1.
Fjæraa Alfredsson, Christina, et al.
(författare)
Altered sensitivity to ellagic acid in neuroblastoma cells undergoing differentiation with 12-0-tetradecanoylphorbol-13-acetate and all-trans retinoic acid
2015
Ingår i: Biomedicine and Pharmacotherapy. - : Elsevier. - 0753-3322 .- 1950-6007. ; 76, s. 39-45
Tidskriftsartikel (refereegranskat) abstract
Ellagic acid has previously been reported to induce reduced proliferation and activation of apoptosis in several tumor cell lines including our own previous data from non-differentiated human neuroblastoma SH-SY5Y cells. The aim of this study was now to investigate if in vitro differentiation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate or the vitamin A derivative all-trans retinoic acid altered the sensitivity to ellagic acid in SH-SY5Y cells. The methods used were cell counting and LDH-assay for evaluation of cell number and cell death, flow cytometric analysis of SubG(1)-and TUNEL-analysis for apoptosis and western blot for expression of apoptosis-associated proteins. In vitro differentiation was shown to reduce the sensitivity to ellagic acid with respect to cell detachment, loss of viability and activation of apoptosis. The protective effect was phenotype-specific and most prominent in all-trans retinoic acid-differentiated cultures. Differentiation-dependent up-regulation of Bcl-2 and integrin expression is introduced as possible protective mechanisms. The presented data also point to a positive correlation between proliferative activity and sensitivity to ellagic-acid-induced cell detachment. In conclusion, the presented data emphasize the need to consider degree of neuronal differentiation and phenotype of neuroblastoma cells when discussing a potential pharmaceutical application of ellagic acid in tumor treatment.
2.
Anclair, Malin, et al.
(författare)
Parental fears following their child's brain tumor diagnosis and treatment.
2009
Ingår i: Journal of pediatric oncology nursing : official journal of the Association of Pediatric Oncology Nurses. - : SAGE Publications. - 1043-4542 .- 1532-8457. ; 26:2, s. 68-74
Tidskriftsartikel (refereegranskat) abstract
The objective of this study is to portray the illness-related threats experienced by parents of children after the diagnosis of central nervous system (CNS) tumor. Parents were asked to rate the extent to which they experienced a set of specific fears related to their child's brain tumor and its treatment. Outcomes for parents of CNS tumor patients (n = 82) were compared with those of reference parents of patients treated for acute lymphoblastic leukemia (n = 208). The fears about an illness recurrence and the late effects of treatment were most prominent among parents of CNS tumor patients. For 7 out of 11 kinds of fear, parents of CNS tumor patients expressed a stronger fear than the reference group. More than a quarter of the parents of children treated for CNS tumors feared a complete decline of the child. Parents of CNS tumor patients experience relatively heightened cancer related fears in several domains. The fear of devastating consequences felt by one fourth of parents signals the need of individualized psychological support and information at diagnosis and follow-up to facilitate parental coping with the posttreatment situation.
3.
Elmabsout, Ali Ateia, et al.
