New amide and diterpene alkaloids with anticholinesterase activity from Delphinium cyphoplectrum roots.

BACKGROUND: The cholinergic hypothesis posits a robust correlation between the onset of Alzheimer's disease and a pronounced deficit in acetylcholine, a pivotal neurotransmitter crucial for the central cholinergic nervous system's function, pivotal for memory and learning. Diterpene alkaloids exhibit intricate and distinctive chemical structures that facilitate their passage through the blood-brain barrier. Moreover, their potent pharmacological attributes render them promising candidates for addressing central nervous system disorders. OBJECTIVES: This investigation aims to scrutinize the alkaloidal composition of Delphinium cyphoplectrum (Ranunculaceae) roots, further exploring their anticholinesterase inhibitory activity and mode of inhibition. METHOD: Innovative chromatography techniques were repetitively employed to purify the alkaloids. Acetylcholinesterase (AChE) inhibition assays were conducted using Ellman's tests. The mode of inhibition was meticulously characterized through Michaelis-Menten, and Lineweaver-Burk plots. Conducting molecular docking studies, we employed the AUTO DOCK 4.2 software package. RESULTS: Eight alkaloids were identified including five C19-diterpene alkaloids (6,14,16,18-tetramethoxy-1,7,8-trihydroxy-4-methylaconitane (1), 6,16,18-trimethoxy-1,7,8,14-tetrahydroxy-4-methylaconitane (2), 6,8,16,18-tetramethoxy-1,7,14-trihydroxy-4-methylaconitane (3), 6,14,16-trimethoxy-1,7,8,18-tetrahydroxy-4-methylaconitane (4), and 14-O-acetyl-8,16-dimethoxy-1,6,7,18-tetrahydroxy-4-methylaconitane (5)), an epoxy C18-diterpene alkaloid (6,8,16-trimethoxy-1,7,14-trihydroxy-3,4-epoxyaconitane (6)), a known (pyrrolidin-2-one (7) and an undescribed amide alkaloid (1-(2'-hydroxylethylamine)-3,5,5,-trimethyl-1,5-dihydro-2H-pyrrol-2-one (8). All diterpene alkaloids underwent assessment for acetylcholinesterase (AChE) inhibition assay and displayed noteworthy AChE activity, surpassing that of the reference drug (with IC50 values of 13.7, 21.8, 23.4, 28.2, 40.4, and 23.9 for compounds 1-6, respectively, in comparison to 98.4 for Rivastigmine). Analysis of Michaelis-Menten and Lineweaver-Burk plots represents an uncompetitive mode of inhibition for compound 1 on AChE. Notably, computational docking simulations indicated that all diterpene alkaloids were accommodated within the same enzymatic cleft as the reference ligand, and displaying superior free binding energy values (from - 10.32 to -8.59 Kcal.mol-1) in contrast to Rivastigmine (-6.31 Kcal.mol-1). CONCLUSION: The phytochemical analysis conducted on the roots of Delphinium cyphoplectrum yielded the identification of eight alkaloidal compounds including one C18-diterpene, five C19-diterpene, one pyrrolidine and one amide alkaloids. AChE inhibition assay and molecular simulations unveiled remarkable significant potency attributed to the C19-diterpene alkaloids by the order of 1 > 2 > 3,6 > 4 > 5. Presence of hydroxyl group on C-1, C-7, C-8, C-14, and C-18 increased the effect. The best in vitro activity was recorded for compound 1 able to bind to Asp72 in the narrow region of PAS, while interacting by pi-sigma with Phe330 at the hydrophobic region of the gorge involving the acyl and choline binding site. This observation underscores the substantial promise of this category of natural products in the realm of drug discovery for Alzheimer's Disease, offering a compelling avenue for further research and therapeutic development.

Dihydropyrimidine derivatives as MDM2 inhibitors.

One of the chief pathways to regulate p53 levels is MDM2 protein, which negatively controls p53 by direct inhibition. Many cancers overproduce MDM2 protein to interrupt p53 functions. Therefore, impeding MDM2's binding to p53 can reactivate p53 in tumor cells may suggest an effective approach for tumor therapy. Here, some Monastrol derivatives were designed in silico as MDM2 inhibitors, and their initial cytotoxicity was evaluated in vitro on MFC-7 and MDA-MB-231 cells. A small library of Monastrol derivatives was created, and virtual screening (VS) was performed on them. The first-ranked compound, which was extracted from VS, and the other six compounds 5a-5f were selected to carry out the single-docking and docking with explicit waters. The compound with the best average results was then subjected to molecular dynamic (MD) simulation. Compounds 5a-5f were chemically synthesized and evaluated in vitro for their initial cytotoxicity on MFC-7 and MDA-MB-231 cells by MTT assay. The best compound was compound 5d with ΔGave = -10.35 kcal/mol. MD simulation revealed a median potency in comparison with Nutlin-3a. The MTT assay confirmed the docking and MD experiments. 5d has an IC50 of 60.09 µM on MCF-7 cells. We attempted to use Monastrol scaffold as a potent inhibitor of MDM2 rather than an Eg5 inhibitor using in silico modification. The results obtained from the in silico and in vitro evaluations were noteworthy and warranted much more effort in the future.

