Anticancer activity of Inula graveolens by induction of ROS-independent apoptosis and suppression of IL6-IL8 in cervical cancer cells / Actividad anticancerígena de Inula graveolens por inducción de apoptosis independiente de ROS y supresión de IL6-IL8 en células de cáncer de cuello uterino

Cancer is an important disease that causing to deaths in the world. Cervical cancer is one of the most common among women and must be treated quickly to prevent cell proliferation. Natural products have been used for cancer treatment due to their antiproliferative and apoptosis induction properties. In the present study, we aimed to determine the antiproliferative and apoptosis induction activities of methanol extract of Inula graveolens (IGME) in the human cerival cancer cell line (HeLa). Antiproliferative activity of IGME was evaluated by using MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) method. Apoptosis induction activity was determined by AnnexinV/propodium iodide staining with FACS. ROS induction was determined by DCFH-DA (2',7'-Dichlorofluorescin diacetate) staining. Interleukin 6 and 8 levels (IL-6 and IL-8) were determined by the Elisa method. IGME exhibited antiproliferative effect and induced apoptosis in the HeLa cells. IL-6, IL-8 and intracellular ROS levels were decreased after treatment of IGME. Consequently, IGME was found to have antiproliferative and apoptosis induction activities as an indicator of the anticancer activity.

El cáncer es una enfermedad importante que causa muertes en el mundo. El cáncer de cuello uterino es uno de los más comunes entre las mujeres y debe tratarse rápidamente para evitarla proliferación celular. Los productos naturales se han utilizado para el tratamiento del cáncer debido a sus propiedades anti proliferativas y de inducción de la apoptosis. En el presente estudio, nuestro objetivo fue determinar las actividades de inducción de apoptosis y antiproliferativas del extracto de metanol de Inula graveolens (IGME) en la línea celular de cáncer de cuello uterino humano (HeLa). La actividad antiproliferativa de IGME se evaluó utilizando el método MTT (bromuro de 3-[4,5-dimetiltiazol-2-il]-2,5-difenil-tetrazolio). La actividad de inducción de apoptosis se determinó mediante tinción con anexina V/yoduro de propodio con FACS. La inducción de ROS se determinó mediante tinción con DCFH-DA (diacetato de 2',7'-diclorofluoresceina). Los niveles de interleucina 6 y 8 (IL-6 e IL-8) se determinaron por el método Elisa. IGME exhibió un efecto antiproliferativo e indujo apoptosis en las células HeLa. Los niveles de IL-6, IL-8 y ROS intracelulares disminuyeron después del tratamiento con IGME. En consecuencia, se encontró que IGME tenía actividades antiproliferativas y de inducción de apoptosis como indicador de la actividad anticancerígena.

Wound healing activity of Salvia huberi ethanolic extract in streptozocin-induced diabetic rats.

Objective: The aim of this study was to examine the in vivo wound healing potential of Salvia huberi Hedge (endemic to Turkey) on excision and incision wound models in diabetic rats. Method: Male Wistar albino rats, 3-4 months old and weighing 180-240g were used. The animals were randomly divided into five groups including Control, Vehicle and Fito reference, and two different concentrations (0.5% and 1% weight/weight (w/w)) of ethanol extract of Salvia huberi were investigated in both wound models on streptozocin-induced diabetic rats using macroscopic, biomechanical, biochemical, histopathological, genotoxic and gene expression methods over both seven and 14 days. Fito cream (Tripharma Drug Industry and Trade Inc., Turkey) was used as the reference drug. Results: A total of 60 rats were used in this study. Salvia huberi ointments at 0.5% and 1% (w/w) concentrations and Fito cream showed 99.3%, 99.4% and 99.1% contraction for excision wounds, and 99.9%, 97.0% and 99% contraction for incision wounds, respectively. In Salvia huberi ointments and Fito cream groups, re-epithelialisation increased dramatically by both day 7 and day 14 (p<0.05). By day 14, low hydroxyproline and malondialdehyde (MDA) levels, and high glutathione (GSH) levels were observed in the Salvia huberi ointment groups. After two application periods, damaged cell percent and genetic damage index values and micronucleus frequency of Salvia huberi ointment treatment groups were lower than Control and Vehicle groups (p<0.001). A growth factor expression reached a high level by day 7 in the Control group; in Salvia huberi-treated groups it was decreased. Conclusion: The study showed that application of Salvia huberi ointments ameliorated the healing process in diabetic rats with excisional and incisional wounds and may serve as a potent healing agent.

