These are representative sample records from Science.gov related to your search topic.
For comprehensive and current results, perform a real-time search at Science.gov.
1

Cytokine mRNA expression in intestinal tissue of interleukin-2 deficient mice with bowel inflammation  

PubMed Central

Background—Mice deficient in interleukin-2 (IL-2) develop inflammatory bowel disease resembling ulcerative colitis in humans. Recent studies provided evidence that ?? T cells, particularly CD4 T cells, rather than B cells, are involved in the pathogenesis of bowel inflammation of IL-2 deficient mice. ?Aim—To analyse the pattern of expression of cytokine mRNA in intestinal tissue of normal and IL-2 deficient mice. ?Methods—Expression of ?-actin, IL-1?, IL-1?, IL-6, IL-10, tumour necrosis factor ? (TNF-?), interferon ? (IFN-?) and transforming growth factor ?1 (TGF-?1) mRNA was analysed in colon and small intestinal tissue of both IL-2 deficient (IL-2?/?) mice and normal (wild type) litter mates (IL-2+/+) at different ages by using qualitative, as well as semiquantitative, competitive reverse transcription polymerase chain reaction (RT-PCR). Results were correlated with the phase of progression of the disease, as determined by histology. ?Results—IL-2?/? mice had expressed low levels of IL-1?, IL-1?, IL-6, TNF-?, and IFN-? mRNA in the colon by 1.5 weeks of age. In advance of the development of histologically and clinically detectable bowel inflammation, expression of IL-1?, IL-1?, IL-6, TNF-?, IFN-?, and IL-10, but not TGF-?1, mRNA increased in the colon of IL-2 deficient mice. In contrast, IL-2+/+ mice expressed TGF-?1 mRNA in colon tissue at 13 and 23 weeks of age, but not IL-1?, IL-1?, IL-6, TNF-?, IL-10, or IFN-? mRNA. Levels of expression of cytokine mRNA in tissue from the small intestine were comparable in IL-2?/? and IL-2+/+ mice. ?Conclusions—Bowel inflammation in IL-2 deficient mice is preceded by an increase in IL-1?, IL-1?, TNF-?, and IFN-? mRNA expression in colon tissue. Low levels of TGF-?1, but high levels of IL-1?, IL-1?, IL-6, TNF-?, IFN-?, and IL-10 mRNA expression correlate with the manifestation of severe colitis, and suggest that T cells and macrophages are involved in bowel inflammation of IL-2 deficient mice. ?? Keywords: cytokine; mRNA expression; interleukin-2 deficient mice; bowel inflammation PMID:9462212

Autenrieth, I; Bucheler, N; Bohn, E; Heinze, G; Horak, I

1997-01-01

2

Expression of the type 1 human immunodeficiency virus Nef protein in T cells prevents antigen receptor-mediated induction of interleukin 2 mRNA.  

PubMed Central

Stable transformants of the Jurkat T-cell line have been obtained that express either of two distinct forms of the type 1 human immunodeficiency virus nef gene: the nef-1-encoded protein (Nef-1) contains alanine, glycine, and valine at positions 15, 29, and 33, respectively; the protein specified by nef-2 (Nef-2) has threonine, arginine, and alanine at the corresponding positions. When Jurkat cells or their Nef-2-expressing transformants are treated with phorbol 12-myristate 13-acetate (PMA) plus either phytohemagglutinin (PHA) or antibodies against CD3 epsilon, T-cell receptor beta chain, or CD2, there is a prompt increase in interleukin 2 (IL-2) mRNA and intracellular calcium and in the IL-2 receptor alpha chain on the cell surface. Although cells expressing Nef-1 also induce calcium mobilization and the production of IL-2 receptor alpha chain, the formation of IL-2 mRNA is blocked in response to these stimuli. Moreover, Nef-1-expressing cells transfected with a plasmid in which the IL-2 promoter is fused to the chloramphenicol acetyltransferase (CAT) gene fail to induce CAT following treatment with PMA and PHA. By contrast, the parental and Nef-2-containing cells induce CAT normally. Nef-1-expressing cells can produce IL-2 mRNA in response to a combination of PMA and ionomycin, although much less efficiently than the parental Jurkat cells or Nef-2-expressing cells. These findings, and others described herein, suggest that the virally encoded Nef protein interferes with a signal emanating from the T-cell receptor complex that induces IL-2 gene transcription. Images PMID:2052609

Luria, S; Chambers, I; Berg, P

1991-01-01

3

Inhibition of the serine/threonine protein phosphatases PP1 and PP2A in lymphocytes: effect on mRNA levels for interleukin-2, IL-2R alpha, krox-24, p53, hsc70 and cyclophilin.  

PubMed

Lymphocyte activation requires signal transduction mediated by reversible phosphorylation. Changing profiles of phosphorylated intermediates relate to the progressive series of transduction pathways in cells moving from G0 to G1, and thereafter through the cell cycle. We have previously shown that transient inhibition of the serine/threonine protein phosphatases PP1 and PP2A by okadaic acid enhances early mitogenic stimulation. Thus target proteins of PP1/PP2A may be involved in regulation of early mitogenic signalling, with the phosphorylated form(s) being associated with signal enhancement. Later, pathways require dephosphorylation of these proteins, since continuous treatment with okadaic acid blocks lymphocyte progression through the cell cycle. Delayed addition of okadaic acid showed that this blockade occurs between 8 and 24 hr. Here we have furthered these observations to the level of gene induction by measuring messenger RNA (mRNA) levels for the following proteins: interleukin-2 (IL-2) and IL-2R alpha; p53, a tumour suppressor protein; the transcription factor krox-24; and two mediators of protein folding, namely cyclophilin and the heat-shock protein hsc70. An external standard was used to quantitate the mRNA levels per cell. We found that 24 hr exposure to okadaic acid has a general suppressive effect on concanavalin A (Con A)-stimulated gene induction. However, at 4 hr okadaic acid enhanced IL-2 mRNA levels induced by Con A. Moreover, in unstimulated lymphocytes, okadaic acid caused the induction of krox-24, indicating a role for PP1 and PP2A in the regulation of this gene in resting cells. PMID:1328040

Richards, F M; Milner, J; Metcalfe, S

1992-08-01

4

A JNK-mediated autophagy pathway that triggers c-IAP degradation and necroptosis for anticancer chemotherapy  

PubMed Central

Killing cancer cells through the induction of apoptosis is one of the main mechanisms of chemotherapy. However, numerous cancer cells have primary or acquired apoptosis resistance, resulting in chemoresistance. In this study, using a novel chalcone derivative chalcone-24 (Chal-24), we identified a novel anticancer mechanism through autophagy-mediated necroptosis (RIP1- and RIP3-dependent necrosis). Chal-24 potently killed different cancer cells with induction of necrotic cellular morphology while causing no detectable caspase activation. Blocking the necroptosis pathway with either necrostatin-1 or by knockdown of RIP1 and RIP3 effectively blocked the cytotoxicity of Chal-24, suggesting that Chal-24-induced cell death is associated with necroptosis. Chal-24 robustly activated JNK and ERK and blockage of which effectively suppressed Chal-24-induced cytotoxicity. In addition, Chal-24 strongly induced autophagy that is dependent on JNK-mediated phosphorylation of Bcl-2 and Bcl-xL and dissociation of Bcl-2 or Bcl-xL from Beclin1. Importantly, suppression of autophagy, with either pharmacological inhibitors or siRNAs targeting the essential autophagy components ATG7 and Beclin1, effectively attenuated Chal-24-induced cell death. Furthermore, we found that autophagy activation resulted in c-IAP1 and c-IAP2 degradation and formation of the Ripoptosome that contributes to necroptosis. These results thus establish a novel mechanism for killing cancer cells that involves autophagy-mediated necroptosis, which may be employed for overcoming chemoresistance. PMID:23831571

He, Weiyang; Wang, Qiong; Srinivasan, Balasubramanian; Xu, Jennings; Padilla, Mabel T.; Li, Zi; Wang, Xia; Liu, Yushi; Gou, Xin; Shen, Han-Ming; Xing, Chengguo; Lin, Yong

2014-01-01

5

A JNK-mediated autophagy pathway that triggers c-IAP degradation and necroptosis for anticancer chemotherapy.  

PubMed

Killing cancer cells through the induction of apoptosis is one of the main mechanisms of chemotherapy. However, numerous cancer cells have primary or acquired apoptosis resistance, resulting in chemoresistance. In this study, using a novel chalcone derivative chalcone-24 (Chal-24), we identified a novel anticancer mechanism through autophagy-mediated necroptosis (RIP1- and RIP3-dependent necrosis). Chal-24 potently killed different cancer cells with induction of necrotic cellular morphology while causing no detectable caspase activation. Blocking the necroptosis pathway with either necrostatin-1 or by knockdown of RIP1 and RIP3 effectively blocked the cytotoxicity of Chal-24, suggesting that Chal-24-induced cell death is associated with necroptosis. Chal-24 robustly activated JNK and ERK and blockage of which effectively suppressed Chal-24-induced cytotoxicity. In addition, Chal-24 strongly induced autophagy that is dependent on JNK-mediated phosphorylation of Bcl-2 and Bcl-xL and dissociation of Bcl-2 or Bcl-xL from Beclin-1. Importantly, suppression of autophagy, with either pharmacological inhibitors or small interfering RNAs targeting the essential autophagy components ATG7 and Beclin-1, effectively attenuated Chal-24-induced cell death. Furthermore, we found that autophagy activation resulted in c-IAP1 and c-IAP2 degradation and formation of the Ripoptosome that contributes to necroptosis. These results thus establish a novel mechanism for killing cancer cells that involves autophagy-mediated necroptosis, which may be employed for overcoming chemoresistance. PMID:23831571

He, W; Wang, Q; Srinivasan, B; Xu, J; Padilla, M T; Li, Z; Wang, X; Liu, Y; Gou, X; Shen, H-M; Xing, C; Lin, Y

2014-06-01

6

Sodium fluoride induces apoptosis in mouse embryonic stem cells through ROS-dependent and caspase- and JNK-mediated pathways  

SciTech Connect

Sodium fluoride (NaF) is used as a source of fluoride ions in diverse applications. Fluoride salt is an effective prophylactic for dental caries and is an essential element required for bone health. However, fluoride is known to cause cytotoxicity in a concentration-dependent manner. Further, no information is available on the effects of NaF on mouse embryonic stem cells (mESCs). We investigated the mode of cell death induced by NaF and the mechanisms involved. NaF treatment greater than 1 mM reduced viability and DNA synthesis in mESCs and induced cell cycle arrest in the G{sub 2}/M phase. The addition of NaF induced cell death mainly by apoptosis rather than necrosis. Catalase (CAT) treatment significantly inhibited the NaF-mediated cell death and also suppressed the NaF-mediated increase in phospho-c-Jun N-terminal kinase (p-JNK) levels. Pre-treatment with SP600125 or z-VAD-fmk significantly attenuated the NaF-mediated reduction in cell viability. In contrast, intracellular free calcium chelator, but not of sodium or calcium ion channel blockers, facilitated NaF-induced toxicity in the cells. A JNK specific inhibitor (SP600125) prevented the NaF-induced increase in growth arrest and the DNA damage-inducible protein 45?. Further, NaF-mediated loss of mitochondrial membrane potential was apparently inhibited by pifithrin-? or CAT inhibitor. These findings suggest that NaF affects viability of mESCs in a concentration-dependent manner, where more than 1 mM NaF causes apoptosis through hydroxyl radical-dependent and caspase- and JNK-mediated pathways. -- Highlights: ? The mode of NaF-induced cell death and the mechanisms involved were examined. ? NaF induced mainly apoptotic death of mouse embryonic stem cells (mESCs). ? NaF induced mitochondrial-mediated and caspase-dependent apoptosis. ? JNK- and p53-mediated pathways are involved in NaF-mediated apoptosis in the cells. ? ROS are the up-stream effector in NaF-mediated activation of JNK and p53 in mESCs.

Nguyen Ngoc, Tam Dan [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of)] [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Son, Young-Ok [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States)] [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Lim, Shin-Saeng [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of) [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Department of Bioactive Material Sciences and Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Shi, Xianglin [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States)] [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Kim, Jong-Ghee [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of)] [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Heo, Jung Sun [Department of Maxillofacial Biomedical Engineering and Institute of Oral Biology, School of Dentistry, Kyung Hee University, Seoul 130-701 (Korea, Republic of)] [Department of Maxillofacial Biomedical Engineering and Institute of Oral Biology, School of Dentistry, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Choe, Youngji [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of)] [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Jeon, Young-Mi, E-mail: young@jbnu.ac.kr [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of)] [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Lee, Jeong-Chae, E-mail: leejc88@jbnu.ac.kr [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of) [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Department of Bioactive Material Sciences and Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of)

2012-03-15

7

A Humanized Antibody that Binds to the Interleukin 2 Receptor  

Microsoft Academic Search

The anti-Tac monoclonal antibody is known to bind to the p55 chain of the human interleukin 2 receptor and to inhibit proliferation of T cells by blocking interleukin 2 binding. However, use of anti-Tac as an immunosuppressant drug would be impaired by the human immune response against this murine antibody. We have therefore constructed a ``humanized'' antibody by combining the

Cary Queen; William P. Schneider; Harold E. Selick; Philip W. Payne; Nicholas F. Landolfi; James F. Duncan; Nevenka M. Avdalovic; Michael Levitt; Richard P. Junghans; Thomas A. Waldmann

1989-01-01

8

Bilateral carpal tunnel syndrome associated with interleukin 2 therapy.  

PubMed Central

We report the development of synchronous bilateral carpal tunnel syndrome in a woman with metastatic colorectal cancer, undergoing treatment with recombinant interleukin 2. A carpal tunnel decompression was carried out on the hand which was more severely affected, with a gradual recovery in median nerve function. To the best of our knowledge, this is the first reported case of carpal tunnel syndrome in association with recombinant interleukin 2. PMID:1437960

Heys, S. D.; Mills, K. L.; Eremin, O.

1992-01-01

9

Nicotine exaggerates LPS-induced airway hyperreactivity via JNK-mediated up-regulation of Toll-like receptor 4.  

PubMed

Tobacco smokers often display increased airway hyperreactivity (AHR) when faced with bacterial infections. The present study uses a murine organ-culture model to dissect the mechanisms involved in this exaggerated smooth muscle response. Nicotine simulates the effects of smoking, and LPS represents bacterial infection. Contractile responses of isolated murine tracheal segments were analyzed in myographs after organ culture with increasing concentrations of LPS and/or nicotine for 4 days with or without specific MAPK inhibitors. Nicotine's effect on the expression of cell surface Toll-like receptors (TLRs), MCP-1, COX-2, and TNF-? were examined by real-time PCR. Increased protein expression was verified by immunohistochemistry. LPS concentration-dependently increased contractile responses to bradykinin and des-Arg(9)-bradykinin. A combination of nicotine and low-dose LPS caused powerful synergistic contractions along with increased kinin receptor expression. Specific kinin B1 and B2 receptor inhibitors blocked this reaction. Nicotine increased mRNA and protein expression of TLR4 and -6 in the epithelium and smooth muscle layer, with MCP-1 and COX-2 mRNA increasing in parallel. Specific inhibition of JNK attenuated nicotine's effects. In conclusion, long-term exposure to nicotine up-regulated the expression of TLR4 and -6 via a JNK-related pathway, causing an exaggeration of the LPS-induced local airway inflammation and increased AHR. This might offer a mechanistic explanation to the increased AHR seen in tobacco smokers confronted with bacterial infections. PMID:24669857

Xu, Yuan; Zhang, Yaping; Cardell, Lars-Olaf

2014-09-01

10

Leflunomide or A77 1726 protect from acetaminophen-induced cell injury through inhibition of JNK-mediated mitochondrial permeability transition in immortalized human hepatocytes  

SciTech Connect

Leflunomide, a disease-modifying anti-rheumatic drug, protects against T-cell-mediated liver injury by poorly understood mechanisms. The active metabolite of leflunomide, A77 1726 (teriflunomide) has been shown to inhibit stress-activated protein kinases (JNK pathway), which are key regulators of mitochondria-mediated cell death. Therefore, we hypothesized that leflunomide may protect from drugs that induce the mitochondrial permeability transition (mPT) by blocking the JNK signaling pathway. To this end, we exposed cultured immortalized human hepatocytes (HC-04) to the standard protoxicant drug acetaminophen (APAP), which induces CsA-sensitive mPT-mediated cell death. We determined the effects of leflunomide on the extent of APAP-induced hepatocyte injury and the upstream JNK-mediated mitochondrial signaling pathways. We found that leflunomide or A77 1726 concentration-dependently protected hepatocytes from APAP (1 mM)-induced mitochondrial permeabilization and lethal cell injury. This was not due to proximal inhibition of CYP-catalyzed APAP bioactivation to its thiol-reactive metabolite. Instead, we demonstrate that leflunomide (20 {mu}M) inhibited the APAP-induced early (3 h) activation (phosphorylation) of JNK1/2, thus inhibiting phosphorylation of the anti-apoptotic protein Bcl-2 and preventing P-Bcl-2-mediated induction of the mPT. This greatly attenuated mitochondrial cytochrome c release, which we used as a marker for mitochondrial permeabilization. The specific JNK2 inhibitor SP600125 similarly protected from APAP-induced cell death. In conclusion, these findings are consistent with our hypothesis that leflunomide protects from protoxicant-induced hepatocyte injury by inhibiting JNK signaling and preventing mPT induction.

Latchoumycandane, Calivarathan [Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597 (Singapore); Seah, Quee Ming [Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597 (Singapore); Tan, Rachel C.H. [Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597 (Singapore); Sattabongkot, Jetsumon [Armed Forces Research Institute of Medical Sciences, Bangkok 10400 (Thailand); Beerheide, Walter [Siam Life Science Ltd., Bangkok 10500 (Thailand); Boelsterli, Urs A. [Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597 (Singapore) and Department of Pharmacy, Faculty of Science, National University of Singapore, Singapore 117597 (Singapore)]. E-mail: phcbua@nus.edu.sg

2006-11-15

11

Effects of interleukin-2 and interleukin-2-activated cells on in vitro myelopoiesis.  

