Sakineh Kazemi Noureini

Crocin, the main pigment of Crocus sativus L., has been shown to have antiproliferative effects on cancer cells, but the involved mechanisms are only poor understood. This study focused on probable effect of crocin on the immortality of hepatic cancer cells. Cytotoxicity of crocin (<TEX>$IC_{50}$</TEX> 3 mg/ml) in hepatocarcinoma HepG2 cells was determined after 48 h by neutral red uptake assay and MTT test. Immortality was investigated through quantification of relative telomerase activity with a quantitative real-time PCR-based telomerase repeat amplification protocol (qTRAP). Telomerase activity in 0.5 <TEX>${\mu}g$</TEX> protein extract of HepG2 cells treated with 3 mg/ml crocin was reduced to about 51% as compared to untreated control cells. Two mechanisms of inhibition, i.e. interaction of crocin with telomeric quadruplex sequences and down regulation of hTERT expression, were examined using FRET analysis to measure melting temperature of a synthetic telomeric oligonucleotide in the presence of crocin and quantitative real-time RT-PCR, respectively. No significant changes were observed in the <TEX>$T_m$</TEX> telomeric oligonucleotides, while the relative expression level of the catalytic subunit of telomerase (hTERT) gene showed a 60% decrease as compared to untreated control cells. In conclusion, telomerase activity of HepG2 cells decreases after treatment with crocin, which is probably caused by down-regulation of the expression of the catalytic subunit of the enzyme.

Corona Virus Disease 2019 (COVID-19) as the infectious disease caused the pandemic disease around the world through infection by SARS-CoV-2 virus. The common diagnosis approach is Quantitative RT-PCR (qRT-PCR) which is time consuming and labor intensive. In the present study a novel colorimetric aptasensor was developed based on intrinsic catalytic activity of chitosan film embedded with ZnO/CNT (ChF/ZnO/CNT) on 3,3',5,5'-tetramethylbenzidine (TMB) substrate. The main nanocomposite platform was constructed and functionalized with specific COVID-19 aptamer. The construction subjected with TMB substrate and H2O2 in the presence of different concentration of COVID-19 virus. Separation of aptamer after binding with virus particles declined the nanozyme activity. Upon addition of virus concentration, the peroxidase like activity of developed platform and colorimetric signals of oxidized TMB decreased gradually. Under optimal conditions the nanozyme could detect the virus in the linear range of 1-500 pg mL and LOD of 0.05 pg mL. Also, a paper-based platform was used for set up the strategy on applicable device. The paper-based strategy showed a linear range between 50 and 500 pg mL with LOD of 8 pg mL. The applied paper based colorimetric strategy showed reliable results for sensitive and selective detection of COVID-19 virus with the cost-effective approach.

Natural bioproducts are invaluable resources in drug discovery. Isoquinoline alkaloids of Chelidonium majus constitute a structurally diverse family of natural products that are of great interest, one of them being their selectivity for human telomeric G-quadruplex structure and telomerase inhibition.The study focuses on the mechanism of telomerase inhibition by stabilization of telomeric G-quadruplex structures by berberine, chelerythrine, chelidonine, sanguinarine and papaverine. Telomerase activity and mRNA levels of hTERT were estimated using quantitative telomere repeat amplification protocol (q-TRAP) and qPCR, in MCF-7 cells treated with different groups of alkaloids. The selectivity of the main isoquinoline alkaloids of Chelidonium majus towards telomeric G-quadruplex forming sequences were explored using a sensitive modified thermal FRET-melting measurement in the presence of the complementary oligonucleotide CT22. We assessed and monitored G-quadruplex topologies using circular dichroism (CD) methods, and compared spectra to previously well-characterized motifs, either alone or in the presence of the alkaloids. Molecular modeling was performed to rationalize ligand binding to the G-quadruplex structure.The results highlight strong inhibitory effects of chelerythrine, sanguinarine and berberine on telomerase activity, most likely through substrate sequestration. These isoquinoline alkaloids interacted strongly with telomeric sequence G-quadruplex. In comparison, chelidonine and papaverine had no significant interaction with the telomeric quadruplex, while they strongly inhibited telomerase at transcription level of hTERT. Altogether, all of the studied alkaloids showed various levels and mechanisms of telomerase inhibition.We report on a comparative study of anti-telomerase activity of the isoquinoline alkaloids of Chelidonium majus. Chelerythrine was most effective in inhibiting telomerase activity by substrate sequesteration through G-quadruplex stabilization.Understanding structural and molecular mechanisms of anti-cancer agents can help in developing new and more potent drugs with fewer side effects. Isoquinolines are the most biologically active agents from Chelidonium majus, which have shown to be telomeric G-quadruplex stabilizers and potent telomerase inhibitors.

