scholarly journals Novel Tandem Biotransformation Process for the Biosynthesis of a Novel Compound, 4-(2,3,5,6-Tetramethylpyrazine-1)-4′-Demethylepipodophyllotoxin

2011 ◽  
Vol 77 (9) ◽  
pp. 3023-3034 ◽  
Author(s):  
Ya-Jie Tang ◽  
Wei Zhao ◽  
Hong-Mei Li
Keyword(s):  
Cell Line ◽  
Alternaria Alternata ◽  
Water Solubility ◽  
Tumor Cell Line ◽  
Gibberella Fujikuroi ◽  
Hydroxyl Group ◽  
Epithelial Cell Line ◽  
The Novel ◽  
Content Type ◽  
Biotransformation Process

ABSTRACTAccording to the structure of podophyllotoxin and its structure-function relationship, a novel tandem biotransformation process was developed for the directional modification of the podophyllotoxin structure to directionally synthesize a novel compound, 4-(2,3,5,6-tetramethylpyrazine-1)-4′-demethylepipodophyllotoxin (4-TMP-DMEP). In this novel tandem biotransformation process, the starting substrate of podophyllotoxin was biotransformed into 4′-demethylepipodophyllotoxin (product 1) with the demethylation of the methoxyl group at the 4′ position byGibberella fujikuroiSH-f13, which was screened out from Shennongjia prime forest humus soil (Hubei, China). 4′-Demethylepipodophyllotoxin (product 1) was then biotransformed into 4′-demethylpodophyllotoxone (product 2) with the oxidation of the hydroxyl group at the 4 position byAlternaria alternataS-f6, which was screened out from the gatheredDysosma versipellisplants in the Wuhan Botanical Garden, Chinese Academy of Sciences. Finally, 4′-demethylpodophyllotoxone (product 2) and ligustrazine were linked with a transamination reaction to synthesize the target product 4-TMP-DMEP (product 3) byAlternaria alternataS-f6. Compared with podophyllotoxin (i.e., a 50% effective concentration [EC50] of 529 μM), the EC50of 4-TMP-DMEP against the tumor cell line BGC-823 (i.e., 0.11 μM) was significantly reduced by 5,199 times. Simultaneously, the EC50of 4-TMP-DMEP against the normal human proximal tubular epithelial cell line HK-2 (i.e., 0.40 μM) was 66 times higher than that of podophyllotoxin (i.e., 0.006 μM). Furthermore, compared with podophyllotoxin (i.e., logP= 0.34), the water solubility of 4-TMP-DMEP (i.e., logP= 0.66) was significantly enhanced by 94%. For the first time, the novel compound 4-TMP-DMEP with superior antitumor activity was directionally synthesized from podophyllotoxin by the novel tandem biotransformation process developed in this work.

A New Human Breast Tumor Cell Line

1975 ◽  
Vol 33 ◽  
pp. 388-389
Author(s):  
R.E. Nordquist ◽  
R.M. Wasik ◽  
P.J. Riggs ◽  
P.L. Munson ◽  
F.B. Schafer
Keyword(s):  
Electron Microscopy ◽  
Cell Line ◽  
Calf Serum ◽  
Tumor Cell Line ◽  
Electron Microscopic Examination ◽  
Epithelial Cell Line ◽  
Electron Microscopic ◽  
Fixed Tissue ◽  
Excised Tissue ◽  
Caucasian Female

An infiltrating ductal cell carcinoma was removed from the breast of a postmenopausal Caucasian female. The excised tissue was divided into three parts; one part for electron microscopy, one part for tissue culture and the remainder frozen for immunological studies.The tissue for culture was minced finely with sterile razor blades and cultured in Falcon flasks containing Eagel's MEM supplemented with 10% heat denatured fetal calf serum. The tissue for electron microscopy was fixed in 6.25% glutaraldehyde in 0.1 M PO4 buffer plus 5% sucrose and postfixed in 1% OsO4 in the same buffer. The fixed tissue was dehydrated in graded ethanol and embedded in Spurr.The tissue which was cultured began to grow out after approximately six weeks and became a continuous epithelial cell line which was designated BOT-2 (Breast Original Tumor). Electron microscopic examination revealed that these cells had epithelial characteristics, i.e. the presence of tonofilaments and well formed desmosomes.

scholarly journals Differential Response of the Human Renal Proximal Tubular Epithelial Cell Line HK-2 to Shiga Toxin Types 1 and 2