PubMed Central

Lymphokine-activated killer (LAK) cells from human peripheral blood mononuclear cells cultured with recombinant interleukin-2 (IL-2) have been used clinically in adoptive immunotherapy for cancer patients. To study the influence of LAK cells and IL-2 on haematopoiesis, an in vitro assay system for colony formation of granulocyte-macrophage progenitor cells (GM-CFC) was used. LAK cells from cultures of either human peripheral blood (PB) or human bone marrow (BM) mononuclear cells were both inhibitory to allogeneic BM-derived GM-CFC. Inhibitory activity could be transferred with supernatants from co-cultures of LAK cells and BM targets, but also from the IL-2 activated PB- or BM-derived cells alone. The inhibitory activity from the initially non-cytotoxic/non-inhibitory BM population was rapidly induced by IL-2 activation, and preceded the generation of cytotoxic LAK cells in the culture. These experiments show that inhibition of haematopoietic progenitor cells by IL-2 is not dependent on generation of cytotoxic LAK cells, but rather the result of IL-2-induced cytokine production. We conclude that the synergistic action of interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) may contribute to inhibition, but that also other cytokines are responsible for the observed inhibition of BM-derived GM-CFC. PMID:2118847

Clerigue, M; Pisa, P; Tsai, L; Hanson, M

1990-01-01

12

Ginsenoside Rb3 Protects Cardiomyocytes against Ischemia-Reperfusion Injury via the Inhibition of JNK-Mediated NF-?B Pathway: A Mouse Cardiomyocyte Model  

PubMed Central

Ginsenoside Rb3 is extracted from the plant Panax ginseng and plays important roles in cardiovascular diseases, including myocardial ischemia-reperfusion (I/R) injury. NF-?B is an important transcription factor involved in I/R injury. However, the underlying mechanism of ginsenoside Rb3 in myocardial I/R injury remains poorly understood. In the current study, a model of myocardial I/R injury was induced via oxygen and glucose deprivation (OGD) followed by reperfusion (OGD-Rep) in mouse cardiac myoblast H9c2 cells. Our data demonstrate that ginsenoside Rb3 suppresses OGD-Rep-induced cell apoptosis by the suppression of ROS generation. By detecting the NF-?B signaling pathway, we discover that the protective effect of ginsenoside Rb3 on the OGD-Rep injury is closely related to the inhibition of NF-?B activity. Ginsenoside Rb3 inhibits the upregulation of phospho-I?B-? and nuclear translocation of NF-?B subunit p65 which are induced by ORD-Rep injury. In addition, the extract also inhibits the OGD-Rep-induced increase in the expression of inflammation-related factors, such as IL-6, TNF-?, monocyte chemotactic protein-1 (MCP-1), MMP-2 and MMP-9. However, LPS treatment alleviates the protective roles of ginsenoside Rb3 and activates the NF-?B pathway. Finally, the upstream factors of NF-?B were analyzed, including the Akt/Foxo3a and MAPK signaling pathways. We find that ginsenoside Rb3 pretreatment only decreases the phosphorylation of JNK induced by OGD-Rep injury, an indicator of the MAPK pathway. Importantly, an inhibitor of phospho-JNK, SP600125, protects against OGD-Rep induced apoptosis and inhibited NF-?B signaling pathway, similar to the roles of ginsenoside Rb3. Taken together, our results demonstrate that the protective effect of ginsenoside Rb3 on the OGD-Rep injury is attributed to the inhibition of JNK-mediated NF-?B activation, suggesting that ginsenoside Rb3 has the potential to serve as a novel therapeutic agent for myocardial I/R injury. PMID:25084093

Xie, Zulong; Yang, Shuang; Xu, Wei; Hou, Jingbo; Yu, Bo

2014-01-01

13

Expression of Tac antigen component of bovine interleukin-2 receptor in different leukocyte populations infected with Theileria parva or Theileria annulata.  

PubMed Central

The Tac antigen component of the bovine interleukin-2 receptor was expressed as a Cro-beta-galactosidase fusion protein in Escherichia coli and used to raise antibodies in rabbits. These antibodies were used for flow cytofluorimetric analysis to investigate the expression of Tac antigen in a variety of Theileria parva-infected cell lines and also in three Theileria annulata-infected cell lines. Cells expressing Tac antigen on their surface were found in all T. parva-infected cell lines tested whether these were of T- or B-cell origin. T cells expressing Tac antigen could be CD4- CD8-, CD4+ CD8-, CD4- CD8+, or CD4+ CD8+. Tac antigen expression was observed both in cultures which had been maintained in the laboratory for several years and in transformed cell lines which had recently been established by infection of lymphocytes in vitro with T. parva. Northern (RNA) blot analysis demonstrated Tac antigen transcripts in RNA isolated from all T. parva-infected cell lines. Three T. annulata-infected cell lines which were not of T-cell origin were also tested. Two of them expressed Tac antigen on their surface. Abundant Tac antigen mRNA was detected in these T. annulata-infected cell lines, but only trace amounts were demonstrated in the third cell line, which contained very few Tac antigen-expressing cells. In all cell lines tested, whether cloned or uncloned, a proportion of the cells did not express detectable levels of Tac antigen on their surface. This was also the case for a number of other leukocyte surface markers. In addition, we showed that the interleukin-2 receptors were biologically functional, because addition of recombinant interleukin-2 to cultures stimulated cell proliferation. Recombinant interleukin-2 treatment also resulted in increased amounts of steady-state Tac antigen mRNA. The relevance of interleukin-2 receptor expression on Theileria-infected cells is discussed. Images PMID:1979317

Dobbelaere, D A; Prospero, T D; Roditi, I J; Kelke, C; Baumann, I; Eichhorn, M; Williams, R O; Ahmed, J S; Baldwin, C L; Clevers, H

1990-01-01

14

Effect of spaceflight on lymphocyte proliferation and interleukin-2 production  

NASA Technical Reports Server (NTRS)

In this study, inguinal lymp node lymphocytes from rats flown on the Cosmos 2044 mission were tested for proliferation and interleukin-2 (IL-2) production. Cells cultured with mitogenic lectins, phorbol ester, and calcium ionophore, or T-cell mitogen and lymphokine, were assayed for DNA synthesis by (H-3) thymidine incorporation. Lymphocytes incubated with a T-cell mitogen alone also were tested for IL-2 production. Proliferation of lymphocytes from flight rats was not significantly different from controls for any of the mitogens tested. Furthermore, lymph node lymphocytes from control and flown rats produced similar amounts of IL-23. Thus microgravity may act on lymphocytes in a tissue-specific manner, a new finding that could impact on the evaluation of spaceflight effects on immunocompetence.

Nash, Patricia V.; Konstantinova, Irina V.; Fuchs, Boris B.; Rakhmilevich, Alexandr L.; Lesniak, A. T.; Mastro, Andrea M.

1992-01-01

15

Enhancement of interleukin-2 immunotherapy with L-arginine.  

PubMed Central

Nutrient substrates have been shown to enhance cell-mediated immunity, but their role as adjuvants to immunotherapy has not been previously determined. This study evaluated L-arginine as an essential substrate for optimal generation of lymphokine-activated killer (LAK) cells. This experiment also assessed supplemental dietary L-arginine as a means to potentiate the host antitumor response to interleukin-2 (IL-2) in a murine neuroblastoma (NRB) model. A/J mice received 1% arginine or isonitrogenous 1.7% glycine in addition to a regular diet 14 days before subcutaneous inoculation with C1300 NRB cells. Twenty-four hours later, animals received low (1 x 10(6) U/kg three times a day) or high (3 x 10(6) U/kg three times a day) doses of IL-2 or saline intraperitoneally for 4 days. On days 4 and 10 post-C1300 NRB inoculation, mice were killed for assessment of natural killer cell and tumor specific cytotoxicity. Remaining animals were followed for tumor incidence, tumor growth, and duration of host survival. Interleukin-2 therapy in mice receiving dietary arginine compared with those receiving glycine resulted in significantly augmented natural killer cell cytotoxicity (day 4) and generation of specific tumoricidal mechanisms (day 10). The addition of dietary arginine to low-dose IL-2 therapy significantly diminished C1300 NRB engraftment (p less than 0.05) and growth (p less than 0.001) and prolonged the duration of host survival (p less than 0.05) compared with the glycine treatment group. In vitro studies demonstrated that L-arginine is an essential substrate for optimal generation of LAK cells. Thus, supplemental dietary L-arginine enhances lymphocyte cytotoxic mechanisms and potentiates IL-2 immunotherapy. PMID:1546902

Lieberman, M D; Nishioka, K; Redmond, H P; Daly, J M

1992-01-01

16

Does interleukin-2 restore lymphocyte responses suppressed by Trypanosoma cruzi?  

PubMed Central

There has been disagreement about the ability of exogenous interleukin-2 (IL-2) to restore responsiveness to lymphocytes from either Trypanosoma cruzi-infected animals or normal individuals co-cultured with this parasite. The discrepancy has been attributed to the use of different strains of mice or T. cruzi isolates, or to the use of lymphoid cells from different organs. As T. cruzi inhibits the expression of IL-2 receptors by activated lymphocytes in vitro, we were able to test whether restoration of responsiveness by exogenous IL-2 might depend on the level of suppression present in the system. Human or mouse lymphocytes stimulated with phytohaemagglutinin (PHA) exhibited gradual decreases in IL-2 receptor expression, [3H]thymidine incorporation and IL-2 secretion as the concentration of T. cruzi in the culture increased. Exogenous IL-2 afforded a degree of restoration of both IL-2 receptor expression and [3H]thymidine uptake which was substantial at the lower, but very small--if any--at the higher, parasite concentrations tested. Trypanosoma cruzi could not have competed with the lymphocytes for IL-2 because it did not bind significant amounts of this cytokine. These results suggested that the controversy about the corrective effects of IL-2 may be more apparent than real, reflecting variations in the extent of immunosuppression present in different model systems of T. cruzi-associated immunosuppression. PMID:8288318

Kierszenbaum, F; Mejia Lopez, H; Sztein, M B

1993-01-01

17

Intravenous ascorbic acid as an adjuvant to interleukin-2 immunotherapy  

PubMed Central

Interleukin-2 (IL-2) therapy has been demonstrated to induce responses in 10-20% of advanced melanoma and renal cell carcinoma patients, which translates into durable remissions in up to half of the responsers. Unfortunately the use of IL-2 has been associated with severe toxicity and death. It has been previously observed and reported that IL-2 therapy causes a major drop in circulating levels of ascorbic acid (AA). The IL-2 induced toxicity shares many features with sepsis such as capillary leakage, systemic complement activation, and a relatively non-specific rise in inflammatory mediators such as TNF-alpha, C-reactive protein, and in advanced cases organ failure. Animal models and clinical studies have shown rapid depletion of AA in conditions of sepsis and amelioration associated with administration of AA (JTM 9:1-7, 2011). In contrast to other approaches to dealing with IL-2 toxicity, which may also interfere with therapeutic effects, AA possesses the added advantage of having direct antitumor activity through cytotoxic mechanisms and suppression of angiogenesis. Here we present a scientific rationale to support the assessment of intravenous AA as an adjuvant to decrease IL-2 mediated toxicity and possibly increase treatment efficacy. PMID:24884532

2014-01-01

18

Modeling interleukin-2-based immunotherapy in AIDS pathogenesis.  

PubMed

In this paper, we sought to identify the CD4(+) T-cell dynamics in the course of HIV infection in response to continuous and intermittent intravenous courses of interleukin-2 (IL-2), the principal cytokine responsible for progression of CD4(+) T-lymphocytes from the G1 to the S phase of the cell cycle. Based on multivariate regression models, previous literature has concluded that the increase in survival of CD4(+) T-cell appears to be the critical mechanism leading to sustained CD4(+) T-cell levels in HIV-infected patients receiving intermittent IL-2 therapy. Underscored by comprehensive mathematical modeling, a major finding of the present work is related to the fact that, rather than due to any increase in survival of CD4(+) T-cells, the expressive, selective and sustained CD4(+) T-cell expansions following IL-2 administration may be related to the role of IL-2 in modulating the dynamics of Fas-dependent apoptotic pathways, such as activation-induced cell death (AICD) or HIV-specific apoptotic routes triggered by viral proteins. PMID:23806696

Joly, Marcel; Odloak, Darci

2013-10-21

19

Focus on FOCIS: interleukin 2 treatment associated autoimmunity.  

PubMed

A patient from the University of Pittsburgh is presented who developed autoimmunity during IL-2 based combination therapy. IL-2 was originally described as a "T cell growth factor" capable of expanding previously activated T cells, enhancing the cytotoxicity of antigen-specific cytotoxic T cells and NK cells. High dose Interleukin 2 (HDIL2) is now FDA-approved for therapy of patients with metastatic melanoma and renal cell carcinoma, based on its ability to induce durable responses in 5-10% of patients. The antitumor effect of HDIL2 is incompletely understood, but it appears that this regimen alters the balance of immigrant T effector cells in relation to T suppressor cells. It promotes a less immunosuppressive tumor microenvironment, inducing tumor regression in a subset of patients that is yet to be defined. The antitumor activity of IL-2, as for other agents that promote durable antitumor activity against melanoma such as interferon alpha and anti-CTLA4 antibody, is frequently associated with development of autoimmunity as observed in this patient. We present studies investigating the mechanisms for the therapeutic benefit of HDIL2 in melanoma. PMID:18405863

Moschos, Stergios J; Mandic, Maja; Kirkwood, John M; Storkus, Walter J; Lotze, Michael T

2008-05-01

20

Interleukin-2 induces early multisystem organ edema mediated by neutrophils.  

PubMed Central

Interleukin-2 (IL-2), an agent known to activate neutrophils (PMN) with thromboxane (Tx)B2 release, produces pulmonary edema within 6 hours of intravenous infusion. This study tests the role of PMN in mediating the edema. Anesthetized rats received 10(6)U recombinant human IL-2 (n = 15) or vehicle (n = 14) as a constant intravenous infusion during a period of 1 hour. At this time there was leukopenia 3.63 +/- 0.43 (x10(3)/mm3) relative to vehicle-infused control rats 6.12 +/- 0.86 and a decline in PMN, 2.19 +/- 0.14 relative to control value of 3.33 +/- 0.05 (both p less than 0.05). After 6 hours edema, as measured by increase in the wet to dry weight (W/d) ratio, was present in the lungs (4.93 +/- 0.20 relative to control 4.06 +/- 0.10), heart (4.09 +/- 0.11 versus 3.76 +/- 0.08), liver (3.50 +/- 0.10 versus 3.18 +/- 0.10), and kidney (4.25 +/- 0.07 versus 4.00 +/- 0.07) (all p less than 0.05). There was increased lung permeability demonstrated by bronchoalveolar lavage fluid protein concentration of 1970 +/- 210 micrograms/mL relative to control 460 +/- 90 micrograms/mL (p less than 0.05). Interleukin-2 resulted in lung PMN sequestration of 53 +/- 7 PMN/10 high-power fields (HPF) relative to 23 +/- 2 PMN/10 HPF in controls (p less than 0.05) and increased plasma TxB2 levels to 1290 +/- 245 pg/mL relative to control 481 +/- 93 pg/mL (p less than 0.05). Pretreatment of other rats (n = 8) with selective anti-rat neutrophil antiserum 18 hours before the experiment led to a peripheral PMN count 10% of baseline and prevented edema in the lungs (W/d ratio 4.20 +/- 0.16) and heart (3.67 +/- 0.07) (both p less than 0.05) but not liver or kidney. Protein in lung lavage was reduced to 760 +/- 220 micrograms/mL (p less than 0.05). The protection afforded by leukopenia was associated with lack of PMN sequestration and prevention of the increase in plasma Tx levels (484 +/- 120 pg/mL, p less than 0.05). These data indicate that the rapid induction of lung and heart edema with a 1-hour infusion of IL-2 in the rat is mediated, in large part, by activated PMNs. PMID:1867524

Welbourn, R; Goldman, G; Kobzik, L; Paterson, I; Shepro, D; Hechtman, H B

1991-01-01

21

Myelostimulatory activity of recombinant human interleukin-2 in mice  

SciTech Connect

In a series of studies designed to extend our understanding of interleukin-2 (IL-2) and to study the effect of biologic response modifiers on bone marrow, we observed that administering recombinant human (rH) IL-2 to normal mice resulted in an increase in the frequency of colony-forming units-culture (CFU-C) in bone marrow. In addition, rH IL-2 was able to accelerate host recovery from cyclophosphamide (CTX)- or radiation-induced bone marrow depression and peripheral blood leukopenia. Not only can rH IL-2 accelerate, in a dose-dependent manner, the return of bone marrow, peripheral blood cellularity, and CFU-C frequency to normal levels following cytoreduction by CTX or irradiation, but it also significantly increases CFU-C frequency to greater than normal levels. Furthermore, rH IL-2 can significantly prolong survival of animals receiving a lethal dose of irradiation or CTX. Thus, multiple mechanisms are responsible for the synergistic therapeutic activity associated with rH IL-2 and CTX. rH IL-2 does not act only as an immunomodulatory agent in the presence or absence of suppressor T cells, but also accelerates host recovery from cytoreductive agents, resulting in decreased leukopenia and perhaps resistances to secondary infection. Thus, rH IL-2 plus chemotherapy may increase therapeutic activity against neoplastic disease, not only by adding immune stimulation to the direct antitumor effect of the drug but also by allowing delivery of higher, more effective doses of chemotherapy.

Talmadge, J.E.; Schneider, M.; Keller, J.; Ruscetti, F.; Longo, D.; Pennington, R.; Bowersox, O.; Tribble, H.

1989-05-01

22

Fluorinated 4-quinolones induce hyperproduction of interleukin 2.  