To investigate the potential effects of chelidonine, the main alkaloid of Chelidonium majus, on telomerase activity and its regulation in HepG2 cells.Cytotoxicity of chelidonine for HepG2 cells was determined by neutral red assay. A modified polymerase chain reaction (PCR)-based telomerase repeat amplification protocol was used to estimate relative telomerase activity in chelidonine-treated cells in comparison with the untreated control cells. Relative expression level of the catalytic subunit of telomerase (hTERT) gene and P-glycoprotein (pgp) were estimated using semi-quantitative real-time reverse transcription-PCR (RT-PCR). Cell senescence in treated cells was demonstrated using a beta-galactosidase test.Cytotoxicity of chelidonine in HepG2 cells was not dose-dependent and tended to reach plateau immediately after the living cells were reduced in number to slightly higher than 50%. However, 12 micromol/L concentration of chelidonine was considered as LD(50), where the maximal attainable effects were realized. Real-time RT-PCR data showed that the expression of pgp increased three-fold in chelidonine treated HepG2 cells in comparison with the untreated controls. Morphologically, treated HepG2 cells showed apoptotic features after 24 h and a small fraction of cells appeared with single blister cell death. The relative expression level of Bcl-2 dropped to less than 50% of control cells at a sub-apoptotic concentration of chelidonine and subsequently increased to higher than 120% at LD(50). Telomerase activity was reduced considerably after administration of very low doses of chelidonine, whereas higher concentrations of chelidonine did not remarkably enhance the effect. Real-time RT-PCR experiments indicated a drastic decrease in expression level of hTERT subunit of telomerase under treatment with chelidonine. Repeated treatment of cells with very low doses of chelidonine caused a decline in growth rate by 4 wk and many of the cells appeared to be aged with large volume and dark staining in the beta-galactosidase assay.Chelidonine reduces telomerase activity through down-regulation of hTERT expression. Senescence induction might not be directly caused by reducing telomerase activity as it occurs after a few population doublings.

Zoological Journal of the Linnean SocietyVolume 158, Issue 3 p. 641-660 Molecular phylogeny of the Eremias persica complex of the Iranian plateau (Reptilia: Lacertidae), based on mtDNA sequences E. RASTEGAR POUYANI, Corresponding Author E. RASTEGAR POUYANI Department of Biology, Tarbiat Moalem University of Sabzevar, PO Box 397 Sabzevar, Iran E-mail: rastegarpouyani45@gmail.com; erastegar@yahoo.comSearch for more papers by this authorN. RASTEGAR POUYANI, N. RASTEGAR POUYANI Department of Biology, Razi University, Kermanshah, IranSearch for more papers by this authorS. KAZEMI NOUREINI, S. KAZEMI NOUREINI Department of Biology, Tarbiat Moalem University of Sabzevar, PO Box 397 Sabzevar, IranSearch for more papers by this authorU. JOGER, U. JOGER State Natural History Museum, Pockelsstr. 10, 38106 Braunschweig, GermanySearch for more papers by this authorM. WINK, M. WINK Institute of Pharmacy and Molecular Biotechnology, Department of Biology, University of Heidelberg, Im Neuenheimer Feld 364, 69120 Heidelberg, GermanySearch for more papers by this author E. RASTEGAR POUYANI, Corresponding Author E. RASTEGAR POUYANI Department of Biology, Tarbiat Moalem University of Sabzevar, PO Box 397 Sabzevar, Iran E-mail: rastegarpouyani45@gmail.com; erastegar@yahoo.comSearch for more papers by this authorN. RASTEGAR POUYANI, N. RASTEGAR POUYANI Department of Biology, Razi University, Kermanshah, IranSearch for more papers by this authorS. KAZEMI NOUREINI, S. KAZEMI NOUREINI Department of Biology, Tarbiat Moalem University of Sabzevar, PO Box 397 Sabzevar, IranSearch for more papers by this authorU. JOGER, U. JOGER State Natural History Museum, Pockelsstr. 10, 38106 Braunschweig, GermanySearch for more papers by this authorM. WINK, M. WINK Institute of Pharmacy and Molecular Biotechnology, Department of Biology, University of Heidelberg, Im Neuenheimer Feld 364, 69120 Heidelberg, GermanySearch for more papers by this author First published: 26 February 2010 https://doi.org/10.1111/j.1096-3642.2009.00553.xCitations: 7Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Abstract The Persian racerunner Eremias persica Blanford, 1875 is confined to the Iranian plateau, and forms one of the most widespread but rarely studied species of the family Lacertidae. With many local populations inhabiting a variety of habitats, and exhibiting considerable morphological, genetic, and ecological variations, it represents a species complex. We analysed sequences of mitochondrial cytochrome b and 12S ribosomal RNA (rRNA) genes derived from 13 geographically distant populations belonging to the E. persica complex. Using our knowledge of palaeogeographical events, a molecular clock was calibrated to assess the major events in fragmentation, radiation, and intraspecific variation. The sequence data strongly support a basal separation of the highland populations of western Iran from those of the open steppes and deserts, occurring in the east. The subsequent radiation, fragmentation, and evolution of these major assemblages have led to several discernable geographical lineages across the wide area of the Iranian plateau. The results indicate a middle-Miocene origin for the clade as a whole. The first split, isolating the western and eastern clades, appears to have occurred 11–10 Mya. Further fragmentations and divergence within the major clades began about 8 Mya, with an evolutionary rate of 1.6% sequence divergence per million years among the lineages in the genes studied (combined data set). Molecular and morphological data strongly support a taxonomic revision of this species complex. At least four of the discovered clades should be raised to species, and two to subspecies, rank. © 2009 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 158, 641–660. Citing Literature Volume158, Issue3March 2010Pages 641-660 RelatedInformation