2011 ◽  
Vol 79 (9) ◽  
pp. 3527-3540 ◽  
Author(s):  
Erin K. Lentz ◽  
Dinorah Leyva-Illades ◽  
Moo-Seung Lee ◽  
Rama P. Cherla ◽  
Vernon L. Tesh
Keyword(s):  
Epithelial Cell ◽  
Cell Line ◽  
Shiga Toxin ◽  
Renal Damage ◽  
Diarrheal Disease ◽  
Shigella Dysenteriae ◽  
Parp Cleavage ◽  
Epithelial Cell Line ◽  
Cytotoxic Action ◽  
Content Type

ABSTRACTShiga toxins (Stxs) are expressed by the enteric pathogensShigella dysenteriaeserotype 1 and certain serotypes ofEscherichia coli. Stx-producing bacteria cause bloody diarrhea with the potential to progress to acute renal failure. Stxs are potent protein synthesis inhibitors and are the primary virulence factors responsible for renal damage that may follow diarrheal disease. We explored the use of the immortalized human proximal tubule epithelial cell line HK-2 as anin vitromodel of Stx-induced renal damage. We showed that these cells express abundant membrane Gb3and are differentially susceptible to the cytotoxic action of Stxs, being more sensitive to Shiga toxin type 1 (Stx1) than to Stx2. At early time points (24 h), HK-2 cells were significantly more sensitive to Stxs than Vero cells; however, by 72 h, Vero cell monolayers were completely destroyed while some HK-2 cells survived toxin challenge, suggesting that a subpopulation of HK-2 cells are relatively toxin resistant. Fluorescently labeled Stx1 B subunits localized to both lysosomal and endoplasmic reticulum (ER) compartments in HK-2 cells, suggesting that differences in intracellular trafficking may play a role in susceptibility to Stx-mediated cytotoxicity. Although proinflammatory cytokines were not upregulated by toxin challenge, Stx2 selectively induced the expression of two chemokines, macrophage inflammatory protein-1α (MIP-1α) and MIP-1β. Stx1 and Stx2 differentially activated components of the ER stress response in HK-2 cells. Finally, we demonstrated significant poly(ADP-ribose) polymerase (PARP) cleavage after exposure to Stx1 or Stx2. However, procaspase 3 cleavage was undetectable, suggesting that HK-2 cells may undergo apoptosis in response to Stxs in a caspase 3-independent manner.

scholarly journals Avibactam and Inhibitor-Resistant SHV β-Lactamases

2015 ◽  
Vol 59 (7) ◽  
pp. 3700-3709 ◽  
Author(s):  
Marisa L. Winkler ◽  
Krisztina M. Papp-Wallace ◽  
Magdalena A. Taracila ◽  
Robert A. Bonomo
Keyword(s):  
Alternative Pathway ◽  
Proton Acceptor ◽  
Hydroxyl Group ◽  
The Novel ◽  
Content Type ◽  
Class A ◽  
Important Amino Acid ◽  
Class D ◽  
Continued Use ◽  
Lactamase Inhibitor

ABSTRACTβ-Lactamase enzymes (EC 3.5.2.6) are a significant threat to the continued use of β-lactam antibiotics to treat infections. A novel non-β-lactam β-lactamase inhibitor with activity against many class A and C and some class D β-lactamase variants, avibactam, is now available in the clinic in partnership with ceftazidime. Here, we explored the activity of avibactam against a variety of characterized isogenic laboratory constructs of β-lactamase inhibitor-resistant variants of the class A enzyme SHV (M69I/L/V, S130G, K234R, R244S, and N276D). We discovered that the S130G variant of SHV-1 shows the most significant resistance to inhibition by avibactam, based on both microbiological and biochemical characterizations. Using a constant concentration of 4 mg/liter of avibactam as a β-lactamase inhibitor in combination with ampicillin, the MIC increased from 1 mg/liter forblaSHV-1to 256 mg/liter forblaSHV S130Gexpressed inEscherichia coliDH10B. At steady state, thek2/Kvalue of the S130G variant when inactivated by avibactam was 1.3 M−1s−1, versus 60,300 M−1s−1for the SHV-1 β-lactamase. Under timed inactivation conditions, we found that an approximately 1,700-fold-higher avibactam concentration was required to inhibit SHV S130G than the concentration that inhibited SHV-1. Molecular modeling suggested that the positioning of amino acids in the active site of SHV may result in an alternative pathway of inactivation when complexed with avibactam, compared to the structure of CTX-M-15–avibactam, and that S130 plays a role in the acylation of avibactam as a general acid/base. In addition, S130 may play a role in recyclization. As a result, we advance that the lack of a hydroxyl group at position 130 in the S130G variant of SHV-1 substantially slows carbamylation of the β-lactamase by avibactam by (i) removing an important proton acceptor and donator in catalysis and (ii) decreasing the number of H bonds. In addition, recyclization is most likely also slow due to the lack of a general base to initiate the process. Considering other inhibitor-resistant mechanisms among class A β-lactamases, S130 may be the most important amino acid for the inhibition of class A β-lactamases, perhaps even for the novel diazabicyclooctane class of β-lactamase inhibitors.

