scholarly journals Comparison between the effects of the microsomal Ca2+-translocase inhibitors thapsigargin and 2,5-di-(t-butyl)-1,4-benzohydroquinone on cellular calcium fluxes

1991 ◽  
Vol 277 (2) ◽  
pp. 553-556 ◽  
Author(s):  
J Llopis ◽  
S B Chow ◽  
G E N Kass ◽  
A Gahm ◽  
S Orrenius
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Cell Types ◽  
Jurkat Cells ◽  
Bivalent Cation ◽  
Calcium Fluxes ◽  
Cellular Calcium ◽  
Entry Pathway

The effects of two inhibitors of the microsomal Ca(2+)-ATPase, thapsigargin and 2,5-di-(t-butyl)-1,4-benzohydroquinone, were compared in hepatocytes and in a T-cell line (JURKAT). Both compounds mobilized the same intracellular Ca2+ pool, which contained the Ins(1,4,5)P3-sensitive store, in hepatocytes and in JURKAT cells. The mobilization of the internal Ca2+ store with either compound activated Mn2+ entry in JURKAT cells, but not in hepatocytes. This suggests different properties of the bivalent-cation entry pathway between these cell types.

scholarly journals A 275 basepair fragment at the 5' end of the interleukin 2 gene enhances expression from a heterologous promoter in response to signals from the T cell antigen receptor.

1987 ◽  
Vol 165 (2) ◽  
pp. 395-407 ◽  
Author(s):  
D B Durand ◽  
M R Bush ◽  
J G Morgan ◽  
A Weiss ◽  
G R Crabtree
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Antigen Receptor ◽  
Calcium Ionophore ◽  
Jurkat Cells ◽  
T Cell Antigen Receptor ◽  
Linked Gene ◽  
Cell Antigen Receptor ◽  
Cell Antigen ◽  
Flanking Region

Using a transient transfection assay, we have defined the sequences required for the activation of the IL-2 gene in response to signals from the T cell antigen receptor. To do so we have transfected the human T cell line Jurkat with hybrid DNA constructs in which fragments from the IL-2 gene are linked to an indicator gene. The indicator gene product, as well as IL-2 production from the endogenous IL-2 gene were assayed after activation of the transfected Jurkat cells by various stimuli. We have demonstrated that a 275 bp fragment stretching from 52 to 326 bp upstream of the IL-2 gene transcription initiation site is required for expression of the linked indicator gene. This IL-2 gene fragment has several of the characteristics of a transcriptional enhancer element, in that it functions in both orientations and will enhance the expression from the promoter of an unrelated gene. Such enhancement occurred only after activation of Jurkat cells through the T cell antigen receptor. More specifically, this 275 bp fragment activated the expression of a linked gene after binding of a monoclonal antibody to the Jurkat T cell antigen receptor in the presence of PMA. In addition, calcium ionophore, which circumvents antigen receptor binding in T cell activation, induced the expression of the linked gene through this 275 bp sequence, in the presence of PMA. Finally, in a mutant Jurkat cell line lacking T3/antigen receptor complexes at the cell surface, no expression due to the IL-2 5' flanking region was seen after exposure to antibody to the T cell antigen receptor plus PMA or to PHA plus PMA. In contrast, calcium ionophore plus PMA did induce the expression of a linked gene through the IL-2 5' flanking region in the mutant Jurkat cell line. The responsiveness of the transfected hybrid genes containing the IL-2 regulatory region paralleled the expression of the endogenous IL-2 gene, as determined by IL-2 bioassays. We conclude that the 275 bp IL-2 sequence (-326 to -52 bp) is a target for the signal pathway originating at the T cell antigen receptor. Definition of this 275 bp target sequence should now permit the isolation of the molecules that bind to and activate the IL-2 gene.

scholarly journals Infection of monocyte-derived macrophages with human immunodeficiency virus type 1 (HIV-1). Monocyte-tropic and lymphocyte-tropic strains of HIV-1 show distinctive patterns of replication in a panel of cell types.

