Significant Increase In Cell Proliferation Research Articles

Osteoblast cell behavior on polyetheretherketone dental implant surfaces treated with different grit size aluminum oxide particles: An in vitro analysis

The hydrophobic and bioinert nature of polyetheretherketone (PEEK) implants needs to be addressed for successful osseointegration. The purpose of this in vitro study was to evaluate the osteoblast cell behavior on PEEK implant surfaces treated with airborne-particle abrasion using different grit size aluminum oxide (Al2O3) particles. Disk-shaped specimens (n=96) were prepared from medical grade PEEK rods and were distributed into 4 groups (n=24) of untreated PEEK (PEEK 0), airborne-particle abrasion using 50-μm Al2O3 particles (PEEK 50), airborne-particle abrasion using 110-μm Al2O3 particles (PEEK 110), and airborne-particle abrasion using 150-μm Al2O3 particles (PEEK 150). The surface characteristics were assessed using water contact angle (WCA) measurements and scanning electron microscopy (SEM). MG-63 osteoblast cells were cultured, and the biocompatibility of PEEK was assessed using a CellTiter-blue cell viability assay and florescence staining at day 1, 3, and 7. The specimens were stained with Alizarin red to assess the osteoblast cell differentiation on day 10 and 14. The Levene test was used to test the homogeneity of variances. One-way and Welch ANOVA with post hoc corrections were used to assess the overall statistical significance of differences among the groups (α=.05). The lowest mean WCA was demonstrated in PEEK 150 (49.25 ±5.51) and the highest in PEEK 0 (89.14 ±4.24) (P<.001). SEM images of PEEK 150 illustrated a more complex structure with a large area of globular outcroppings throughout the surface. PEEK 150 showed the highest cell metabolic activity at each time point with florescence staining showing a substantial cell confluence at day 3 and 7. Although PEEK 150 did not show a significant increase in cell proliferation, the number of cells attached was significantly higher than other groups (P<.05). PEEK 110 and 150 also showed a substantial increase in the extent of mineralization. Airborne-particle abrasion using moderate Al2O3 grit size (110- or 150-μm) improved the hydrophilicity and osteoblast cell behavior on PEEK implants.

Carboxymethyl cellulose-agarose hydrogel in poly(3-hydroxybutyrate-co-3-hydroxyvalerate) nanofibers: A novel tissue engineered skin graft

Healing chronic and critical-sized full-thickness wounds is a major challenge in the healthcare sector. Scaffolds prepared using electrospinning and hydrogels serve as effective treatment options for wound healing by mimicking the native skin microenvironment. Combining synthetic nanofibers with tunable hydrogel properties can effectively overcome limitations in skin scaffolds made only with nanofibers or hydrogels. In this study, a biocompatible hybrid scaffold was developed for wound healing applications using poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) nanofibers embedded with hydrogel made of 2 % carboxymethyl cellulose (CMC) blended with 1 % agarose. Hybrid scaffolds, characterized for surface morphology, swellability, porosity, and degradation, were found to be suitable for wound healing. Furthermore, the incorporation of CMC-agarose hydrogel into nanofibers significantly enhanced their mechanical strength compared to PHBV nanofibers alone (p < 0.05). Extract cytotoxicity and direct cytotoxicity tests showed that the hybrid scaffolds developed in this study are cytocompatible (>75 % viability). Furthermore, human adult dermal fibroblasts (HDFa) and human adult immortalized keratinocytes (HaCaT) adhesion, viability, and proliferation studies revealed that the hybrid scaffolds exhibited a significant increase in cell proliferation over time, similar to PHBV nanofibers. Finally, the developed hybrid scaffolds were evaluated in rat full-thickness wounds, demonstrating their ability to promote full-thickness wound healing with reepithelialization and epidermis closure.

Influence of Different Photobiomodulation Parameters on Multi-Potent Adipose Tissue Mesenchymal Cells In Vitro.

