Tart cherry (Prunus cerasus L.) dietary supplement modulates visceral adipose tissue CB1 mRNA levels along with other adipogenesis-related genes in rat models of diet-induced obesity.

PURPOSE: There is increasing evidence for the involvement of dietary bioactive compounds in the cross-talk modulation of endocannabinoid system and some of the key regulators of transcriptional control for adipogenesis. METHODS: We aimed to characterize the expression of cannabinoid CB1/CB2 receptors and fatty acid amide hydrolase (FAAH) along with selected adipogenesis-related genes (PPARγ, SREBP-1c and PREF-1), adipocyte-secreted factors (leptin and adiponectin), mitochondrial bioenergetic modulators (PGC-1A and UCP-2), and transient receptor potential vanilloid subtype 1 (TRPV1) and 2 (TRPV2) channels in visceral adipose tissue of rats fed with a high-fat diet (HFD) containing either tart cherry seeds alone or tart cherry seeds and juice for 17 weeks. The visceral adipose tissue was weighed and checked the expression of different markers by qRT-PCR, Western blot and immunohistochemistry. RESULTS: Tart cherry supplements were able to downregulate the HFD-induced mRNA expression of CB1 receptor, SREBP-1c, PPARγ, leptin, TRPV1 and TRPV2 resulting in potential anti-adipogenic effects. CONCLUSION: The present study points out that the intake of bioactive constituents of tart cherry may attenuate the effect of adipogenesis by acting directly on the adipose tissue and modulating the interplay between CB1, PPARγ and TRPV channel gene transcription.

Effects of Prunus cerasus L. Seeds and Juice on Liver Steatosis in an Animal Model of Diet-Induced Obesity.

The accumulation of adipose tissue increases the risk of several diseases. The fruits-intake, containing phytochemicals, is inversely correlated with their development. This study evaluated the effects of anthocyanin-rich tart cherries in diet-induced obese (DIO) rats. DIO rats were exposed to a high-fat diet with the supplementation of tart cherry seeds powder (DS) and seed powder plus juice (DJS). After 17 weeks, the DIO rats showed an increase of body weight, glycaemia, insulin, and systolic blood pressure. In the DS and DJS groups, there was a decrease of systolic blood pressure, glycaemia, triglycerides, and thiobarbituric reactive substances in the serum. In the DJS rats, computed tomography revealed a decrease in the spleen-to-liver attenuation ratio. Indeed, sections of the DIO rats presented hepatic injury characterized by steatosis, which was lower in the supplemented groups. In the liver of the DIO compared with rats fed with a standard diet (CHOW), a down-regulation of the GRP94 protein expression and a reduction of LC3- II/LC3-I ratio were found, indicating endoplasmic reticulum stress and impaired autophagy flux. Interestingly, tart cherry supplementation enhanced both unfolded protein response (UPR) and autophagy. This study suggests that tart cherry supplementation, although it did not reduce body weight in the DIO rats, prevented its related risk factors and liver steatosis.

Isofuranodiene synergizes with temozolomide in inducing glioma cells death.

BACKGROUND: Glioblastoma multiforme (GBM) is the most common and deadly brain form of tumor. GBM exhibits high resistance to the standard treatment consisting of temozolomide (TMZ) combined with radiotherapy. Isofuranodiene (IFD) is a bioactive sesquiterpene occurring in the essential oils obtained from Alexanders (Smyrnium olusatrum L., Apiaceae). This compound has shown a broad spectrum of antitumoral activities in different human cancer cell lines both in vitro and in vivo. However, the mechanism of action of IFD on GBM and its potential effects in combination with chemotherapeutic drugs, have not been fully elucidated. PURPOSE: The aim of the present study was to evaluate the anticancer effects of IFD itself and in combination with TMZ in GBM. METHODS: Sulforhodamine B-based proliferation assay, cell cycle analysis and Annexin V/PI staining were carried out to determine the IFD effects on three human GBM cell lines, U87, T98, U251 and in normal human astrocyte. Modulation of protein expression levels was determined by western blot analysis. Reactive oxygen species (ROS) production was evaluated by cytofluorimetry. Moreover, the effects on cell viability of the IFD and TMZ co-administration was evaluated through the calculation of combination index (CI). RESULTS: IFD exerted cytotoxic effects against the GBM cell lines, but not in normal cells (normal human astrocytes). This compound induced a cell cycle blockage and a necrotic cell death depending on the increase of intracellular ROS levels. Furthermore, the synergism between IFD and TMZ was demonstrated in GBM cell lines. CONCLUSION: This study demonstrated the glioma selectivity of IFD and its cytotoxic properties suggesting a new strategy for the treatment of GBM in order to overcome the TMZ resistance and to reduce its side effects.