1989 ◽  
Vol 170 (4) ◽  
pp. 1149-1163 ◽  
Author(s):  
R Collman ◽  
N F Hassan ◽  
R Walker ◽  
B Godfrey ◽  
J Cutilli ◽  
...  
Keyword(s):  
T Cell ◽  
Host Range ◽  
Cell Line ◽  
Cell Types ◽  
Primary Cells ◽  
Primary Monocyte ◽  
Single Cell Type ◽  
Different Strains ◽  
Different Cell Types ◽  
Hiv 1

To characterize the host range of different strains of HIV-1, we have used four types of cells, primary monocyte-derived macrophages (MDM), primary PBL, a promonocyte cell line (U937), and a CD4+ T cell line (SUP-T1). These cells were infected with three prototype strains of HIV-1, a putative lymphocyte-tropic strain (IIIB), and two putative monocyte-tropic strains (SF162 and DV). Infections were monitored by assays for infectious virus, for cell-free and cell-associated viral antigen (p24), and for the proportion of cells infected by immunohistochemical staining. It was concluded that: (a) the use of four different cell types provides a useful biological matrix for distinguishing the tropism of different strains of HIV-1; this matrix yields more information than the infection of any single cell type. (b) A monocyte-tropic strain of HIV-1, such as strain SF162, shows a reciprocal host range when compared with a lymphocyte-tropic strain such as IIIB; strain SF162 replicates well in primary MDM but not in U937 or SUP-T1 cells, while strain IIIB replicates well in both U937 and SUP-T1 cells but not in MDM. (c) Both lymphocyte-tropic and monocyte-tropic strains of HIV-1 replicate well in PBL. (d) The promonocyte cell line, U937, and the T cell line, SUP-T1, differ markedly from primary cells, such as MDM and PBL, in their ability to support the replication of different strains of HIV-1; these cell lines cannot be used as surrogates for primary cells in host range studies of HIV-1 strains.

A yeast-based screening system identified bakkenolide B contained in Petasites japonicus as an inhibitor of interleukin-2 production in a human T cell line

2021 ◽  
Author(s):  
Shota Uesugi ◽  
Mayuka Hakozaki ◽  
Yuko Kanno ◽  
Honoka Takahashi ◽  
Yui Kudo ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Screening Method ◽  
Interleukin 2 ◽  
Jurkat Cells ◽  
Yeast Cells ◽  
Wild Plant ◽  
Human T Cell ◽  
Petasites Japonicus ◽  
Human T Cell Line

Abstract Ca2+ signaling is related to various diseases such as allergies, diabetes, and cancer. We explored Ca2+ signaling inhibitors in natural resources using a yeast-based screening method, and found bakkenolide B from the flower buds of edible wild plant, Petasites japonicus, using the YNS17 strain (zds1Δ erg3Δ pdr1/3Δ). Bakkenolide B exhibited growth-restoring activity against the YNS17 strain and induced Li+ sensitivity of wild-type yeast cells, suggesting that it inhibits the calcineurin pathway. Additionally, bakkenolide B inhibited interleukin-2 production at gene and protein levels in Jurkat cells, a human T cell line, but not the in vitro phosphatase activity of human recombinant calcineurin, an upstream regulator of interleukin-2 production. Furthermore, bakkenolide A showed weak activity in YNS17 and Jurkat cells compared with bakkenolide B. These findings revealed new biological effects and the structure-activity relationships of bakkenolides contained in Petasites japonicus as inhibitors of interleukin-2 production in human T cells.

scholarly journals STRL33, A Novel Chemokine Receptor–like Protein, Functions as a Fusion Cofactor for Both Macrophage-tropic and T Cell Line–tropic HIV-1

1997 ◽  
Vol 185 (11) ◽  
pp. 2015-2023 ◽  
Author(s):  
Fang Liao ◽  
Ghalib Alkhatib ◽  
Keith W.C. Peden ◽  
Geetika Sharma ◽  
Edward A. Berger ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Cell Lines ◽  
Chemokine Receptor ◽  
Chemokine Receptors ◽  
Sequence Similarity ◽  
Jurkat Cells ◽  
Productive Infection ◽  
Lymphoid Tissues ◽  
Hiv 1