Evaluation of the flavonol-rich fraction of Rosa damascena in an animal model of liver fibrosis by targeting the expression of fibrotic cytokines, antioxidant/oxidant ratio and collagen cross-linking.

INTRODUCTION: The flavonoid-rich fraction of Rosa damascena (FRFRD) contains antioxidant and active compounds. Therefore, this study aimed to investigate the role of FRFRD, rich in quercetin and kaempferol, in liver fibrosis induced by CCl4. MATERIALS AND METHODS: The FRFRD fraction was separated and standardized by High-Performance Liquid Chromatography (HPLC) based on the levels of quercetin and kaempferol. Liver fibrosis was induced over CCl4 over 12 weeks in 30 male Wistar rats, and three concentrations of FRFRD were administered to them during the last four weeks. Subsequently, after evaluation of liver serum markers and fibrotic parameters, the relative expression of transforming growth factor-beta-1 (TGF-ß1), platelet-derived growth factor (PDGF), and lysyl oxidase homolog 2 (Loxl2) genes were assessed, along with the measurement of lysyl oxidase activity and oxidative markers. RESULTS: Fibrotic markers demonstrated progressive recovery of liver damage in the treated group compared to the non-treatment group (p < 0.01). These results were accompanied by a significant decrease in the expression of TGF-ß1, PDGF, and Loxl2 genes, as well as, a reduction in lysyl oxidase activity (p < 0.001). The antioxidant effects of the treatment were observed through a significant decrease in malondialdehyde (MDA) levels and an increase in catalase enzyme (CAT) and glutathione peroxidase (GPx) activity in the treatment group compared to the fibrotic group (p < 0.01). CONCLUSION: The flavonoid-rich fraction of Rosa damascena ameliorates liver damage by affecting collagen cross-linking and lowering oxidative and inflammatory levels.

SAR131675 exhibits anticancer activity on human ovarian cancer cells through inhibition of VEGFR-3/ERK1/2/AKT signaling pathway.

Vascular endothelial growth factor receptor-3 (VEGFR-3) is known to participate in tumorigenesis and lymphangiogenesis, and as such, has the potential to serve as a molecular target for cancer therapy. SAR131675 is a highly selective VEGFR-3 antagonist that has an inhibitive effect on lymphatic cell growth. However, the anticancer effects and underlying mechanisms of SAR131675 in ovarian cancer remain poorly understood. In this study, we investigated the pathological role of VEGFR-3, and the effects of SAR131675 on proliferation, cell cycle, migration, and apoptosis in ovarian cancer cells. Our results showed that the mRNA and protein of VEGFR-3 were expressed in OVCAR3 and SKOV3 ovarian cancer cells, and this receptor was activated following stimulation with 50 ng/ml VEGF-C Cys156Ser (VEGF-CS), a selective ligand for VEGFR-3. Enhancing VEGFR-3 phosphorylation by treatment of ovarian cancer cells with VEGF-CS resulted in increased levels of phosphorylated extracellular signal-regulated kinases 1/2 (ERK1/2) and AKT. Moreover, our data demonstrated that SAR131675 inhibited VEGF-CS-mediated proliferation, colony formation, and migration of cancer cells in a dose-dependent manner. In addition, inhibition of VEGFR-3 activation with SAR131675 significantly increased cell cycle arrest and promoted apoptosis in both OVCAR3 and SKOV3 cells. Mechanistically, SAR131675 effectively suppressed the VEGF-CS-induced phosphorylation of VEGFR-3 and its downstream effectors including activated ERK1/2 and AKT in ovarian cancer cells. Our results reveal an anticancer activity of SAR131675 on the growth and migration of ovarian cancer cells, which may be through inhibiting VEGFR-3/ERK1/2/AKT pathway. SAR131675 may serve as an effective targeted drug for ovarian cancer.

MiR-1236: Key controller of tumor development and progression: Focus on the biological functions and molecular mechanisms.