Objective: Investigating the effect of different parameters of photobiomodulation (PBM) with low-power laser on multi-potent mesenchymal stem cells (MSCs) derived from adipose tissue in terms of proliferation and cell death. Methods: MSCs were submitted to PBM applications with combinations of the following physical parameters: control group (no intervention), wavelengths of 660 and 830 nm; energy of 0.5, 2, and 4 J; and power of 40 and 100 mW. MSC analysis was performed using MetaXpress® software at 24, 48, and 72 h. Results: Irradiation promoted a significant increase in cell proliferation (p < 0.05), with 830 nm laser, 100 mW, with energy of 0.5, 2, and 4 J in relation to the control group at all times. PBM with 660 nm, power of 40 mW, and energy of 0.5, 2, and 4 J produced greater cell death at 24 h compared with the control group. At the time of 72 h, there was no significant difference concerning cell death. Conclusions: According to the results found, we can conclude that both wavelengths were effective; however, the 830 nm laser was more effective in terms of cell proliferation compared with the 660 nm laser. The 660 nm wavelength showed a significant increase in cell death when compared with the 830 nm laser.

Gingival-derived mesenchymal stem cell therapy regenerated the radiated salivary glands: functional and histological evidence in murine model

BackgroundRadiotherapy in head and neck cancer management causes degeneration of the salivary glands (SG). This study was designed to determine the potential of gingival mesenchymal stem cells (GMSCs) as a cell-based therapy to regenerate irradiated parotid SG tissues and restore their function using a murine model.MethodsCultured isolated cells from gingival tissues of 4 healthy guinea pigs at passage 3 were characterized as GMSCSs using flow cytometry for surface markers and multilineage differentiation capacity. Twenty-one Guinea pigs were equally divided into three groups: Group I/Test, received single local irradiation of 15 Gy to the head and neck field followed by intravenous injection of labeled GMSCs, Group II/Positive control, which received the same irradiation dose followed by injection of phosphate buffer solution (PBS), and Group III/Negative control, received (PBS) injection only. Body weight and salivary flow rate (SFR) were measured at baseline, 11 days, 8-, 13- and 16-weeks post-irradiation. At 16 weeks, parotid glands were harvested for assessment of gland weight and histological and immunohistochemical analysis.ResultsThe injected GMSCs homed to degenerated glands, with subsequent restoration of the normal gland histological acinar and tubular structure associated with a significant increase in cell proliferation and reduction in apoptotic activity. Subsequently, a significant increase in body weight and SFR, as well as an increase in gland weight at 16 weeks in comparison with the irradiated non-treated group were observed.ConclusionThe study provided a new potential therapeutic strategy for the treatment of xerostomia by re-engineering radiated SG using GMSCs.

45S5/PEEK Coatings by Cold Gas Spray with In Vitro Bioactivity, Degradation, and Cellular Proliferation

This study evaluated the biological response of cold-sprayed coatings composed of bioactive glass 45S5 and polyetheretherketone (PEEK). The functional coatings were produced by cold gas spray (CGS) technology, a technique that allows the deposition of powders at significantly low temperatures, avoiding heat damage to polymeric surfaces. By CGS, blends with different ratios of bioactive glass and PEEK powders have been deposited onto PEEK substrates to improve the response of the bio-inert polymer. The bioactivity of the coatings when immersed in a simulated body fluid solution was evaluated by observation with scanning electron microscopy (SEM) and x-ray diffraction (XRD). Results verify that bioactive glass particles in the composite coatings enhance their bioactivity. A degradation test was performed with Tris–HCl solution. From the results obtained by inductively coupled plasma optical emission spectroscopy (ICP-OES) and the weight loss of the samples, it was noticed that the degradation was directly related to the amount of glass in the coatings. Finally, the ability of bone-forming cells to adhere and proliferate on the coatings was evaluated. These experiments showed that the presence of glass particles does not cause a significant increase in cell proliferation. Combining a bioactive material with PEEK leads to forming a final component that provides suitable bioactivity to the final implant.