Extract of Glycyrrhiza glabra root attenuates nociception in experimental pain models: the role of BKCa channels / Extracto de raíz de Glycyrrhiza glabra atenúa la nocicepción en modelos experimentales de dolor: el rol de los canales BKCa

Abstract: The aim of this study was to evaluate the functional interaction of Glycyrrhiza glabra root extract (GGRE) on the large conductance Ca 2+ - activated K + (BKCa) channels expressed in the peripheral nervo us system by using nociception and inflammation models in rodents in vivo . Besides toxicity studies and open field tests, nociception and inflammation tests were performed on rodents. Different doses of GGRE were given orally to rats and mice. Naloxone, in domethacin, morphine, NS1619 and iberiotoxin (IbTX) were administered. GGRE had both anti - nociceptive and anti - inflammatory activity in rats and mice. GGRE exhibited an analgesic effect by decreasing the time - course of the pain threshold or reaction time i n some nociceptive tests. Furthermore, GGRE reduced level of pro - inflammatory cytokines, including TNF - α and IL - 1ß. As a conclusion, GGRE can alleviate the pain sensation of the afferent nerves and can reduce inflammation and associated pain by activating B KCa channels and reducing the levels of TNF - α, IL1ß

Resumen: El objetivo de este estudio fue evaluar la interacción funcional del extracto de raíz de Glycyrr hiza glabra (GGRE) en los canales de K + (BKCa) activados por Ca 2+ de gran conductancia expresados en el sistema nervioso periférico mediante el uso de modelos de nocicepción e inflamación en roedores in vivo . Además de los estudios de toxicidad y las prueb as de campo abierto, se realizaron pruebas de nocicepción e inflamación en roedores. Se administraron por vía oral diferentes dosis de GGRE a ratas y ratones. Se administraron naloxona, indometacina, morfina, NS1619 e iberiotoxina (IbTX). GGRE tenía activi dad tanto antinociceptiva como antiinflamatoria en ratas y ratones. GGRE mostró un efecto analgésico al disminuir la evolución temporal del umbral del dolor o el tiempo de reacción en algunas pruebas nociceptivas. Además, GGRE redujo el nivel de citocinas proinflamatorias, incluidas TNF - α e IL - 1ß. Como conclusión, GGRE puede aliviar la sensación de dolor de los nervios aferentes y puede reducir la inflamación y el dolor asociado activando los canales BKCa y reduciendo los niveles de TNF - α, IL1ß.

Sideritis perfoliata inhibits cell proliferation, induces apoptosis and exhibits cellular antioxidant activity in cervical cancer cells / Sideritis perfoliata inhibe la proliferación celular, induce apoptosis y exhibe actividad antioxidante celular en células de cáncer de cuello uterino

In this study, it was aimed to determine the antioxidant and anticancer activities of Sideritis perfoliata methanolic extract (SPE) on cervical cancer cells (HeLa). Different doses (25, 50,100 and 200 µg/mL) of SPE were used to determine proliferation of HeLa cells by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) staining method. Induction of apoptosis was determined by Annexine-V and propidium iodide staining method. Interleukin (IL) 6-8 levels were measured by ELISA method. Antioxidant activities of SPE were determined by DPPH, DNA (plasmid pBR322) protecting and cellular antioxidant activity tests. Some phytochemicals of SPE were also screened by LC-MS-MS. It was determined that SPE reduced the proliferation of HeLa cells and also induced apoptosis. IL6-8 levels importantly decreased at 200 µg/mL. SPE exhibited moderately antioxidant activities in tests used. Among the phenolics identified, vanillic acid had the highest amount. As a result, it was determined to have the anticancer activity of SPE by decreasing cell proliferation, inducing apoptosis and decreasing IL6-8 in HeLa cells.

En este estudio, se tuvo como objetivo determinar las actividades antioxidantes y anticancerígenas del extracto metanólico de Sideritis perfoliata (SPE) en las células de cáncer de cuello uterino (HeLa). Se utilizaron diferentes dosis (25, 50, 100 y 200 µg/mL) de SPE para determinar la proliferación de células HeLa mediante el método de tinción con bromuro de 3-[4,5-dimetiltiazol-2-il] -2,5-difenil-tetrazolio (MTT). La inducción de apoptosis se determinó mediante el método de tinción con anexina-V y yoduro de propidio. Los niveles de interleucina (IL) 6-8 se midieron mediante el método ELISA. Las actividades antioxidantes de SPE se determinaron mediante pruebas de DPPH, protección de ADN (plásmido pBR322) y actividad antioxidante celular. Algunos fitoquímicos de SPE también se analizaron mediante LC-MS-MS. Se determinó que SPE redujo la proliferación de células HeLa y también indujo apoptosis. Los niveles de IL6-8 disminuyeron de manera importante a 200 µg/mL. SPE mostró actividades moderadamente antioxidantes en las pruebas utilizadas. Entre los fenólicos identificados, el ácido vainílico tuvo la mayor cantidad. Como resultado, se determinó que tenía la actividad anticancerígena de SPE al disminuir la proliferación celular, inducir apoptosis y disminuir la IL6-8 en las células HeLa.