Cancer cells are often immortal through up-regulation of the hTERT gene, which encodes the catalytic subunit of a special reverse transcriptase to overcome end-replication problem of chromosomes.This study demonstrates that papaverine, an isoquinoline alkaloid from the Papaveraceae, can overcome telomerase dependent immortality of HepG-2 cells that was used as a model of hepatocarcinoma.Although this alkaloid does not directly interact with telomeric sequences, papaverine inhibits telomerase through down-regulation of hTERT, which was analysed using thermal FRET and qRT-PCR, respectively.The IC 50 values for the reduction of both telomerase activity and hTERT expression was 60 µM, while IC 50 for cytotoxicity was 120 µM.Repeated treatments of the cells with very low non-toxic concentrations of papaverine resulted in growth arrest and strong reduction of population doublings after 40 days.This treatment induced senescent morphology in HepG-2 cells, which was evaluated by beta-galactosidase staining.Altogether, papaverine can be regarded as a promising model compound for drug design targeting cancer development.

Plant metabolites are valuable sources of novel therapeutic compounds. In an anti-telomerase screening study of plant secondary metabolites, the aporphine alkaloid boldine (1,10-dimethoxy-2,9-dihydroxyaporphine) exhibited a dose and time dependent cytotoxicity against hepatocarcinoma HepG-2 cells. Here we focus on the modes and mechanisms of the growth-limiting effects of this compound. Telomerase activity and expression level of some related genes were estimated by real-time PCR. Modes of cell death also were examined by microscopic inspection, staining methods and by evaluating the expression level of some critically relevant genes. The growth inhibition was correlated with down-regulation of the catalytic subunit of telomerase (hTERT) gene (p < 0.01) and the corresponding reduction of telomerase activity in sub-cytotoxic concentrations of boldine (p < 0.002). However, various modes of cell death were stimulated, depending on the concentration of boldine. Very low concentrations of boldine over a few passages resulted in an accumulation of senescent cells so that HepG-2 cells lost their immortality. Moreover, boldine induced apoptosis concomitantly with increasing the expression of bax/bcl2 (p < 0.02) and p21 (p < 0.01) genes. Boldine might thus be an interesting candidate as a potential natural compound that suppresses telomerase activity in non-toxic concentrations.

In a preliminary study screening anti-proliferative natural alkaloids, a very potent benzophenanthridine, chelidonine showed strong cytotoxicity in cancer cells. While several modes of death have been identified, most of anti-cancer attempts have focused on stimulation of cells to undergo apoptosis. Chelidonine seems to trigger multiple mechanisms in MCF-7 breast cancer cells. It induces both apoptosis and autophagy modes of cell death in a dose dependent manner. Alteration of expression levels of bax/bcl2, and dapk1a by increasing concentration of chelidonine approves switching the death mode from apoptosis induced by very low to autophagy by high concentrations of this compound. On the other hand, submicromolar concentrations of chelidonine strongly suppressed telomerase at both enzyme activity and hTERT transcriptional level. Long exposure of the cells to 50 nanomolar concentration of chelidonine considerably accelerated senescence. Altogether, chelidonine may provide a promising chemistry from nature to treat cancer.

In a preliminary screening study of natural alkaloids, boldine, an aporphine alkaloid, showed an interesting dose and time dependent anti-proliferative effect in several cancer cell lines. Cytotoxicity of boldine in human fibroblasts was considerably lower than the telomerase positive embryonic kidney HEK293 and breast cancer MCF-7 and MDA-MB-231 cells. Whether boldine can inhibit telomerase was investigated here using a modified quantitative real-time telomere repeat amplification protocol (q-TRAP). This test showed that boldine inhibits telomerase in cells treated with sub-cytotoxic concentrations. Telomerase inhibition occurs via down-regulation of hTERT, the catalytic subunit of the enzyme. Boldine changed the splicing variants of hTERT towards shorter non-functional transcripts as well. A direct interaction of boldine with the enzyme may also be involved, though thermal FRET method did not detect any substantial interaction between boldine and synthetic telomere sequences. This study advocates boldine as a valuable candidate for telomerase-targeted cancer care. This study suggests that derivatives of boldine could be potent anti-cancer drugs.