Assessment of normal human breast cells (MCF10A cells)–Surface interactions on porous poly-di-methyl-siloxane (PDMS) structures for potential biomedical applications

Biocompatibility stands out as a crucial and fundamental requirement before approval of biomaterials for medical use. The study aimed to evaluate the interaction between normal human breast cells (MCF10A cells) and porous poly-di-methyl-siloxane (PDMS) structures for potential biomedical applications. Preparation and characterization of the PDMS substrate were carried out, followed by the assessment of cell proliferation and fluorescence imaging using an Alamar blue assay and fluorescence microscopy, respectively. The results revealed that initially (at 4 hours post-incubation), there was no notable difference in cell proliferation among the various groups (non-porous PDMS, PDMS_0-150, PDMS_150-250, and PDMS_250500). However, at 48 and 96 hours, a significant increase in cell proliferation was observed in the PDMS_250–500 μm group compared to other groups (P&lt;0.05). Furthermore, the results of the fluorescence microscopy corroborated a substantial enhancement in cell growth and attachment as the porosity of the PDMS substrate increased. However, cells seeded on non-porous PDMS surfaces exhibited a significant decline (P&lt;0.05) in cell growth in both the Alamar blue assay and fluorescence imaging. These findings hold great promise for the creation of surfaces and materials that are specifically designed to influence biological reactions and show potential for a range of biomedical uses.

Effect of curcumin nanoparticles on proliferation and migration of mouse airway smooth muscle cells and airway inflammatory infiltration.

Curcumin (CUR) possesses the capability to inhibit various inflammatory factors, exert anti-inflammatory effects, and alleviate asthma attacks; however, its hydrophobicity and instability significantly impede its clinical application. In this study, we synthesized CUR-loaded nanoparticles (CUR-NPs) and evaluated their impact on the proliferation, migration, and inflammatory infiltration of mouse airway smooth muscle cells (ASMCs), while investigating their underlying mechanisms. To achieve this objective, ASMCs were isolated from BALB/c mice and subjected to TGF-β1-induced cell proliferation and migration. Our findings demonstrate that CUR-NPs effectively regulate the release of CUR within cells with superior intracellular uptake compared to free CUR. The CCK-8 assay results indicate that the blank carrier does not exhibit any cytotoxic effects on cells, thus rendering the impact of the carrier itself negligible. The TGF-β1 group exhibited a significant increase in cell proliferation, whereas treatment with CUR-NPs significantly suppressed TGF-β1-induced cell proliferation. The findings from both the cell scratch assay and transwell assay demonstrated that TGF-β1 substantially enhanced cell migration, while CUR-NPs treatment effectively attenuated TGF-β1-induced cell migration. The Western blot analysis demonstrated a substantial increase in the expression levels of TGF-β1, p-STAT3, and CTGF in ASMCs following treatment with TGF-β1 when compared to the control group. Nevertheless, this effect was effectively counteracted upon administration of CUR-NPs. Furthermore, an asthma mouse model was successfully established and CUR-NPs were administered through tail vein injection. The serum levels of TGF-β1 and the expression levels of TGF-β1, p-STAT3, and CTGF proteins in the lung tissue of mice in the model group exhibited significant increases compared to those in the control group. However, CUR-NPs treatment effectively attenuated this change. Our research findings suggest that CUR-NPs possess inhibitory effects on ASMC proliferation, migration, and inflammatory infiltration by suppressing activation of the TGF-β1/p-STAT3/CTGF signaling pathway, thereby facilitating inhibition of airway remodeling.

Low-Intensity Nanosecond Pulsed Electric Field Accelerates Osteogenic Transformation of Human Dermal Fibroblasts by Enhancing Cell Pluripotency.

Autologous human fibroblasts have the potential to differentiate into the osteogenic lineage under specific conditions and can be utilized for bone regeneration. However, their efficiency is currently unsatisfactory. Recently, low-intensity nanosecond pulsed electric field (nsPEF) stimulation has been demonstrated to enhance cell pluripotency by activating epigenetic regulatory pathways. In this study, human dermal fibroblasts were exposed to different intensities of nsPEF to assess whether these exposures resulted in changes in proliferation rate, calcium salt deposition, and expression of differentiation-related markers in different experimental groups. The results showed a significant increase in cell proliferation, pluripotency, bone marker expression, and osteogenic differentiation efficiency when stimulating cells with 5 kV/cm of nsPEF. However, cell proliferation and differentiation significantly decreased at 25 kV/cm. Additionally, the proliferation and efficiency of osteogenic differentiation were reduced when the nsPEF intensity was increased to 50 kV/cm. Treatment with a 5 kV/cm of nsPEF led to increased and concentrated expression of Yes-Associated Protein (YAP) in the nucleus. These observations suggest that human dermal fibroblasts possess a heightened potential to differentiate into osteogenic cells when activated with nsPEF at 5 kV/cm. Consequently, the nsPEF strengthening strategy shows promise for fibroblast-based tissue-engineered bone repair research.

