Effect of Ylang-Ylang (Cananga odorata Hook. F. & Thomson) Essential Oil on Acute Inflammatory Response In Vitro and In Vivo

The aim of this study is to evaluate the phytochemical profile, oral acute toxicity, and the effect of ylang-ylang (Cananga odorata Hook. F. & Thomson) essential oil (YEO) on acute inflammation. YEO was analyzed by gas chromatography/mass spectrometry. For in vitro tests, YEO was assessed using cytotoxicity, neutrophil chemotaxis induced by N-formyl methionyl leucyl phenylalanine (fMLP), and phagocytic activity tests. YEO was orally administered in zymosan-induced peritonitis, carrageenan-induced leukocyte rolling, and adhesion events in the in situ microcirculation model and in carrageenan-induced paw edema models. YEO (2000 mg/kg) was also tested using an acute toxicity test in Swiss mice. YEO showed a predominance of benzyl acetate, linalool, benzyl benzoate, and methyl benzoate. YEO did not present in vitro cytotoxicity. YEO reduced the in vitro neutrophil chemotaxis induced by fMLP and reduced the phagocytic activity. The oral treatment with YEO reduced the leukocyte recruitment and nitric oxide production in the zymosan-induced peritonitis model, reduced rolling and adherent leukocyte number induced by carrageenan in the in situ microcirculation model, and reduced carrageenan-induced edema and mechanical hyperalgesia. YEO did not present signs of toxicity in the acute toxicity test. In conclusion, YEO affected the leukocyte activation, and presented antiedematogenic, anti-hyperalgesic, and anti-inflammatory properties.     

The Assessment of Meloxicam Phototoxicity in Human Normal Skin Cells: In Vitro Studies on Dermal Fibroblasts and Epidermal Melanocytes

Meloxicam (MLX), which belongs to the oxicam nonsteroidal anti-inflammatory drug derivatives, is an inhibitor of the cyclooxygenase-2 (COX-2) enzyme. Cutaneous adverse effects caused by interaction between UVA radiation and exogenous factors can manifest as phototoxic reactions. Phototoxicity may be a reason for the accumulation of genetic and molecular changes in long-lived cells with low proliferation potential, leading to tumor development. There are several potentially phototoxic drugs, the active component of which is meloxicam. The research aimed to evaluate the influence of MLX and UVAR on skin cells—fibroblasts and melanocytes homeostasis. The obtained results indicated that co-treatment with MLX and UVAR inhibited skin cell proliferation, proportionally to the drug concentration. The observation was confirmed by cytometric analysis of the cell number and viability. The phototoxic effect of MLX was revealed in morphological changes. It was stated that MLX with UVAR lowered the mitochondrial transmembrane potential and changed the cell cycle profile. Additionally, MLX and UVAR caused the disruption of redox homeostasis by lowering the intracellular level of reduced thiols. The presented study revealed that the phototoxic activity of MLX is associated with oxidative stress induction and disruptions in cell homeostasis. The differences in the phototoxic effects of MLX at the cellular level may be related to the different content of melanin pigments.     

In Vitro Assessment of the Cytotoxic and Antiproliferative Profile of Natural Preparations Containing Bergamot,Orange and Clove Essential Oils

Medicinal plants and essential oils (EOs), in particular, were intensively studied in recent years as viable alternatives for antiproliferative chemical synthetic agents. In the same lines, the present study focuses on investigating the effects of natural preparations (emulsions) based on EOs obtained from Citrus bergamia Risso (bergamot-BEO), Citrus sinensis Osbeck (orange-OEO), and Syzygium aromaticum Merill et L. M. Perry (clove-CEO) on different healthy (human immortalized keratinocytes—HaCaT and primary human gingival fibroblasts—HGF) and human tumor cell lines (human melanoma—A375 and oral squamous carcinoma—SCC-4) in terms of the cells’ viability and cellular morphology. The obtained results indicate that the CEO emulsion (ECEO) induced a dose-dependent cytotoxic in both healthy (HaCaT and HGF) and tumor (A375 and SCC-4) cells. OEO emulsion (EOEO) increased cell viability percentage both for HaCaT and A375 cells and had an antiproliferative effect at the highest concentration in HGF and SCC-4 cells. BEO emulsion (EBEO) decreased the viability percentage of SCC-4 tumor cells. By associating OEO with CEO as a binary mixture in an emulsified formulation, the inhibition of tumor cell viability increases. The E(BEO/OEO) binary emulsion induced an antiproliferative effect on oral health and tumor cells, with a minimal effect on skin cells. The non-invasive tests performed to verify the safety of the test compound’s emulsions at skin level indicated that these compounds do not significantly modify the physiological skin parameters and can be considered safe for human skin.     