This report focuses on the application of zinc oxide nanoparticles (ZnO NPs) carrying phycomolecule ligands as a novel plant growth promoter aimed at increasing the crop productivity of purslane (&lt;em&gt;Portulaca oleracea&lt;/em&gt; L.). Experiments were performed under controlled greenhouse conditions using a completely randomized design with nine replications. Purslane seeds were treated with four concentrations of ZnO NPs (0, 10, 100, and 500 mg L&lt;sup&gt;−1&lt;/sup&gt;) and four concentrations of bulk ZnO (0, 10, 100, and 500 mg L&lt;sup&gt;−1&lt;/sup&gt;). The ultrastructural characteristics of the leaves of the plants treated with of 500 mg L&lt;sup&gt;−1&lt;/sup&gt; ZnO NPs were determined using transmission electron microscopy (TEM). The results indicated that the treatment with ZnO NPs increased the content of chlorophyll &lt;em&gt;a&lt;/em&gt; and chlorophyll &lt;em&gt;b&lt;/em&gt;, carotenoids, and total phenolic and flavonoid compounds significantly more than the treatment with bulk ZnO. Our findings also showed that the application of high concentrations of ZnO NPs is the most effective strategy to considerably induce the antioxidant capacity and enzymes of purslane plants. Furthermore, the seed germination percentage and sprout growth rates were significantly higher in the plants treated with 500 mg L&lt;sup&gt;−1&lt;/sup&gt; of ZnO NPs (100% ±0.00), compared to the control plants (93.33% ±1.66). The TEM images revealed the concentration of ZnO NPs and cell membrane rupture, as well as a deformation in the shape of chloroplasts and a decrease in their number in the plants treated with 500 mg L&lt;sup&gt;−1&lt;/sup&gt; ZnO NPs, compared to the control plants. Owing to their toxicity, high concentrations of ZnO NPs lead to oxidative stress in plants. Thus, our findings provide a new alternative strategy for increasing crop productivity, i.e., the application of ZnO NPs as a novel plant growth booster, in comparison with the bulk ZnO treatment.

The industrial world exposes living organisms to a variety of metal pollutants. Here we investigated whether such elements affect G-rich sequences susceptible to fold into G-quadruplex (GQ) structures. Thermal stability and conformation of these oligoncleotides was studied at various molar ratios of a variety of heavy metal salts using thermal FRET, transition-FRET (t-FRET) and circular dichroism. Metal ions affected the thermal stability of the GQs to different extents; some metals had no effect on Tm while other metals caused small to moderate changes in Tm at 1:1 or 1:10 molar ratio. While most of the metals had no major effect, Al3+, Cd2+, Pb2+, Hg2+ and Zn2+ altered the thermal stability and structural features of the GQs. Some metals such as Pb2+ and Hg2+ exhibit differential interactions with telomere, c-myc and c-kit GQs. Overall, toxic heavy metals affect G-quadruplex stability in a sequence and topology dependent manner. This study provides new insight into how heavy metal exposure may affect gene expression and cellular responses.

To identify the altered gene expression patterns in squamous cell carcinoma of esophagus (ESCC) in relation to adjacent normal esophageal epithelium.Total RNA was extracted using SV total RNA isolation kit from snap frozen tissues of ESCC samples and normal esophageal epithelium far from the tumor. Radio-labeled cDNA were synthesized from equal quantities of total RNAs of tumor and normal tissues using combinations of 24 arbitrary 13-mer primers and three different anchoring oligo-dT primers and separated on sequencing gels. cDNA with considerable different amounts of signals in tumor and normal tissue were reamplified and cloned. Using southern blot, the clones of each band were controlled for false positive results caused by probable heterogeneity of cDNA population with the same size. Clones that confirmed differential expression by slot blot selected for sequencing and northern analysis. Corresponding full-length gene sequences was predicted using human genome project data, related transcripts were translated and used for various protein/motif searches to speculate their probable functions.The 97 genes showed different levels of cDNA in tumor and normal tissues of esophagus. The expression of mal gene was remarkably down regulated in all 10 surveyed tumor tissues. Akr1c2, a member of the aldo-keto reductase 1C family, which is involved in metabolism of sex hormones and xenobiotics, was up-regulated in 8 out of 10 inspected ESCC samples. Rab11a, RPL7, and RPL28 showed moderate levels of differential expression. Many other cDNAs remained to further studies.The mal gene which is switched-off in all ESCC samples can be considered as a tumor suppressor gene that more studies in its regulation may lead to valuable explanations in ESCC development. Akr1c2 which is up-regulated in ESCC probably plays an important role in tumor development of esophagus and may be proposed as a potential molecular target in ESCC treatments. Differential display technique in spite of many disadvantages is still a valuable technique in gene function exploration studies to find new candidates for improved ones like gene chips.