Electrospun Core-Sheath Nanofibers with a Cellulose Acetate Coating for the Synergistic Release of Zinc Ion and Drugs.

Precisely modulating the synergistic release behavior of multiple bioactive substances has emerged as a formidable challenge in recent years. In this work, we successfully prepared core-sheath nanofibers, where a thin cellulose acetate (CA) coating enrobed the core. Curcumin (Cur) was encapsulated in the core layer as a model drug, while zinc oxide (ZnO) nanoparticles were loaded on the sheath layer. The prepared fiber exhibited a straight cylindrical morphology containing nanoparticles, and the distinct core-sheath nanostructure was demonstrated through transmission electron microscopy (TEM). X-ray diffraction (XRD) and Fourier transform infrared (FTIR) were conducted to study the physical state and compatibility among CA, Cur, and ZnO. Drug release data indicated that core-sheath nanofibers were able to decelerate the rate of drug release, and the thickness of the sheath layer increased in the presence of ZnO particles. Most remarkably, these core-sheath nanofibers exhibited the remarkable ability to sustain the release of drugs and zinc ion (Zn2+), the two-day synergistically release behavior leading to a significant increase in cell proliferation. This material preparation strategy for the synergistic and controlled release of two bioactive substances is instructive for the exploration of innovative and versatile drug delivery systems.

Melatonin Inhibits Gastric Cancer Cell Proliferation by Suppressing Exosome miR-27b-3p Expression.

In addition to its established role in regulating circadian rhythms and reducing inflammation, melatonin has been demonstrated to possess anti-cancer properties. In this study, we investigated the effects of exosomal miRNAs released by melatonin-treated GC cells on gastric cancer. To identify the potential exosomal miRNAs involved in the treatment of gastric cancer, we performed exosome small RNA sequencing (sRNA-seq) to screen significant changes in 34 exosomal miRNAs in AGS cells before and after melatonin treatment. CCK-8, wound healing, and transwell invasion assays were used to examine the effects of miRNAs on cancer characteristics. Furthermore, a dual-luciferase reporter gene assay was performed to identify the miRNA targets. Exosomal miR-27b-3p was down-regulated by approximately 1.37-fold following melatonin treatment. The CCK-8 assay revealed a significant increase in cell proliferation in the miR-27b-3p mimic group compared to that in the miR-27b-3p mimic NC group. In the wound healing assay, cells treated with miR-27b-3p mimics displayed significantly more rapid wound closure than that observed in the miR-27b-3p mimic NC group. The transwell invasion assay revealed a substantial increase in the number of invading cells in the miR-27b-3p mimic group compared to that in the miR-27b-3p mimic NC group. Additional analysis revealed that miR-27b-3p directly targets ADAMTS5 and that its up-regulation results in increased proliferation, invasion, and metastasis of GC cells. Melatonin suppressed the progression of gastric cancer by regulating the exosomal miR-27b-3p-ADAMTS5 pathway. Thus, melatonin represents a promising potential therapeutic agent for patients with gastric cancer.

Efficient improvement of the proliferation, differentiation, and anti-arthritic capacity of mesenchymal stem cells by simply culturing on the immobilized FGF2 derived peptide, 44-ERGVVSIKGV-53