Combating with the cancer, as one of the leading causes of morbidity and mortality worldwide, scientific community extensively evidenced microRNA 1236 (miR-1236) roles in the pathogenesis of malignant tumors. It has been mentioned that miR-1236 target genes and signal pathways that are key controller of tumor development and progression. Consistently, increasing evidence reports that miR-1236 participates in cancer cell growth, migration, invasion, apoptosis, and drug resistance, as well as tumor diagnosis, and prognosis. MiR-1236 is also implicated in epithelial-mesenchymal transition (EMT), which is a significant indicator of the metastatic process. Moreover, miR-1236 itself is regulated by several newly discovered long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs). Current review aimed to summarize and discuss different dimensions of miR-1236 involvement in the fundamental cellular and molecular mechanisms of tumor progressions. We believe that miR-1236 may serve as a non-invasive diagnostic marker and potential therapeutic target for cancer.

VEGFR3 suppression through miR-1236 inhibits proliferation and induces apoptosis in ovarian cancer via ERK1/2 and AKT signaling pathways.

Vascular endothelial growth factor receptor 3 (VEGFR3) is expressed in cancer cell lines and exerts a critical role in cancer progression. However, the signaling pathways of VEGFR3 in ovarian cancer cell proliferation remain unclear. This study aimed to demonstrate the signaling pathways of VEGFR3 through the upregulated expression of miR-1236 in ovarian cancer cells. We found that the messenger RNA and protein of VEGFR3 were expressed in the ovarian cancer cell lines, but downregulated after microRNA-1236 (miR-1236) transfection. The inhibition of VEGFR3, using miR-1236, significantly reduced cell proliferation, clonogenic survival, migration, and invasion ability in SKOV3 and OVCAR3 cells (p < 0.01). The flow cytometry results indicated that the rate of apoptotic cells in SKOV3 (38.65%) and OVCAR3 (41.95%) cells increased following VEGFR3 inhibition. Moreover, VEGFR3 stimulation (using a specific ligand, VEGF-CS) significantly increased extracellular signal-regulated kinase 1/2 (ERK1/2) and protein kinase B (AKT) phosphorylation (p < 0.01), whereas VEGFR3 suppression reduced p-ERK1/2 (67.94% in SKOV3 and 93.52% in OVCAR3) and p-AKT (59.56% in SKOV3 and 78.73% in OVCAR3) compared to the VEGF-CS treated group. This finding demonstrated that miR-1236 may act as an endogenous regulator of ERK1/2 and AKT signaling by blocking the upstream regulator of VEGFR3. Overall, we demonstrated the important role of the miR-1236/VEGFR3 axis in ovarian cancer cell proliferation by regulating the ERK1/2 and AKT signaling that might be an effective strategy against ovarian cancer.

Molecular mechanisms of miR-1236 in the assessment of tumor lymphangiogenesis in human ovarian cancer patients.

BACKGROUND: Tumor lymphangiogenesis is a critical component in the progression of cancers and specific microRNAs have been reported to be implicated in this process. Recent studies revealed the involvement of miR-1236 in lymphangiogenic signaling by targeting vascular endothelial growth factor receptor 3 (VEGFR3). However, the prognostic importance of miR-1236 and its clinical relevance for lymphangiogenesis in ovarian cancer (OC) remains unclear. METHODS: The study included 52 ovarian tumors and 28 normal ovarian tissues. Quantitative real-time PCR was utilized to analyze the VEGFR3, VEGF-C, LYVE-1 and PROX1 mRNA expression as well as miR-1236. VEGFR3 protein expression was measured by immunohistochemistry staining. Immunohistochemistry for the podoplanin marker (D2-40) was performed to measure lymphatic vessel density (LVD). In addition, diagnostic evaluation based on the receiver-operating characteristic (ROC) curve was performed. The influence of miR-1236 on overall survival was evaluated by Kaplan-Meier method. RESULTS: Here, we show that miR-1236 expression was significantly decreased in ovarian tumors compared with control tissues (p < 0.001) and correlated with advanced clinical stage, lymph node metastasis, distant metastasis and patient survival (All P < 0.05). Moreover, in ovarian tumors, LVD as well as the gene expression of VEGFR3, VEGF-C and LYVE-1, but not PROX1, were found to be remarkably higher compared with control tissues. We also detected a more robust positive staining for VEGFR3 in OC tissues than in control tissues. Furthermore, our results demonstrated an inverse association of miR-1236 expression with LVD, VEGFR3, LYVE-1 and PROX1 expression in OC tissues. The ROC curve analysis indicated that miR-1236 expression has the potential to be used as a diagnostic and prognostic biomarker in OC. Survival analysis further verified a lowered overall survival rate in patients with low miR-1236 expression than in those with high expression. CONCLUSIONS: Our results provide evidence for the translational involvement of miR-1236 in the lymphangiogenesis of OC by regulating lymphangiogenesis-related factors and support the clinical importance of miR-1236 as a new diagnostic and prognostic biomarker for OC.