Journal of Zoological Systematics and Evolutionary ResearchVolume 50, Issue 3 p. 220-229 Molecular phylogeny and intraspecific differentiation of the Eremias velox complex of the Iranian Plateau and Central Asia (Sauria, Lacertidae) Molecular phylogeny and intraspecific differentiation of the Eremias velox complex of the Iranian Plateau and Central Asia (Sauria, Lacertidae) Eskandar R. Pouyani, Eskandar R. Pouyani Department of Biology, Faculty of Science, Hakim Sabzevari University, Sabzevar, Iran Department of Biology, Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, GermanySearch for more papers by this author Sakineh K. Noureini, Sakineh K. Noureini Department of Biology, Faculty of Science, Hakim Sabzevari University, Sabzevar, IranSearch for more papers by this author Nasrullah R. Pouyani, Nasrullah R. Pouyani Department of Biology, Faculty of Science, Razi University, Kermanshah, IranSearch for more papers by this authorUlrich Joger, Ulrich Joger State Natural History Museum, Braunschweig, GermanySearch for more papers by this authorMichael Wink, Michael Wink Department of Biology, Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, GermanySearch for more papers by this author Eskandar R. Pouyani, Eskandar R. Pouyani Department of Biology, Faculty of Science, Hakim Sabzevari University, Sabzevar, Iran Department of Biology, Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, GermanySearch for more papers by this author Sakineh K. Noureini, Sakineh K. Noureini Department of Biology, Faculty of Science, Hakim Sabzevari University, Sabzevar, IranSearch for more papers by this author Nasrullah R. Pouyani, Nasrullah R. Pouyani Department of Biology, Faculty of Science, Razi University, Kermanshah, IranSearch for more papers by this authorUlrich Joger, Ulrich Joger State Natural History Museum, Braunschweig, GermanySearch for more papers by this authorMichael Wink, Michael Wink Department of Biology, Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, GermanySearch for more papers by this author First published: 14 May 2012 https://doi.org/10.1111/j.1439-0469.2012.00662.xCitations: 10 Corresponding author: Eskandar R. Pouyani ([email protected]; [email protected]) Contributing authors: Sakineh K. Noureini ([email protected]), Nasrullah R. Pouyani ([email protected]), Ulrich J. ([email protected]), Michael Wink ([email protected]) Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Abstracten The Central Asian racerunner, Eremias velox, is a widely distributed lizard of the Eurasian lacertid genus Eremias. Nucleotide sequences of mitochondrial genes, cyt b and 12S rDNA from 13 geographically distant localities in Iran and Central Asia, were analysed. Phylogenetic analyses of the sequence data unambiguously recovered five major clades within the E. velox complex with a high level of genetic divergence, indicating long periods of isolation. The basal position of the Iranian clades in the phylogenetic trees suggests that the E. velox clade originated on the Iranian plateau in the Middle Miocene. According to our calibrations, the northern Iranian clade diverged first some 10–11 Ma and that the Central Asian lineages split from the northeastern Iranian lineage approximately 6 Ma, most likely as a result of uplifting of the Kopet-Dagh Mountains in the northern margin of the Iranian plateau. Topology of the phylogenetic trees, combined with the degree of the genetic distances among the independent lineages recovered in this study, provide a solid foundation for a fundamental revision of the taxonomic status of the major clades within this species complex. Zusammenfassungde Eremias velox (Gattung Eremias) aus Zentralasien gehört zu den weit verbreiteten Eidechsen Eurasiens. Nucleotidsequenzen mitochondrialer DNA (Cytochrom b und 12 S rDNA) von 13 geographisch unterschiedlichen Standorten aus dem Iran und Zentralasien wurden genauer analysiert. Die phylogenetische Auswertung fand 5 genetische Linien innerhalb des E. velox– Komplexes, die sich durch große genetische Distanzen unterscheiden und auf lange Zeiten der Isolation hinweisen. Innerhalb der Gesamtphylogenie liegen die iranischen Kladen basal. Dies deutet darauf hin, dass der E. velox Komplex im mittleren Miozän auf dem iranischen Plateau entstand. Aufgrund von Kalkulationen über eine molekulare Uhr wird angenommen, dass die nordiranischen Linien von 10–11 Millionen Jahren entstanden, während sich die zentralasiatischen Taxa vor 6 Ma abzweigten. Dies wurde vermutlich durch die Entstehung der Kopet-Dagh-Berge am Nordrand des iranischen Plateaus ausgelöst. 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Sulfur mustard (SM) is a vesicant chemical warfare agent, and a very potent alkylating agent. SM exerts its cytotoxicity via direct alkylation of biomacromolecules, and overproduction of reactive oxygen species (ROS). Previous studies have shown that SM-induced oxidative stress has adverse effects on antioxidant defense system, and damages lipids and proteins. The aim of this study was to investigate the effect of SM-induced oxidative stress on DNA damage, and cellular senescence in SM-exposed victims. For this purpose, MDA levels as a measure of oxidative stress in the serum, 8-oxo-dG content of the genomic DNA, and OGG1 expression as two biomarkers of oxidative DNA damage, as well as, telomere length, and p16INK4a expression as two biomarkers of cellular senescence were measured in the peripheral blood leukocytes of 215 males who were exposed to SM 20 to 25 years ago, and 53 unexposed healthy males as the control group. Our results indicated that the levels of 8-oxo-dG, and OGG1 mRNA expression were significantly higher in SM-exposed individuals. Furthermore, a significant increase in the expression of p16INK4a was observed in SM-exposed patients, and leukocyte telomere length (LTL) was also significantly shorter in severe/very severe cases of SM-exposed patients when compared with unexposed controls. In conclusion, our data indicate that oxidative DNA damage is higher in SM-exposed patients, and their immune system has subjected to cellular senescence.