IntroductionThe stem cell microenvironment has been evidenced to robustly affect its biological functions and clinical grade. Natural or synthetic growth factors, especially, are essential for modulating stem cell proliferation, metabolism, and differentiation via the interaction with specific extracellular receptors. Fibroblast growth factor-2 (FGF-2) possesses pleiotropic functions in various tissues and organs. It interacts with the FGF receptor (FGFR) and activates FGFR signaling pathways, which involve numerous biological functions, such as angiogenesis, wound healing, cell proliferation, and differentiation. ObjectivesHere, we aim to explore the molecular functions, mode of action, and therapeutic activity of yet undetermined function, FGF-2-derived peptide, FP2 (44-ERGVVSIKGV-53) in promoting the proliferation, differentiation, and therapeutic application of human Wharton’s jelly mesenchymal stem cells (hWJ-MSCs) in comparison to other test peptides, canofin1 (FP1), hexafin2 (FP3), and canofin3 (FP4) with known functions. MethodsThe immobilization of test peptides that are fused with mussel adhesive proteins (MAP) on the culture plate was carried out via EDC/NHS chemistry. Cell Proliferation assay, colony-forming unit, western blotting analysis, gene expression analysis, RNA-Seq. analysis, osteogenic, and chondrogenic differentiation capacity were applied to test the activity of the test peptides. We additionally utilized three-dimensional (3D) structural analysis and artificial intelligence (AI)-based AlphaFold2 and CABS-dock programs for receptor interaction prediction of the peptide receptor. We also verified the in vivo therapeutic capacity of FP2-cultured hWJ-MSCs using an osteoarthritis mice model. ResultsCulture of hWJ-MSC onto an FP2-immobilized culture plate showed a significant increase in cell proliferation (n = 3; *p < 0.05, **p < 0.01) and the colony-forming unit (n = 3; *p < 0.05, **p < 0.01) compared with the test peptides. FP2 showed a significantly upregulated phosphorylation of FRS2α and FGFR1 and activated the AKT and ERK signaling pathways (n = 3; *p < 0.05, **p < 0.01, ***p < 0.001). Interestingly, we detected efficient FP2 receptor binding that was predicted using AI-based tools. Treatment with an AKT inhibitor significantly abrogated the FP2-mediated enhancement of cell differentiation (n = 3; *p < 0.05, **p < 0.01, ***p < 0.001). Intra-articular injection of FP2-cultured MSCs significantly mitigated arthritis symptoms in an osteoarthritis mouse model, as shown through the functional tests (n = 10; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001), modulation of the expression level of the pro-inflammatory and anti-inflammatory genes, and improved osteochondral regeneration as demonstrated by tissue sections. ConclusionOur study identified the FGF-2-derived peptide FP2 as a promising candidate peptide to improve the therapeutic potential of hWJ-MSCs, especially in bone and cartilage regeneration.

Ellagic acid loaded nanospheres/biodegradable PVA‐sodium alginate hydrogel for wound healing application

AbstractIn the present study, a novel polyvinyl alcohol‐sodium alginate hydrogel was prepared using ellagic acid loaded nanospheres in its structure. Ellagic acid was loaded into polymeric nanospheres using a double‐emulsion solvent evaporation technique. Obtained nanospheres were dispersed in a polyvinyl alcohol‐sodium alginate hydrogel, prepared using freeze–thaw method followed by freeze‐drying. Ellagic acid loaded nanospheres and the final hydrogel preparation were characterized using scanning electron microscopy, dynamic light scattering, and Fourier transformation infrared spectroscopy, and investigated for ellagic acid content, and release profile. MTT assays were conducted to measure the viability of cells after treatment with the hydrogel, and wound healing activity was investigated through in vivo studies on open wounds of an animal model. Results showed formation of nanospheres with average diameter of 241.5 nm and prolonged‐release profile of ellagic acid during 48 h. MTT assays showed a significant increase in cell proliferation upon treatment of NIH3T3 cells with the hydrogel. The hydrogel also significantly increased the rate of wound closure through improved epithelialization and production of collagen fibers. It can be concluded that the hydrogel containing ellagic acid‐loaded nanospheres prepared in this study can be designated as a biocompatible dressing to accelerate the wound healing process.