Cycloartane triterpenoid from Euphorbia macrostegia modulates ER stress signaling pathways to induce apoptosis in MDA-MB231 and MCF-7 breast cancer cell lines.

Unfolded protein response (UPR) is involved in breast cancer (BC) progression and drug resistance. Many natural products (NPs) could modulate UPR and used for therapeutic purposes. Herein, we aimed to investigate the molecular mechanism of Cycloart-23E-ene-3ß, 25-diol (Cycloart-E25), cytotoxicity, as a NP extracted from Euphorbia macrostegia and focused on endoplasmic-reticulum stress (ERS) and UPR signaling pathways. Reactive oxygen species (ROS) were probed by DCFDA fluorescence dye. Apoptosis was assayed by annexin V/propidium iodide (PI), immunoblotting of anti- and proapoptotic, Bcl-2 and Bax proteins, and mitochondrial transmembrane potential (ΔΨm) changes. Thioflavin T (ThT) staining and immunoblotting of UPR signaling components (CHOP, PERK, ATF6, BiP, and XBP1) were recruited for the assessment of ERS. Our results indicated that Cycloart-E25 noticeably increases ROS levels in both MB-231 MDA-MB-231 and MCF-7 cell lines, p>0.05. Flow cytometry assessments revealed an increase in the cell population undergoing apoptosis. Also, the Bax/Bcl-2 ratio increased in a dose-dependent manner following Cycloart-E25 treatment, significantly, p>0.05. Mitochondrial involvement could be deduced by significant decreases in ΔΨm, p>0.05. Cycloart-E25 potently induces protein aggregation and upregulated CHOP, PERK, ATF6, BiP, and XBP1 factors in both MDA-MB-231 MB-231 and MCF-7 cell lines, indicating the involvement of ERS in Cycloart-E25-mediated apoptosis. In conclusion, Cycloart-E25 increased the accumulation of misfolded proteins and upregulated UPR components. Therefore, induction of ERS may be involved in the trigger of apoptosis in BC cell lines. Cycloart-E25 induced apoptosis in breast cancer cell lines through ERS. More assessments are needed to confirm its in vivo anti-tumoral effects.

Preparation and characterisation of a new form of silymarin as a potential antidiabetic agent in the adult male rat.

Silymarin is used for a wide variety of biological applications including, antidiabetic activities. However, the effectiveness of Silymarin is affected by its poor aqueous solubility and low systemic bioavailability after oral administration. The present study aimed to formulate a new, simple, and inexpensive form of silymarin solution. A new form of silymarin solution (NFSM) characterised by small particle size (227.5 nm), high entrapment efficiency (>82%), and appropriate zeta potential(-24.7mv). Moreover, the antidiabetic effects of NESM were evaluated relative to native Silymarin (SM). Oral administration of NFSM for 14 days in diabetic rats significantly decreased fasting blood glucose, oxidative stress levels, and improved lipid profile compared with SM. Also, NFSM significantly increased serum insulin levels, the gene expression of insulin and Pdx1, restored and improved the structure of the liver, and pancreas histologically. Our results concluded that NFSM may be an efficient carrier for oral delivery of silymarin for the management of diabetes and aggravated antioxidant status.

Caveolae-dependent endocytosis mediates the cellular uptake of CdTe quantum dots in ovarian cancer cell lines.

Background and purpose: Quantum dots (QDs) are semiconductor nanocrystals that are widely used in biology due to their good optical properties. QDs, especially cadmium-based QDs, play an important role in the diagnosis and treatment of cancer due to their intrinsic fluorescence. The aim of the present study was the evaluation of the cellular uptake mechanisms of CdTe QDs in ovarian cancer cell lines. Experimental approach: In this study, we used CdTe QDs coated with thioglycolic acid. The ovarian cancer cell lines SKOV3 and OVCAR3 were treated with different concentrations of QDs, triamterene, chlorpromazine, and nystatin, and cell viability was evaluated through the MTT test. To find the way of cellular uptake of CdTe QDs, we used the MTT test and interfering compounds in endocytic pathways. Intrinsic fluorescence and cellular internalization of CdTe QDs were assessed using flow cytometry and fluorescence microscopy imaging. Findings / Results: The viability of CdTe QDs-treated cells dose-dependently decreased in comparison to untreated cells. To evaluate the cellular uptake pathways of CdTe QDs, in most cases, a significant difference was observed when the cells were pretreated with nystatin. The results of flow cytometry showed the cellular uptake of CdTe QDs was dose- and time-dependent. Conclusion and implications: Nystatin had a measurable effect on the cellular uptake of CdTe QDs. This finding suggests that caveola-mediated endocytosis has a large portion on the internalization of CdTe QDs. According to the results of this study, CdTe QDs may have potential applications in cancer research and diagnosis.