(författare)
Cloning and Functional Studies of a Splice Variant of CYP26B1 Expressed in Vascular Cells
2012
Ingår i: Plos One. - San Francisco, USA : Public Library of Science (PLoS). - 1932-6203. ; 7:5
Tidskriftsartikel (refereegranskat) abstract
Background: All-trans retinoic acid (atRA) plays an essential role in the regulation of gene expression, cell growth and differentiation and is also important for normal cardiovascular development but may in turn be involved in cardiovascular diseases, i.e. atherosclerosis and restenosis. The cellular atRA levels are under strict control involving several cytochromes P450 isoforms (CYPs). CYP26 may be the most important regulator of atRA catabolism in vascular cells. The present study describes the molecular cloning, characterization and function of atRA-induced expression of a spliced variant of the CYP26B1 gene. Methodology/Principal Findings: The coding region of the spliced CYP26B1 lacking exon 2 was amplified from cDNA synthesized from atRA-treated human aortic smooth muscle cells and sequenced. Both the spliced variant and full length CYP26B1 was found to be expressed in cultured human endothelial and smooth muscle cells, and in normal and atherosclerotic vessel. atRA induced both variants of CYP26B1 in cultured vascular cells. Furthermore, the levels of spliced mRNA transcript were 4.5 times higher in the atherosclerotic lesion compared to normal arteries and the expression in the lesions was increased 20-fold upon atRA treatment. The spliced CYP26B1 still has the capability to degrade atRA, but at an initial rate one-third that of the corresponding full length enzyme. Transfection of COS-1 and THP-1 cells with the CYP26B1 spliced variant indicated either an increase or a decrease in the catabolism of atRA, probably depending on the expression of other atRA catabolizing enzymes in the cells. Conclusions/Significance: Vascular cells express the spliced variant of CYP26B1 lacking exon 2 and it is also increased in atherosclerotic lesions. The spliced variant displays a slower and reduced degradation of atRA as compared to the full-length enzyme. Further studies are needed, however, to clarify the substrate specificity and role of the CYP26B1 splice variant in health and disease.
4.
Eriksson, Anders, et al.
(författare)
Demonstration of functionally different interactions between phospholipase C-gamma and the two types of platelet-derived growth factor receptors
1995
Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 270:13, s. 7773-7781
Tidskriftsartikel (refereegranskat) abstract
Phosphorylated tyrosine residues in receptor tyrosine kinases serve as binding sites for signal transduction molecules. We have identified two autophosphorylation sites, Tyr-988 and Tyr-1018, in the platelet-derived growth factor (PDGF) alpha-receptor carboxyl-terminal tail, which are involved in binding of phospholipase C-gamma (PLC-gamma). The capacities of the Y988F and Y1018F mutant PDGF alpha-receptors, expressed in porcine aortic endothelial cells, to bind PLC-gamma are 60 and 5% of that of the wild-type receptor, respectively. Phosphorylated but not unphosphorylated peptides containing Tyr-1018 are able to compete with the intact receptor for binding to immobilized PLC-gamma SH2 domains; a phosphorylated Tyr-988 peptide competes 10 times less efficiently. The complex between PLC-gamma and the PDGF alpha-receptor is more stable than that of PLC-gamma and the PDGF beta-receptor. However, PDGF stimulation results in a smaller fraction of tyrosine-phosphorylated PLC-gamma and a smaller accumulation of inositol trisphosphate in cells expressing the alpha-receptor as compared with cells expressing the beta-receptor. We conclude that phosphorylated Tyr-988 and Tyr-1018 in the PDGF alpha-receptor carboxyl-terminal tail bind PLC-gamma, but this association leads to only a relatively low level of tyrosine phosphorylation and activation of PLC-gamma.
5.
Erlandsson, Ann, et al.
(författare)
Studies of the interactions between the anticytokeratin 8 monoclonal antibody TS1, its antigen and its anti-idiotypic antibody αTS1
2003
Ingår i: Journal of Molecular Recognition. - : Wiley. - 0952-3499 .- 1099-1352. ; 16:3, s. 157-163
Tidskriftsartikel (refereegranskat) abstract
The monoclonal antibody TS1 against cytokeratin 8 and its antiidiotype αTS1 have been used for immunotargeting and therapy of carcinomas in experimental tumor model systems. The interaction surfaces between mab TS1, the cytokeratin 8 epitope, and its anti-idiotypic antibody, αTS1, were studied in detail in order to make future veneering of the interactions possible. The V-genes of TS1 and αTS1 were cloned and sequenced and the CDRs and the framework residues were identified. Amino acids participating in the interactions were identified following chemical modifications of residues in non-protected and protected molecules of cytokeratin 8, αTS1 and TS1. From the sequences, the three-dimensional structures were generated using computer modelling of the antibody variable regions. Several charged amino acid, histidine and tyrosine residues were displayed in the antibody surfaces implicated in the interactions and chemical modification confirmed the importance of these amino acids. The cytokeratin 8 epitope has previously been identified by Johansson et al. and it displays negatively charged amino acid residues which could be identified in the chemical modification. It was also revealed that the TS1 binding to cytokeratin 8 and αTS1 respectively are partly overlapping; a histidine identified in TS1 is probably involved only in the interaction with αTS1. Furthermore, the chemical modification demonstrated that exchanging aspartic–glutamic acids to asparagine–glutamine residues in TS1 increased the binding of TS1 to cytokeratin 8, indicating that there is at least one acidic amino acid that is an obstacle in the TS1–CK8 binding. The detailed assembly of the interaction surfaces will facilitate the future use of site directed mutagenesis to improve the TS1–CK8 association rate and the clearing of TS1 with αTS1 in vivo.