Cytotoxic Activity Evaluation on Breast Cells of Guest-host Complexes Containing Artemisinin

2019 ◽  
Vol 70 (8) ◽  
pp. 2843-2846 ◽  
Author(s):  
Denisa Circioban ◽  
Ioana Zinuca Pavel ◽  
Adriana Ledeti ◽  
Ionut Ledeti ◽  
Corina Danciu ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Water Solubility ◽  
Breast Adenocarcinoma ◽  
Breast Epithelial Cell ◽  
Epithelial Cell Line ◽  
Potential Candidate ◽  
In Vivo Testing ◽  
Breast Cells

Artemisinin is a sesquiterpene lactone with vastly proved anti-cancer effects and a low toxicity profile. However, the compound has poor water solubility, bioavailability and a short half-life. As such, the present paper aims to evaluate the cytotoxic effect on breast cells of three guest-host inclusion complexes containing artemisinin as the active compound and different cyclodextrins as hosts. These were tested using two different concentrations (i.e. 12.5 mM and 25 mM) and three cell lines, namely two human breast adenocarcinoma cell lines (MCF7 and MDA-MB-231) and one human non-tumorigenic breast epithelial cell line (MCF10A) employing the colorimetric microculture tetrazolium assay. After a 72h stimulation period, the most promising results were obtained for the complex containing artemisinin and Heptakis(2,3,6-tri-O-methyl)-b-cyclodextrin, the cell viability decrease being significant for the estrogen positive MCF7 cell line (80.0 � 2.3 %), making the complex a potential candidate for further in vivo testing.

scholarly journals Modulation of Porcine β-Defensins 1 and 2 upon Individual and Combined Fusarium Toxin Exposure in a Swine Jejunal Epithelial Cell Line

2013 ◽  
Vol 79 (7) ◽  
pp. 2225-2232 ◽  
Author(s):  
Murphy Lam-Yim Wan ◽  
Chit-Shing Jackson Woo ◽  
Kevin J. Allen ◽  
Paul C. Turner ◽  
Hani El-Nezami
Keyword(s):  
Epithelial Cell ◽  
Mrna Expression ◽  
Cell Line ◽  
Defense Mechanisms ◽  
Fumonisin B1 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Epithelial Cell Line ◽  
Content Type ◽  
Antimicrobial Effects ◽  
Food And Feed

ABSTRACTDefensins are small antimicrobial peptides (AMPs) that play an important role in the innate immune system of mammals. Since the effect of mycotoxin contamination of food and feed on the secretion of intestinal AMPs is poorly understood, the aim of this study was to elucidate the individual and combined effects of four commonFusariumtoxins, deoxynivalenol (DON), nivalenol (NIV), zearalenone (ZEA), and fumonisin B1 (FB1), on the mRNA expression, protein secretion, and corresponding antimicrobial effects of porcine β-defensins 1 and 2 (pBD-1 and pBD-2) using a porcine jejunal epithelial cell line, IPEC-J2. In general, upregulation of pBD-1 and pBD-2 mRNA expression occurred following exposure toFusariumtoxins, individually and in mixtures (P< 0.05). However, no significant increase in secreted pBD-1 and pBD-2 protein levels was observed, as measured by enzyme-linked immunosorbent assay (ELISA). Supernatants from IPEC-J2 cells exposed to toxins, singly or in combination, however, possessed significantly less antimicrobial activity againstEscherichia colithan untreated supernatants. When single toxins and two-toxin combinations were assessed, toxicity effects were shown to be nonadditive (including synergism, potentiation, and antagonism), suggesting interactive toxin effects when cells are exposed to mycotoxin combinations. The results show thatFusariumtoxins, individually and in mixtures, activate distinct antimicrobial defense mechanisms possessing the potential to alter the intestinal microbiota through diminished antimicrobial effects. Moreover, by evaluating toxin mixtures, this improved understanding of toxin effects will enable more effective risk assessments for common mycotoxin combinations observed in contaminated food and feed.