New Cytotoxic Premyrsinane-Type Diterpenes from Euphorbia aleppica Against Breast Cancer Cells.

Euphorbia is used in traditional medicine to remove warts, possibly due to its cytotoxic or antiviral effects. This study investigated its phytochemistry and bioactive compounds. Euphorbia aleppica from the Euphorbiaceae family was collected from Kuhdasht, Lorestan, Iran. Plant material was dried and ground. Extraction was performed by maceration using a dichloromethane-acetone solvent. After removing fatty contents, fractionation was done by open column chromatography. Based on the initial H-NMR spectra, fractions containing diterpenoid compounds were identified. The Sephadex column and HPLC performed isolation. The HPLC was done with a regular YMC silica column using a hexane: Ethyl acetate (70: 30) solvent. The selected sub-fractions were identified by one and two-dimensional corelative NMR spectra. Accurate mass spectra confirmed the molecular formula of the obtained structures. Cytotoxicity was assessed using a standard MTT assay against breast cancer cells. The NMR and mass analysis identified compound 1 as a newly described and compound 2 as a pre-defined compound as 3, 7, 15ß-triacetyl-5α-tigliate-13(17)-α-epoxy-14-oxopremyrsinane and 3, 7, 14, 15, 17-pentaacetyl-5-tigliate-13(17)-epoxypremyrsinane, respectively. Compound 1 showed moderate cytotoxicity, and compound 2 exhibited a potent cytotoxic effect dose-dependently against MCF-7 and MDA-MB 231 breast cancer cells, probably because of 14-O-acetyl and 17-O-acetylated hemiacetal groups.

Phytochemical analysis of Artemisia kopetdaghensis: Sesquiterpene lactones with proapoptotic activity against prostate cancer cells.

Phytochemical investigation of the aerial parts of Artemisia kopetdaghensis resulted in the isolation and characterization of three undescribed eudesmane-type sesquiterpene lactones, persianolide A, 4-epi-persianolide A, and 3α,4-epoxypersianolide A, together with three previously described eudesmane-type sesquiterpene lactones, 11-epi-artapshin, 1ß,8α-dihydroxy-11α,13-dihydrobalchanin, and 1ß-hydroxy-11-epi-colartin. The abundantly obtained 11-epi-artapshin was oxidized to undescribed 11α,13-dihydroeudesma-12,6α-olide-1,8-dione and 8ß-hydroxy-11α,13-dihydroeudesma-12,6α-olide-1-one and acetylated to the undescribed 1,8-O-diacetyl-11α,13-dihydroeudesma-12,6α-olide. Structures were elucidated based on extensive spectral data analyses, including 1D and 2D NMR and HRESIMS. The absolute configuration was determined using calculated and experimental ECD spectral data. Compounds were subsequently subjected to the MTT assay to evaluate their cytotoxicity against prostate cancer cells (DU-145 and LNCaP). Related factors associated with the sequence of apoptosis were tested by ELISA, western blotting, and biochemical assay. Results suggested that 11-epi-artapshin hinders the growth of DU-145 cells through mitochondria-mediated apoptosis initiated by stimulation of ROS build-up, ΔΨm depletion, regulation of the Bax/Bcl-2 ratio, and activation of caspase 3, respectively.

Long-term GABA administration improves FNDC5, TFAM, and UCP3 mRNA expressions in the skeletal muscle and serum irisin levels in chronic type 2 diabetic rats.