Thyme extract increases mucociliary-beating frequency in primary cell lines from chronic obstructive pulmonary disease patients.

Chronic obstructive pulmonary disease (COPD) is a respiratory disorder characterized by a progressive and irreversible airflow limitation. COPD is associated to a chronic inflammatory response with infiltration of inflammatory cells in the surface epithelium of large airways and abnormalities in structure and functions of cilia. Thyme (Thymus vulgaris L.) is a traditional medicinal plant of the Mediterranean area used to treat respiratory disorders. We previously evidenced that thyme extract reduce IL-1beta and IL-8, by downregulating the activated NF-κB levels, suggesting its potential therapeutically use in COPD. Cilia beating frequency (CBF) is dramatically impaired in COPD and different pharmacological agents can modulate cilia function. Herein we evaluated the effect of a commercial thyme extract in modulating CBF by measuring its activity in stimulating cAMP, Ca2+ levels and CBF in a MucilAir 3D human COPD airway epithelia reconstituted in vitro system using salmeterol, YM976, isoproterenol and GSK1016790 A as positive controls. Results showed that thyme extract increased cAMP levels starting from 12 h post-treatment, decreased extracellular Ca2+ levels and increased the CBF in airway epithelia from COPD donors. Overall, this work demonstrated that thyme extract is effective in stimulating CBF by inducing an increase of cAMP and Ca2+ levels, thus supporting its therapeutical use in the treatment of COPD.

Capsaicin triggers autophagic cell survival which drives epithelial mesenchymal transition and chemoresistance in bladder cancer cells in an Hedgehog-dependent manner.

Bladder cancer (BC) is a common urologic tumor characterized by high risk of recurrence and mortality. Capsaicin (CPS), used as an intravesical drug for overactive bladder, was demonstrated to induce cell death in different cancer cells including BC cells.Here we found that treatment of high-grade BC cells with high dose of CPS triggers autophagy. Infact, the CPS treatment alters the redox homeostasis by inducing production of radicals, mitochondrial depolarization, alterations of ADP/ATP ratio and activation of AMPK pathway stimulating the autophagic process in BC cells. The inhibition of autophagy, by using the specific inhibitor bafilomycin A or Beclin 1 knock-down, enhanced the CPS-induced cell death, demonstrating that CPS-induced autophagy acts as a pro-survival process in BC cells. By using PCR arrays and FACS analysis, we found that the CPS-treated BC cells displayed typical mesenchymal features of the epithelial mesenchymal transition (EMT) as elongated shape and over-expression of vimentin, α5 and ß1 integrin subunits, integrin-like kinase and the anti-apoptotic Bcl-2 proteins. Moreover, we demonstrated that CPS treatment stimulates upregulation of Dhh/Ptch2/Zeb2 members of the Hedgehog signaling pathway, increases CD24, VEGFA and TIMP1 and decreases CD44 and ALCAM mRNA expression levels. By PTCH2 knock-down we found that the Hedgehog signaling pathway is involved in the CPS-induced autophagy and EMT phenotype.Finally, we also showed that the CPS-resistant EMT-positive BC cells displayed an increased drug-resistance to the cytotoxic effects of mitomycin C, gemcitabine and doxorubicine drugs commonly used in BC therapy.

Capsaicin promotes a more aggressive gene expression phenotype and invasiveness in null-TRPV1 urothelial cancer cells.