6.
Fagerström, Sofia, et al.
(författare)
Protein kinase C-dependent tyrosine phosphorylation of p130cas in differentiating neuroblastoma cells
1998
Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 273:4, s. 2336-2343
Tidskriftsartikel (refereegranskat) abstract
The cell signaling docking protein p130cas became tyrosine-phosphorylated in SH-SY5Y human neuroblastoma cells during induced differentiation with 12-O-tetradecanoylphorbol-13-acetate (TPA) and serum or a combination of basic fibroblast growth factor (bFGF) and insulin-like growth factor-I (IGF-I). The differentiating cells develop a neuronal phenotype with neurites and growth cones and sustained activation of protein kinase C (PKC) and pp60c-src. The TPA-induced p130cas phosphorylation increased within 5 min of stimulation and persisted for at least 4 days, whereas bFGF/IGF-I-induced p130cas phosphorylation was biphasic. However, the increase in tyrosine phosphorylation of p130cas was not restricted to differentiation inducing stimuli. The phosphorylation was blocked by the specific PKC inhibitor GF 109203X, and transient transfection with active PKC-epsilon induced p130cas tyrosine phosphorylation. pp60c-src, known to directly phosphorylate p130cas in other cell systems, was not activated after stimulation with TPA or bFGF/IGF-I for up to 30 min, and the initial p130cas phosphorylation was resistant to the Src family kinase inhibitor herbimycin A. However, in long term stimulated cells, herbimycin A blocked the induced phosphorylation of p130cas. Also, overexpression of src induced phosphorylation of p130cas. p130cas protein and phosphorylated p130cas were present in growth cones isolated from differentiated SH-SY5Y cells. Inhibition of PKC activity in differentiating cells with GF 109203X leads to a rapid retraction of growth cone filopodia, and p130cas phosphorylation decreased transiently (within minutes). Growth cones isolated from these cells were virtually devoid of phosphorylated p130cas. These data suggest a function for p130cas as a PKC downstream target in SH-SY5Y cells and possibly also in their growth cones.
7.
Fagerström, Sofia, et al.