In this study, we aimed to investigate whether the anti-diabetic effects of γ-aminobutyric acid (GABA) and insulin can be mediated through the regulation of gene expression related to irisin production and mitochondrial biogenesis in type 2 diabetic mellitus (T2DM) rats. Four groups (n = 6) were used in this study: control, T2DM, T2DM + insulin, and T2DM + GABA groups. After T2DM induction for 3 months (high-fat diet + 35 mg/kg streptozotocin) and treatment with GABA or insulin for 3 months, circulating levels of FBG, triglyceride, LDL, Ox-LDL, and insulin as well as hepatic and serum irisin levels were measured. The mRNA expressions of fibronectin type III domain-containing protein 5 (FNDC5), mitochondrial transcription factor A (TFAM), and mitochondrial uncoupling protein 3 (UCP3) were also evaluated in the skeletal muscle of all groups. GABA therapy improved the FBG and insulin levels in diabetic rats. Insulin treatment significantly reduced FBG and failed to maintain glucose close to the control level. Insulin or GABA therapy significantly decreased the levels of LDL, Ox-LDL, and HOMA-IR index. Circulating irisin levels were markedly decreased in insulin-treated group, while irisin levels did not show significant changes in GABA-treated group compared with control group. GABA or insulin therapy increased mRNA expressions of TFAM and UCP3 in diabetic rats. GABA therapy also led to a significant increase in FNDC5 mRNA. Our findings suggest that the anti-diabetic effect of GABA may be mediated, in part, by a decrease in Ox-LDL levels and an increase in the levels of irisin as well as FNDC5, TFAM, and UCP3 gene expression in T2DM rats.

Phytochemical analysis of Daphne pontica L. stems with their pro-apoptotic properties against DU-145 and LNCaP prostate cancer cells.

BACKGROUND: Daphne pontica is an endemic plant grown wild in the North part of Iran, with anticancer activities. OBJECTIVES: This study aims to analyze the phytochemistry and screen the cytotoxic activity of new bioactive compounds against a panel of cancer cells, in addition to proapototic properties against prostate cancer cells. METHOD: Purification procedure was done using repeated column chromatographies by MPLC and HPLC systems. The structures were elucidated by the NMR and exact mass spectroscopy, stereochemistry by NOESY, and absolute configuration by electronic circular dichroism (ECD) spectra. Cytotoxicity was done against DU 145, LNCaP, HeLa, MCF-7, and MDA-MB 231 cells by standard MTT assay. An annexin V/PI assay was performed to measure the type of death following treatment with these compounds for 48 h, followed by the caspase-3 activity test. RESULTS: In this study, one new dilignan named lignopontin A (9), in addition to 13 known compounds including two phenolic acids (3, 5), one flavanone (6), one bis flavonoid (1), one cumarin glycoside (2), one mono (4) and two dicumarins (10, 11), two lignans (7, 8), and three daphnane diterpenoids (12-14) were isolated for the first time from D. pontica stems. Complete spectral data of compound 12, named as 6,7α-epoxy-5ß-hydroxy-9,13,14-ortho-(4,2E)-pentadeca-2,4-diene-1-yl)-resiniferonol, and compound 14, named as 6,7α-epoxy-5ß-hydroxy-9,3,14-ortho-(2,4E)-pentadeca-2,4-di-1-yl)-resiniferonol-12ß-yl-acetate are reported for the first time. In the MTT assay of newly described compounds against a panel of cancer cells, compounds 9, 12, and 14 possessed moderate to potent cytotoxicity against prostate, breast, and cervical cancer cells in a dose-dependent manner. Flow cytometry analysis against prostate cancer cells indicated that the cytotoxicity of compounds 12 and 14 was due to their ability to induce apoptosis. In the case of compound 9, in Du 145 cells, cell death was mainly through apoptosis. In contrast, LNCaP cells showed both apoptosis and necrotic cell death, predominated by necrosis at the higher concentrations. Caspase-3 activity confirmed apoptosis observed in these compounds through the caspase pathway in prostate cancer cells. CONCLUSION: D. pontica is a new source of dimeric phenolic compounds, including bisflavonoids, phenylpropanoid-cumarin adduct, and dilignans, as well as daphnane diterpenoids with resiniferonol core with long-chain orthoester moieties. In cytotoxicity screening, compounds 9, 12, and 14 inhibited the growth of DU-145 and LNCaP cells in a dose-dependent manner with IC50 varied from 0.9 - 27.3 and 25.2 - 87.4 µM, respectively. Among them, 9 exhibited selective growth inhibition against DU 145 treated cells. LNCaP cells demonstrated the highest sensitivity to treatment with compound 12.

Nisin induces apoptosis in cervical cancer cells via reactive oxygen species generation and mitochondrial membrane potential changes.