The chemokine receptors CXCR4, CCR2B, CCR3, and CCR5 have recently been shown to serve along with CD4 as coreceptors for HIV-1. The tropisms of HIV-1 strains for subgroups of CD4+ cells can be explained, at least partly, by the selective use of G protein–coupled receptors (GPCRs). We have identified a novel human gene, STRL33, located on chromosome 3 that encodes a GPCR with sequence similarity to chemokine receptors and to chemokine receptor–like orphan receptors. STRL33 is expressed in lymphoid tissues and activated T cells, and is induced in activated peripheral blood lymphocytes. When transfected into nonhuman NIH 3T3 cells expressing human CD4, the STRL33 cDNA rendered these cells competent to fuse with cells expressing HIV-1 envelope glycoproteins (Envs). Of greatest interest, STRL33, in contrast with CXCR4 or CCR5, was able to function as a cofactor for fusion mediated by Envs from both T cell line–tropic and macrophage-tropic HIV-1 strains. STRL33-transfected Jurkat cell lines also supported enhanced productive infection with HIV-1 compared with control Jurkat cells. Despite the sequence similarities between STRL33 and chemokine receptors, STRL33-transfected cell lines did not respond to any in a panel of chemokines. Based on the pattern of tissue expression of the STRL33 mRNA, and given the ability of STRL33 to function with Envs of differing tropisms, STRL33 may play a role in the establishment and/or progression of HIV-1 infection.

scholarly journals Differentiation of BC3H1 smooth muscle cells changes the bivalent cation selectivity of the capacitative Ca2+ entry pathway

1996 ◽  
Vol 316 (3) ◽  
pp. 759-764 ◽  
Author(s):  
Lisa M. BROAD ◽  
David A. POWIS ◽  
Colin W. TAYLOR
Keyword(s):  
Smooth Muscle ◽  
Cell Types ◽  
Bivalent Cation ◽  
Fura 2 ◽  
Entry Pathway ◽  
Cation Selectivity ◽  
Differentiated Cells ◽  
Undifferentiated Cells ◽  
Rapid Removal ◽  
Permeation Properties

Differentiation of BC3H1 cells leads to expression of a variety of proteins characteristic of smooth muscle and to changes in the behaviour of intracellular Ca2+ stores. Treatment of both differentiated and undifferentiated cells with thapsigargin (2 μM) emptied their intracellular Ca2+ stores, and in the presence of extracellular Ca2+ caused an increase in cytosolic [Ca2+] that rapidly reversed after its removal. The amplitudes of these capacitative Ca2+ entry signals were 101±8 nM (n = 42) in differentiated cells and 188±16 nM (n = 35) in undifferentiated cells. Mn2+ entry in thapsigargin-treated cells, measured by recording the quenching of cytosolic fura 2 fluorescence, was 374±26% (n = 34) and 154±7% (n = 41) of control rates in differentiated and undifferentiated cells, respectively. Empty stores caused Ba2+ entry to increase to 282±20% (n = 8) of its basal rate in differentiated cells and to 187±20% (n = 8) in undifferentiated cells. Rates of Ca2+ extrusion, measured after rapid removal of extracellular Ca2+ from cells in which capacitative Ca2+ entry had been activated, were similar in differentiated (t½ = 23±2 s, n = 7) and undifferentiated (23±1 s, n = 6) cells. The different relationships between capacitative Ca2+ and Mn2+ signals are not, therefore, a consequence of more active Ca2+ extrusion mechanisms in differentiated cells, nor are they a consequence of different fura 2 loadings in the two cell types. We conclude that during differentiation of BC3H1 cells, the cation selectivity of the capacitative pathway changes, becoming relatively more permeable to Mn2+ and Ba2+. The change may result either from expression of a different capacitative pathway or from modification of the permeation properties of a single pathway.