(författare)
Protein kinase C-epsilon is implicated in neurite outgrowth in differentiating human neuroblastoma cells
1996
Ingår i: Cell growth & differentiation. - : American Association of Cancer Research. - 1044-9523. ; 7:6, s. 775-785
Tidskriftsartikel (refereegranskat) abstract
A combination of basic fibroblast growth factor (bFGF) and insulin-like growth factor-I (IGF-I) or 16 nM 12-O-tetradecanoylphorbol-13-acetate (TPA) and serum induces human SH-SY5Y neuroblastoma cells to undergo differentiation and acquire a neuronal phenotype. Nerve growth factor (NGF) added to SH-SY5Y cells stably transfected with the NGF-receptor TRK-A (SH-SY5Y/trk) induces a similar differentiated phenotype. SH-SY5Y cells express protein kinase C (PKC)-alpha, PKC-beta I, PKC-epsilon, and PKC-zeta protein, and phorbol ester- or growth factor-induced differentiation results in a sustained activation of PKC. The specific PKC inhibitor GF 109203X blocked TPA- and bFGF-IGF-I-induced neurite outgrowth in wild-type SH-SY5Y cells and NGF-induced neurite outgrowth in SH-SY5Y/trk cells. When added to differentiated cells, GF 109203X caused rapid retraction of growth cone filopodia. In TPA- and bFGF-IGF-I-treated cells, addition of GF 109203X also blocked induced expression of growth associated protein-43 and neuropeptide tyrosine while the increase in expression of these two genes was only slightly affected by the inhibitor in NGF-treated SH-SY5Y/trk cells. Thus, a portion of the NGF-induced phenotypic changes appears not to be mediated via PKC-dependent signaling. A high concentration of TPA (1.6 microM) down regulated PKC-alpha and PKC-beta I almost completely and PKC-epsilon partially in wild-type SH-SY5Y and SH-SY5Y/trk cells. Cells with down-regulated PKC-alpha and PKC-beta I after 1.6 microM TPA treatment still differentiated with growth factors. In these cells, the PKC-epsilon level was restored, and the PKC-epsilon protein was enriched in the growth cones. The 1.6 microM TPA-induced down-regulation of PKC-epsilon was counteracted by bFGF and NGF but not by platelet-derived growth factor or IGF-I. These data indicate that PKC activity is vital for neurite formation, and that the cells can differentiate under conditions when PKC-alpha and PKC-beta I are extensively down regulated. The close correlation between differentiation and presence of PKC-epsilon protein suggests an important function for this isoform during this process.
8.
Hedbrant, Alexander, 1987-
(författare)
Cancer and Inflammation : Role of Macrophages and Monocytes
2015
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Macrophages are cells of the innate immune system that can be found in large quantities in cancer tumors and affect cancer progression by regulating growth and invasiveness of cancer cells. There are two main phenotypes of macrophages denoted M1 and M2. In this thesis, the M1 and M2 phenotype of human macrophages were characterized, and effects of the macrophages on the growth and invasiveness of colon and lung cancer cells were studied.Macrophages of the M1 phenotype, but not the M2 phenotype, inhibited growth of both colon and lung cancer cells, and the inhibition for some of the cancer cell lines was induced by cell cycle arrest in the G1/G0 and/or G2/M cell cycle phases. In the colon cancer cell line, the macrophage induced cell cycle arrest was found to attenuate the cytotoxic effect of the chemotherapeutic drug 5-FU. Macrophages were also shown to express high levels of proteases (matrix metalloproteinase-2 and 9) and high levels of proteins of the urokinase-type plasminogen activator (uPA) system, in comparison to the lung cancer cell lines studied. Expression of these has been found to predict poor outcome in lung cancer, and the results suggest macrophages to be important contributors of these in lung tumors. Furthermore, the M1 phenotype was found to express higher levels of the uPA receptor than the M2 phenotype.Prostaglandin E2 (PGE2) is a potent inflammatory molecule expressed by e.g. macrophages and monocytes, and inhibition of its expression has been shown to reduce the risk of colon cancer. Green tea and black tea was found to be potent inhibitors of PGE2 formation in human monocytes, and the inhibitory effects of green tea was likely due to its content of the polyphenol epigallocatechin gallate. Rooibos tea also inhibited PGE2 formation, but was less potent than green and black tea. The primary mechanism for the inhibition was via inhibition of expression of enzymes in the PGE2 formation pathway, and primarily microsomal prostaglandin synthase-1.
9.
Hedbrant, Alexander, Ph.D, 1987-, et al.