In Vitro and Human Pilot Studies of Different Topical Formulations Containing Rosa Species for the Treatment of Psoriasis

The aim of this study was to evaluate the phytochemical profile and antioxidant properties of the extracts from three Rosa species (R. canina, R. damascena, R. cairo), to develop and investigate topical formulations with lyophilized forms of extracts for the treatment of psoriasis. Phytochemical screening and in vitro total antioxidant capacity (DPPH, FRAP, CUPRAC, SOD) of studied samples were examined and compared. Lyophilized extracts of roses were dissolved in Transcutol HP and different formulations of creams were prepared. Franz diffusion method was used to evaluate the drug release and biocompatibility was tested on HaCaT cells. Rosa damascene had the best results regarding all the analyses that were conducted. After the evaluation of topical products, the formulation with Rosa damascena extract in a self-emulsifying drug delivery system was tested on a human clinical study that involved 20 patients. At the end of the clinical study an improvement in the quality of life of the patients was observed and erythema, induration and scaling were reduced. The present study indicates that our examined extracts exhibited great phenolic content, antioxidant capacity and safety profile of topical formulation and therefore can be used as a reliable source of natural antioxidants and may be used as a complementary treatment to improve the quality life of patients with psoriasis or may be tested on another diseases.     

Influence of In Vitro Human Digestion Simulation on the Phenolics Contents and Biological Activities of the Aqueous Extracts from Turkish Cistus Species

Oxidative stress is one of the significant precursors of various metabolic diseases such as diabetes, Parkinson’s disease, cardiovascular diseases, cancer, etc. Various scientific reports have indicated that secondary plant metabolites play an important role in preventing oxidative stress and its harmful effects. In this respect, this study was planned to investigate the phenolic profile and antioxidant and antidiabetic potentials of the aqueous extracts from Turkish Cistus species by employing in vitro methods. In vitro digestion simulation procedure was applied to all extracts to estimate the bioavailability of their phenolic contents. Total phenolic, flavonoid, phenolic acid and proanthocyanidin contents were determined for all phases of digestion. In addition, changes in the quantity of the assigned marker flavonoids (tiliroside, hyperoside and quercitrin) were monitored by High-Performance Thin Layer Chromatography (HPTLC) analysis. The antioxidant activity potentials of the extracts were studied by various methods to reveal their detailed activity profiles. On the other hand, in vitro α-amylase and α-glucosidase enzymes and advanced-glycation end product (AGE) inhibitory activities of the extracts were determined to evaluate the antidiabetic potentials of extracts. The results showed that aqueous extracts obtained from the aerial parts of Turkish Cistus species have rich phenolic contents and potential antioxidant and antidiabetic activities; however, their bioactivity profiles and marker flavonoid concentrations might significantly be affected by human digestion. The results exhibited that total phenolic contents, antioxidant activities and diabetes-related enzyme inhibitions of the bioavailable samples were lower than non-digested samples in all extracts.     

Impact of Different [Tc(N)PNP]-Scaffolds on the Biological Properties of the Small cRGDfK Peptide: Synthesis,In Vitro and In Vivo Evaluations