PubMed Central

The fluorinated 4-quinolones are a "new" group of antibiotics with a broad antibacterial spectrum. They are already widely used in clinical practice. Previous studies have shown that these drugs increase the uptake of [3H]thymidine into DNA of mitogen-stimulated lymphocytes but inhibit cell growth and immunoglobulin secretion. This study shows that the 4-quinolones strongly (up to 100 times) increase the recovery of interleukin 2 (IL-2) in culture supernatants of phytohemagglutinin (PHA)-stimulated normal human lymphocytes and also prolong the kinetics of IL-2 production. The effect was significant at clinically achievable concentrations (5 micrograms/ml). In addition to hyperproduction of IL-2, the level of RNA hybridizing with a human IL-2 cDNA probe was also intensely elevated (16-32 times) in PHA-stimulated lymphocytes cultured with ciprofloxacin (80 micrograms/ml). The mechanism responsible for 4-quinolone-mediated effects on T cells is at present unclear, but evidence is presented that suggests the effect is not exerted at the level of protein kinase C activation. Ciprofloxacin at 80 micrograms/ml also decreased the expression of IL-2 receptors measured by immunofluorescence with CD 25 antibodies and a radiolabeled IL-2 binding assay. At the same concentration of ciprofloxacin, there was a very low expression of the transferrin receptor and the cell size increased very little in human lymphocytes after PHA stimulation. The enhanced IL-2 production by 4-quinolones may contribute to side effects reported when these drugs are used for treatment of patients. Images PMID:2539601

Riesbeck, K; Andersson, J; Gullberg, M; Forsgren, A

1989-01-01

23

Interleukin-2/Anti-Interleukin-2 Immune Complex Expands Regulatory T Cells and Reduces Angiotensin II-Induced Aortic Stiffening  

PubMed Central

Adaptive immune function is implicated in the pathogenesis of vascular disease. Inhibition of T-lymphocyte function has been shown to reduce hypertension, target-organ damage, and vascular stiffness. To study the role of immune inhibitory cells, CD4+CD25+Foxp3+ regulatory T cells (Tregs), on vascular stiffness, we stimulated the proliferation of Treg lymphocytes in vivo using a novel cytokine immune complex of Interleukin-2 (IL-2) and anti-IL-2 monoclonal antibody clone JES6-1 (mAbCD25). Three-month-old male C57BL/6J mice were treated with IL-2/mAbCD25 concomitantly with continuous infusion of angiotensin type 1 receptor agonist, [Val5]angiotensin II. Our results indicate that the IL-2/mAbCD25 complex effectively induced Treg phenotype expansion by 5-fold in the spleens with minimal effects on total CD4+ and CD8+ T-lymphocyte numbers. The IL-2/mAbCD25 complex inhibited angiotensin II-mediated aortic collagen remodeling and the resulting stiffening, analyzed with in vivo pulse wave velocity and effective Young's modulus. Furthermore, the IL-2/mAbCD25 complex suppressed angiotensin II-mediated Th17 responses in the lymphoid organs and reduced gene expression of IL-17 as well as T cell and macrophage infiltrates in the aortic tissue. This study provides data that support the protective roles of Tregs in vascular stiffening and highlights the use of the IL-2/mAbCD25 complex as a new potential therapy in angiotensin II-related vascular diseases. PMID:25258681

Eberson, Lance S.; Secomb, Timothy W.; Larmonier, Nicolas; Larson, Douglas F.

2014-01-01

24

Chemical Modification of Recombinant Interleukin 2 by Polyethylene Glycol Increases Its Potency in the Murine Meth A Sarcoma Model  

NASA Astrophysics Data System (ADS)

Recombinant human interleukin 2 purified from Escherichia coli has limited solubility at neutral pH and a short circulatory half-life. This recombinant interleukin 2 was chemically modified by an active ester of polyethylene glycol. The modified interleukin 2 was purified by hydrophobic interaction chromatography and characterized by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and isoelectric focusing. This conjugate was compared to unmodified recombinant interleukin 2 in vitro and in vivo. Covalent attachment of the hydrophilic polymer polyethylene glycol enhanced the solubility of interleukin 2, decreased its plasma clearance, and increased its antitumor potency in the Meth A murine sarcoma model.

Katre, Nandini V.; Knauf, Michael J.; Laird, Walter J.

1987-03-01

25

White blood cell and lymphocyte populations following interleukin-2 administration in the spontaneously hypertensive rat.  

PubMed

The immune system has been linked to the pathogenesis of hypertension in the spontaneously hypertensive rat (SHR). Recently interleukin-2 has been reported to inhibit the development of hypertension in the SHR, but no measures of different lymphocyte populations were made. To test the effect of interleukin-2 we repeated the protocol in the report by injecting forty two day old, male SHR and WKY rats, and in addition, analyzed lymphocyte subpopulations. Untreated, age matched rats of the same strain were used as a control. At three and four months of age blood was drawn from all animals. Monoclonal antibodies were used to fluorescently label different lymphocyte subpopulations. The populations examined were the total T-cells, T-nonhelper cells, T-helper cells and B-cells. Total numbers of lymphocytes and white blood cells were also examined. Blood pressures were measured in conscious, restrained animals at two and four months of age. The results showed no attenuation of blood pressure in the interleukin-2 treated SHR at either age. The interleukin-2 treated SHR had a decrease in the percentage of B-cells and an increase in the percentage of T-nonhelper cells relative to the control SHR. Both treated and untreated SHR had increased numbers of white blood cells and lymphocytes compared to both groups of WKY. We conclude that the interleukin-2 used was active but failed to have any effect on blood pressure or absolute numbers of white blood cells and lymphocytes in the treated animals. PMID:1424219

Fannon, L D; Phillips, M I

1992-01-01

26

In Vitro Interleukin-1 and 2 Production and Interleukin 2 Receptor Expression in the Rhesus Monkey  

NASA Technical Reports Server (NTRS)

Anti-human monoclonal antibodies were used to detect and quantify interleukins-1 and 2 and interleukin-2 receptor expression in peripheral blood mononuclear cells from a rhesus monkey. Interleukin-1 production could be induced by phorbol esters (PMA) and was potentiated by phytohemagglutinin (PHA). Interleukin-2 secretion could also be induced by the combination of PHA and PMA, but only weakly with PHA alone. Interleukin-2 receptor expression was present in a subpopulation of unstimulated lymphocytes and could be enhanced by PHA or PMA. These data show once again that the rhesus monkey immune system is cross-reactive with the human one and that rhesus macaque could be a good model to study interleukin therapy.

Schmitt, Didier A.; Sonnenfeld, Gerald; Husson, David; Tkaczuk, Jean; Andre, Eric; Schaffar, Laurance

1996-01-01

27

Interleukin2 receptor ? chain regulates the size and content of the peripheral lymphoid compartment  

Microsoft Academic Search

Interleukin-2 receptor ? chain (IL-2R?) expression occurs at specific stages of early T and B lymphocyte development and is induced upon activation of mature lymphocytes. Young mice that lack IL-2R? have phenotypically normal development of T and B cells. However, as adults, these mice develop massive enlargement of peripheral lymphoid organs associated with polyclonal T and B cell expansion, which,

Dennis M. Willerford; Jianzhu Chen; Judith A. Ferry; Laurie Davidson; Averil Ma; Frederick W. Alt

1995-01-01

28

Deregulated T Cell Activation and Autoimmunity in Mice Lacking Interleukin 2 Receptor beta  

Microsoft Academic Search

In mice lacking the interleukin-2 receptor beta chain (IL-2Rbeta), T cells were shown to be spontaneously activated, resulting in exhaustive differentiation of B cells into plasma cells and the appearance of high serum concentrations of immunoglobulins G1 and E as well as autoantibodies that cause hemolytic anemia. Marked infiltrative granulocytopoiesis was also apparent, and the animals died after about 12

Haruhiko Suzuki; Thomas M. Kundig; Caren Furlonger; Andrew Wakeham; Emma Timms; Toshifumi Matsuyama; Rudolf Schmits; John J. L. Simard; Pamela S. Ohashi; Henrik Griesser; Tadatsugu Taniguchi; Christopher J. Paige; Tak W. Mak

1995-01-01

29

Mycobacterial induction of activated killer cells: possible role of tyrosine kinase activity in interleukin-2 receptor alpha expression.  

PubMed Central

Mycobacterium avium is an intracellular opportunistic pathogen commonly seen in AIDS patients. M. avium-infected monocytes have been recently shown to be lysed by interleukin-2 (IL-2)-activated killer cells. Since some bacterial products can directly augment natural killer activity, we examined the ability of these microorganisms to induce killer cell activity. Coculture of M. avium with large granular lymphocytes (LGL) was found to augment the ability of LGL to lyse both tumor cells and bacterially infected autologous monocytes. The induction of tumoricidal activity by M. avium was only partially neutralized by the presence of anti-IL-2 antibodies, indicating that both IL-2-dependent and IL-2-independent mechanisms are responsible for activation of killer cells. Furthermore, only the direct interaction between bacterium and LGL could induce the expression of both IL-2 receptor alpha protein and mRNA, an effect which was abrogated by the presence of genistein, a tyrosine kinase inhibitor. Thus, M. avium was seen to induce killer cells, an activity that is concomitant with the up-regulation of IL-2 receptor alpha, or Tac antigen, expression and which involves signal transduction mechanisms mediated by tyrosine kinase activity. Images PMID:1612749

Blanchard, D K; McMillen, S; Hoffman, S L; Djeu, J Y

1992-01-01

30

Phase I trial of interleukin-2 and high-dose arginine butyrate in metastatic colorectal cancer  

Microsoft Academic Search

Introduction:   Interleukin-2 (IL-2) and sodium butyrate allow rats to be cured of peritoneal carcinomatosis from colon cancer. We performed\\u000a a phase I trial of IL-2 and high-dose arginine butyrate (ArgB) in patients with advanced metastatic colorectal cancer. Patients and methods: From April to July 1997, six patients were included in the trail; they had a median age of 52 years,

J. Y. Douillard; J. Bennouna; F. Vavasseur; R. Deporte-Fety; P. Thomare; F. Giacalone; K. Meflah

2000-01-01

31

RESPONSE OF RESTING HUMAN PERIPHERAL BLOOD NATURAL KILLER CELLS TO INTERLEUKIN 2  

Microsoft Academic Search

Resting T cells can be activated to proliferate by antigens and mitogenic lectins, but their proliferation depends on a hormone-like lymphokine, T cell growth factor or interleukin 2 (IL-2) ~ (1-2). Activation of T cells by a mitogenic stimulus induces both secretion of minute quantities of IL-2 and the appearance of a high-affinity receptor for IL-2 on the T cell

GIORGIO TRINCHIERI; MICHIKO MATSUMOTO-KOBAYASHI; STEVEN C. CLARK; JASBIR SEEHRA; LUCILLE LONDON; BICE PERUSSIA

32

Molecular signatures induced by interleukin-2 on peripheral blood mononuclear cells and T cell subsets  

Microsoft Academic Search

: Experimentally, interleukin-2 (IL-2) exerts complex immunological functions promoting the proliferation, survival and activation of T cells on one hand and inducing immune regulatory mechanisms on the other. This complexity results from a cross talk among immune cells which sways the effects of IL-2 according to the experimental or clinical condition tested. Recombinant IL-2 (rIL-2) stimulation of peripheral blood mononuclear

Ping Jin; Ena Wang; Maurizio Provenzano; Sara Deola; Silvia Selleri; Jiaqiang Ren; Sonia Voiculescu; David Stroncek; Monica C Panelli; Francesco M Marincola

2006-01-01

33

Comparative production of human interleukin-2 fused with green fluorescent protein in several recombinant expression systems  

Microsoft Academic Search

The selection of an optimal recombinant expression system is important for successful protein production. Here, we compared production of human interleukin-2 (hIL-2)-green fluorescent protein (GFP) fusion proteins in several expression systems such as bacteria Escherichia coli, yeast Pichia pastoris, insect Spodoptera frugiperda Sf-9 cells, insect Tricoplusia ni larvae, and insect Drosophila melanogaster S2 cells. Due to the highly hydrophobic nature

Hyung Joon Cha; Hwa Sung Shin; Hye Jung Lim; Hye Sook Cho; Nimish N. Dalal; Minh Q. Pham; William E. Bentley

2005-01-01

34

Interleukin1 Costimulatory Activity on the Interleukin2 Promoter Via AP1  

Microsoft Academic Search

Interleukin-1 (IL-1) is a major regulator of inflammation and immunity. IL-1 induces T lymphocyte growth by acting as a second signal (together with antigen) in enhancing the production of interleukin-2 (IL-2). An IL-1--responsive element in the promoter region of the human IL-2 gene was similar to the binding site for the transcription factor AP-1. IL-1 enhanced expression of c-jun messenger

Kathrin Muegge; Thomas M. Williams; Jeffrey Kant; Michael Karin; Robert Chiu; Albrecht Schmidt; Ulrich Siebenlist; Howard A. Young; Scott K. Durum

1989-01-01

35

Neuropsychological and neurophysiological assessment of the central effects of interleukin-2 administration.  

PubMed

Neuropsychiatric disturbances may occur following interleukin-2 (IL2) administration. We studied the effects of IL2 infusion on cerebral functions in 7 patients with neuropsychological tests and event-related evoked potentials (P300). We observed a failure in the cognitive performances, an increase in latency, and a decrease in amplitude of P300. These effects followed IL2 administration and were reversible. PMID:8343265

Caraceni, A; Martini, C; Belli, F; Mascheroni, L; Rivoltini, L; Arienti, F; Cascinelli, N

1993-01-01

36

Interleukin2 production by persons with the generalized lymphadenopathy syndrome or the acquired immune deficiency syndrome  

Microsoft Academic Search

We measured production of interleukin-2 (IL-2) by phytohemagglutinin-stimulated peripheral blood mononuclear cells from 27 heterosexual persons, 43 asymptomatic homosexual men, 34 homosexual men with generalized lymphadenopathy syndrome (GLS), and 21 patients with acquired immune deficiency syndrome (AIDS). Asymptomatic heterosexual and homosexual subjects produced comparable amounts of IL-2, but 8 of 11 AIDS patients with opportunistic infections and two of three

Charles H. Kirkpatrick; Kathleen C. Davis; Charles R. Horsburgh; David L. Cohn; Kent Penley; Franklyn N. Judson

1985-01-01

37

Distribution of interleukin-2, -4, -10, tumour necrosis factor-alpha and transforming growth factor-beta mRNAs in oral lichen planus.  

PubMed

In the present study, MRNA for the cytokines interleukin-2 (IL-2), IL-4, IL-10 tumour necrosis factor-alpha (TNF-alpha) and transforming growth factor beta-1 (TGF-beta-1) were investigated in oral lichen planus (OLP) lesions using in situ hybridization with 35S-labelled oligonucleotide probes on frozen tissue sections. In addition, the expression of interferon-gamma (IFN-gamma), IL-10 and IL-4 mRNAs was analysed in cultured lesional T lymphocytes from oral lichen planus by polymerase chain reaction. Cells expressing mRNA for IL-2, IL-4, IL-10, TNF-alpha and TGF-beta 1 were found in all the biopsies studied. Approximately 1-2% of the total number of infiltrating cells in the lesions were positive for each of the different cytokine mRNAs. Most biopsies contained basement membrane-oriented, mRNA-positive cells. In the cultured T-cell lines, message for IFN-gamma was detected in all the patients, IL-10 in all but one, and IL-4 in just one of the seven patients investigated. The results suggest that mRNA for both pro- and anti-inflammatory cytokines, i.e., mixed T-helper 1 (TH1) and TH2 cytokine profiles, are generated simultaneously by a limited number of cells in chronic lesions of OLP. PMID:10401528

Simark-Mattsson, C; Bergenholtz, G; Jontell, M; Eklund, C; Seymour, G J; Sugerman, P B; Savage, N W; Dahlgren, U I

1999-06-01

38

Inhibition of G-Protein ?? Signaling Enhances T Cell Receptor-Stimulated Interleukin 2 Transcription in CD4+ T Helper Cells  

PubMed Central

G-protein-coupled receptor (GPCR) signaling modulates the expression of cytokines that are drug targets for immune disorders. However, although GPCRs are common targets for other diseases, there are few GPCR-based pharmaceuticals for inflammation. The purpose of this study was to determine whether targeting G-protein ?? (G??) complexes could provide a useful new approach for modulating interleukin 2 (IL-2) levels in CD4+ T helper cells. Gallein, a small molecule inhibitor of G??, increased levels of T cell receptor (TCR)-stimulated IL-2 mRNA in primary human naïve and memory CD4+ T helper cells and in Jurkat human CD4+ leukemia T cells. G?1 and G?2 mRNA accounted for >99% of G? mRNA, and small interfering RNA (siRNA)-mediated silencing of G?1 but not G?2 enhanced TCR-stimulated IL-2 mRNA increases. Blocking G?? enhanced TCR-stimulated increases in IL-2 transcription without affecting IL-2 mRNA stability. Blocking G?? also enhanced TCR-stimulated increases in nuclear localization of nuclear factor of activated T cells 1 (NFAT1), NFAT transcriptional activity, and levels of intracellular Ca2+. Potentiation of IL-2 transcription required continuous G?? inhibition during at least two days of TCR stimulation, suggesting that induction or repression of additional signaling proteins during T cell activation and differentiation might be involved. The potentiation of TCR-stimulated IL-2 transcription that results from blocking G?? in CD4+ T helper cells could have applications for autoimmune diseases. PMID:25629163

Yost, Evan A.; Hynes, Thomas R.; Hartle, Cassandra M.; Ott, Braden J.; Berlot, Catherine H.

2015-01-01

39

Genetically engineered Newcastle disease virus expressing interleukin-2 and TNF-related apoptosis-inducing ligand for cancer therapy.  