Silymarin mediated osmotic responses and damage in HepG2 cell suspensions and monolayers

Maintenance of cells within a volume range compatible with their functional integrity is a critical determinant of cell survival after cryopreservation, and quantifying this osmotically induced damage is a part of the rational design of improved cryopreservation protocols. The degree that cells tolerate osmotic stress significantly impacts applicable cryoprotocols, but there has been little research on the time dependence of this osmotic stress. Additionally, the flavonoid silymarin has been shown to be hepatoprotective. Therefore, here we test the hypotheses that osmotic damage is time-dependent and that flavonoid inclusion reduces osmotic damage. In our first experiment, cells were exposed to a series of anisosmotic solutions of graded hypo- and hypertonicity for 10–40 min, resulting in a conclusion that osmotically induced damage is time dependent. In the next experiment, adherent cells preincubated with silymarin at the concentration of 10−4 mol/L and 10−5 mol/L showed a significant increase in cell proliferation and metabolic activity after osmotic stress compared to untreated matched controls. For instance, when adherent cells preincubated with 10−5 mol/L silymarin were tested, resistance to osmotic damage and a significant increase (15%) in membrane integrity was observed in hypo-osmotic media and a 22% increase in hyperosmotic conditions. Similarly, significant protection from osmotic damage was observed in suspended HepG2 cells in the presence of silymarin. Our study concludes that osmotic damage is time dependent, and the addition of silymarin leads to elevated resistance to osmotic stress and a potential increase in the cryosurvival of HepG2 cells.

Efficient dual crosslinking of protein–in–polysaccharide bioink for biofabrication of cardiac tissue constructs

Myocardial infarction (MI) is a lethal cardiac disease that causes most of the mortality across the world. MI is a consequence of plaque in the arterial walls of heart, which eventually result in occlusion and ischemia to the myocardial tissues due to inadequate nutrient and oxygen supply. As an efficient alternative to the existing treatment strategies for MI, 3D bioprinting has evolved as an advanced tissue fabrication technique where the cell–laden bioinks are printed layer–by–layer to create functional cardiac patches. In this study, a dual crosslinking strategy has been utilized towards 3D bioprinting of myocardial constructs by using a combination of alginate and fibrinogen. Herein, pre-crosslinking of the physically blended alginate–fibrinogen bioinks with CaCl2 enhanced the shape fidelity and printability of the printed structures. Physicochemical properties of the bioinks such as rheology, fibrin distribution, swelling ratio and degradation behaviour, were determined post-printing for only ionically crosslinked & dual crosslinked constructs and found to be ideal for bioprinting of cardiac constructs. Human ventricular cardiomyocytes (AC 16) exhibited a significant increase in cell proliferation on day 7 and 14 in AF–DMEM–20 mM CaCl2 bioink when compared to A–DMEM–20 mM CaCl2 (p < 0.05). Furthermore, myocardial patches containing neonatal ventricular rat myocytes (NVRM) showed >80 % viability and also expressed sarcomeric alpha actinin & connexin 43. These results indicate that the dual crosslinking strategy was cytocompatible and also possess the potential to be used for biofabrication of thick myocardial constructs for regenerative medicine applications.

The Role of ROR1 in Chemoresistance and EMT in Endometrial Cancer Cells.

Background and Objectives: Receptor tyrosine kinase-like orphan receptor type 1 (ROR1) plays a critical role in embryogenesis and is overexpressed in many malignant cells. These characteristics allow ROR1 to be a potential new target for cancer treatment. The aim of this study was to investigate the role of ROR1 through in vitro experiments in endometrial cancer cell lines. Materials and Methods: ROR1 expression was identified in endometrial cancer cell lines using Western blot and RT-qPCR. The effects of ROR1 on cell proliferation, invasion, migration, and epithelial-mesenchymal transition (EMT) markers were analyzed in two endometrial cancer cell lines (HEC-1 and SNU-539) using either ROR1 silencing or overexpression. Additionally, chemoresistance was examined by identifying MDR1 expression and IC50 level of paclitaxel. Results: The ROR1 protein and mRNA were highly expressed in SNU-539 and HEC-1 cells. High ROR1 expression resulted in a significant increase in cell proliferation, migration, and invasion. It also resulted in a change of EMT markers expression, a decrease in E-cadherin expression, and an increase in Snail expression. Moreover, cells with ROR1 overexpression had a higher IC50 of paclitaxel and significantly increased MDR1 expression. Conclusions: These in vitro experiments showed that ROR1 is responsible for EMT and chemoresistance in endometrial cancer cell lines. Targeting ROR1 can inhibit cancer metastasis and may be a potential treatment method for patients with endometrial cancer who exhibit chemoresistance.