scholarly journals Untargeted Characterization of Chestnut (Castanea sativa Mill.) Shell Polyphenol Extract: A Valued Bioresource for Prostate Cancer Cell Growth Inhibition

Molecules ◽  
2020 ◽  
Vol 25 (12) ◽  
pp. 2730 ◽  
Author(s):  
Nunzio Antonio Cacciola ◽  
Andrea Cerrato ◽  
Anna Laura Capriotti ◽  
Chiara Cavaliere ◽  
Maria D’Apolito ◽  
...  
Keyword(s):  
Cell Line ◽  
Phenolic Content ◽  
Tumor Cell Line ◽  
Castanea Sativa ◽  
Epithelial Cell Line ◽  
Spectrometric Analysis ◽  
Cancer Cell Growth ◽  
Normal Prostate ◽  
Industrial Processing ◽  
By Products

Chestnut seeds are used for fresh consumption and for the industrial preparation of derivatives, such as chestnut flour. During industrial processing, large amounts of by-products are generally produced, such as leaves, flowers, shells and burs. In the present study, chestnut shells were extracted by boiling water in order to obtain polyphenol-rich extracts. Moreover, for the removal or non-phenolic compounds, a separation by preparative reverse phase chromatography in ten fractions was carried out. The richest fractions in terms of phenolic content were characterized by means of untargeted high-resolution mass spectrometric analysis together with a dedicated and customized data processing workflow. A total of 243 flavonoids, phenolic acids, proanthocyanidins and ellagitannins were tentatively identified in the five richest fractions. Due its high phenolic content (450.03 µg GAE per mg of fraction), one tumor cell line (DU 145) and one normal prostate epithelial cell line (PNT2) were exposed to increasing concentration of fraction 3 dry extract for 24, 48 and 72 h. Moreover, for DU 145 cell lines, increase of apoptotic cells and perturbation of cell cycle was demonstrated for the same extract. Those outcomes suggest that chestnut industrial by-products could be potentially employed as a source of bioresources.

scholarly journals Cancer stemness and metastatic potential of the novel tumor cell line K3: an inner mutated cell of bone marrow-derived mesenchymal stem cells

Oncotarget ◽  
2017 ◽  
Vol 8 (24) ◽  
pp. 39522-39533 ◽  
Author(s):  
Hui Qian ◽  
Xiaoqing Ding ◽  
Jiao Zhang ◽  
Fei Mao ◽  
Zixuan Sun ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Tumor Cell ◽  
Cell Line ◽  
Tumor Cell Line ◽  
Metastatic Potential ◽  
The Novel ◽  
Cancer Stemness

scholarly journals Role of Host Cell-Derived Amino Acids in Nutrition of Intracellular Salmonella enterica

2015 ◽  
Vol 83 (12) ◽  
pp. 4466-4475 ◽  
Author(s):  
Jasmin Popp ◽  
Janina Noster ◽  
Kim Busch ◽  
Alexander Kehl ◽  
Gero zur Hellen ◽  
...  
Keyword(s):  
Amino Acids ◽  
Host Cell ◽  
Cell Line ◽  
Hela Cells ◽  
Salmonella Enterica ◽  
Epithelial Cell Line ◽  
Intracellular Replication ◽  
Content Type ◽  
Raw264.7 Macrophages ◽  
Cell Line Hela

