29(±0.33) × 105 cells/cm2 in the control. So, the proposed method can be used to construct highly porous cell scaffolds for cellular engineering.

α-Glucosidase Inhibitory Activity of the Extracts and Major Phytochemical Components of Smilax glabra Roxb

Background: A therapeutic approach to treat diabetes is to decrease postprandial hyperglycemia. α-Glucosidase inhibitors from plant sources offer an attractive strategy for the control of hyperglycemia. Smilax glabra Roxb is a medicinal plant found in Asia, including Vietnam, which is used in the treatment of chronic diseases. However, the antidiabetic activity and the identification of α-glucosidase inhibitors from this plant have not been intensively investigated. This research was carried out to determine the α-glucosidase inhibitory activity of the extracts and that of the major phytochemical components of Smilax glabra Roxb. This could lead to further studies on the role of these compounds in hyperglycemia control, as well as identify their potential future applications. Methods: Column chromatography combined with crystallization procedures were used to isolate active fractions and two major compounds. The chemical structures of these compounds were determined by analysis of their NMR spectroscopic data, as well as MS data and comparisons made with the literature data. The α-glucosidase inhibitory activity was determined spectrophotometrically using p-nitrophenyl α-D-glucopyranoside as a substrate. The in vitro cytotoxicity of the isolated compounds and fractions was determined using the MTT assay. Results: The two major compounds, astilbin and 5-O-caffeoylshikimic acid together with two very active fractions, F7 and F8, were isolated from the rhizome. The two major compounds had α- glucosidase inhibitory activities with IC50 values of ca. 125 µg/mL and 38 µg/mL, respectively which are about 4 and 13 folds higher activity than the reference compound acarbose (IC50 of ca. 525 µg/mL). Fractions F7 and F8 showed very promising inhibitory activities towards α-glucosidase with IC50 values of 5.5 and 5.8 µg/mL, respectively. Cytotoxicity data on mouse fibroblast NIH3T3 cells indicated that the active compounds and fractions were not toxic at concentrations that are greater than their respective IC50 values. The α-glucosidase inhibitory activity of 5-Ocaffeoylshikimic acid and that of the two active fractions are reported here for the first time. Conclusion: The two major isolated compounds and fractions, F7 and F8, significantly contribute to the α-glucosidase inhibitory activity of S. glabra Roxb extract. Further work is needed to clarify their modes of action and potential application.

PEG-PCL Nanocapsules Containing Curcumin: Validation of HPLC Method for Analyzing Drug-loading Efficiency, Stability Testing and Cytotoxicity on NIH-3T3 Cell Line

Curcumin (CUR) shows potential use for treating cancer. However, CUR has low solubility and reduced bioavailability, which limit its clinical effect. Therefore, the development of nanocarriers can overcome these problems and can ensure the desired pharmacological effect. In addition, it is mandatory to prove the quality, the efficacy, and the safety for a novel nanomedicine to be approved. In that sense, this paper aimed (a) to prepare CUR-loaded polyethylene glycol-poly(ε-caprolactone) nanocapsules; (b) to validate an analytical method by high performance liquid chromatography (HPLC) for quantifying CUR in these nanoformulations; (c) to evaluate the physicochemical stability of these formulations; and to investigate their cytotoxicity on NIH-3T3 mouse fibroblast cells. The HPLC method was specific to CUR in the loaded nanocapsules, linear (r = 0.9994) in a range of 10.0 to 90.0 µg.mL-1 with limits of detection and quantification of 0.160 and 0.480 µg.mL-1, respectively. Precision was demonstrated by a relative standard deviation lower than 5%. Suitable accuracy (102.37 ± 0.92%) was obtained. Values of pH, particle size, polydispersity index, and zeta potential presented no statistical difference (p > 0.05) for CUR-loaded nanoparticles. No cytotoxicity was observed against NIH-3T3 mouse embryo fibroblast cell line using both the tetrazolium salt and sulforhodamine B assays. In conclusion, a simple and inexpensive HPLC method was validated for the CUR quantification in the suspensions of nanocapsules. The obtained polymeric nanocapsules containing CUR showed suitable results for all the performed assays and can be further investigated as a feasible novel approach for cancer treatment.

A novel bilayer zein/MMT nanocomposite incorporated with H. perforatum oil for wound healing.

Recently, layered structures composed of nanofibers have gained attention as a novel material to mimic skin tissue in wound healing applications. The aim of this study is to develop a novel hybrid bilayer material composed of zein based composite film and nanofiber layers as a wound dressing material. The upper layer was composed of H. perforatum oil incorporated zein film including MMT and the bottom layer was comprised of 3D electrospun zein/MMT nanofibers to induce wound healing with the controlled release of H. perforatum oil. The bilayer composites were characterized in terms of mechanical test, WVP, water uptake and surface wettability. Antimicrobial activity of the wound dressings against microorganisms were investigated by disc diffusion method. In vitro cytotoxicity of monolayer film and bilayer structure was performed using WST-1 assay on HS2 keratinocyte and 3T3 cell lines. Results indicated that the prepared monolayer films showed appropriate mechanical and gas barrier properties and surface wettability for wound healing. Controlled release of H. perforatum oil was obtained from fabricated membranes up to 48 h. Bilayer membranes showed antimicrobial activity against E. coli, S. aureus, and C. albicans and did not show any toxic effect on NIH3T3 mouse fibroblast and HS2 keratinocyte cell lines. In vitro scratch assay results indicated that H. perforatum oil had a wound healing effect by inducing fibroblast migration. The proliferation study supported these results by increasing fibroblast proliferation on H. perforatum oil loaded bilayer membranes.

Functionally modified magnetic nanoparticles for effective siRNA delivery to prostate cancer cells in vitro.

Different types of siRNA delivery vehicles including nanoparticles have been synthesized and utilized for prostate cancer gene therapy. However, one of the most common limitations being faced is the toxicity of cationic polymers toward the cells. In the current study, magnetic nanoparticles were prepared and conjugated with cationic polymer, polyethylenimine. Then polyethylene glycol was conjugated with polyethylenimine to improve the biocompatibility of nanoparticles. The transmission electron microscopy size of nanoparticles was found to be 15.82 (±9.07) nm, while hydrodynamic size was about 79.20 (±0.68) nm. Zeta potential analysis of polyethylenimine and polyethylene glycol-coated nanoparticles was +31.4 (±0.62) and +5.65 (±0.76) mV, respectively. Fourier transform infrared spectroscopy and 1H nuclear magnetic resonance confirmed the presence of polyethylene glycol and polyethylenimine polymers in magnetic nanoparticles. Cell viability test in mouse fibroblast NIH 3T3 and prostate cancer PC3 cells showed an increased in biocompatibility of functionally modified polyethylene glycol–polyethylenimine–Fe3O4 nanoparticles. siRNA targeting a disintegrin and metalloproteinase 10 (ADAM10) was successfully loaded into the polyethylene glycol–polyethylenimine–Fe3O4 nanoparticles and delivered to PC3 cells. The results clearly demonstrated a significant decrease in cell viability, which increased within a certain siRNA concentration. The inhibitory concentration (IC50) value for ADAM10 siRNA was calculated to be 15.83 nM after 72 h. Confocal microscopy confirmed the delivery of siRNA-loaded nanoparticles intracellularly to the tumor cells cytosol. This magnetic system can be used as a powerful platform to inhibit cancer cells progression.