Human C1q Tumor Necrosis Factor 8 (CTRP8) defines a novel tryptase+ mast cell subpopulation in the prostate cancer microenvironment

The adipokine C1q Tumor Necrosis Factor 8 (CTRP8) is the least known member of the 15 CTRP proteins and a ligand of the relaxin receptor RXFP1. We previously demonstrated the ability of the CTRP8-RXFP1 interaction to promote motility, matrix invasion, and drug resistance. The lack of specific tools to detect CTRP8 protein severely limits our knowledge on CTRP8 biological functions in normal and tumor tissues. Here, we have generated and characterized the first specific antiserum to human CTRP8 which identified CTRP8 as a novel marker of tryptase+ mast cells (MCT) in normal human tissues and in the prostate cancer (PC) microenvironment. Using human PC tissue microarrays composed of neoplastic and corresponding tumor-adjacent prostate tissues, we have identified a significantly higher number of CTRP8+ MCT in the peritumor versus intratumor compartment of PC tissues of Gleason scores 6 and 7. Higher numbers of CTRP8+ MCT correlated with the clinical parameter of biochemical recurrence. We showed that the human MC line ROSAKIT WT expressed RXFP1 transcripts and responded to CTRP8 treatment with a small but significant increase in cell proliferation. Like the cognate RXFP1 ligand RLN-2 and the small molecule RXFP1 agonist ML-290, CTRP8 reduced degranulation of ROSAKIT WT MC stimulated by the Ca2+-ionophore A14187. In conclusion, this is the first report to identify the RXFP1 agonist CTRP8 as a novel marker of MCT and autocrine/paracrine oncogenic factor within the PC microenvironment.

MicroRNA-370-5p inhibits pigmentation and cell proliferation by downregulating mitogen-activated protein kinase kinase kinase 8 expression in sheep melanocytes

In mammals, microRNAs (miRNAs) play key roles in multiple biological processes by regulating the expression of target genes. Studies have found that the levels of miR-370-5p expression differ significantly in the skins of sheep with different hair colors; however, its function remains unclear. In this study, we investigated the roles of miR-370-5p in sheep melanocytes and found that the overexpression of miR-370-5p significantly inhibited cell proliferation (P<0.01), tyrosinase activity (P=0.001) and significantly reduced (P<0.001) melanin production. Functional prediction revealed that the 3´-untranslated region (UTR) of MAP3K8 has a putative miR-370-5p binding site, and the interaction between these two molecules was confirmed using luciferase reporter assays. In situ hybridization assays revealed that MAP3K8 is expressed in the cytoplasm of melanocytes. The results of quantitative RT-PCR and Western blotting analyses revealed that overexpression of miR-370-5p in melanocytes significantly inhibits (P<0.01) MAP3K8 expression via direct targeting of its 3´ UTR. Inhibition of MAP3K8 expression by siRNA-MAP3K8 transfection induced a significant inhibition (P<0.01) of melanocyte proliferation and significant reduction (P<0.001) in melanin production, which is consistent with our observations for miR-370-5p. Target gene rescue experiments indicated that the expression of MAP3K8 in melanocytes co-transfected with miR-370-5p and MAP3K8-cDNA (containing sites for the targeted binding to miR-370-5p) was significantly rescued (P≤0.001), which subsequently promoted significant increases in cell proliferation (P<0.001) and melanin production (P<0.01). Collectively, these findings indicate that miR-370-5p plays a functional role in inhibiting sheep melanocyte proliferation and melanogenesis by downregulating the expression of MAP3K8.

Extracellular Vesicles Released from Cancer Cells Promote Tumorigenesis by Inducing Epithelial to Mesenchymal Transition via β-Catenin Signaling

Lung cancer is the leading cause of cancer-related deaths globally, in part due to a lack of early diagnostic tools and effective pharmacological interventions. Extracellular vesicles (EVs) are lipid-based membrane-bound particles released from all living cells in both physiological and pathological states. To understand the effects of lung-cancer-derived EVs on healthy cells, we isolated and characterized EVs derived from A549 lung adenocarcinoma cells and transferred them to healthy human bronchial epithelial cells (16HBe14o). We found that A549-derived EVs carry oncogenic proteins involved in the pathway of epithelial to mesenchymal transition (EMT) that are regulated by β-catenin. The exposure of 16HBe14o cells to A549-derived EVs resulted in a significant increase in cell proliferation, migration, and invasion via upregulating EMT markers such as E-Cadherin, Snail, and Vimentin and cell adhesion molecules such as CEACAM-5, ICAM-1, and VCAM-1, with concomitant downregulation of EpCAM. Our study suggests a role for cancer-cell-derived EVs to induce tumorigenesis in adjacent healthy cells by promoting EMT via β-catenin signaling.