Nisin, an antimicrobial peptide produced by Lactococcus lactis, is widely used as a safe food preservative and has recently attracted the attention of researchers as a potential anticancer agent. The cytotoxicity of nisin against human cervical cancer cell lines (HeLa), human ovarian carcinoma cell lines (OVCAR-3 and SK-OV-3), and human umbilical vein endothelial cells (HUVECs) was evaluated using an MTT assay. The apoptotic effect of nisin was identified by Annexin-V/propidium iodide assay, which was further confirmed by western blotting analysis, mitochondrial membrane potential (ΔΨm) analysis, and reactive oxygen species (ROS) assay. The MTT assay showed concentration-dependent cytotoxicity of nisin towards cancer cell lines, with IC50 values of 11.5-23 µM, but less toxicity against normal endothelial cells. Furthermore, the treatment of cervical cancer cells with 12 µM nisin significantly (P < 0.05) increased the Bax/Bcl-2 ratio (4.9 fold), reduced ΔΨm (70%), and elevated ROS levels (1.7 fold). These findings indicate that nisin may have anticancer and apoptogenic activities through mitochondrial dysfunction and oxidative stress damage in cervical cancer cells.

Beneficial effects of MgSO4 on TFAM, UPC3 and FNDC5 mRNA expressions in skeletal muscle of type 2 diabetic rats: a possible mechanism to improve insulin resistance.

BACKGROUND: Hypomagnesemia has been associated with development of type 2 diabetes mellitus (T2DM) and its complications. Irisin has beneficial effects on glucose uptake and improves hepatic glucose and lipid metabolism. In this study, we aimed to evaluate the effects of long-term treatment of MgSO4 and insulin on insulin resistance, dyslipidemia, serum and hepatic irisin levels, skeletal muscle gene expression of fibronectin type III domain-containing protein 5 (FNDC5), mitochondrial transcription factor A (TFAM) and mitochondrial uncoupling protein 3 (UCP3) in T2DM rats. METHODS AND RESULTS: Twenty-four rats were divided into four groups: Control group, diabetic control (DC) using a high-fat diet + streptozotocin, insulin-treated diabetic group (DC + Ins), MgSO4-treated diabetic group (DC + Mg). At the end of therapies, serum concentrations of FBG, TG, insulin, Ox-LDL, along with serum and hepatic irisin levels were measured. FNDC5, TFAM, and UCP3 mRNA expressions were measured in the skeletal muscle by Real-time PCR. In comparison with DC group, MgSO4 therapy resulted in decreased FBG, TG, Ox-LDL, improved serum insulin and irisin levels, and increased mRNA expressions of FNDC5, UCP3 and TFAM. Insulin therapy significantly decreased FBG, Ox-LDL, FNDC5 and serum irisin levels compared with the control group. While, insulin therapy markedly increased TFAM and UCP3 compared with the DC group. CONCLUSIONS: In conclusion, MgSO4 can improve insulin resistance and hyperlipidemia partly through decreasing Ox-LDL, increasing serum irisin levels as well as increasing FNDC5, TFAM, and UCP3 mRNA expressions in T2DM rats. These findings can be considered in the management of diabetes treatment.

Mitochondrial Pro-Apoptotic Properties of Sinularia compressa from Persian Gulf against Breast Cancer Cells and Its Chemical Composition.

Locals in the Persian Gulf islands traditionally use Sinularia compressa to treat cancer. Therefore, this study deals with the cytotoxic activity of the soft coral Sinularia compressa chloroform extract (SCE), its pro-apoptotic activity, and the determination of its secondary metabolites. Cytotoxicity was done against MCF-7 and MDA-MB-231 and MCF­10A cells. Apoptosis induction was checked by flow cytometry. The DCFDA and JC-1 probes were used to assess the production of reactive oxygen species (ROS) and the mitochondrial transmembrane potential. Caspase-9, Bax, and Bcl-2 proteins were determined with ELISA Kit, and by western blot analysis. SCE exhibited cytotoxic activity with an IC50 value of 32.51 ± 0.70 µg/ml against MCF-7, and 8.53 ± 0.97 µg/ml against MDA-MB-231 cancer cells. The induction of the intrinsic apoptosis pathway was found by ROS generation, attenuation of Bcl-2 and induction of Bax proteins. It was supported by activation of caspase-9, increased apoptotic cells, as well as decrease of ΔΨm. In the acute toxicity, there was no detectable sign of hepatic or renal toxicity in the SCE 100 mg/kg. GC mass and NMR identified bioactive compounds as one monoterpene, one sesquiterpene, five fatty acids, one phthalate, and two steroidal compounds.

SAR131675 Receptor Tyrosine Kinase Inhibitor Induces Apoptosis through Bcl- 2/Bax/Cyto c Mitochondrial Pathway in Human Umbilical Vein Endothelial Cells.