Telomerase, the enzyme responsible for cell immortality, is an important target in anti-cancer drug discovery. Boldine, an abundant aporphine alkaloid of Peumus boldus, is known to inhibit telomerase at non-toxic concentrations. Cytotoxicity of N-benzylsecoboldine hydrochloride (BSB), a synthetic derivative of boldine, was determined using the MTT method in MCF7 and MDA-MB231 cells. Aliquots of cell lysates were incubated with various concentrations of BSB in qTRAP (quantitative telomere repeat amplification protocol)-ligand experiments before substrate elongation by telomerase or amplification by hot-start Taq polymerase. The crystal structure of TERT, the catalytic subunit of telomerase from Tribolium castaneum, was used for docking and molecular dynamics analysis. The qTRAP-ligand data gave an IC50 value of about 0.17 &plusmn; 0.1 &micro;M for BSB, roughly 400 times stronger than boldine, while the LD50 in the cytotoxicity assays were 12.5 and 21.88 &micro;M, respectively, in cells treated for 48 h. Although both compounds interacted well with the active site, MD analysis suggests a second binding site with which BSB interacts via two hydrogen bonds, much more strongly than boldine. Theoretical analyses also evaluated the IC50 for BSB as submicromolar. BSB, with greater hydrophobicity and flexibility than boldine, represents a promising structure to inhibit telomerase at non-toxic concentrations.

Telomeres, the specialized dynamic structures at chromosome ends, regularly shrink with every replication. Thus, they function as an internal molecular clock counting down the number of cell divisions. However, most cancer cells escape this limitation by activating telomerase, which can maintain telomere length. Previous studies showed that the benzylisoquinoline alkaloid chelidonine stimulates multiple modes of cell death and strongly down-regulates telomerase. It is still unknown if down-regulation of telomerase by chelidonine boosts substantial telomere shortening. The breast cancer cell line MCF7 was sequentially treated with very low concentrations of chelidonine over several cell passages. Telomere length and telomerase activity were measured by a monochrome multiplex quantitative PCR and a q-TRAP assay, respectively. Changes in population size and doubling time correlated well with telomerase inhibition and telomere shortening. MCF7 cell growth was arrested completely after three sequential treatments with 0.1 μM chelidonine, each ending after 48 h, while telomere length was reduced to almost 10% of the untreated control. However, treatment with 0.01 μM chelidonine did not have any apparent consequence. In addition to dose and time dependent telomerase inhibition, chelidonine changed the splicing pattern of hTERT towards non-enzyme coding isoforms of the transcript. In conclusion, telomere length and telomere stability are strongly affected by chelidonine in addition to microtubule formation.

Rosa damascena Mill., farmakolojik aktiviteleri yüksek olan

The study utilizes monodisperse platinum nanoparticles (Pt NPs) biosynthesized from Punica granatum crusts as anti-tumor agents on the human breast cancer cell line, MCF-7. The obtained Pt NPs were fully characterized using the UV–vis spectrum (UV–vis), transmission electron microscopy (TEM), X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM) and Fourier transformation infrared spectroscopy (FTIR). Effectiveness of the Pt NPs was determined by cell viability, propidium iodide staining test, flow cytometry and comet tests on the MCF-7 cancer cell line. Cell survival percentage was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The biosynthesized monodisperse platinum nanoparticles inhibited MCF-7 proliferation with an IC50 of 17.84 μg/ml after 48 h of incubation. Propidium iodide staining demonstrated that the monodisperse Pt NPs induced apoptosis by means of molecular DNA fragmentation.

Background: Traditional medicine is inspiring in drug development research. Epilobium parviflorum extracts have shown promising therapeutic effects on prostate cancer cells. The similarities between breast and prostate cancers at molecular and metabolic levels prompted us to explore its effects on human breast cancer. Methods: The root, aerial parts and flowers of the plant were, collected and dried separately at ambient temperature and away from direct sunlight. The aquatic and methanolic extracts of each part was prepared. The effect of each extract on the growth of MCF-7 breast carcinoma cells and HEK293 normal cell line was evaluated, using MTT assay. Each experiment was repeated at least three times independently. The IC50 values for each treatment time point were analyzed, using ANOVA at P&lt;0.05. Results: While none of the extracts had considerable toxicity on normal HEK293 cells, some of them showed varying levels of toxicity on the MCF-7 cells. The methanolic extracts were more cytotoxic than the aqueous counterpart. The roots’ methanolic extract showed the strongest cytotoxicity on the MCF-7 cells in a dose and exposure time dependent manner. The IC50 after 48 hours of treatment was determined at 73µg/ml. Conclusion: This study is the first to demonstrate that Epilobium parviflorum had a strong growth inhibiting property on MCF-7 cell line, as a potential model to treat human breast cancer cells. The most cytotoxic effect was noted for the methanolic root extract. Determination of the effective biochemical constituents of the extract against cancer cells is the focus of our future research.