In vitro scolicidal effects of Sideritis perfoliata extract against Echinococcus granulosus.

BACKGROUND: Cystic echinococcosis, caused by helminths within the genus Echinococcus, is mainly localised in the liver and lungs of affected hosts. Surgery has been the best choice for the treatment of hydatidosis and using effective scolicidal agents during hydatid surgery is required to prevent secondary infection. Several plant extracts have been shown to exert scolicidal efficacy. This study was designed to investigate the in vitro scolicidal activity of methanol extract of Sideritis perfoliata against the protoscolices of hydatid cysts. METHODS: The protoscolices were collected from a liver of a sheep slaughtered in Adiyaman city slaughter, Turkey. Three concentrations of the aerial part extract of S perfoliata (0.1, 0.2 and 0.4 mg/mL) were assessed at three different exposure periods. All tests were carried in duplicate. The viability of protoscolices was assessed by the eosin exclusion test (0.1% eosin staining). RESULTS: Scolicidal effect of S perfoliata extract at exposure periods of 10, 20 and 30 minutes was 29.6%, 32.5% and 43.6% at the concentration of 0.1%, 37.8%, 50% and 58.1% at concentration of 0.2 mg/mL, and 57.9%, 71.8% and 79.1% at the concentration of 0.4 mg/mL, respectively; indicating a longer time is required to display protoscolicidal effects. LC-MS/MS analysis showed that some phenolic acids, such as fumaric acid (260.13 mg/L), syringic acid (27.92 mg/L) and caffeic acid (26.84 mg/L), and a flavonoid, luteolin (11.23 mg/L) were detected in high concentrations in the extract. CONCLUSIONS: This study has demonstrated that the methanol extract of S perfoliata has high scolicidal activity in vitro. However, research on the in vivo efficacy of S perfoliata extract and its potential side effects is required.

Biochemical, Histopathologic, and Genotoxic Effects of Ethanol Extract of Salvia hypargeia (Fisch. & Mey.) on Incisional and Excisional Wounded Diabetic Rats.

Purpose: Nonhealing wounds are a serious problem of diabetic patients. Salvia species are traditionally used for the treatment of wounds. The aim of the study was to investigate the effects of ointment prepared with ethanol extract obtained from the aerial parts of Salvia hypargeia, an endemic plant from Turkey, on diabetic rat incisional and excisional skin wounds. Materials and Methods: Male Wistar albino rats (n: 60) were divided into five groups. Diabetes was induced and two concentrations (0.5% and 1%) of the extract were used for ointments and applied on wounds for 7 and 14 days. Fito cream was chosen as a reference drug. Results: In excisional wounds, healing ratios of 0.5% (63.4% and 99.3%) and 1% (65.5% and 99.9%) S. hypargeia groups were higher compared to control (35.9% and 75.1%), and in incisional wounds, healing ratios of 0.5% (78.1% and 98.5%) and 1% (84.4% and 99.4%) S. hypargeia groups were higher compared to control (30.5% and 72.9%) (p < .01). Hydroxyproline (0.31 ± 0.3 and 0.34 ± 0.2) levels were lower and GSH (10.7 ± 3.1 and 7.6 ± 0.9) levels were higher in 0.5% and 1% S. hypargeia groups on the 14th day (p < .01). Histopathological results revealed re-epithelialization and formation of granulation tissue in all S. hypargeia groups. Genotoxicologic results indicated, GDI, DCP values, and MN frequency of 0.5% and 1% S. hypargeia groups did not reach to significant levels both on the 7 and 14 days. Conclusions: S. hypargeia may have a potential for therapeutic use in treatment and management of diabetic wounds with a successful topical application.

Pharmacological properties and therapeutic potential of saffron (Crocus sativus L.) in osteosarcoma.

OBJECTIVES: In this comprehensive study, we aimed to investigate pharmacological properties and therapeutic significance of saffron in osteosarcoma cancer cells. METHODS: Plant materials were obtained from Safranbolu district of Karabuk, Turkey. For the determination of anticancer properties, thiazolyl blue tetrazolium bromide (MTT) cell viability, colony formation, wound closure, DNA ladder assays and gene expression analysis by real-time PCR were performed. Also, cellular inflammation, total antioxidant and oxidants status were determined. KEY FINDINGS: Dichloromethane and hexane extracts of saffron were significantly inhibited cell proliferation and interfered with colony forming and migration capabilities of U2-OS osteosarcoma cancer cells. Also, both extracts induced the activation of tumour suppressor CDKN2B gene and altered cellular morphology resembling the induction of apoptosis. However, DNA fragmentation was not observed after extract treatments. Saffron was also found to have no significant effect on cellular inflammation. Unexpectedly, both dichloromethane and hexane extracts of saffron had no marked effect on cellular total antioxidant and oxidant status. Lastly, vanillic acid, resveratrol, caffeic acid and 4-hydroxybenzoic acid were found to be highly rich in our extracts. CONCLUSIONS: Findings of this study demonstrated significant antiproliferative and antitumorigenic properties of saffron in osteosarcoma.