PubMed

Recombinant Newcastle disease virus (rNDV) have shown oncolytic therapeutic efficacy in preclinical studies and are currently in clinical trials. In this study, we have evaluated the possibility to enhance the cancer therapeutic potential of NDV by means of inserting both interleukin-2 (IL-2) and tumor necrosis factor-related apoptosis inducing ligand (TRAIL) delivered by rNDV. We demonstrated that rNDV expressing TRAIL (rNDV-TRAIL) or both human IL-2 and TRAIL (rNDV-IL-2-TRAIL) significantly enhanced inherent anti-neoplastic of rNDV by inducing apoptosis. And we showed that apoptosis-related genes mRNA expression was increased after treated with rNDV-TRAIL or rNDV-IL-2-TRAIL compared with rNDV and rNDV-IL-2. We also demonstrated that both rNDV-IL-2 and rNDV-IL-2-TRAIL induced proliferation of the CD4(+) and CD8(+) in treated mice and elicited expression of TNF-? and IFN-? antitumor cytokines. These mice treated with oncolytic agents exhibited significant reduction in tumor development compared with mice treated with the parental virus. In addition, experiments in both hepatocellular carcinoma and melanoma-bearing mice demonstrated that the genetically engineered rNDV-IL-2-TRAIL exhibited prolonged animals' survival compared with rNDV, rNDV-IL-2, and rNDV-TRAIL. In conclusion, the immunotherapy and oncolytic virotherapy properties of NDV can be enhanced by the introduction of IL-2 and TRAIL genes, whose products initiated a broad cascade of immunological affects and induced tumor cells apoptosis in the microenvironment of the immune system. PMID:24971746

Bai, Fu-Liang; Yu, Yin-Hang; Tian, Hui; Ren, Gui-Ping; Wang, Hui; Zhou, Bing; Han, Xiao-Hui; Yu, Qing-Zhong; Li, De-Shan

2014-09-01

40

Anti-GD2 Antibody with GM-CSF, Interleukin-2, and Isotretinoin for Neuroblastoma  

PubMed Central

BACKGROUND Preclinical and preliminary clinical data indicate that ch14.18, a monoclonal antibody against the tumor-associated disialoganglioside GD2, has activity against neuroblastoma and that such activity is enhanced when ch14.18 is combined with granulocyte–macrophage colony-stimulating factor (GM-CSF) or interleukin-2. We conducted a study to determine whether adding ch14.18, GM-CSF, and interleukin-2 to standard isotretinoin therapy after intensive multimodal therapy would improve outcomes in high-risk neuroblastoma. METHODS Patients with high-risk neuroblastoma who had a response to induction therapy and stem-cell transplantation were randomly assigned, in a 1:1 ratio, to receive standard therapy (six cycles of isotretinoin) or immunotherapy (six cycles of isotretinoin and five concomitant cycles of ch14.18 in combination with alternating GM-CSF and interleukin-2). Event-free survival and overall survival were compared between the immunotherapy group and the standard-therapy group, on an intention-to-treat basis. RESULTS A total of 226 eligible patients were randomly assigned to a treatment group. In the immunotherapy group, a total of 52% of patients had pain of grade 3, 4, or 5, and 23% and 25% of patients had capillary leak syndrome and hypersensitivity reactions, respectively. With 61% of the number of expected events observed, the study met the criteria for early stopping owing to efficacy. The median duration of follow-up was 2.1 years. Immunotherapy was superior to standard therapy with regard to rates of event-free survival (66±5% vs. 46±5% at 2 years, P = 0.01) and overall survival (86±4% vs. 75±5% at 2 years, P = 0.02 without adjustment for interim analyses). CONCLUSIONS Immunotherapy with ch14.18, GM-CSF, and interleukin-2 was associated with a significantly improved outcome as compared with standard therapy in patients with high-risk neuroblastoma. PMID:20879881

Yu, Alice L.; Gilman, Andrew L.; Ozkaynak, M. Fevzi; London, Wendy B.; Kreissman, Susan G.; Chen, Helen X.; Smith, Malcolm; Anderson, Barry; Villablanca, Judith G.; Matthay, Katherine K.; Shimada, Hiro; Grupp, Stephan A.; Seeger, Robert; Reynolds, C. Patrick; Buxton, Allen; Reisfeld, Ralph A.; Gillies, Steven D.; Cohn, Susan L.; Maris, John M.; Sondel, Paul M.

2011-01-01

41

Selection of internalization-deficient cells by interleukin-2-Pseudomonas exotoxin chimeric protein: the cytoplasmic domain of the interleukin-2 receptor beta chain does not contribute to internalization of interleukin-2.  

PubMed

To study the structural basis of ligand-induced receptor-mediated internalization of interleukin-2 (IL-2), a strategy has been developed to generate variant T cells that are deficient in internalization of this cytokine. IL-2 receptor (IL-2R) alpha- and beta-bearing EL4 cells, that express high-affinity IL-2R and internalize IL-2, were treated with low doses of IL-2-Pseudomonas exotoxin chimeric protein (IL-2-PE40). This treatment resulted in isolation of a variant (CX1) that was unable to express high-affinity IL-2R or internalize IL-2. Transfection of CX1 with the IL-2R beta cDNA led to surface expression of IL-2R beta and high-affinity IL-2R as well as the ability to internalize IL-2. This finding indicates that the absence of the beta subunit was the sole defect in CX1 responsible for its failure to internalize IL-2. By transfecting CX1 with mutated beta cDNA, several CX1 transfectants were produced that expressed a beta-subunit that lacked all amino acids of the intracytoplasmic region. These transfectants expressed high-affinity IL-2R and internalized IL-2 at a rate comparable to cells expressing wild-type beta-chain. These results demonstrate that internalization of IL-2 is independent of any signals contained in the intracytoplasmic tail of the beta subunit and raise the possibility that such signals may be entirely contained within the gamma subunit. PMID:8258333

Furse, R K; Malek, T R

1993-12-01

42

HLA class I and II, interferon, interleukin 2, and the interleukin 2 receptor expression on labial biopsy specimens from patients with Sjögren's syndrome.  

PubMed Central

Labial biopsy specimens from eight patients with primary Sjögren's syndrome (SS), 10 patients with secondary SS, and three healthy controls were studied with monoclonal antibodies identifying HLA class I and class II antigens; interferon-alpha, beta, and gamma; interleukin 2 (IL2); and the IL2 receptor (Tac) among others. In the normal biopsy specimens there was evidence of HLA class I and, to a lesser extent, class II antigens in both ducts and acini, though this was much less marked than in the Sjögren's biopsy specimens. Interferon-gamma staining, but not interferon-alpha or beta, was also considerably enhanced in the biopsy specimens from the patients with Sjögren's syndrome. These data support the view that in Sjögren's syndrome the release of interferon-gamma may be involved in the induction of class II determinants. Our observations were broadly similar in both primary and secondary Sjögren's syndrome except that patients with primary Sjögren's syndrome tended to have more diffusely scattered T lymphocytes. Images PMID:2444171

Rowe, D; Griffiths, M; Stewart, J; Novick, D; Beverley, P C; Isenberg, D A

1987-01-01

43

Different interleukin 2 receptor beta-chain tyrosines couple to at least two signaling pathways and synergistically mediate interleukin 2-induced proliferation.  

PubMed Central

One of the earliest events induced by interleukin 2 (IL-2) is tyrosine phosphorylation of cellular proteins, including the IL-2 receptor beta chain (IL-2Rbeta). Simultaneous mutation of three tyrosines (Y338, Y392, and Y510) in the IL-2Rbeta cytoplasmic domain abrogated IL-2-induced proliferation, whereas mutation of only Y338 or of Y392 and Y510 inhibited proliferation only partially. While Y392 and Y510 were critical for IL-2-induced activation of signal transducers and activators of transcription (STAT proteins), Y338 was required for Shc-IL-2Rbeta association and for IL-2-induced tyrosine phosphorylation of Shc. Thus, activation of both Jak-STAT and Shc-coupled signaling pathways requires specific IL-2Rbeta tyrosines that together act in concert to mediate maximal proliferation. In COS-7 cells, overexpression of Jak1 augmented phosphorylation of Y338 as well as Y392 and Y510, suggesting that the role for this Jak kinase may extend beyond the Jak-STAT pathway. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8700888

Friedmann, M C; Migone, T S; Russell, S M; Leonard, W J

1996-01-01

44

Pharmacokinetics of recombinant human interleukin-2 in advanced renal cell carcinoma patients following subcutaneous application  

PubMed Central

Aims The aim of the study was to investigate the pharmacokinetics of recombinant human interleukin-2 (rhIL-2) in patients with metastatic renal cell carcinoma following different subcutaneous (s.c.) administration regimens. Methods RhIL-2 was administered subcutaneously to 10 patients according to two different dosing regimens: group A received 20×106 IU m?2 once daily and group B 10×106 IU m?2 twice daily (every 12 h). Additionally, in all patients the influence of soluble interleukin-2 receptor (sIL-2R) on the pharmacokinetics of rhIL-2 was investigated. Results The mean area under the serum concentration-time curve to 24 h (AUC(0,24 h)) was 627 IU ml?1 h in treatment group A and 1130 IU ml?1 h (P=0.029) in treatment group B. In both study groups Cmax and AUC(0,12 h) were not significantly different. Seventy-two hours after the beginning of s.c. rhIL-2 therapy the sIL-2R increased significantly (P=0.016), and sIL-2R levels over 1200 pmol l?1 seemed to reduce the AUC. Conclusions In patients with metastatic renal cell cancer administration of 20×106 IU m?2 of rhIL-2 s.c. in two daily doses (10×106 IU m?2 every 12 h) provides better bioavailability and is preferable to the single dose administration. PMID:9690943

Kirchner, G I; Franzke, A; Buer, J; Beil, W; Probst-Kepper, M; Wittke, F; Övermann, K; Lassmann, S; Hoffmann, R; Kirchner, H; Ganser, A; Atzpodien, J

1998-01-01

45

L-Arginine modulates neonatal lymphocyte proliferation through an interleukin-2 independent pathway.  

PubMed

In cases of arginine depletion, lymphocyte proliferation, cytokine production and CD3? chain expression are all diminished. In addition to myeloid suppressor cells, polymorphonuclear cells (PMN) also exert T-cell immune suppressive effects through arginase-induced l-arginine depletion, especially during pregnancy. In this study, we investigated how arginase/l-arginine modulates neonatal lymphocyte proliferation. Results showed that the neonatal plasma l-arginine level was lower than in adults (48·1 ± 11·3 versus 86·5 ± 14·6 ?m; P = 0·003). Neonatal PMN had a greater abundance of arginase I protein than adult PMN. Both transcriptional regulation and post-transcriptional regulation were responsible for the higher arginase I expression of neonatal PMN. Exogenous l-arginine enhanced neonate lymphocyte proliferation but not that of adult cells. The RNA-binding protein HuR was important but was not the only modulation factor in l-arginine-regulated neonatal T-cell proliferation. l-Arginine-mediated neonatal lymphocyte proliferation could not be blocked by interleukin-2 receptor blocking antibodies. These results suggest that the altered arginase/l-arginine cascade may be one of the mechanisms that contribute to altered neonatal immune responses. Exogenous l-arginine could enhance neonate lymphocyte proliferation through an interleukin-2-independent pathway. PMID:24697328

Yu, Hong-Ren; Kuo, Ho-Chang; Huang, Li-Tung; Chen, Chih-Cheng; Tain, You-Lin; Sheen, Jiunn-Ming; Tiao, Mao-Meng; Huang, Hsin-Chun; Yang, Kuender D; Ou, Chia-Yo; Hsu, Te-Yao

2014-10-01

46

Gene expression for interleukin-2 and tumor necrosis factor-alpha in the spleen of old rats under physiological condition and during septic shock. Possible pharmacological modulation.  

PubMed

Older individuals are more susceptible to infectious agents than younger and this is related to the disrepair of the immune defence mechanisms associated with aging. In this study we evaluated the activity of a new biological response modifier (BRM), pidotimod ((R)-3-[(S)-(5-oxo-2-pyrrolidinyl)carbonyl]-thiazolidine-4-carboxylic acid, PGT/1A, CAS 121808-62-6) in relation to the expression of some cytokine genes. We utilized 24 month-old Sprague-Dawley rats (n = 24), randomly divided into 4 groups: controls (n = 6), pidotimod-treated (n = 6; 200 mg/kg i.p., for 10 days), infected (n = 6; i.p. infection of E. coli CH 198) and pidotimod-treated + infected (n = 6). Poly(A+)RNA purified from the spleens of the animals killed 48 h after the infection was probed with Interleukin-2 (IL-2) and Tumor Necrosis Factor-alpha (TNF-alpha) cDNA clones. Northern blot analysis showed a slight signal of the IL-2 steady state mRNA in the groups of control, pidotimod-treated and infected animals, with an increase (20%) evident only in pidotimod + infected rats, 48 h after E. coli injection. On the contrary, the TNF-alpha mRNA levels were easily detectable in controls and infected rats and lower (20%, 40%) following the drug treatment, independent of i.p. infection. These results account for the BRM activity of pidotimod. PMID:7531977

Annoni, G; Arosio, B; Santambrogio, D; Cullurà, D; Gagliano, N; Uslenghi, C

1994-12-01

47

T-cell subpopulations, expression of interleukin-2 receptor, and production of interleukin-2 and gamma interferon in human American cutaneous leishmaniasis.  

PubMed Central

Leukocyte subpopulations, the expression of the interleukin-2 (IL-2) receptor, and the production of IL-2 and gamma interferon (IFN-gamma) were studied in the peripheral blood mononuclear cells of American cutaneous leishmaniasis patients that had been stimulated in vitro with either leishmanial antigen or mitogen (phytohemagglutinin M). The 75 patients examined were classified as having either the localized (LCL; 66 patients), mucocutaneous (MCL; 5 patients), or the rare diffuse (DCL; 4 patients) form of the disease. Patients with DCL, who are characterized by their defective cell-mediated immune response to leishmanial antigen, failed to express the IL-2 receptor and did not produce IFN-gamma when exposed to the antigen but did so when stimulated by phytohemagglutinin M. Both LCL and MCL patients showed strong proliferative responses to leishmanial antigen; these were by far the greatest in MCL patients. Both groups had significantly increased IL-2 receptor expression and IFN-gamma production after exposure to either antigen or mitogen, and these were highest in the MCL patients. Concerning the leukocyte subpopulations evaluated (CD2, CD4, CD8, CD20, MO2), the most significant findings were a decrease of both CD4+ cells and the CD4/CD8 ratio in MCL patients compared with the other groups. Considering IL-2 production, in response to phytohemagglutinin M both MCL and LCL patients showed amounts of IL-2 comparable to those of the controls. Our results help explain the anergy of T cells from DCL patients to leishmanial antigen, which could lead to a defective production of IFN-gamma and possibly contribute to their incapacity to kill the Leishmania parasite. Concerning MCL patients, the significantly increased expression of IL-2 receptor, decreased expression of the CD4 (helper-inducer of suppression) phenotype, and elevated IFV-gamma production might partially explains the state of hypersensitivity and mucosal damage exhibited by these patients. PMID:3133391

Castes, M; Cabrera, M; Trujillo, D; Convit, J

1988-01-01

48

Discovery of small-molecule interleukin-2 inhibitors from a DNA-encoded chemical library.  

PubMed

Libraries of chemical compounds individually coupled to encoding DNA tags (DNA-encoded chemical libraries) hold promise to facilitate exceptionally efficient ligand discovery. We constructed a high-quality DNA-encoded chemical library comprising 30,000 drug-like compounds; this was screened in 170 different affinity capture experiments. High-throughput sequencing allowed the evaluation of 120?million DNA codes for a systematic analysis of selection strategies and statistically robust identification of binding molecules. Selections performed against the tumor-associated antigen carbonic anhydrase?IX (CA?IX) and the pro-inflammatory cytokine interleukin-2 (IL-2) yielded potent inhibitors with exquisite target specificity. The binding mode of the revealed pharmacophore against IL-2 was confirmed by molecular docking. Our findings suggest that DNA-encoded chemical libraries allow the facile identification of drug-like ligands principally to any protein of choice, including molecules capable of disrupting high-affinity protein-protein interactions. PMID:22588840

Leimbacher, Markus; Zhang, Yixin; Mannocci, Luca; Stravs, Michael; Geppert, Tim; Scheuermann, Jörg; Schneider, Gisbert; Neri, Dario

2012-06-18

49

Local administration trials of interleukin-2 for head and neck cancer.  

PubMed

Recombinant interleukin-2 (rIL-2) has been administered locally in 20 patients with head and neck cancer. Two complete responses in lower lip cancer and 1 partial response in lingual cancer have been obtained. Immunohistological study reveals that tumor infiltrating lymphocytes (TILs), including activated T lymphocytes and natural killer cells, are increased after rIL-2 use. Intraarterial chemotherapy, performed subsequently to the local use of rIL-2 results in a dramatic decrease in tumor size within a short time, and a high frequency of CR cases is observed. Local use of rIL-2 is beneficial for treatment of head and neck cancers, and induction immunochemotherapy combining locally used rIL-2 and arterially infused anticancer drugs plays an important role in a multidisciplinary treatment for these cancers. PMID:1741710

Saito, T; Yoda, J; Tabata, T

1991-01-01

50

Influence of tumour physico-chemical conditions on interleukin-2-stimulated lymphocyte proliferation.  

PubMed Central

The proliferative response of murine lymphocytes to interleukin-2 (IL-2) was examined under physico-chemical conditions present in solid tumours, namely low oxygen and glucose concentrations and acidic pH. Lymphocytes were cultured for four days in 30 U ml-1 IL-2 to simulate serum IL-2 concentrations attainable with high-dose systemic IL-2 therapy. Lymphocyte proliferation was significantly (P < 0.05) reduced by low oxygen concentrations (both anoxia [0% O2] and hypoxia [10%, low glucose (6 mg dl-1), or acidic pH (6.7 or 6.4). Moderate glucose concentration (32 mg dl-1), or neutral pH (7.0) did not impair proliferation. This study indicates that impairment of lymphocyte proliferation by tumour physico-chemical conditions may be a factor in the relatively poor success rate of IL-2/LAK cell immunotherapy. PMID:1419598

Loeffler, D. A.; Juneau, P. L.; Masserant, S.

1992-01-01

51

Interleukin-2 can induce suppression of human natural killer cell cytotoxicity.  

PubMed Central

By interaction with monocytes, interleukin-2 (IL-2) suppressed natural killer (NK) cell cytotoxicity (NKCC) of human, Percoll-fractionated, low-density mononuclear cells. The NK-suppressive effect of IL-2 was independent of de novo formation of prostaglandins or protein since it was unaffected by treatment with indomethacin and cycloheximide, respectively. A monoclonal antibody to the p55 (beta) moiety of the IL-2 receptor (anti-Tac/anti-CD25) blocked IL-2-induced NKCC suppression but did not affect the NK-enhancing effect of the lymphokine. We conclude that IL-2 exerts a monocyte-dependent, IL-2 beta-receptor mediated suppressive influence on human NK cells. PMID:2509116

Hellstrand, K; Hermodsson, S

1989-01-01

52

Interleukin-2 and blood brain barrier in cats: pharmacokinetics and tolerance following intrathecal and intravenous administration.  