Telomerase activation is one of the main pathways to immortalize cancer cells. In many kinds of cancer cells, this special reverse transcriptase stabilizes and elongates telomeres and prevents telomere erosion that naturally occurs in every cell division. Esophageal cancer is the fifth most frequent cause of cancer death worldwide, and is highly associated with alcohol, smoking, cultural habits, and environmental factors. Telomerase has been suggested as a tumor marker and a molecular target for drug design in several kinds of cancers. In this work telomerase activation was inspected among Iranian patients with Esophageal Squamous Cell Carcinoma (ESCC), and detected in 90% of samples of different stages. This may be an indication that telomerase activation happens in an initial step in the development of ESCC. Although there is no correlation between telomerase activity and the progress of ESCC, it could be considered as a good tumor marker in ESCC. Telomerase activity te! sts are suggested for screening purposes in high risk areas for ESCC, which can be easily done on a small amount of scrapped samples of esophageal mucosa. It is also possible that ESCC results from incomplete differentiation or a failure in telomerase gene switching off that normally occurs during the differentiation of esophageal epithelial cells.

AbstractHuman MRP1 protein plays a vital role in cancer multidrug resistance. Coumarins show promising pharmacological properties. Virtual screening, ADMET, molecular docking and molecular dynamics (MD) simulations were utilized as pharmacoinformatic tools to identify potential MRP1 inhibitors among coumarin derivatives. Using in silico ADMET, 50 hits were further investigated for their selectivity toward the nucleotide-binding domains (NBDs) of MRP1 using molecular docking. Accordingly, coumarin, its symmetrical ketone derivative Lig. No. 4, and Reversan were candidates for focused docking study with the NBDs domains compared with ATP. The result indicates that Lig. No. 4, with the best binding score, interacts with NBDs via hydrogen bonds with residues: GLN713, LYS684, GLY683, CYS682 in NBD1, and GLY1432, GLY771, SER769 and GLN1374 in NBD2, which mostly overlap with ATP binding residues. Moreover, doxorubicin (Doxo) was docked to the transmembrane domains (TMDs) active site of MRP1. Doxo interaction with TMDs was subjected to MD simulation in the NBDs free and occupied with Lig. No. 4 states. The results showed that Doxo interacts more strongly with TMD residues in inward facing feature of TMDs helices. However, when Lig. No. 4 exists in NBDs, Doxo interactions are different, and TMD helices show more outward-facing conformation. This result may suggest a partial competitive inhibition mechanism for the Lig. No. 4 on MRP1 compared with ATP. So, it may inhibit active complex formation by interfering with ATP entrance to NBDs and locking MRP1 conformation in outward-facing mode. This study suggests a valuable coumarin derivative that can be further investigated for potent MRP1 inhibitors.Communicated by Ramaswamy H. SarmaHIGHLIGHTSVirtual screening scored a symmetrical ketone derivative IUPAC ([2-[(1E, 4E)-5-(2-acetyloxyphenyl)-3-oxopenta-1, 4-dienyl] phenyl] acetate); PubChem CID 5468558 (Lig. No. 4 in this study) as the best candidate among coumarins to inhibit MRP1.This compound binds to NBD1 and NBD2 of ABC transporters via hydrogen bonds shared with residues that are also involved in the ATP binding.This result, if not at all, suggest a partial competitive inhibition mechanism for Lig. No. 4 on the MRP1 protein.Molecular dynamics simulation study reveals different doxorubicin binding modes in interaction with MRP1 transmembrane domain in free and occupied NBDs with Lig. No. 4.Lig. No. 4 is a valuable candidate for further drug development studies to suppress drug resistance.Keywords: Multidrug resistanceABC transporter membrane proteinsMRP1coumarin derivativesmolecular dockingmolecular dynamicscancer AcknowledgmentThis research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.Ethical approvalEthical considerations were not applicable as the study has focused on a computational model only.Author contributionsAll authors have substantially contributed to this study. Parisa Shahpouri: conducting the research and investigation process, molecular docking; writing the original draft to fulfill her MSc degree. Havva Mehralitabar: molecular docking, molecular dynamics simulation, analysis, visualization, discussion and literature review. Sakineh Kazemi Noureini: conceptualization; oversight and leadership responsibility for the research activity planning and execution; review and editing of the manuscript. Mitra Kheirabadi: advice, training and data validation; review and editing of the manuscript.Disclosure statementNo potential conflict of interest was reported by the author(s).Additional informationFundingThe author(s) reported there is no funding associated with the work featured in this article.