The facultative intracellular pathogenSalmonella entericaresides in a specific membrane-bound compartment termed theSalmonella-containing vacuole (SCV). Despite being segregated from access to metabolites in the host cell cytosol,Salmonellais able to efficiently proliferate within the SCV. We set out to unravel the nutritional supply ofSalmonellain the SCV with focus on amino acids. We studied the availability of amino acids by the generation of auxotrophic strains for alanine, asparagine, aspartate, glutamine, and proline in a macrophage cell line (RAW264.7) and an epithelial cell line (HeLa) and examined access to extracellular nutrients for nutrition. Auxotrophies for alanine, asparagine, or proline attenuated intracellular replication in HeLa cells, while aspartate, asparagine, or proline auxotrophies attenuated intracellular replication in RAW264.7 macrophages. The different patterns of intracellular attenuation of alanine- or aspartate-auxotrophic strains support distinct nutritional conditions in HeLa cells and RAW264.7 macrophages. Supplementation of medium with individual amino acids restored the intracellular replication of mutant strains auxotrophic for asparagine, proline, or glutamine. Similarly, a mutant strain deficient in succinate dehydrogenase was complemented by the extracellular addition of succinate. Complementation of the intracellular replication of auxotrophicSalmonellaby external amino acids was possible if bacteria were proficient in the induction ofSalmonella-induced filaments (SIFs) but failed in a SIF-deficient background. We propose that the ability of intracellularSalmonellato redirect host cell vesicular transport provides access of amino acids to auxotrophic strains and, more generally, is essential to continuously supply bacteria within the SCV with nutrients.

Essential role of biliary glycoprotein (CD66a) in morphogenesis of the human mammary epithelial cell line MCF10F

1999 ◽  
Vol 112 (23) ◽  
pp. 4193-4205 ◽  
Author(s):  
J. Huang ◽  
J.D. Hardy ◽  
Y. Sun ◽  
J.E. Shively
Keyword(s):  
Breast Cancer ◽  
Cell Death ◽  
Epithelial Cells ◽  
Cell Line ◽  
Myoepithelial Cell ◽  
Mammary Epithelial Cells ◽  
Tumor Cell Line ◽  
Mammary Epithelial ◽  
Cell Phenotype ◽  
Epithelial Cell Line

Normal mammary epithelial cells express the cell surface protein biliary glycoprotein (BGP or CD66a) in a polarized manner, suggesting that this protein may play a role in the formation of mammary acini. In order to test this hypothesis, we interrupted the expression of BGP in the mammary epithelial line MCF10F when cultured in or on Matrigel, a source of extracellular matrix (ECM). When analyzed by immunofluorescence confocal microscopy, the BGP staining is confined to the lumenal surface and colocalizes with actin. Sequential scanning electron microscopy demonstrates that the MCF10F cells migrate to form clusters, followed by apoptotic cell death within the center, resulting in lumen formation. Transmission electron micrographs reveal the presence of tight junctions and desmosomes between the cells, microvilli along the lumenal surface, and typical apoptotic bodies within the lumen. When the MCF10F cells are transfected with the BGP antisense gene and grown in Matrigel, they exhibit reduced acini formation (12% and 20%) compared to untransfected cells (52%) or to cells transfected with vector only (62%). Acini formation is also significantly reduced when MCF10F cells grown in Matrigel are treated with anti-BGP antibody (18% at 100 microgram/ml), or recombinant soluble BGP (18% at 0.4 microM). In contrast, the BGP-negative MCF7 breast tumor cell line, which does not form acini when grown in matrigel, exhibits &gt;60% cell death with the occasional formation of acini, when transfected with the BGP sense gene and grown in Matrigel. These results support the hypothesis that BGP plays a role in the normal differentiation program of mammary epithelial cells, indicating that its expression is essential to the formation of the lumen. Furthermore, and as shown by others, the differentiation program depends on the presence of ECM. The lack of expression of BGP in the MCF7 breast cancer cell line suggests that the downregulation of BGP expression confers a growth advantage to these cells in ECM. In addition, we found that the MCF10F cells could be separated into a BGP-positive epithelial fraction (MCF10F-e), and a BGP-negative myoepithelial fraction (MCF10F-m). When the myoepithelial cell-enriched fraction is grown on Matrigel, web-like structures are formed. These cells have a typical spindle shape cell morphology and express keratin, alpha-smooth muscle actin and vimentin, markers of the myoepithelial cell phenotype. When MCF10F-m cells are treated with IFNgamma, they express CEA (carcinoembryonic antigen) but not BGP. Since breast carcinomas, especially in situ carcinomas, express CEA, this finding may suggest a heretofore unappreciated relationship between myoepithelial cells and breast cancer.

Activated β-catenin in the novel human parathyroid tumor cell line sHPT-1

2007 ◽  
Vol 352 (2) ◽  
pp. 532-536 ◽  
Author(s):  
P. Björklund ◽  
G. Åkerström ◽  
G. Westin
Keyword(s):  
Tumor Cell ◽  
Cell Line ◽  
Tumor Cell Line ◽  
The Novel ◽  
Parathyroid Tumor
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