Alteration of membrane potential of head and neck cancer cells using a piezoelectric nanofiber interface.

Membrane potential (Vmem) is an electrical property associated with any cell, specific to its origin and function. These transmembrane voltage gradients have been established to control not only neural signaling via gap junctions, but also cell proliferation, migration, differentiation, and orientation in both excitable and non-excitable cells. The bioelectric state of cancer cells is different from that of healthy cells, causing a disruption in cell signaling pathways, affecting all three processes of carcinogenesis - initiation, promotion, and progression. More positively charged or depolarized Vmem correspond to more undifferentiated, proliferative, and stem-like cells. Understanding of the underlying bioelectric mechanisms and their disruption in cancer can be advanced by developing molecular tools capable of measuring and altering Vmem. Current gold standards include bio actuators, patch clamps, conductive polymers, and treatment with pharmacological drugs such as ivermectin. Biocompatible piezoelectric (PE) materials can alter the Vmem of cells by contact, without the use of transgenes. Here we introduce a PE polyvinylidene fluoride-trifluoroethylene (PVDF-TrFE) biomaterial interface capable of generating quantified electrical stimuli in response to mechanical deformation, thus altering resting Vmem of cells. PVDF-TrFE nanofibers seeded with Cal27 - oral squamous carcinoma cells (OSCC) were subjected to 1 cm mechanical stretch deformations at a frequency of 1 Hz for 3 minutes every 24 h. Results indicated significant increase in cell proliferation for treatment groups on days 3, 5, and 7 of culture as compared to controls. This platform can be further evolved to alter Vmem of cancer spheroids in 3D to better mimic the bioelectric state of cells in the tumor microenvironment (TME) and study subsequent alterations in signaling pathways.

Anthocyanin/Honey-Incorporated Alginate Hydrogel as a Bio-Based pH-Responsive/Antibacterial/Antioxidant Wound Dressing.

Infection is a major problem that increases the normal pH of the wound bed and interferes with wound healing. Natural biomaterials can serve as a suitable environment to acquire a great practical effect on the healing process. In this context, anthocyanin-rich red cabbage (Brassica oleracea var. capitata F. rubra) extract and honey-loaded alginate hydrogel was fabricated using calcium chloride as a crosslinking agent. The pH sensitivity of anthocyanins can be used as an indicator to monitor possible infection of the wound, while honey would promote the healing process by its intrinsic properties. The mechanical properties of the hydrogel film samples showed that honey acts as a plasticizer and that increasing the incorporation from 200% to 400% enhances the tensile strength from 3.22 to 6.15 MPa and elongation at break from 0.69% to 4.75%. Moreover, a water absorption and retention study showed that the hydrogel film is able to absorb about 250% water after 50 min and retain 40% of its absorbed water after 12 h. The disk diffusion test showed favorable antibacterial activity of the honey-loaded hydrogel against both Gram-positive and Gram-negative Staphylococcus aureus and Escherichia coli, respectively. In addition, the incorporation of honey significantly improved the mechanical properties of the hydrogel. 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay proved the antioxidant activity of the honey and anthocyanin-containing hydrogel samples with more than 95% DPPH scavenging efficiency after 3 h. The pH-dependent property of the samples was investigated and recorded by observing the color change at different pH values of 4, 7, and 9 using different buffers. The result revealed a promising color change from red at pH = 4 to blue at pH = 7 and purple at pH = 9. An in vitro cell culture study of the samples using L929 mouse fibroblast cells showed excellent biocompatibility with significant increase in cell proliferation. Overall, this study provides a promising start and an antibacterial/antioxidant hydrogel with great potential to meet wound-dressing requirements.