PubMed

Single bolus doses of glycosylated human interleukin-2 (n IL-2) in the range of 2.8 x 10(3) to 2.0 x 10(6) IU/kg were administered to anesthesized cats via the cephalic vein (n = 10) or using suboccipital puncture (n = 8). CSF (cerebrospinal fluid) and blood samples were collected by repeated puncture. The n IL-2 concentration in four cats was determined on the basis of its biologic activity using 3H-thymidine incorporation into human ConA-blasts and by radioimmunoassay. In additional experiments radioactivity was determined in cerebrospinal fluid and serum after intravenous and intrathecal (i.th.) application of 5.8 x 10(3) - 3.2 x 10(3) IU/kg of 14C-acetyl-n IL-2 in regular time intervals. CSF and serum concentration time-profiles show a biexponential decline in the plasma elimination phase with half-lives of 4 min (alpha-phase) and 90 min (beta-phase) after intravenous and 20-120 min (alpha-phase) and 2-16 hours (beta-phase) after intrathecal application. There is a trend towards longer terminal elimination half-lives with increasing doses. Interleukin-2 is able to penetrate the blood brain barrier from the circulation into the cerebrospinal fluid and vice versa. Due to a slow rate of penetration and rapid elimination from blood only traces of n IL-2 (2-8 IU/ml) are detected in CSF after i.v. injection of 2 x 10(6) IU/kg, whereas concentrations between 400 and 1600 IU/ml are maintained in CSF for several hours following i.th. administration of 2-10 x 10(5) IU/kg.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1421012

Martin, R; Schwuléra, U; Menke, G; Rudolph, W; Buch, K; Fasold, H; Lissner, R; Thrun, A; Krauseneck, P; Bogdahn, U

1992-01-01

53

Histamine, leukotriene C4 and interleukin-2 increase antibody uptake into a human carcinoma xenograft model.  

PubMed Central

Systemically administered radiolabelled anti-tumour antibody is ineffective in treating the majority of patients with liver metastasis from colorectal carcinoma. We have assessed whether agents which increase capillary permeability can increase tumour uptake of antibody isotope conjugate. We developed a xenograft model of colorectal carcinoma using an antibody directed against carcinoembryonic antigen (CEA). Tumours were grown subcutaneously in the hind limbs of athymic rats to derive their circulation from the femoral artery. Cannulae were placed in the common iliac artery and iliolumbar vein. Antibody was delivered systemically, regionally and regionally with histamine, leukotriene C4 and interleukin-2. Regionally administered anti-CEA antibody resulted in a significantly greater (P = 0.004) tumour to normal tissue ratio (1.66, s.d. = 0.68) compared to systematically administered antibody (1.25, s.d. = 0.73). The addition of vasoactive drugs produced an approximately 3-fold increase with an increase to a mean tumour:liver ratio of 3.24 (s.d. = 1.39) for histamine (P less than 0.001 compared to systemic delivery), 3.21 (s.d. = 1.13, P less than 0.001) for leukotriene C4 and 3.80 (s.d. = 1.53, P less than 0.001) for interleukin-2. The addition of histamine significantly (P = 0.004) increased the mean tumour to liver ratio (1.73, s.d. = 0.44) of non-specific antibody uptake compared with either systemic (1.12, s.d. = 0.24) or regional delivery (1.25, s.d. = 0.54) of non-specific antibody alone. Increasing tumour capillary permeability can produce a significant clinically useful increase in tumour uptake of antibody-isotope conjugate. Images Figure 1 PMID:1931608

Hennigan, T. W.; Begent, R. H.; Allen-Mersh, T. G.

1991-01-01

54

Live Attenuated Influenza Virus Expressing Human Interleukin2 Reveals Increased Immunogenic Potential in Young and Aged Hosts  

Microsoft Academic Search

Despite the reported efficacy of commercially available influenza virus vaccines, a considerable proportion of the human population does not respond well to vaccination. In an attempt to improve the immunogenicity of live influenza vaccines, an attenuated, cold-adapted (ca) influenza A virus expressing human interleukin-2 (IL-2) from the NS gene was generated. Intranasal immunization of young adult and aged mice with

Boris Ferko; Christian Kittel; Julia Romanova; Sabine Sereinig; Hermann Katinger; Andrej Egorov

2006-01-01

55

Simulated microgravity inhibits the genetic expression of interleukin-2 and its receptor in mitogen-activated T lymphocytes  

Microsoft Academic Search

Experiments conducted in space in the last two decades have shown that T lymphocyte activation in vitro is remarkably reduced in microgravity. The data indicate that a failure of the expression of the interleukin-2 receptor (measured as protein secreted in the supernatant) is responsible of the loss of activity. To test such hypothesis we have studied the genetic expression of

Isabelle Walther; Proto Pippia; Maria Antonia Meloni; Franco Turrini; Franca Mannu; Augusto Cogoli

1998-01-01

56

Effects of Interleukin 2 Receptor Blockers on Patient and Graft Survival in Renal-Transplanted Children  

PubMed Central

Background: Monoclonal antibodies block interleukin-2 receptors on alloantigen-reactive T-Lymphocytes and induce selective immunosuppression. It is postulated that induction therapy with these agents in pediatric transplantation may decrease acute rejection and improve graft survival with no significant side effect or increase in the incidence of viral infections. Objectives: The aim of this study was to examine the effects of interleukin 2 receptor blockers on patient and graft survival in renal-transplanted children. Patients and Methods: One hundred and eighty six children aged 7-13 years who received renal transplantation in university-affiliated hospital between 2003 and 2012 were enrolled in the study. All patients received prednisolone, cyclosporine and mycophenolate mofetil or azathioprine as basic immunosuppressive therapy. Patients were divided into two groups according to receiving induction therapy with IL2-receptor blockers. We investigated for acute rejection episodes, Cytomegalovirus (CMV) and BK virus infection and one and three year’s survival of the patients and the grafts Results: From 186 renal-transplanted children included in this study, 36 patients were in treated group (group 1) and 150 patients in control group (group 2). The mean age of the patients was 10.4 ± 2 years and 55.6% were males. In first six months of transplantation, eight patients in group one had one episode of acute rejection and no one had two episodes. Early acute rejection rate was 8.36 (22%). In the control group, 37 patients had one episode and three patients had two episodes of acute rejection (rejection rate 28.6%). Therefore, early acute rejection rates were lower in group one. Late acute rejection rates did not show any difference in group 1 and group 2 (27.7% vs. 27.3% respectively). There was lower prevalence of steroid-resistance rejection in group 1 patients (5.5%) compared with 6.6% in group 2, but it did not reach statistical significance. None of the patients in IL2-R blocker group died at one year follow-up (patient survival 100%). However, in control group, four (2.6%) patients died toward the end of first year (patient survival 97.4%). When patients in group 1 and group 2 were age and sex matched with equal number the difference was significant (P < 0.05). Conclusions: Induction therapy with IL2-R blockers reduced the rate of early acute rejection, but had no effect on late acute rejections. Patient and graft survival were better in treated group, but did not reach statistical significance. A longer period of follow-up may be required to discern a clear advantage for induction therapy with these agents.

Sharifian, Mostafa; Arad, Banafsheh; Simfroosh, Naser; Basiri, Abbas; Otukesh, Hassan; Esfandiar, Nasrin

2014-01-01

57

The protein pheromone Er-1 of the ciliate Euplotes raikovi stimulates human T-cell activity: Involvement of interleukin-2 system  

SciTech Connect

Water-soluble protein signals (pheromones) of the ciliate Euplotes have been supposed to be functional precursors of growth factors and cytokines that regulate cell–cell interaction in multi-cellular eukaryotes. This work provides evidence that native preparations of the Euplotes raikovi pheromone Er-1 (a helical protein of 40 amino acids) specifically increases viability, DNA synthesis, proliferation, and the production of interferon-?, tumor necrosis factor-?, interleukin (IL)-1?, IL-2, and IL-13 in human Jurkat T-cells. Also, Er-1 significantly decreases the mRNA levels of the ? and ? subunits of IL-2 receptor (IL-2R), while the mRNA levels of the ? subunit appeared to be not affected. Jurkat T-cell treatments with Er-1 induced the down-regulation of the IL-2R? subunit by a reversible and time-dependent endocytosis, and increased the levels of phosphorylation of the extracellular signal-regulated kinases (ERK). The cell-type specificity of these effects was supported by the finding that Er-1, although unable to directly influence the growth of human glioma U-373 cells, induced Jurkat cells to synthesize and release factors that, in turn, inhibited the U-373 cell proliferation. Overall, these findings imply that Er-1 coupling to IL-2R and ERK immuno-enhances T-cell activity, and that this effect likely translates to an inhibition of glioma cell growth. -- Highlights: ? Euplotes pheromone Er-1 increases the growth of human Jurkat T-cells. ? Er-1 increases the T-cell production of specific cytokines. ? Er-1 activates interleukin-2 receptor and extracellular signal-regulated kinases. ? The immuno-enhancing effect of Er-1 on Jurkat cells translates to an inhibition of human glioma cell growth.

Cervia, Davide, E-mail: d.cervia@unitus.it [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy) [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy); Department of Biomedical and Clinical Sciences, “Luigi Sacco” University Hospital, University of Milan, Milano (Italy); Catalani, Elisabetta; Belardinelli, Maria Cristina [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy)] [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy); Perrotta, Cristiana [Department of Biomedical and Clinical Sciences, “Luigi Sacco” University Hospital, University of Milan, Milano (Italy)] [Department of Biomedical and Clinical Sciences, “Luigi Sacco” University Hospital, University of Milan, Milano (Italy); Picchietti, Simona [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy)] [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy); Alimenti, Claudio [Department of Environmental and Natural Sciences, University of Camerino, Camerino (Italy)] [Department of Environmental and Natural Sciences, University of Camerino, Camerino (Italy); Casini, Giovanni; Fausto, Anna Maria [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy)] [Department for Innovation in Biological, Agro-food and Forest systems (DIBAF), University of Tuscia, Viterbo (Italy); Vallesi, Adriana [Department of Environmental and Natural Sciences, University of Camerino, Camerino (Italy)] [Department of Environmental and Natural Sciences, University of Camerino, Camerino (Italy)

2013-02-01

58

Only high-affinity receptors for interleukin 2 mediate internalization of ligand  

SciTech Connect

Interleukin 2 (IL-2) receptors are expressed on activated T cells and in select T-cell leukemias. Recently, it has been demonstrated that at least two classes of receptor for IL-2 exist with markedly different affinities for ligand. All known biological actions of IL-2 have been correlated with occupancy of high-affinity sites; the function of the low-affinity sites remains unknown. Receptor-mediated endocytosis is the primary means of internalization of cell-surface receptors and their ligands. The internalization of IL-2 bound to high- and low-affinity receptor sites was studied in a human T-cell lymphotrophic virus type 1 (HTLV-1)-infected human T-cell leukemia cell line and in a cloned murine cytotoxic T-cell line (CTLL). Internalization of IL-2 occurred only when bound to high-affinity sites. In addition, an anti-receptor antibody (anti-Tac), which binds equally well to high- and low-affinity sites, demonstrated no detectable internalization. The implications of these findings as they relate to IL-2 receptor structure and function are discussed.

Weissman, A.M.; Harford, J.B.; Svetlik, P.B.; Leonard, W.L.; Depper, J.M.; Waldmann, T.A.; Greene, W.C.; Klausner, R.D.

1986-03-01

59

Activity of outpatient intravenous interleukin-2 and famotidine in metastatic clear cell kidney cancer.  

PubMed

Outpatient daily intravenous infusions of interleukin-2 (IL-2) have been developed to maintain anticancer activity and decrease toxicity of this agent against kidney cancer. Lymphokine activated killer cell (LAK) numbers are increased with these IL-2 schedules. Famotidine may enhance the LAK activity by increasing IL-2 internalization by the IL-2 receptor on lymphocytes. Fifteen patients with metastatic clear cell kidney cancer received IL-2 18 million IU/M² intravenously over 15-30 minutes preceded by famotidine 20 mg IV daily for 3 days for 6 consecutive weeks as outpatients. Cycles were repeated every 8 weeks. Patient characteristics were seven males/eight females, median age 59 (range: 28-70), median Eastern Cooperative Oncology Group (ECOG) performance status-1; common metastatic sites were lungs (14), lymph nodes (9), liver (4), bone (4), and pancreas (4). Prior systemic therapies were oral tyrosine kinase inhibitor (8), IL-2 (6), and mTor inhibitor (2). Most common toxicities were rigors, arthralgia/myalgia, nausea/emesis, fever, and hypotension. All episodes of hypotension were reversible with intravenous fluid. No patients required hospitalization due to toxicity. One complete response (7%) and four partial responses (26%) were seen (total response rate=33%; 95% confidence interval: 15%-59%). Responses occurred in the lungs, liver, lymph nodes, and bone. Outpatient intravenous IL-2 with famotidine has activity in metastatic clear cell kidney cancer. PMID:24251758

Quan, Walter D Y; Quan, Francine Marie

2014-03-01

60

Chromatin remodeling of the interleukin-2 gene: distinct alterations in the proximal versus distal enhancer regions.  

PubMed Central

Known transcription factor-DNA interactions in the minimal enhancer of the murine interleukin-2 gene (IL-2) do not easily explain the T cell specificity of IL-2 regulation. To seek additional determinants of cell type specificity, in vivo methodologies were employed to examine chromatin structure 5' and 3' of the 300 bp IL-2 proximal promoter/enhancer region. Restriction enzyme accessibility revealed that until stimulation the IL-2 proximal promoter/enhancer exists in a closed conformation in resting T and non-T cells alike. Within this promoter region, DMS and DNase I genomic footprinting also showed no tissue-specific differences prior to stimulation. However, DNase I footprinting of the distal -600 to -300 bp region revealed multiple tissue-specific and stimulation-independent DNase I hypersensitive sites. Gel shift assays detected T cell-specific complexes binding within this region, which include TCF/LEF or HMG family and probable Oct family components. Upon stimulation, new DNase I hypersensitive sites appeared in both the proximal and distal enhancer regions, implying that there may be a functional interaction between these two domains. These studies indicate that a region outside the established IL-2 minimal enhancer may serve as a stable nucleation site for tissue-specific factors and as a potential initiation site for activation-dependent chromatin remodeling. PMID:9611237

Ward, S B; Hernandez-Hoyos, G; Chen, F; Waterman, M; Reeves, R; Rothenberg, E V

1998-01-01

61

Cytomegalovirus immediate early genes prevent the inhibitory effect of cyclosporin A on interleukin 2 gene transcription.  

PubMed Central

The use of cyclosporin A (CsA) as an immunosuppressive agent has markedly improved the clinical outcome in solid organ transplantation. However, posttransplantation infection remains a significant problem and may contribute to subsequent organ rejection. In this study the effect of cytomegalovirus (CMV) immediate early (IE) gene products on interleukin 2 (IL-2) gene transcription in the absence and presence of CsA was investigated using a transient transfection system. Jurkat T cells were transfected with plasmids expressing the CMV IE gene products or with a control plasmid. The presence of the CMV IE2 gene product abolished the inhibitory effect of CsA on IL-2 promoter activation and gene transcription. This effect was noted regardless of the time of CsA addition relative to the time of stimulation and was independent of CsA concentration. CsA had no effect on the CMV or the IL-2 receptor promoters. These studies suggest that the CMV IE gene products may play a role in graft rejection after solid organ transplantation. Images PMID:1331182

Geist, L J; Monick, M M; Stinski, M F; Hunninghake, G W

1992-01-01

62

[A case of retroperitoneal angiosarcoma effectively treated with recombinant interleukin-2].  

PubMed

Angiosarcoma is rare and highly malignant vascular neoplasm, and primary retroperitoneal angiosarcoma is extremely rare. Preoperative diagnosis is very difficult because there are no specific imaging features, and definitively effective treatment has not yet been established. We recently treated a patient with primary retroperitoneal angiosarcoma in which a prompt and exact diagnosis was difficult to obtain. One month after surgery, local recurrence appeared, but salvage immunotherapy using recombinant interleukin-2 (rIL-2) showed good efficacy, and the patient obtained complete response. Here we report this rare case of angiosarcoma. A 60-year-old woman with abdominal pain was diagnosed with a left retroperitoneal mass on CT scan. The tumor was about 9 cm in diameter and positioned above the left kidney. Further study using MRI, 131I-MIBG scintigraphy, and enhanced CT suggested chronic expanding hematoma and the patient underwent surgical resection. Histopathological diagnosis was primary retroperitoneal angiosarcoma based on positive staining for VIII factor, CD31, CD34, and p53. One month after surgery, FDG-PET revealed local recurrence adjacent to the psoas major. We initiated salvage immunotherapy using rIL-2. The patient was treated effectively and achieved complete response. She is alive and well 19 months after surgery and rIL-2 treatment. PMID:24261193

Noguchi, Go; Ota, Junichi; Ishigaki, Hanako; Onuki, Tatsuaki; Kato, Yoshitake; Moriyama, Masatoshi

2012-11-01

63

Loss of Neuronal Phenotype and Neurodegeneration: Effects of T Lymphocytes and Brain Interleukin-2  

PubMed Central

Loss of neuronal phenotype and reversal of neuronal atrophy have been demonstrated in different models of central nervous system (CNS) injury. These processes may be generalizable to different types of brain neurons and circuitry. The idea that some injured neurons may lose their phenotype and/or atrophy with the potential to rejuvenate is a remarkable and potentially promising form of neuronal plasticity that is not well understood. In this paper, we present some of our laboratory’s basic neuroimmunology research showing that peripheral T cells entering the CNS, and brain-derived interleukin-2 (IL-2), play significant roles in these intriguing processes. Our findings suggest, for example, that T cell immunosenesence could be involved in related processes of brain aging and contribute to neurodegenerative disease. Neuroimmunological approaches may provide new insights into yet undiscovered factors and brain mechanisms that regulate changes in neuronal integrity associated with aging and disease. Such findings could have important implications for discovering more effective strategies for treating patients with neurotrauma and neurodegenerative diseases (e.g., Alzheimer’s disease). PMID:24058743

Meola, Danielle; Huang, Zhi; Ha, Grace K; Petitto, John M

2013-01-01

64

Loss of Neuronal Phenotype and Neurodegeneration: Effects of T Lymphocytes and Brain Interleukin-2.  