(författare)
Green, Black and Rooibos Tea Inhibit Prostaglandin E2 Formation in Human Monocytes by Inhibiting Expression of Enzymes in the Prostaglandin E2 Pathway
2022
Ingår i: Molecules. - : MDPI. - 1431-5157 .- 1420-3049. ; 27:2
Tidskriftsartikel (refereegranskat) abstract
The formation of prostaglandin E2 (PGE2) is associated with adverse inflammatory effects. However, long-term treatment with nonsteroidal anti-inflammatory drugs (NSAIDs) comes with risk of severe side effects. Therefore, alternative ways to inhibit PGE2 are warranted. We have investigated the effects of tea extracts and the polyphenols epigallocatechin gallate (EGCG) and quercetin on PGE2 formation, determined by immunoassay, and protein expression, determined by immunoblotting, of cytosolic phospholipase A2 (cPLA2), cyclooxygenase 2 (COX-2) and microsomal PGE synthase-1 (mPGES-1) in human monocytes. Green and black tea extracts, and with a lower potency, Rooibos tea extract, inhibited lipopolysaccharide (LPS) and calcium ionophore-induced PGE2 formation. In addition, all tea extracts inhibited the LPS-induced expression of mPGES-1, and the green and black tea extracts also inhibited, to a lesser extent, COX-2 expression. The tea extracts only marginally reduced cPLA2 expression and had no effect on COX-1 expression. EGCG, present in green and black tea, and quercetin, present in all three teas, also inhibited PGE2 formation and expression of mPGES-1, COX-2 and cPLA2. Cell-based and cell-free assays were also performed to evaluate direct effects on the enzymatic activity of COX and PGE synthases. Mainly, the cell-free assay demonstrated partial inhibition by the tea extracts and polyphenols. However, the inhibition required higher doses compared to the effects demonstrated on protein expression. In conclusion, green and black tea, and to a lesser extent Rooibos tea, are potent inhibitors of PGE2 formation in human monocytes, and mediate their effects by inhibiting the expression of the enzymes responsible for PGE2 formation, especially mPGES-1.
10.
Holm, Patrik, 1975-
(författare)
Functional mapping and in vivo metabolism of the monoclonal antibody TS1 and its single-chain fragment : Its interaction with the antigen and the anti-idiotype
2006
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Antibodies are proteins capable of specific interactions to a wide range of molecules. These interactions are facilitated by the complementary determining regions (CDR).Carcinomas are the most common of human cancers and they release significant amount of cytokeratins (CK) in the necrotic areas of the tumors. The CKs stay in the tumor, since they have low solubility. The antibody studied in this thesis, the anti-CK 8 antibody TS1, has shown to be effective in tumor targeting and is proposed to be useful in therapy.Single-chain antibodies (scFv) are recombinant antibodies which are much smaller than the intact IgG. This is an advantage when used in tumor therapy, since they can penetrate the tumors more easily than the larger IgG. Moreover, they are expressed by one single gene which make them easy to modify, for example by site-directed mutagenesis.The anti-idiotypic antibody αTS1 can be used to clear the TS1 form the circulation and thereby clear the body from non-tumor bound TS1 in therapy. To be able to modify the binding of an antibody to its antigen and or anti-idiotype, these interactions must be studied. In this study this is accomplished by chemical modifications of the IgGs TS1 and αTS1 and the antigen CK 8. Guided by these results, amino acid residues were mutated by using site-directed mutagenesis in the TS1-218 scFv and the effects were studied. From mutational study results, the functional epitope could be mapped and it was found that there are mainly tyrosines, but also charged residues, serine and a tryptophan that are important for both interactions. The binding of TS1-218 to both αTS1 and CK 8 could be improved by changing the negatively charged side-chains by mutations to their corresponding amide or alanine.Both the IgG and scFv versions of TS1 were administered in vivo. The IgG αTS1 was used to clear the TS1 from the circulation by forming immune complexes. The immune complexes, consisting of four or more antibodies, were mainly metabolized by the liver. The scFv TS1-218 could localize to the tumor in a tumor xenograft mouse model, although a higher uptake would be desired in a therapeutic strategy. The scFv was cleared rapidly by the kidneys, but the clearance could be slowed by pre-formed immune complexes with anti-TS1 scFv in vitro, prior to administration in vivo.