Background: The [^99mTc][Tc(N)(PNP)] system, where PNP is a bisphosphinoamine, is an interesting platform for the development of tumor ‘receptor-specific’ agents. Here, we compared the reactivity and impact of three [Tc(N)(PNP)] frameworks on the stability, receptor targeting properties, biodistribution, and metabolism of the corresponding [^99mTc][Tc(N)(PNP)]-tagged cRGDfK peptide to determine the best performing agent and to select the framework useful for the preparation of [^99mTc][Tc(N)(PNP)]-housing molecular targeting agents. Methods: cRGDfK pentapeptide was conjugated to Cys and labeled with each [Tc(N)(PNP)] framework. Radioconjugates were assessed for their lipophilicity, stability, in vitro and in vivo targeting properties, and performance. Results: All compounds were equally synthetically accessible and easy to purify (RCY ≥ 95%). The main influences of the synthon on the targeting peptide were observed in in vitro cell binding and in vivo. Conclusions: The variation in the substituents on the phosphorus atoms of the PNP enables a fine tuning of the biological features of the radioconjugates. ws[^99mTc][Tc(N)(PNP3OH)]– and [^99mTc][Tc(N)(PNP3)]– are better performing synthons in terms of labeling efficiency and in vivo performance than the [^99mTc][Tc(N)(PNP43)] framework and are therefore more suitable for further radiopharmaceutical purposes. Furthermore, the good labeling properties of the ws[^99mTc][Tc(N)(PNP3OH)]– framework can be exploited to extend this technology to the labeling of temperature-sensitive biomolecules suitable for SPECT imaging.     

Rosmarinus officinalis L. Leaf Extracts and Their Metabolites Inhibit the Aryl Hydrocarbon Receptor (AhR) Activation In Vitro and in Human Keratinocytes: Potential Impact on Inflammatory Skin Diseases and Skin Cancer

Aryl hydrocarbon receptor (AhR) activation by environmental agents and microbial metabolites is potentially implicated in a series of skin diseases. Hence, it would be very important to identify natural compounds that could inhibit the AhR activation by ligands of microbial origin as 6-formylindolo[3,2-b]carbazole (FICZ), indirubin (IND) and pityriazepin (PZ) or the prototype ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Five different dry Rosmarinus officinalis L. extracts (ROEs) were assayed for their activities as antagonists of AhR ligand binding with guinea pig cytosol in the presence of [³H]TCDD. The methanolic ROE was further assayed towards CYP1A1 mRNA induction using RT-PCR in human keratinocytes against TCDD, FICZ, PZ, and IND. The isolated metabolites, carnosic acid, carnosol, 7-O-methyl-epi-rosmanol, 4′,7-O-dimethylapigenin, and betulinic acid, were assayed for their agonist and antagonist activity in the presence and absence of TCDD using the gel retardation assay (GRA). All assayed ROE extracts showed similar dose-dependent activities with almost complete inhibition of AhR activation by TCDD at 100 ppm. The methanol ROE at 10 ppm showed 99%, 50%, 90%, and 85% inhibition against TCDD, FICZ, IND, and PZ, respectively, in human keratinocytes. Most assayed metabolites exhibited dose-dependent antagonist activity. ROEs inhibit AhR activation by TCDD and by the Malassezia metabolites FICZ, PZ, and IND. Hence, ROE could be useful for the prevention or treatment of skin diseases mediated by activation of AhR.     

The Analytical Strategy of “Ion Induction and Deduction Based on Net-Hubs” for the Comprehensive Characterization of Naringenin Metabolites In Vivo and In Vitro Using a UHPLC-Q-Exactive Orbitrap Mass Spectrometer

Naringenin (5,7,4′-trihydroxyflavanone), belonging to the flavanone subclass, is associated with beneficial effects such as anti-oxidation, anticancer, anti-inflammatory, and anti-diabetic effects. Drug metabolism plays an essential role in drug discovery and clinical safety. However, due to the interference of numerous endogenous substances in metabolic samples, the identification and efficient characterization of drug metabolites are difficult. Here, ultra-high-performance liquid chromatography (UHPLC) coupled with high-resolution mass spectrometry was used to obtain mass spectral information of plasma (processed by three methods), urine, feces, liver tissue, and liver microsome samples. Moreover, a novel analytical strategy named “ion induction and deduction” was proposed to systematically screen and identify naringenin metabolites in vivo and in vitro. The analysis strategy was accomplished by the establishment of multiple “net-hubs” and the induction and deduction of fragmentation behavior. Finally, 78 naringenin metabolites were detected and identified from samples of rat plasma, urine, feces, liver tissue, and liver microsomes, of which 67 were detected in vivo and 13 were detected in vitro. Naringenin primarily underwent glucuronidation, sulfation, oxidation, methylation, ring fission, and conversion into phenolic acid and their composite reactions. The current study provides significant help in extracting target information from complex samples and sets the foundation for other pharmacology and toxicology research.     