BACKGROUND: Tyrosine Kinase Inhibitors (TKIs) can be used to inhibit cancer cell proliferation by targeting the vascular endothelial growth factor receptor (VEGFR) family. SAR131675 is a highly selective receptor tyrosine kinase inhibitor to VEGFR3 that reveals the inhibitory effect on proliferation in human lymphatic endothelial cells. However, the molecular mechanisms underlying this process are generally unclear. OBJECTIVE: This study was performed to investigate the possible involvement of the Bcl-2/Bax/Cyto c apoptosis pathway in Human Umbilical Vein Endothelial Cells (HUVECs). In addition, the role of Reactive Oxygen Species (ROS) and mitochondrial membrane potential was evaluated. METHODS: The effect of SAR131675 on HUVEC cell viability was evaluated by MTT assay. The activity of SAR131675 in inducing apoptosis was carried out through the detection of Annexin V-FITC/PI signal by flow cytometry. To determine the mechanisms underlying SAR131675 induced apoptosis, the mitochondrial membrane potential, ROS generation, the activity of caspase-3, and expression of apoptosis-related proteins such as Bcl-2, Bax, and cytochrome c were evaluated in HUVECs. RESULTS: SAR131675 significantly inhibited cell viability and induced apoptosis in HUVECs in a dose-dependent manner. Moreover, SAR131675 induced mitochondrial dysfunction, ROS generation, Bcl-2 down-regulation, Bax upregulation, cytochrome c release, and caspase-3 activation, which displays features of mitochondria-dependent apoptosis signaling pathway. CONCLUSION: Our present data demonstrated that SAR131675-induced cytotoxicity in HUVECs associated with the mitochondria apoptotic pathway. These results suggest that further studies are required to fully elucidate the role of TKIs in these cellular processes.

Novel 16,17-epoxy-23-methylergostane derivative from Sinularia variabilis, a soft coral from the Persian Gulf, with apoptotic activities against breast cancer cell lines.

The steroidal and terpenoidal composition of Sinularia variabilis was investigated by chromatography methods. One new (1), and four known [gorgasta-5-en-3ß-ol (2), ergosta-5-en-3ß-ol (3), ergosta-5, 22(Z)-dien-3ß-ol (4), 5,8-epidioxy-5α, 8α-ergosta-6, 22E-dien-3ß-ol (5)] steroids, in addition to one known diterpenoidal alkaloid [sinulasulfone (6)] isolated for the first time from S. variabilis. If we named the 23-methylergostane core structure as sinustane, new compound (1) was elucidated as 16α,17α-epoxysinusta-5-en-3ß-ol-20ß-yl sulfate based on NMR and HR Mass data. It was submitted for cytotoxic activity evaluation against MCF-7 and MDA-MB-231 cell lines using MTT assay. Apoptosis induction was checked by flow cytometry (annexin V/propidium iodide) staining. To determine the production of reactive oxygen species, and the mitochondrial transmembrane potential (ΔΨm), the DCFDA, and JC-1 probes were used in this study.

Galectin-9 inhibits cell proliferation and induces apoptosis in Jurkat and KE-37 acute lymphoblastic leukemia cell lines via caspase-3 activation.

BACKGROUND AND PURPOSE: Acute lymphoblastic leukemia (ALL) is a type of cancer of blood and bone marrow characterized by abnormal proliferation of lymphoid progenitor cells. Galectin-9 is a tandem-repeat type galectin expressed in various tumor cells. It seems that the connection between galectin-9 and T cell immunoglobulin mucin-3 receptor acts as a negative regulator of cancer cells proliferation. EXPERIMENTAL APPROACH: In this research, the effects of galectin-9 were investigated using MTS cell proliferation colorimetric, colony-forming, annexin V-FITC/PI, and caspase-3 assays in the Jurkat and KE-37 cell lines of ALL. Furthermore, the western blotting technique was used to evaluate the levels of apoptotic proteins such as Bax and Bcl-2 in these cell lines. FINDINGS/RESULTS: Our results indicated that galectin-9 can considerably reduce the cell growth and colony formation ability of both Jurkat and KE-37 cell lines in a concentration-dependent manner. Besides, galectin-9 induced apoptosis in a concentration-dependent manner in ALL cells by a mechanism associated with Bax/Bcl-2 expression and activation of the caspase-3 activation. CONCLUSION AND IMPLICATIONS: Galectin-9 inhibited the growth and proliferation of cell lines with increased programmed cell death, therefore it can be considered as a potential factor in the progression of ALL therapeutics that needs more research in this context.