G-rich sequences have the potential to fold into G-quadruplexes (GQs). G-quadruplexes, particularly those positioned in the regulatory regions of proto-oncogenes, have recently garnered attention in anti-cancer drug design. A thermal FRET assay was employed to conduct preliminary screening of various alkaloids, aiming to identify stronger interactions with a specific set of G-rich double-labeled oligonucleotides in both K + and Na + buffers. These oligonucleotides were derived from regions associated with Kit, Myc, Ceb, Bcl2, human telomeres, and potential G-quadruplex forming sequences found in the Nrf2 and Trf2 promoters. Palmatine generally increased the stability of different G-rich sequences into their folded GQ structures, more or less in a concentration dependent manner. The thermal stability and interaction of palmatine was further studied using transition FRET (t-FRET), CD and UV-visible spectroscopy and molecular dynamics simulation methods. Palmatine showed the strongest interaction with T RF2 in both K+ and Na+ buffers even at equimolar concentration ratio. T-FRET studies revealed that palmatine has the potential to disrupt double-strand formation by the T RF2 sequence in the presence of its complementary strand. Palmatine exhibits a stronger interaction with G-rich strand DNA, promoting its folding into G-quadruplex structures. It is noteworthy that palmatine exhibits the strongest interaction with T RF2, which is the shortest sequence among the G-rich oligonucleotides studied, featuring only one nucleotide for two of its loops. Palmatine represents a suitable structure for drug design to develop more specific ligands targeting G-quadruplexes. Whether palmatine can also affect the expression of the T RF2 gene requires further studies.Communicated by Ramaswamy H. Sarma.

Introduction: Breast cancer is the second and the most common leading cause of death in the world that has attracted special attention in drug design and discovery research projects.Here we investigated the cytotoxic effects of two closely related plant secondary metabolites from the family of benzyl isoquinolinic alkaloids on MCF-7 and MDA-MB231 cell lines.Materials and Methods: Cytotoxicity of the derivatives in both MCF-7 and MDA-MB231 cell lines were measured by using MTT test.Cells were seeded on 96 well plates and were treated with different concentrations of the alkaloid.After 48 hours, MTT was added at final concentration of 0.5 mg/ml and incubated for 4 hours.The purple dye developed by living cells was dissolved in DMSO and measured by Powerwave XS2 and Biotek plate reader.IC50 values were calculated by Gen5 software of the machine using at least 3 independent repeats of the experiment.Results: MTT tests showed that with increasing concentration of these alkaloids, the percentage of cell survival significantly decreased.The obtained data indicated that Palmatine had an IC50= 186μM in the MCF-7 cell line and 91μM in MDA-MB231 cell line.On the other hand, Corydaline with IC50= 55μM in MCF-7 and 42μM in MDA-MB231 has a stronger cytotoxicity than Palmatine.Conclusions: Comparing the MTT results, we find that corydaline was stronger than palmatine and it was slightly more cytotoxic in MDA-MB231 than MCF-7.

The expression of telomerase is essential for cells to be immortalized, and most immortal cell lines possessed telomerase activity. Using the cell fusion technique, it has been shown that mortal and telomerase-negative phenotypes of normal cells are dominant over immortal and telomerase-positive phenotypes, suggesting that the normal cells possessed dominant repressor-type activity for telomerase expression. Several telomerase-negative immortal human cell lines were reported, in which telomerase-independent mechanisms was supposed to maintain telomere length. We aimed at seeing whether the telomerase-negative phenotype of these immortal cells is dominant over telomerase-positive phenotype of other immortal cells in correlation with cellular mortality. Results showed that, when telomerase-positive and -negative immortal parental cell lines belonging to the different complementation groups were fused, telomerase-negative mortal hybrid clones arose, i.e. telomerase-negative phenotype was dominant as well as mortal phenotype. However, when immortal hybrid cells arose from telomerase-positive and -negative immortal parents belonging to either the same or different complementation groups, they were all telomerase-positive, i.e. telomerase-negative phenotype appeared to be recessive. Telomerase-negative immortal hybrid was never established from any combinations between telomerase-negative and -positive immortal parental cells.

Introduction Boldine is an aporphine alkaloid that is found in high amount in boldo tree and Lindera aggregate.It is used as homeopathic and herbal medicine that has various antioxidant and anti-cancer.Our previous study has reported anti-telomerase effect of this alkaloid.In order to enhance this valuable inhibitory effect, several new synthetic substitutes have been synthesized in Prof. Bruce Cassel's Lab. Materials and Methods: LD50 values of the derivatives in both MCF-7 and MDA-MB231 cell lines were measured by MTT test.Cells seeded in 96 well plates were treated with six different concentrations of each compound (400μM, 200μM, 100μM, 50μM, 25μM and 12.5μM).MTT solution was added after 48 hours, then incubated for four hours.IC50 values were measured through Gen5 software of Powerwave XS2 Biotek plate reader.Data was collected by at least 3 independent repeats of the experiment.Results: The collected data showed that B2 with LD50= 16.25 μM in MCF-7 and 21.88 μM in MDA-MB231 was the most potent anti-proliferative derivative.The IC50 values for the other three compounds, B3, B4 and B5 in the MCF-7 cell line were 156.4μM, 610μM and 252.5μM, and in the MDA-MB231 cell line were 107.5μM, 363.8μM, 217.5μM, respectively.Also in microscopic observations, the survival percentage of both cell lines in presence of B2 was significantly reduced dose dependently.Conclusions: B2, the most potent anti-proliferative derivative of boldine, showed stronger cytotoxic effects in MCF-7 than MDA-MB231 cell line.The higher hydrophobicity and more flexible structure of this derivative may play critical role in enhancing the interaction with important molecular targets inside the cell.