Presence of a novel influx pathway for Mg2+ in MDCK cells

1990 ◽  
Vol 259 (3) ◽  
pp. C521-C525 ◽  
Author(s):  
G. A. Quamme ◽  
L. J. Dai
Keyword(s):  
Cell Line ◽  
Mdck Cell ◽  
Mdck Cells ◽  
Cell Types ◽  
Madin Darby Canine Kidney ◽  
Entry Pathway ◽  
Electrical Gradient ◽  
Free Media ◽  
Darby Canine Kidney ◽  
Canine Kidney

Basal free Mg2+ concentration was 0.49 +/- 0.03 mM in normal single Madin-Darby canine kidney (MDCK) cells as measured by fluorescence with the aid of mag-fura-2. Accordingly, Mg2+ may enter the cell down a transmembrane electrical gradient. The present study describes some aspects of Mg2+ entry into the established MDCK cell line. MDCK cells were Mg2(+)-depleted (0.26 +/- 0.01 mM) by culturing in Mg2(+)-free media for 16-20 h. Cells were subsequently exposed to 5 mM MgCl2, and intracellular Mg2+ concentration ([Mg2+]i) was monitored with fluorescence. [Mg2+]i returned to normal basal levels, 0.56 +/- 0.05 mM, with a refill rate of 272 +/- 39 nM/s, n = 4. Mg2+ entry was not changed by 5.0 mM external Ca2+ but was completely inhibited with 5.0 mM La3+. Intracellular Ca2+ concentration was not altered by Mg2+ depletion or during Mg2+ repletion. Mg2+ uptake was inhibited by verapamil (0 +/- 27 nM/s, n = 3), was inhibited less so by diltiazem (141 +/- 34 nM/s, n = 3), and was not affected by nifedipine (300 +/- 53 nM/s, n = 6). These inhibitors were fully reversible on removal, and [Mg2+]i returned to normal levels. These data indicate the presence of a unique Mg2+ entry pathway in MDCK cells that may be important in Mg2+ homeostasis. The model of Mg2+ refill into Mg2(+)-depleted cells may be useful in other cell types.

scholarly journals Two independently regulated Ca2+ entry mechanisms coexist in Jurkat T cells during T cell receptor antigen activation

1993 ◽  
Vol 293 (2) ◽  
pp. 395-398 ◽  
Author(s):  
S C Chow ◽  
G E Kass ◽  
S Orrenius
Keyword(s):  
T Cell ◽  
T Cell Receptor ◽  
Antigen Receptor ◽  
Cell Receptor ◽  
Jurkat Cells ◽  
Receptor Complex ◽  
T Cell Antigen Receptor ◽  
Bivalent Cation ◽  
Cell Antigen Receptor ◽  
Cell Antigen

Receptor-mediated Ca2+ influx was studied in the human leukaemic T cell line, Jurkat. Stimulation of these cells through the T cell antigen-receptor complex with OKT3 (an antibody against the CD3 molecules of the T cell antigen-receptor complex), or inhibition of the endoplasmic reticular Ca(2+)-ATPase with thapsigargin, resulted in Ca2+ mobilization from intracellular stores and the activation of Ca2+ and Mn2+ entry. The rates of thapsigargin-induced Ca2+ and Mn2+ entry in Jurkat cells were 76% and 64% respectively of those observed after treatment of these cells with OKT3. The combined addition of thapsigargin plus OKT3 to Jurkat cells produced an enhanced effect on the sustained increase in the cytosolic free Ca2+ concentration that was greater than that obtained by addition of thapsigargin or OKT3 alone. The rates of Ca2+ and Mn2+ entry were increased to 119% and 112% respectively of the OKT3-induced rates. Taken together, these results suggest that the inositol 1,4,5-trisphosphate-sensitive Ca(2+)-pool-dependent bivalent cation entry only accounts for 57% and 52% respectively of the total OKT3-dependent Ca2+ and Mn2+ entry, and that the rest is mediated by second messenger(s). Thus two separate pathways coexist in regulating Ca2+ entry in Jurkat cells during activation mediated through the T cell receptor.

scholarly journals c-Met-Specific Chimeric Antigen Receptor T Cells Demonstrate Anti-Tumor Effect in c-Met Positive Gastric Cancer