The Lipophilic Purine Nucleoside—Tdp1 Inhibitor—Enhances DNA Damage Induced by Topotecan In Vitro and Potentiates the Antitumor Effect of Topotecan In Vivo

The use of cancer chemotherapy sensitizers is a promising approach to induce the effect of clinically used anticancer treatments. One of the interesting targets is Tyrosyl-DNA Phosphodiesterase 1 (Tdp1), a DNA-repair enzyme, that may prevent the action of clinical Topoisomerase 1 (Top1) inhibitors, such as topotecan (Tpc). Tdp1 eliminates covalent Top1-DNA (Top1c) complexes that appear under the action of topotecan and determines the cytotoxic effect of this drug. We hypothesize that Tdp1 inhibition would sensitize cells towards the effect of Tpc. Herein, we report the synthesis and study of lipophilic derivatives of purine nucleosides that efficiently suppress Tdp1 activity, with IC₅₀ values in the 0.3–22.0 μM range. We also showed that this compound class can enhance DNA damage induced by topotecan in vitro by Comet assay on human cell lines HeLa and potentiate the antitumor effect of topotecan in vivo on a mice ascitic Krebs-2 carcinoma model. Thereby, this type of compound may be useful to develop drugs, that sensitize the effect of topotecan and reduce the required dose and, as a result, side effects.     

In Vitro Evaluation of the Cytotoxic Effect of Streptococcus pyogenes Strains,Protegrin PG-1, Cathelicidin LL-37, Nerve Growth Factor and Chemotherapy on the C6 Glioma Cell Line

Brain cancer treatment, where glioblastoma represents up to 50% of all CNS malignancies, is one of the most challenging calls for neurooncologists. The major driver of this study was a search for new approaches for the treatment of glioblastoma. We tested live S. pyogenes, cathelicidin family peptides and NGF, assessing the oncolytic activity of these compounds as monotherapy or in combination with chemotherapeutics. For cytotoxicity evaluation, we used the MTT assay, trypan blue assay and the xCELLigence system. To evaluate the safety of the studied therapeutic approaches, we performed experiments on normal human fibroblasts. Streptococci and peptides demonstrated high antitumor efficiency against glioma C6 cells in all assays applied, surpassing the effect of chemotherapeutics (doxorubicin, carboplatin, cisplatin, etoposide). A real-time cytotoxicity analysis showed that the cell viability index dropped to 21% 2–5 h after S. pyogenes strain exposure. It was shown that LL-37, PG-1 and NGF also exhibited strong antitumor effects on C6 glioma cells when applied at less than 10⁻⁴ M. Synergistic effects for combinations of PG-1 with carboplatin and LL-37 with etoposide were shown. Combinations of S. pyogenes strain #7 with NGF or LL-37 demonstrated a cytotoxic effect (56.7% and 57.3%, accordingly) on C6 glioma cells after 3 h of exposure.     

Aligned Poly(ε‐caprolactone) Nanofibers Guide the Orientation and Migration of Human Pluripotent Stem Cell‐Derived Neurons,Astrocytes, and Oligodendrocyte Precursor Cells In Vitro

Stem cell transplantations for spinal cord injury (SCI) have been studied extensively for the past decade in order to replace the damaged tissue with human pluripotent stem cell (hPSC)‐derived neural cells. Transplanted cells may, however, benefit from supporting and guiding structures or scaffolds in order to remain viable and integrate into the host tissue. Biomaterials can be used as supporting scaffolds, as they mimic the characteristics of the natural cellular environment. In this study, hPSC‐derived neurons, astrocytes, and oligodendrocyte precursor cells (OPCs) are cultured on aligned poly(ε‐caprolactone) nanofiber platforms, which guide cell orientation to resemble that of spinal cord in vivo. All cell types are shown to efficiently spread over the nanofiber platform and orient according to the fiber alignment. Human neurons and astrocytes require extracellular matrix molecule coating for the nanofibers, but OPCs grow on nanofibers without additional treatment. Furthermore, the nanofiber platform is combined with a 3D hydrogel scaffold with controlled thickness, and nanofiber‐mediated orientation of hPSC‐derived neurons is also demonstrated in a 3D environment. In this work, clinically relevant materials and substrates for nanofibers, fiber coatings, and hydrogel scaffolds are used and combined with cells suitable for developing functional cell grafts for SCI repair.