Capsaicin (CPS) has been found to exhibit either tumor promoting or suppressing effects, many of which are mediated by the specific transient receptor potential vanilloid type-1 (TRPV1). Herein, we provide evidence that CPS treatment induced a more aggressive gene phenotype and invasiveness in 5637 cells-lacking TRPV1 receptor. CPS treatment of 5637 cells induced upregulation of pro-angiogenetic (angiopoietin 1, angiopoietin 2 and vascular endothelial growth factor), pro-invasive and pro-metastatic genes (MMP1, MMP9, TIMP1, TIMP3, granzyme A (GZMA), NM23A and S100A) with a downregulation of apoptotic genes (Fas/CD95 and tumor necrosis factor receptor superfamily member 1A). CPS increased the invasiveness of 5637 cells by triggering IGF (insulin-like growth factor)-1 release, GZMA and MMP9 activation, α-tubulin disassembly and cytoskeleton degradation. Finally, in order to evaluate the relationship between the lack of TRPV1 expression and increased CPS-induced invasiveness, we transfected 5637 cells with the TRPV1 complementary DNA (cDNA) sequence. We found that TRPV1-expressing cells show CPS-mediated calcium level increase, growth inhibition and apoptosis. Moreover, CPS-induced migration and MMP9 activation were reverted, suggesting an inhibitory role played by TRPV1 in urothelial cancer cell invasion and metastasis.

Synthesis and biological evaluation of indazolo[4,3-bc][1,5]naphthyridines (10-aza-pyrazolo[3,4,5-kl]acridines): a new class of antitumor agents.

A series of potential DNA-binding antitumor agents, 2-[omega-(alkylamino)alkyl]-9-methoxy-5-nitro-2,6-dihydroindazolo[4,3-bc][1,5]naphthyridines (2a-f), 10-aza derivatives of PZA, has been prepared by condensation of 9-chloro-2-methoxy-6-nitro-5,10-dihydrobenzo[b][1,5]naphthyridin-10-one (6) with the appropriate (omega-aminoalkyl)hydrazine in tetrahydrofuran/methanol. Compound 6 was obtained by heating at 100 degrees C in H(2)SO(4)5, yielded by the condensation of 2,6-dichloro-3-nitrobenzoic acid (4) and 6-methoxy-3-pyridinamine (3). The non-covalent DNA-binding properties of 2 have been examined using a fluorometric technique. In vitro cytotoxic potencies of these derivatives against human hormone-refractory prostate adenocarcinoma cell line (PC-3) are described and compared to that of parent drug PZA. We selected the most cytotoxic target derivatives 2c,d, the in vitro inactive 2f, and reference compound PZA to investigate whether in vitro treatment with these drugs was able to induce necrotic and/or apoptotic cell death. To this purpose, we evaluated the percentage of apoptotic cells in PC-3 treated with the target compounds 2c,d,f and reference compound PZA, by Annexin V staining and Propidium iodide (PI)/Annexin V, biparametric flow cytometric analysis and agarose gel electrophoresis.

Neonatal capsaicin treatment affects rat thymocyte proliferation and cell death by modulating substance P and neurokinin-1 receptor expression.

Herein we provide evidence that substance P (SP) and its neurokinin-1 receptor (NK-1R) expressed on thymocytes counteract thymus depletion induced by neonatal capsaicin (CPS) treatment by affecting thymocyte proliferation and apoptotic death. SP administration reversed the CPS-mediated inhibitory effects on the total thymocyte number and subset distribution, namely CD4+ and CD4- CD8- cells, through its interaction with NK-1R as shown by concomitant NK-1R (SR140333) antagonist administration. SP-induced enhancement of thymus cellularity parallels its ability of inhibiting the thymocyte apoptotic program. Indeed, exogenously administered SP completely nullified CPS-induced apoptosis, and SR140333 abrogated the SP-mediated protective effect. SP administration also stimulated concanavalin A (Con A)-induced thymocyte proliferation of CPS-treated rats, completely reversing the CPS-induced inhibition. The SP-mediated stimulation of Con A-induced thymocyte proliferation was NK-1R dependent as shown by concomitant administration of SP and SR140333 to CPS-treated rats. Our results also demonstrate that CPS treatment induces a marked decrease of thymocyte PPT-A mRNA level and endogenous SP content as evaluated by quantitative RT-PCR, in situ hybridization and cytofluorimetric analysis. By contrast, NK-1R mRNA levels were increased in thymocytes from CPS-treated rats. Exogenous SP administration augmented PPT-A, SP and NK-1R thymocyte expression in CPS-treated rats, and this enhancement was antagonized by SR140333 administration. Overall, our results strongly suggest that the immunomodulatory effects of neonatal CPS treatment on rat thymocyte functions are dependent on vanilloid-mediated regulation of SP and NK-1R functional expression by neuronal and immune cells.