PubMed

Loss of neuronal phenotype and reversal of neuronal atrophy have been demonstrated in different models of central nervous system (CNS) injury. These processes may be generalizable to different types of brain neurons and circuitry. The idea that some injured neurons may lose their phenotype and/or atrophy with the potential to rejuvenate is a remarkable and potentially promising form of neuronal plasticity that is not well understood. In this paper, we present some of our laboratory's basic neuroimmunology research showing that peripheral T cells entering the CNS, and brain-derived interleukin-2 (IL-2), play significant roles in these intriguing processes. Our findings suggest, for example, that T cell immunosenesence could be involved in related processes of brain aging and contribute to neurodegenerative disease. Neuroimmunological approaches may provide new insights into yet undiscovered factors and brain mechanisms that regulate changes in neuronal integrity associated with aging and disease. Such findings could have important implications for discovering more effective strategies for treating patients with neurotrauma and neurodegenerative diseases (e.g., Alzheimer's disease). PMID:24058743

Meola, Danielle; Huang, Zhi; Ha, Grace K; Petitto, John M

2013-06-01

65

Induction of soluble tumour necrosis factor receptors during treatment with interleukin-2.  

PubMed Central

Interleukin-2 (IL-2) treatment induces other cytokines such as tumour necrosis factor (TNF) TNF may mediate some of the anti-tumour activity of IL-2, but conversely, may contribute to its dose limiting toxicities. Cleaved extracellular domains of the p55 and the p75 TNF receptors (sTNF-R1 and R2) bind to and inhibit the biological activity of TNF in vitro, but may also act as carrier molecules. We have assayed TNF and sTNFR-1 and 2 in the plasma of advanced cancer patients, before and during treatment with IL-2. Plasma levels of TNF in 22 patients were not significantly different from 25 normal controls, but levels of sTNFR-1 and sTNFR-2 were higher (P < 0.001). Levels of TNF and both its soluble receptors were significantly increased in 13 patients receiving IL-2 therapy. Maximum induced levels of sTNFR-1 and sTNFR-2 correlated closely with maximum induced levels of TNF (P < 0.001), but peak levels of sTNFR-1 and two were achieved 24-48 h after peak TNF. Levels of TNF and sTNF-Rs did not correlate with toxicity. Treatment with IL-2 leads not only to induction of TNF but also soluble binding proteins at levels which may modulate its biological activity. PMID:1333789

Miles, D. W.; Aderka, D.; Engelmann, H.; Wallach, D.; Balkwill, F. R.

1992-01-01

66

Platelet activating factor mediates interleukin-2-induced lung injury in the rat.  

PubMed Central

Interleukin-2 was recently shown to cause acute lung injury characterized by microvascular permeability defect, interstitial edema, and leukosequestration. Similar responses can also be produced by platelet activating factor (PAF). Thus, the present study aimed to examine whether PAF plays a key role in the development of IL-2-induced lung injury in the anesthetized rat. Intravenous infusion (60 min) of recombinant human IL-2 at 10(5)-10(6) U/rat (n = 7-9) dose-dependently elevated lung water content (27 +/- 1%, P less than 0.01), myeloperoxidase activity (+84 +/- 23%, P less than 0.05), and serum thromboxane B2 (990 +/- 70%, P less than 0.01), but failed to alter blood pressure, hematocrit, serum tumor necrosis factor-alpha, and circulating leukocytes and platelets. Pretreatment (-30 min) with a potent and specific PAF antagonist, BN 50739 (10 mg/kg, intraperitoneally, n = 6) prevented the pulmonary edema (P less than 0.05) and thromboxane B2 production (P less than 0.01), and attenuated the elevation of lung myeloperoxidase activity (+18 +/- 16%, P less than 0.05) induced by IL-2. These data suggest that PAF is involved in the pathophysiological processes leading to IL-2-induced lung injury, and point to the potential therapeutic capacity of PAF antagonists in preventing pulmonary edema during IL-2 therapy. PMID:1314853

Rabinovici, R; Sofronski, M D; Renz, J F; Hillegas, L M; Esser, K M; Vernick, J; Feuerstein, G

1992-01-01

67

Clinical and immunological effects of single bolus administration of recombinant interleukin-2 in cattle.  

PubMed Central

Recombinant bovine interleukin-2 (rBoIL-2) was administered as a single intramuscular bolus to healthy calves to determine the minimal dose capable of exerting a biological response. Doses ranging from 2.5 to 0.05 micrograms rBoIL-2/kg did not induce pyrexia, diarrhea, or depression, nor did they alter any blood chemistry or hematological parameters commonly associated with IL-2 toxicity. Moreover, the only significant immunological change observed was a reduction in the number of peripheral blood lymphocytes identified with the monoclonal antibodies B7A, BAQ4A (WC1+ cells), CACTB6A (WC2+ cells) and DH59B (monocytes). The decrease in cells associated with these markers did not influence non-MHC restricted cytotoxicity or in vitro lymphocyte proliferative responses to mitogens and IL-2. The treatments had no effect on delayed type hypersensitivity responses to phytohemagglutinin. These results indicate that IL-2 may be involved in the regulation of trafficking patterns of a unique subpopulation of lymphocytes in cattle. PMID:1586889

Campos, M; Hughes, H P; Godson, D L; Sordillo, L M; Rossi-Campos, A; Babiuk, L A

1992-01-01

68

Pentoxifylline inhibits interleukin-2-induced toxicity in C57BL/6 mice but preserves antitumor efficacy.  

PubMed Central

Interleukin 2 (IL-2) mediates the regression of metastatic cancer but clinical use has been limited due to associated toxicities. Tumor necrosis factor (TNF) is an important mediator of IL-2 toxicity and may have a limited role in IL-2 antitumor efficacy. Because pentoxifylline (PTXF) inhibits TNF production, we hypothesized that PTXF would ameliorate IL-2 toxicity without compromising antitumor efficacy. Four groups of female C57BL/6 mice with pulmonary metastases from a 3-methylcholanthrene-induced fibrosarcoma (MCA-105) and four groups of nontumored mice were treated every 6 h for 4 d by intraperitoneal injections of either IL-2 alone, IL-2 and PTXF, PTXF alone, or equal volumes of saline. Upon completion of therapy, we found that PTXF suppressed many of the IL-2-induced effects including TNF production, lymphocytic infiltration of multiple organs, multiple organ edema, hepatic dysfunction, leukopenia, and thrombocytopenia. Tumor response was determined 21 d after cessation of therapy by quantitating the number and surface area of pulmonary metastases. PTXF preserved antitumor efficacy while reducing the morbidity and mortality caused by IL-2 treatment. These data strongly support the use of PTXF in extending the therapeutic index of IL-2 in the treatment of cancer. Images PMID:1644928

Edwards, M J; Heniford, B T; Klar, E A; Doak, K W; Miller, F N

1992-01-01

69

High-dose interleukin 2 promotes bacterial translocation from the gut.  

PubMed Central

Toxicity associated with high-dose recombinant interleukin 2 (rIL-2) therapy simulates a sepsis syndrome, but the mechanism remains unclear. We hypothesised that translocated gut-origin bacteria may be important. Fifty-one male rats were randomised to receive rIL-2 by intraperitoneal injection at doses (IU) of 10(5) (n = 15), 10(4) (n = 8), 10(3) (n = 8) or 10(2) (n = 8) twice daily, or a saline bolus (n = 12). After 5 days, ileal histomorphology was assessed and the mesenteric lymph node complex cultured. Results showed that colonisation of mesenteric lymph nodes with Escherichia coli occurred in all rats treated with 10(5) IU of rIL-2, and in 62%, 37% and 12% of rats treated with decreasing doses of rIL-2. No translocation was observed in control animals. An increase in submucosal lymphatics and occasional mucosal disruption was seen only in the group receiving 10(5) IU. These data show that rIL-2 promotes bacterial translocation and suggests a mechanism that may fuel high-dose rIL-2 toxicity in man. Images Figure 1 PMID:7669573

Reynolds, J. V.; Murchan, P.; Leonard, N.; Gough, D. B.; Clarke, P.; Keane, F. B.; Tanner, W. A.

1995-01-01

70

Definition and spatial location of mouse interleukin-2 residues that interact with its heterotrimeric receptor.  

PubMed Central

The high affinity receptor for interleukin-2 (IL-2) contains three subunits called IL-2R alpha, beta and gamma. A biological and receptor binding analysis based on 1393 different mutant mouse IL-2 (mIL-2) proteins was used to define the function of each of the 149 residues. By this genetic analysis, 44 residues were assigned important functions, 21 of which were structural. The remaining 23 residues consisted of 19 residues, from three separate regions, that were important for IL-2R alpha interaction; three residues, from two separate regions, that were important for IL-2R beta interaction; and a single residue important for IL-2R gamma interaction. We built a model mIL-2 structure based on the homologous human IL-2 (hIL-2) crystal structure. The roles of the 21 residues presumed to be important for structure were consistent with the model. Despite discontinuity in the primary sequence, the residues specific for each IL-2R subunit interaction were clustered and located to three disparate regions of the tertiary mIL-2 structure. The relative spatial locations of these three surfaces are different from the two receptor binding sites known for the structurally related human growth hormone and the significance of this observation is discussed. Images PMID:8262055

Zurawski, S M; Vega, F; Doyle, E L; Huyghe, B; Flaherty, K; McKay, D B; Zurawski, G

1993-01-01

71

Interleukin-2 can prevent and reverse antigen-induced unresponsiveness in cloned human T lymphocytes.  

PubMed Central

The exposure of human T-cell clones to supra-immunogenic concentrations of peptide antigen in the absence of accessory cells induces antigen-specific unresponsiveness. Using this model we have investigated the ability of cytokines to modulate the induction of, or reversal of, T-cell tolerance. Our findings demonstrate that interleukin-2 (IL-2), but not interferon-gamma (IFN-gamma) or interleukin-1 (IL-1), is able to inhibit the induction of T-cell unresponsiveness in a dose-dependent fashion. Moreover, IL-2 was able to reverse established antigen-dependent T-cell unresponsiveness. In order to determine if modulation of IL-2 receptors is able to induce or abrogate unresponsiveness, the T cells were treated with anti-Tac antibody alone or together with tolerizing concentrations of antigen. Anti-Tac antibody was neither able to induce nor inhibit the induction of tolerance. The application of this model in the manipulation of immune responses is discussed here. PMID:2457547

Essery, G; Feldmann, M; Lamb, J R

1988-01-01

72

Effect of interleukin 2 on killer cell inhibitory receptors in patients with rheumatoid arthritis  

PubMed Central

OBJECTIVE—The genes for killer cell inhibitory receptors (KIRs) have been cloned and their functions and responses to other molecules, including cytokines, have been partially clarified. However, the expression of KIRs has not been analysed in patients with autoimmune diseases, such as rheumatoid arthritis (RA), who are highly susceptible to microbial infection. Therefore, KIR expression on lymphocytes in patients with RA, and the regulation of KIR expression by interleukin 2 (IL2) in RA was investigated.?METHODS—CD158a/b expression on peripheral blood mononuclear cells (PBMC) obtained from 25 patients with RA and 14 healthy subjects was analysed by flow cytometry. Additionally, PBMC from the two groups of subjects were cultured in RPMI 1640 medium with or without IL2 for 48 hours, and then their CD158a/b expression was analysed.?RESULTS—The rate of CD158a expression on the CD8+ cells was lower in patients with RA than in healthy subjects, though there was no significant difference in the CD158a/b expression on the CD16+ cells between the two groups. The upregulation of CD16+CD158a/b+ cells in response to IL2 was significantly reduced in patients with RA compared with healthy subjects.?CONCLUSION—The reduced induction of KIR expression in response to IL2 may provide insight into the reason for the high susceptibility of patients with RA to microbial infection.?? PMID:11156551

Kogure, T; Niizawa, A; Hai, L; Fujinaga, H; Shimada, Y; Ochiai, H; Terasawa, K

2001-01-01

73

Inhibiting Autophagy During Interleukin 2 (IL-2) Immunotherapy Promotes Long Term Tumor Regression  

PubMed Central

Administration of high dose interleukin 2 (HDIL-2) has durable antitumor effects in 5-10% patients with melanoma and renal cell carcinoma. However, treatment is often limited by side effects, including reversible, multi-organ dysfunction and characterized by a cytokine-induced ‘systemic autophagic syndrome’. Here we hypothesized that the autophagy inhibitor chloroquine (CQ) would enhance IL-2 immunotherapeutic efficacy and limit toxicity. In an advanced murine metastatic liver tumor model, IL-2 inhibited tumor growth in a dose-dependent fashion, and these anti-tumor effects were significantly enhanced upon addition of CQ. The combination of IL-2 with CQ increased long term survival, decreased toxicity associated with vascular leakage, and enhanced immune cell proliferation and infiltration in the liver and spleen. HDIL-2 alone increased serum levels of IFN-?, IL-6 and IL-18 and also induced autophagy within the liver and translocation of HMGB1 from the nucleus to the cytosol in hepatocytes, effects that were inhibited by combined administration with CQ. In tumor cells, CQ increased autophagic vacuoles and LC3-II levels inhibited oxidative phosphorylation and ATP production and promoted apoptosis, which was associated with increased Annexin V+/PI- cells, cleaved-PARP, cleaved-caspase 3, and cytochrome C release from mitochondria. Taken together, our findings provide a novel clinical strategy to enhance the efficacy of HDIL-2 immunotherapy for cancer patients. PMID:22472122

Liang, Xiaoyan; De Vera, Michael E.; Buchser, William J.; Romo de Vivar Chavez, Antonio; Loughran, Patricia; Stolz, Donna Beer; Basse, Per; Wang, Tao; Van Houten, Bennett; Zeh, Herbert J.; Lotze, Michael T.

2012-01-01

74

Enhancement of a delayed hypersensitivity reaction to a contact allergen, by the systemic administration of interleukin-2.  

PubMed Central

The immunopharmacological effects of interleukin-2 (IL-2) on the sensitization and effector phases of the delayed-type hypersensitivity (DTH) reaction were studied using contact sensitivity to the haptenizing agent dinitrochlorobenzene (DNCB). When administered at the time of priming to DNCB, IL-2 had no effect on the subsequent magnitude of the response. Interleukin-2 was, however, able to increase the magnitude of the response when given at the time of secondary challenge; the degree of change was directly related to the dose of IL-2. The proportions of T cells in the draining lymph node and spleen of IL-2-treated animals decreased by approximately one-third, but there was no alteration to the balance between CD4+ and CD8+ T cells. The results suggest that the increase in DTH observed was due to a pharmacological effect rather than to an increase in T-cell number. PMID:2037319

Zaloom, Y; Walsh, L P; McCulloch, P; Gallagher, G

1991-01-01

75

Production of tumor necrosis factor ? and interferon ? in interleukin-2-treated melanoma patients: Correlation with clinical toxicity  

Microsoft Academic Search

Summary Interleukin-2 (IL-2)-based immunotherapy regimens are accompanied by dose-limiting toxicity consisting of fever, tachycardia, chills and capillary leak syndrome. We hypothesized that the toxicity was caused by the induction and release of endogenous cytokines such as tumor necrosis factor a (TNFa) and interferon ? (IFN?). We measured the serum levels of TNFa and IFN? in IL-2-treated melanoma patients and attempted

James S. Economou; Mary Hoban; Jeffrey D. Lee; Richard Essner; Stephen Swisher; William McBride; Dave B. Hoon; Donald L. Mortonl

1991-01-01

76

Clonal analysis of peripheral blood lymphocytes from three patients with advanced neuroblastoma receiving recombinant interleukin-2 and interferon ?  

Microsoft Academic Search

In this study we have investigated, at the population and the clonal levels, the immunophenotypes and the non-specific cytotoxic functions of peripheral blood lymphocytes from three stage IV neuroblastoma patients receiving treatment with recombinant interleukin-2 (IL-2) and interferon a (IFNa). Both IL-2 alone and the combination of IL-2 and IFNa caused an in vivo expansion of CD56+, CD3- NK cells

Ignazia Prigione; Paola Facchettil; Edoardo Lanino; Alberto Garaventa; Vito Pistoial

1993-01-01

77

Leukocyte orchestration in blood and tumour tissue following interleukin-2 based immunotherapy in metastatic renal cell carcinoma  

Microsoft Academic Search

With the objective of evaluating leukocyte orchestration in situ, serial blood samples and tumour tissue core needle biopsies were obtained at baseline and repeated after 1 month of therapy, among 49 consecutive single-institution patients with metastatic renal cell carcinoma (mRCC). Patients were treated with outpatient low-dose subcutaneous interleukin 2 (IL-2) and interferon a (IFN-a) alone ( n=23) or in combination with

Frede Donskov; Karen Marie Bennedsgaard; Marianne Hokland; Niels Marcussen; Rune Fisker; Hans Henrik Torp Madsen; Kirsten Fode; Hans von der Maase

2004-01-01

78

Low-Dose Interleukin-2 Therapy: A Driver of an Imbalance between Immune Tolerance and Autoimmunity  

PubMed Central

For many years, the role of interleukin-2 (IL-2) in autoimmune responses was established as a cytokine possessing strong pro-inflammatory activity. Studies of the past few years have changed our knowledge on IL-2 in autoimmune chronic inflammation, suggesting its protective role, when administered at low-doses. The disrupted balance between regulatory and effector T cells (Tregs and Teffs, respectively) is a characteristic of autoimmune diseases, and is dependent on homeostatic cytokines, including IL-2. Actually, inherent defects in the IL-2 signaling pathway and/or levels leading to Treg compromised function and numbers as well as Th17 expansion have been attributed to autoimmune disorders. In this review, we discuss the role of IL-2 in the pathogenesis of autoimmune diseases. In particular, we highlight the impact of the dysregulated IL-2 pathway on disruption of the Treg/Th17 balance, reversal of which appears to be a possible mechanism of the low-dose IL-2 treatment. The negative effects of IL-2 on the differentiation of follicular helper T cells (Tfh) and pathogenic Th17 cells, both of which contribute to autoimmunity, is emphasized in the paper as well. We also compare the current IL-2-based therapies of animal and human subjects with immune-mediated diseases aimed at boosting the Treg population, which is the most IL-2-dependent cell subset desirable for sufficient control of autoimmunity. New perspectives of therapeutic approaches focused on selective delivery of IL-2 to inflamed tissues, thus allowing local activity of IL-2 to be combined with its reduced systemic and pleiotropic toxicity, are also proposed in this paper. PMID:25322151

Kosmaczewska, Agata

2014-01-01

79

Fine epitope specificity of antibodies against interleukin-2 explains their paradoxical immunomodulatory effects  