Hepatoprotective properties for Salvia cryptantha extract on carbon tetrachloride-induced liver injury.

The present study was designed to determine the possible hepatoprotective effects of Salvia cryptantha (black weed) plant extract against carbon tetrachloride (CCl4)-induced hepatic injury in rats. Animals were grouped as follows: control group (Group I), CCl4 group (Group II), olive oil group (Group III), CCl4 + S. cryphantha 200 mg/kg group (Group IV), and CCl4 + S. cryptantha 400mg/kg group (Group V). Rats were injected intraperitoneally with CCl4 diluted in olive oil (50% v/v) at a dose of 1ml/kg body weight.  Bax and Caspase3 were determined by immunohistochemical staining, while apoptotic index was evaluated using TUNEL assay. Total mRNA was isolated from liver tissues, and the levels of BCL2, Caspase3, SOD, CAT, and glutathione peroxidase (GPx) were determined by using PCR, while MDA level were determined using a colorimetric assay. The antioxidant and anti-apoptotic gene transcripts were decreased in all of the control and treatment groups, while Caspase3 levels were not statistically different. The S. cryptantha plant extract treatment was also found to improve SOD, GPx, and catalase levels, while reducing the serum levels of MDA. The extract of S. cryptantha supplementation had a protective effect against CCl4-induced liver damage. S. cryptantha extract as a supplement may be useful as a hepato-protective agent to combat the toxic effects caused by CCl4 and other chemicals.

The effects of Nigella sativa (Ns), Anthemis hyalina (Ah) and Citrus sinensis (Cs) extracts on the replication of coronavirus and the expression of TRP genes family.

Extracts of Anthemis hyalina (Ah), Nigella sativa (Ns) and peels of Citrus sinensis (Cs) have been used as folk medicine to fight antimicrobial diseases. To evaluate the effect of extracts of Ah, Ns and Cs on the replication of coronavirus (CoV) and on the expression of TRP genes during coronavirus infection, HeLa-CEACAM1a (HeLa-epithelial carcinoembryonic antigen-related cell adhesion molecule 1a) cells were inoculated with MHV-A59 (mouse hepatitis virus-A59) at moi of 30. 1/50 dilution of the extracts was found to be the safe active dose. ELISA kits were used to detect the human IL-8 levels. Total RNA was isolated from the infected cells and cDNA was synthesized. Fluidigm Dynamic Array nanofluidic chip 96.96 was used to analyze the mRNA expression of 21 TRP genes and two control genes. Data was analyzed using the BioMark digital array software. Determinations of relative gene expression values were carried out by using the 2(-∆∆Ct) method (normalized threshold cycle (Ct) value of sample minus normalized Ct value of control). TCID50/ml (tissue culture infectious dose that will produce cytopathic effect in 50% of the inoculated tissue culture cells) was found for treatments to determine the viral loads. The inflammatory cytokine IL-8 level was found to increase for both 24 and 48 h time points following Ns extract treatment. TRPA1, TRPC4, TRPM6, TRPM7, TRPM8 and TRPV4 were the genes which expression levels changed significantly after Ah, Ns or Cs extract treatments. The virus load decreased when any of the Ah, Ns or Cs extracts was added to the CoV infected cells with Ah extract treatment leading to undetectable virus load for both 6 and 8 hpi. Although all the extract treatments had an effect on IL-8 secretion, TRP gene expression and virus load after CoV infection, it was the Ah extract treatment that showed the biggest difference in virus load. Therefore Ah extract is the best candidate in our hands that contains potential treatment molecule(s).

Phenolic acid contents, essential oil compositions and antioxidant activities of two varieties of Salvia euphratica from Turkey.

This study was designed to evaluate the in vitro antioxidant activities of n-hexane (Hex), dichloromethane (DCM), methanol (MeOH) and essential oils (EO) extracts obtained from Salvia euphratica var. euphratica and Salvia euphratica var. leiocalycina and to determine their essential oil and phenolic acid compositions. The samples were screened for their antioxidant activity by using DPPH and ß-carotene/linoleic acid assays. Methanol extracts of both varieties exhibited strong antioxidant activities. Our results showed that rosmarinic acid was dominant phenolic acid of MeOH extracts (39.4 and 55.8 µg mg⁻¹, respectively). The chemical compositions of essential oils of two varieties were analysed and their main components were determined as eucalyptol (18.4%) and trans-pinocarvyl acetate (24.9%), respectively. It can be said that these varieties could be used as natural antioxidant.