Cancers ◽  
2021 ◽  
Vol 13 (22) ◽  
pp. 5738
Author(s):  
Chung Hyo Kang ◽  
Yeongrin Kim ◽  
Da Yeon Lee ◽  
Sang Un Choi ◽  
Heung Kyoung Lee ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Cell Lines ◽  
Chimeric Antigen Receptor ◽  
Antigen Receptor ◽  
Jurkat Cells ◽  
Car T Cells ◽  
Car T

Chimeric antigen receptor (CAR) technology has been highlighted in recent years as a new therapeutic approach for cancer treatment. Although the impressive efficacy of CAR-based T cell adoptive immunotherapy has been observed in hematologic cancers, limited effect has been reported on solid tumors. Approximately 20% of gastric cancer (GC) patients exhibit a high expression of c-Met. We have generated an anti c-Met CAR construct that is composed of a single-chain variable fragment (scFv) of c-Met antibody and signaling domains consisting of CD28 and CD3ζ. To test the CAR construct, we used two cell lines: the Jurkat and KHYG-1 cell lines. These are convenient cell lines, compared to primary T cells, to culture and to test CAR constructs. We transduced CAR constructs into Jurkat cells by electroporation. c-Met CAR Jurkat cells secreted interleukin-2 (IL-2) only when incubated with c-Met positive GC cells. To confirm the lytic function of CAR, the CAR construct was transduced into KHYG-1, a NK/T cell line, using lentiviral particles. c-Met CAR KHYG-1 showed cytotoxic effect on c-Met positive GC cells, while c-Met negative GC cell lines were not eradicated by c-Met CAR KHYG-1. Based on these data, we created c-Met CAR T cells from primary T cells, which showed high IL-2 and IFN-γ secretion when incubated with the c-Met positive cancer cell line. In an in vivo xenograft assay with NSG bearing MKN-45, a c-Met positive GC cell line, c-Met CAR T cells effectively inhibited the tumor growth of MKN-45. Our results show that the c-Met CAR T cell therapy can be effective on GC.

scholarly journals Combination Therapy with miR-34a and Doxorubicin Synergistically Induced Apoptosis in T-Cell Acute Lymphoblastic Leukemia Cell Line

2021 ◽  
Author(s):  
Shiva Najjary ◽  
Reza Mohammadzadeh ◽  
Behzad Mansoori ◽  
Fatemeh Vahidian ◽  
Ali Mohammadi ◽  
...  
Keyword(s):  
T Cell ◽  
Combination Therapy ◽  
Cell Line ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cell ◽  
Jurkat Cells ◽  
Leukemia Cell Line ◽  
Dapi Staining ◽  
Induced Apoptosis ◽  
In Cells

Abstract Reduced expression of tumor suppressor miRNAs leads to cancer cell development, so restoring the expression of these miRNAs can be an appropriate treatment option for cancer. Due to the heterogeneity of cancer cells, single-drug therapy often results in drug resistance. Therefore, the combination of chemotherapy with miRNA can be a powerful strategy for cancer treatment. In the current investigation, we researched the synergic effect of miR-34a in combination with doxorubicin (DOX) on cell death of acute T-cell lymphoblastic leukemia (T-ALL) Jurkat cell line, as well as the expression of some genes including Caspase-3, Bcl-2, and p53 which are involved in apoptosis. Our outcomes showed that the combination of miR-34a and doxorubicin remarkably reduced the expression of the Bcl-2 gene, the target gene of miR-34a. According to the results of the MTT assay in cells treated with miR-34a and doxorubicin, the survival rate was significantly decreased compared to the untreated cells. Results of the flow cytometry assay and DAPI staining demonstrated an increased apoptosis rate of Jurkat cells in combination therapy. Moreover, cell cycle arrest was observed at the G2/M phase in cells that were treated with miR-34a/doxorubicin. Most importantly, we showed that the transfection of the Jurkat cells with miR-34a increased the sensitivity of these cells to doxorubicin. Furthermore, the combination of miR-34a and doxorubicin drug effectively increased apoptosis of treated cells. Therefore, this method can be used as an impressive treatment for T-ALL.

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