PubMed Central

The functional dichotomy of antibodies against interleukin-2 (IL-2) is thought to depend upon recognition of different cytokine epitopes. Beyond functional studies, the only molecular evidence obtained so far located the epitopes recognized by the immunoenhancing antibodies S4B6 and JES6–5H4 within the predicted interface of IL-2 with the ? receptor subunit, explaining the preferential stimulation of effector cells displaying only ? and ? receptor chains. A consistent functional map of the epitope bound by the immunoregulatory antibody JES6-1A12 has now been delineated by screening the interactions of phage-displayed antigen variants (with single and multiple mutations) and antigen mimotopes. The target determinant resides in a region between the predicted interfaces with ? and ?/? receptor subunits, supporting the dual inhibitory role of the antibody on both interactions. Binding by JES6-1A12 would thus convert complexed IL-2 into a very weak agonist, reinforcing the advantage of T regulatory cells (displaying the high affinity ??? heterotrimeric receptor) to capture the cytokine by competition and expand over effector cells, ultimately resulting in the observed strong tolerogenic effect of this antibody. Detailed knowledge of the epitopes recognized by anti-IL-2 antibodies with either immunoenhancing or immunoregulatory properties completes the molecular scenario underlying their use to boost or inhibit immune responses in multiple experimental systems. The expanded functional mapping platform now available could be exploited to study other interactions involving related molecular pairs with the final goal of optimizing cytokine and anti-cytokine therapies. PMID:24253188

Rojas, Gertrudis; Cabrera Infante, Yanelys; Pupo, Amaury; Carmenate, Tania

2014-01-01

80

Systemic Administration of Interleukin 2 Enhances the Therapeutic Efficacy of Dendritic Cell-Based Tumor Vaccines  

NASA Astrophysics Data System (ADS)

We have reported previously that murine bone marrow-derived dendritic cells (DC) pulsed with whole tumor lysates can mediate potent antitumor immune responses both in vitro and in vivo. Because successful therapy was dependent on host immune T cells, we have now evaluated whether the systemic administration of the T cell stimulatory/growth promoting cytokine interleukin-2 (IL-2) could enhance tumor lysate-pulsed DC-based immunizations to further promote protective immunity toward, and therapeutic rejection of, syngeneic murine tumors. In three separate approaches using a weakly immunogenic sarcoma (MCA-207), the systemic administration of non-toxic doses of recombinant IL-2 (20,000 and 40,000 IU/dose) was capable of mediating significant increases in the potency of DC-based immunizations. IL-2 could augment the efficacy of tumor lysate-pulsed DC to induce protective immunity to lethal tumor challenge as well as enhance splenic cytotoxic T lymphocyte activity and interferon-? production in these treated mice. Moreover, treatment with the combination of tumor lysate-pulsed DC and IL-2 could also mediate regressions of established pulmonary 3-day micrometastases and 7-day macrometastases as well as established 14- and 28-day s.c. tumors, leading to either significant cure rates or prolongation in overall survival. Collectively, these findings show that nontoxic doses of recombinant IL-2 can potentiate the antitumor effects of tumor lysate-pulsed DC in vivo and provide preclinical rationale for the use of IL-2 in DC-based vaccine strategies in patients with advanced cancer.

Shimizu, K.; Fields, R. C.; Giedlin, M.; Mule, J. J.

1999-03-01

81

Administration in vivo of recombinant interleukin 2 protects mice against septic death.  

PubMed Central

Administration in vivo of recombinant interleukin 2 (rIL-2) to mice induces a polyclonal IgM response. When co-administered with a specific antigen, rIL-2 can enhance concentrations of murine IgM antibodies specific for the antigen by fivefold within 7 d of initial treatment. IgM antibodies that are induced after injection of rIL-2 include antibodies specific for J5, a cell wall core lipopolysaccharide (LPS) antigen that is shared by the different members of the Enterobactericeae family. We report here that mice pretreated with rIL-2 or immunized with J5 antigen 7 d before bacterial challenge were protected from septic death that is caused by intraperitoneal challenges with Escherichia coli. Optimal protection was provided by a combined J5 antigen and rIL-2 treatment. Acquisition of the rIL-2 and J5 antigen-induced protection against lethal bacterial infection coincided temporally with maximal serum IgM titers that also contained IgM antibodies specific for the J5 antigen. In passive immunization experiments, the affinity-purified IgM fraction in sera of rIL-2-treated animals was identified as necessary and sufficient for protection. The IgM-depleted serum had no protective effect. The nonspecific augmentation of host-defense mechanisms without the induction of endotoxin manifestations makes rIL-2 a potential candidate to any alternative LPS-containing vaccines for the prevention of bacterial infections by gram-negative organisms since the core LPS antigen is shared among gram-negative bacteria. PMID:3294901

Weyand, C; Goronzy, J; Fathman, C G; O'Hanley, P

1987-01-01

82

Cyclotides Suppress Human T-Lymphocyte Proliferation by an Interleukin 2-Dependent Mechanism  

PubMed Central

Cyclotides are a diverse and abundant group of ribosomally synthesized plant peptides containing a unique cyclic cystine-knotted topology that confers them with remarkable stability. Kalata B1, a representative member of this family of mini-proteins, has been found to inhibit the proliferation of human peripheral blood mononuclear cells. Analysis of T-cell proliferation upon treatment with chemically synthesized kalata B1 mutants revealed a region comprising inter-cysteine loops 1 and 2 of the cyclotide framework to be important for biological activity. Cytokine signaling analysis using an ‘active’ kalata B1 mutant [T20K], and the reference drug cyclosporin A (CsA) demonstrated that treatment of activated T-lymphocytes with these compounds decreased the expression of the interleukin-2 (IL-2) surface receptor as well as IL-2 cytokine secretion and IL-2 gene expression, whereas the ‘inactive’ kalata B1 mutant [V10K] did not cause any effects. The anti-proliferative activity of [T20K] kalata B1 was antagonized by addition of exogenous IL-2. Furthermore, treatment with [T20K] kalata B1 led to an initial reduction of the effector function, as indicated by the reduced IFN-? and TNF-? production, but the levels of both cytokines stabilized over time and returned to their normal levels. On the other hand, the degranulation activity remained reduced. This indicated that cyclotides interfere with T-cell polyfunctionality and arrest the proliferation of immune-competent cells through inhibiting IL-2 biology at more than one site. The results open new avenues to utilize native and synthetically-optimized cyclotides for applications in immune-related disorders and as immunosuppressant peptides. PMID:23840803

Gründemann, Carsten; Thell, Kathrin; Lengen, Karin; Garcia-Käufer, Manuel; Huang, Yen-Hua; Huber, Roman; Craik, David J.; Schabbauer, Gernot; Gruber, Christian W.

2013-01-01

83

Systemic administration of interleukin 2 enhances the therapeutic efficacy of dendritic cell-based tumor vaccines  

PubMed Central

We have reported previously that murine bone marrow-derived dendritic cells (DC) pulsed with whole tumor lysates can mediate potent antitumor immune responses both in vitro and in vivo. Because successful therapy was dependent on host immune T cells, we have now evaluated whether the systemic administration of the T cell stimulatory/growth promoting cytokine interleukin-2 (IL-2) could enhance tumor lysate-pulsed DC-based immunizations to further promote protective immunity toward, and therapeutic rejection of, syngeneic murine tumors. In three separate approaches using a weakly immunogenic sarcoma (MCA-207), the systemic administration of nontoxic doses of recombinant IL-2 (20,000 and 40,000 IU/dose) was capable of mediating significant increases in the potency of DC-based immunizations. IL-2 could augment the efficacy of tumor lysate-pulsed DC to induce protective immunity to lethal tumor challenge as well as enhance splenic cytotoxic T lymphocyte activity and interferon-? production in these treated mice. Moreover, treatment with the combination of tumor lysate-pulsed DC and IL-2 could also mediate regressions of established pulmonary 3-day micrometastases and 7-day macrometastases as well as established 14- and 28-day s.c. tumors, leading to either significant cure rates or prolongation in overall survival. Collectively, these findings show that nontoxic doses of recombinant IL-2 can potentiate the antitumor effects of tumor lysate-pulsed DC in vivo and provide preclinical rationale for the use of IL-2 in DC-based vaccine strategies in patients with advanced cancer. PMID:10051630

Shimizu, K.; Fields, R. C.; Giedlin, M.; Mulé, J. J.

1999-01-01

84

Rational interleukin 2 therapy for HIV positive individuals: daily low doses enhance immune function without toxicity.  

PubMed Central

When administered in high doses to HIV positive (HIV+) individuals, interleukin 2 (IL-2) causes extreme toxicity and markedly increases plasma HIV levels. Integration of the information from the structure-activity relationships of the IL-2 receptor interaction, the cellular distribution of the different classes of IL-2 receptors, and the pharmacokinetics of IL-2 provides for the rationale that low IL-2 doses should circumvent toxicity. Therefore, to identify a nontoxic, but effective and safe IL-2 treatment regimen that does not stimulate viral replication, doses of IL-2 from 62,500 to 250,000 IU/m2/day were administered subcutaneously for 6 months to 16 HIV+ individuals with 200-500 CD4+ T cells/mm3. IL-2 was already detectable in the plasma of most HIV+ individuals even before therapy. Peak plasma IL-2 levels were near saturating for high affinity IL-2 receptors in 10 individuals who received the maximum nontoxic dose, which ranged from 187,500 to 250,000 IU/m2/day. During the 6 months of treatment at this dose range, plasma levels of proinflammatory cytokines remained undetectable, and plasma HIV RNA levels did not change significantly. However, delayed type hypersensitivity responses to common recall antigens were markedly augmented, and there were IL-2 dose-dependent increases in circulating Natural Killer cells, eosinophils, monocytes, and CD4+ T cells. Expanded clinical trials of low dose IL-2 are now warranted, especially in combination with effective antivirals to test for the prevention of immunodeficiency and the emergence of drug-resistant mutants and for the eradication of residual virions. Images Fig. 1 Fig. 2 PMID:8816813

Jacobson, E L; Pilaro, F; Smith, K A

1996-01-01

85

Clinical effects and toxicity of interleukin-2 in patients with cancer.  

PubMed

Interleukin-2 (IL-2) is a 15,000 dalton glycoprotein produced naturally by human T-cells during an immune response. IL-2 has been demonstrated to have substantial activity alone or in combination with the adoptive transfer of lymphokine-activated killer cells in murine tumor models. IL-2 derived from both natural (Jurkat human T-cell tumor) and recombinant (Escherichia coli) sources has been studied in Phase I protocols designed to evaluate toxicity in patients with a variety of solid tumors and to ascertain improvement in clinical parameters and immunologic status. A total of 16 patients (7 with acquired immune deficiency syndrome [AIDS] and 9 with non-AIDS malignancies) were treated with Jurkat derived IL-2. The total maximum dose (1.3 X 10(5) U/kg) was limited only by supply of this reagent. A total of 25 patients have been treated with recombinant IL-2 (RIL-2) alone. Dose-limiting toxicity manifested by marked malaise and weight gain was achieved with doses of RIL-2 of 10(6) U/kg as a single bolus or 3000 U/kg/hr. IL-2 could be administered intraperitoneally with similar toxicity. Minimal renal or hepatic toxicity was demonstrated. Hematologic toxicity was limited to mild anemia (25/25), thrombocytopenia (10/25), and marked reversible eosinophilia (15/25). Pronounced weight gain greater than 2 kg (16/25) occurred in most patients, primarily those who received cumulative doses of greater than 1-3 X 10(5) U/kg of IL-2. The weight gain amounted to as much as 10% to 20% of the pretreatment weight over 3 weeks of treatment and limited our ability to give higher doses. Two partial responses (greater than 50% decrease in cross sectional diameters) were seen in two patients with melanoma metastatic to the lung. PMID:3490903

Lotze, M T; Matory, Y L; Rayner, A A; Ettinghausen, S E; Vetto, J T; Seipp, C A; Rosenberg, S A

1986-12-15

86

DNA array analysis of interleukin-2-regulated immediate/early genes  

PubMed Central

Background Lymphocyte activation culminates in blastogenesis, cell cycle progression, DNA replication and mitosis. These complex cellular changes are programmed almost simultaneously by multiple ligands and receptors that trigger specific signal transduction pathways and transcription factors. Until now, the discovery of the genes regulated by each ligand/receptor pair has been hampered by the technologies available. Results To identify interleukin-2 (IL-2)-responsive genes, human peripheral blood mononuclear cells (PBMC) were pre-activated with anti-CD3, rested, and restimulated with IL-2 for 4 hr. Gene expression was analyzed using Affymetrix U95Av2 oligonucleotide arrays. To determine the most stringent parameters to score a gene as a bona fide IL-2 target, the expression of 19 known IL-2-regulated genes was examined first. All were induced at least 2-fold, with a difference in fluorescent intensity of ? 100 at p < 0.05. An additional 53 unique genes met these criteria. To determine which of these were immediate/early IL-2 targets in T cells, purified T cells were stimulated with IL-2 for 4 hr in the presence of cycloheximide to prevent secondary gene expression. Of the 72 genes identified in PBMCs, 20 were detected as immediate/early IL-2-regulated genes in purified T cells. In addition, 27 unique genes were IL-2-regulated in T cells but not in PBMCs. Conclusions For a successful reductionist approach to the analysis of gene expression in lymphocyte activation, it is necessary to examine purified cell populations and immediate/early gene expression regulated by each ligand/receptor pair involved. This approach should allow the discovery of genes regulated by all of the ligand/receptor pairs involved in lymphocyte activation. PMID:12459040

Beadling, Carol; Smith, Kendall A

2002-01-01

87

Regulation of mitogen-stimulated human T-cell proliferation, interleukin-2 production, and interleukin-2 receptor expression by protein kinase C inhibitor, H-7  

SciTech Connect

Recently published reports suggest that the activation of protein kinase C (PKC) plays an important role in the activation pathway of many cell types. In this study, the authors examined the role of PKC in human T-cell proliferation, IL-2 production, and IL-2R expression, when cultured with the mitogen PHA, the PKC inhibitor H-7, and H-7 control HA1004. H-7 inhibited the PHA-simulated ({sup 3}H)thymidine uptake, IL-2production, and IL-2R expression in a dose-related manner. Further, they found H-7 inhibited T-cell proliferation, IL-2 production, and IL-2mRNA from PHA plus PMA-stimulated cultures. They also found that H-7 inhibited the early-stage activation of PHA-stimulated cells. The presence of exogenous purified human IL-2 or rIL-4 partly reversed the immunosuppression caused by H-7. In contrast, HA1004 had no effect on cell proliferation, IL-2 production, or IL-2R expression. The results demonstrate that PKC activation is one major pathway through which T-cells become activated.

Atluru, D.; Polam, S.; Atluru, S. (Kansas State Univ., Manhattan (United States)); Woloschak, G.E. (Argonne National Lab., IL (United States))

1990-01-01

88

Immunomodulatory effects of interleukin-2 and interleukin-4 in patients with malignancy.  

PubMed

A phase I trial of simultaneously administered recombinant interleukin-2 (rIL-2) and recombinant human IL-4 (rHuIL-4) was conducted to evaluate the toxicity and the clinical and immunologic effects of this cytokine combination. Thirty-nine eligible patients with refractory malignancy were treated at eight different dose levels (1A to 3B): 1-3 of rIL-2 [3.0, 12.0, and 48.0 x 10(6) IU/m(2) i.v. three times weekly (TIW)] and A-C of rHuIL-4 (40, 120, and 400 mu g/m(2) s.c. TIW). The toxicity of these two cytokines was moderate and was comparable with that seen with rIL-2 alone. The maximal tolerated dose (MTD) of the combination was not reached because of lack of sufficient rHuIL-4 but is at least 48.0 x 10(6) IU/m(2) of rIL-2 and 120 mu g/m(2) of rHuIL-4. Two patients with melanoma had partial responses. The immunologic effects included increases in absolute lymphocyte numbers, and the CD3- /CD56+/ CD2+, total CD56+, CD8+, and CD16c+ lymphocyte subsets with increasing rIL-2 dose levels, but not with rHuIL-4. This increase in natural killer (NK) cells in the peripheral blood was accompanied by an increase over baseline in NK lytic activity against K562 targets; however, concomitant increases in lymphokine-activated killer (LAK) activity (Daudi targets) were not seen. The CD3+, CD4+, and CD3+/CD25+/HLA-Dr+ T-cell subsets also increased, and these increases were related to both increasing rIL-2 and rRuIL-4 doses. Finally, in four of six patients, serial tumor biopsies demonstrated increases in major histocompatibility complex (MHC) class I or II antigen expression on tumor cells or increasing T-cell infiltrates during cytokine therapy or both. This trial demonstrated that rIL-2 and rHuIL-4 can be administered simultaneously with acceptable toxicity. The immunologic findings demonstrated the expected rIL-2-associated increases of CD56+ and CD16c+ lymphocytes and NK activity, and interestingly, no development of LAK activity. These findings suggest regulatory effects of rHuIL-4 on rIL-2-related effects in vivo. PMID:8859726

Olencki, T; Finke, J; Tubbs, R; Tuason, L; Greene, T; McLain, D; Swanson, S J; Herzog, P; Stanley, J; Edinger, M; Budd, G T; Bukowski, R M

1996-01-01

89

Extravasation of intravascular fluid mediated by the systemic administration of recombinant interleukin 2.  

PubMed

Adoptive immunotherapy with lymphokine-activated killer cells and recombinant interleukin 2 (IL 2) can produce significant reduction of visceral metastases in tumor-bearing mice and, as shown recently, in humans with disseminated cancer. Because further dose escalations of IL 2 have been prevented by the development of a vascular leak syndrome (VLS) in both mice and humans, we investigated this VLS in mice undergoing the systemic administration of high-dose IL 2. A model for quantitating capillary permeability was used in which 125I-bovine serum albumin was injected i.v., and 2 hr later, tissues were counted in a gamma analyzer. A permeability index (PI) was calculated by dividing the mean counts per minute (cpm) of tissues from IL 2-treated mice by those from control animals. The injection of IL 2 produced increases in vascular permeability that were most pronounced in the thymus, spleen, lungs, liver, and kidneys (PI = 18.0, 10.0, 9.7, 6.7, and 6.3, respectively, on day 6). The development of the VLS was highly dependent on the number of days of IL 2 treatment (for example, the lungs contained 638, 1382, 3350, and 6187 cpm after 0, 1, 3, and 6 days of IL 2, respectively). Moreover, the degree of the VLS was directly related to the dose of IL 2 administered. Measurement of the wet and dry weights of lungs from IL 2-treated mice demonstrated that IL 2 produced a dramatic increase in their water weight (from 0.10 g at base line to 0.22 g after 200,000 U of IL 2 for 6 days). The injection of the IL 2 excipient failed to induce capillary leakage in tissues. Immunosuppression of mice by pretreatment irradiation (500 rad) or by injection of cyclophosphamide or by concurrent use of cortisone acetate markedly reduced or eliminated the development of the VLS. Similarly, the VLS was not observed in nude mice receiving IL 2. Thus, the administration of IL 2 produces a dose-limiting VLS that may be mediated, directly or indirectly, by host lymphoid elements. PMID:3528289

Rosenstein, M; Ettinghausen, S E; Rosenberg, S A

1986-09-01

90

Interleukin-2. A review of its pharmacological properties and therapeutic use in patients with cancer.  

PubMed

Recombinant interleukin-2 (IL-2) products (e.g. aldesleukin, teceleukin) are nonglycosylated, modified forms of the endogenous compound. IL-2 acts as a pleiotropic mediator within the immune system, having a variety of effects via specific cell surface receptors. The interaction of IL-2 with the IL-2 receptor induces proliferation and differentiation of a number of T lymphocyte subsets, and stimulates a cytokine cascade that includes various interleukins, interferons and tumour necrosis factors. Antitumour effects of IL-2 appear to be mediated by its effects on natural killer, lymphokine-activated killer (LAK) and other cytotoxic cells. In vivo and in vitro effects of IL-2 seem to be dependent to a large extent on the environment; many studies have reported conflicting results, perhaps due to diverse populations of effector cells, the availability of other cytokines that have synergistic or inhibitory influences, and the dosage regimens used. The recombinant products appear to be biologically indistinguishable from native IL-2 in vitro and in vivo; the former induce minor antibody formation but this does not appear to alter functional properties. In patients with metastatic renal cell carcinoma, IL-2 therapy achieves average objective response rates of 20% (range 0 to 40%), with a complete response rate of about 5% (range 0 to 19%). Response duration varies considerably but can be durable (lasting for > 12 months), with some patients remaining in complete response for > 60 months. It is unclear at present whether higher dosage regimens improve clinical response, or whether combination therapy with other agents and/or adoptive therapy is beneficial. Survival duration may depend on the risk factors present, with poorer performance status and more than one site of metastases associated with shorter survival times. Patients with metastatic malignant melanoma receiving IL-2 as monotherapy show an average objective response rate of 13% (range 3 to 24%); however, objective response rate averages 30% (range 4 to 59%) when IL-2 is used in combination with other agents. Overall median survival appears to be about 10 months. Preliminary data indicate that IL-2 produces a lower response rate in patients with refractory colorectal carcinoma, ovarian cancer, bladder cancer, acute myeloid leukemia or non-Hodgkin's lymphoma. Adverse effects accompanying high dose, intravenous IL-2 therapy can be severe, with cardiovascular, pulmonary, haematological, hepatic, neurological, endocrine, renal and/or dermatological complications frequently requiring doses to be withheld.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:7693434

Whittington, R; Faulds, D

1993-09-01

91

Opposite effects of interleukin-2 on normal and transfusion-suppressed healing of experimental intestinal anastomoses.  

PubMed Central

OBJECTIVE: This study was done to investigate whether administration of interleukin-2 (IL-2) can abrogate the negative effects of blood transfusions on anastomotic healing. SUMMARY BACKGROUND DATA: Recently, the authors showed that blood transfusion severely impairs anastomotic repair and significantly increases the susceptibility to intra-abdominal septic complications in rats. It has been reported that blood transfusions suppress IL-2 production and that IL-2 may stimulate wound healing. METHODS: Lewis rats underwent resection and anastomosis of both the ileum and colon. Subsequently, they received either 3 mL of saline (control and IL-2 groups) or 3 mL of blood from brown Norway donors (transfusion and transfusion/IL-2 groups) intravenously. From the operation onward, the animals in the IL-2 and transfusion/IL-2 groups received daily injections of 5.4 x 10(5) IU of IL-2 in dextrose solution subcutaneously; the rats in the other groups received only the dextrose solution. The animals were killed 3 or 7 days after the operation and examined for septic complications and anastomotic repair. RESULTS: Transfusion led to an enhanced incidence of anastomotic abscesses, which was almost completely abrogated after IL-2 administration. The anastomotic strength was consistently and significantly reduced after transfusion. Seven days after surgery, the anastomotic strength was completely restored by IL-2 treatment. For instance, the average bursting pressure (+/- the standard deviation) of the ileal anastomoses in the control, transfusion, and transfusion/IL-2 groups were 86 +/- 15, 32 +/- 8,* and 63 +/- 10 mmHg* [symbol: see text] on day 3 and 293 +/- 36, 227 +/- 16,* and 299 +/- 19 mmHg on day 7, respectively (where * = significant vs. control group and [symbol: see text] = significant vs. transfusion group). In addition, IL-2 administration elevated the anastomotic hydroxyproline content, which was significantly decreased by transfusion alone, to the level found in the control group. The administration of IL-2 to control animals resulted unexpectedly in a significant reduction in anastomotic strength. CONCLUSIONS: Exogenous IL-2 reverses the negative effects of blood transfusions on anastomotic repair, but it impairs healing under normal conditions. PMID:8257231

Tadros, T; Wobbes, T; Hendriks, T

1993-01-01

92

Impaired left ventricular filling rate induced by treatment with recombinant interleukin 2 for advanced cancer.  

PubMed Central

BACKGROUND--Immunotherapy with recombinant interleukin 2 (rIL 2) has been extensively used to treat cancer but its use has been hampered by serious side effects including severe hypotension, arrhythmias, and myocardial infarction. OBJECTIVE--To assess the effects of rIL 2 on human left ventricular function. METHODS--Left ventricular (LV) function was monitored in 22 patients (9 women, 13 men) (mean (SD) age 53 (10) years) undergoing a 120 h continuous intravenous infusion of rIL 2 (18 x 10(6) IU/m2/day) for melanoma (4), renal cell (16), ovarian (1), and colon cancer (1). Radionuclide ventriculography was performed before and 1 h after the end of treatment. Ejection fraction (EF), peak emptying rate (PER), peak filling rate (PFR), and regional left ventricular wall motion were analysed. Heart rate (HR), central venous pressure (CVP), systolic (SBP) and diastolic blood pressures (DBP), the electrocardiogram, and myocardial enzyme concentrations were monitored throughout the study. RESULTS--All variables (mean (SD)) were normal before rIL 2 was given. After rIL 2 administration HR increased significantly from 84 (11) to 125 (18) beats/min (p < 0.0001), SBP fell from 128 (11) to 100 (9) mmHg (p < 0.001) and DBP from 76 (9) to 65 (7) mmHg (p < 0.0001). CVP decreased from 3.70 (3.2) to 1.30 (0.45) cm H2O (p < 0.001). EF (65 (7) to 64 (8%) and PER (3.56 (0.60) to 3.86 (0.83) EDV/s) did not change significantly. PFR decreased significantly at the end of the rIL 2 infusion from 2.68 (0.46) to 2.37 (0.43) EDV/s (p < 0.01). Left ventricular segmental hypokinesia developed in 6 patients. Myocardial enzyme concentrations remained normal throughout the study. CONCLUSIONS--The results of this study confirmed that rIL 2 produces important haemodynamic changes, predominantly related to decreased systemic resistance. However, the observed reduction in PFR in most patients suggested that rIL 2 might exert its action at the level of the heart muscle itself. The localised systolic dysfunction in some patients suggested that rIL 2 might also adversely affect myocardial perfusion. PMID:8130026

Fragasso, G.; Tresoldi, M.; Benti, R.; Vidal, M.; Marcatti, M.; Borri, A.; Besana, C.; Gerundini, P. P.; Rugarli, C.; Chierchia, S.

1994-01-01

93

Intrapleural administration of interleukin 2 in pleural mesothelioma: a phase I-II study.  

PubMed Central

Twenty-three patients with pleural mesothelioma stage I-IIA were entered in a study of continuous daily intrapleural infusion of interleukin 2 (IL-2) for 14 days, repeated every 4 weeks. IL-2 was administered according to a groupwise dose escalation schedule (group A, 3 x 10(4); group B, 3 x 10(5); group C, 3 x 10(6); group D, 6 x 10(6); group E, 18 x 10(6); and group F, 36 x 10(6) IU day-1). Each group consisted of at least three patients. Intrapleural administration of IL-2 was associated with acceptable toxicity. All patients were treated on an outpatient basis except for the patients at dose levels E and F. Dose-limiting toxicity was observed at level F, 36 x 10(6) IU daily, and consisted of catheter infection, fever and flu-like symptoms. Intrapleural IL-2 levels were high (> 20,000 IU ml-1) at levels E and F, while serum levels in most patients were not or barely detectable (< 3-30 IU ml-1). Intrapleural IL-2 levels were up to 6000-fold higher than systemic levels. Intrapleural tumour necrosis factor alpha (TNF-alpha) levels varied greatly and did not correlate with IL-2 dosage. Intrapleural mononuclear cells (MNCs) displayed IL-2-induced lymphokine-activated killer (LAK) activity in all patients. Two patients were not evaluable for response owing to catheter-related problems which precluded the delivery of IL-2. Partial response (PR) occurred in 4 of 21 evaluable patients (19%; 95% confidence interval 5-42%) with a median time to progression of 12 months (range 5-37). Stable disease (SD) occurred in seven patients with a median time to progression of 5 months (range 2-7). There were no complete responses (CRs). The median overall survival was 15.6 months (range 3.0-43). No relationship between the dose of IL-2 and response rate was observed. We conclude that IL-2 given intrapleurally is accompanied with acceptable toxicity and has anti-tumour activity against mesothelioma. In view of the refractory nature of the disease IL-2 may be a treatment option for mesothelioma. A formal phase II study is warranted. Based on the observed toxicity, the lack of dose-response relationship and the immunomodulatory effects seen at relatively low-dose IL-2, the recommended dose for a phase II study is 3 x 10(6) IU day-1 using the present treatment schedule. Images Figure 1 PMID:7577483

Goey, S. H.; Eggermont, A. M.; Punt, C. J.; Slingerland, R.; Gratama, J. W.; Oosterom, R.; Oskam, R.; Bolhuis, R. L.; Stoter, G.

1995-01-01

94

Involvement of thromboxane and neutrophils in multiple-system organ edema with interleukin-2.  

PubMed Central

Interleukin-2 (IL-2) produces toxicity characterized by generalized edema within 24 hours. This study tests whether the rate of IL-2 administration modulates the onset of edema and examines thromboxane (Tx) and neutrophils as possible mediators of this event. Recombinant human IL-2, 10(5) U (n = 7), 10(6) U (n = 9), or vehicle (n = 8) were given to anesthetized rats intravenously during a period of 1 hour. At 6 hours edema, as measured by increase in wet to dry weight (w/d) ratio, was present in the heart, liver, and kidney, with 10(5) U IL-2 and in the lung, heart, liver and kidney, with 10(6) U IL-2, relative to values with vehicle-infused controls (all p less than 0.05). With a 1-hour infusion of 10(6) U IL-2, there was an increase in plasma thromboxane (Tx)B2 level to 1290 +/- 245 pg/mL, higher than 481 +/- 93 pg/mL in control rats (p less than 0.05); lung polymorphonuclear leukocyte (PMN) sequestration of 53 +/- 7 PMN/10 higher-power fields (HPF) relative to 23 +/- 2 PMN/10 HPF in controls (p less than 0.05); and increased bronchoalveolar lavage (BAL) fluid protein concentration of 1970 +/- 210 micrograms/mL relative to 460 micrograms/mL in controls (p less than 0.05). When 10(6) U IL-2 was given as a 1-minute intravenous bolus (n = 9), edema was not demonstrated, plasma TxB2 levels were similar to controls, there was no leukosequestration, and BAL protein levels were normal. These data indicate that a constant infusion but not the rapid bolus administration of IL-2 produces in rats multiple-system organ edema, increased plasma TxB2, sequestration of PMNs, and microvascular permeability. These findings may explain the early toxicity seen in patients given high-dose IL-2 in cancer treatment. PMID:2256765

Welbourn, R; Goldman, G; Kobzik, L; Valeri, C R; Shepro, D; Hechtman, H B

1990-01-01

95

Recombinant interleukin 2 stimulates in vivo proliferation of adoptively transferred lymphokine-activated killer (LAK) cells.  

PubMed

We previously reported that the adoptive transfer of lymphokine-activated killer (LAK) cells plus repetitive injections of recombinant interleukin 2 (IL 2) produced a marked reduction in established pulmonary metastases from a variety of murine sarcomas. The requirement for the exogenous administration of IL 2 prompted a subsequent examination of the role of IL 2 in the in vivo function of transferred LAK cells. The in vivo proliferation and migration patterns of lymphoid cells in C57BL/6 mice were examined after i.v. transfer of LAK cells alone, i.p. injection of IL 2 alone, or the combination of LAK cells and IL 2. A model for in vivo labeling of the DNA of dividing cells was used in which mice were injected with 5-[125I]-iodo-2'-deoxyuridine (125IUdR) and, 20 hr later, their tissues were removed and were counted in a gamma analyzer. A proliferation index (PI) was calculated by dividing the mean cpm of organs of experimentally treated mice by the mean cpm of organs of control mice. In animals given LAK cells alone, the lungs and liver demonstrated little if any uptake of 125IUdR above saline-treated controls (PI = 2.5 and 0.8, respectively, on day 5), whereas the same organs of mice receiving 6000 U of IL 2 alone displayed higher radiolabel incorporation (PI = 7.1 and 5.9, respectively). When mice were given LAK cells plus 6000 U of IL 2, their tissues showed an additional increase in 125IUdR uptake. In the spleen, kidneys, and mesenteric lymph nodes, IL 2 treatment alone (6000 U) produced elevated PI values that were not, however, additionally increased if LAK cells were also administered. To separate the stimulatory effects of IL 2 on host lymphocyte proliferation from similar IL 2 effects on injected LAK cells, these studies were repeated in mice immunosuppressed by 500 rad total body irradiation. Pre-irradiation of the host sufficiently reduced endogenous lymphoid expansion stimulated by IL 2 so as to allow the demonstration that IL 2 also induced the proliferation of the transferred LAK cells. A variety of studies confirmed that the injected LAK cells were actively proliferating in tissues in vivo under the influence of IL 2. Substitution of "normal" LAK cells with fresh and cultured (without IL 2) splenocytes, or irradiated LAK cells did not result in increased 125IUdR uptake in tissues. Histologic studies corroborated the findings of the 125IUdR incorporation assays and revealed extensive lymphoid proliferation in irradiated mice receiving LAK cells plus IL 2.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3900213

Ettinghausen, S E; Lipford, E H; Mulé, J J; Rosenberg, S A

1985-11-01

96

Systemic administration of recombinant interleukin 2 stimulates in vivo lymphoid cell proliferation in tissues.  

PubMed

Recent work in our laboratory has demonstrated that the repeated injections of high doses of recombinant interleukin 2 (IL 2) can dramatically reduce the number of established pulmonary and hepatic metastases and the growth of intradermal tumors in a variety of murine tumor models. We have thus undertaken studies to define the mechanisms underlying these in vivo effects of IL 2. Using an in vivo DNA-labeling technique in which we employed 5-[125I]iodo-2'-deoxyuridine (125IUdR), we examined the in vivo cell proliferation in the tissues of mice treated with IL 2. A proliferation index (PI) was calculated by dividing the raw counts per minute (cpm) of tissues in IL 2-treated mice by the cpm in corresponding tissues of control animals. At an IL 2 dose of 6000 U given i.p. three times a day, the highest 125IUdR incorporation was seen in the lungs, liver, spleen, kidneys, and mesenteric lymph nodes (PI = 6.9, 6.9, 5.1, 7.1, 24.6, respectively, at 5 days). The amount of lymphoid proliferation in these organs was a direct function of the dose of IL 2 administered. Other tissues including thymus, intestines, skin, and hind limb showed no significant increase in 125IUdR uptake even after host treatment with the highest doses of IL 2. Blood and brain demonstrated intermediate incorporation of the radiolabel. Preirradiation of the host largely eliminated the proliferative response to IL 2. Histologic studies of normal and irradiated mice receiving IL 2 corroborated the result of the 125IUdR findings. In normal IL 2-treated mice, large collections of activated lymphoid cells were seen, most prominently in the lungs, liver, and kidneys, whereas markedly decreased lymphoid proliferation was evident histologically in preirradiated mice. A fluorescein-labeled monoclonal antibody directed against the Thy-1.2 surface determinant was used to identify these dividing cells in frozen tissue sections as T lymphoid cells. Activated lymphocytes isolated from the lungs, liver, spleen, and mesenteric lymph nodes of IL 2-treated mice demonstrated significant lysis of a fresh murine sarcoma target in short-term 51Cr-release assays. These studies demonstrate that the systemic administration of recombinant IL 2 causes in vivo activation and proliferation of host lymphoid cells and has important implications for the adoptive immunotherapy of tumors. PMID:3891854

Ettinghausen, S E; Lipford, E H; Mulé, J J; Rosenberg, S A

1985-08-01

97

Time-dependent effects of striatal interleukin-2 on open field behaviour in rats  

Microsoft Academic Search

There is evidence that immune messengers like cytokines can modulate emotional and motivated behaviours and are involved in psychiatric conditions like anxiety, and depression. Previously, we showed that cytokine gene expression (interleukin (IL)-2 mRNA) in specific brain tissues (striatum, prefrontal cortex) correlated with anxiety-like behaviour (open arm time) in an elevated plus-maze in rats. In a subsequent experiment, a single

Britta D. Karrenbauer; Ying-Jui Ho; Verena Ludwig; Jeanette Löhn; Rainer Spanagel; Rainer K. W. Schwarting; Cornelius R. Pawlak

2009-01-01

98

Induction of lymphokine-activated killer cells with low-dose interleukin 2 and interferon-? in oral cancer patients  

Microsoft Academic Search

Lymphokine-activated killer (LAK) cells were induced with low-dose recombinant interleukin 2 (rIL-2) and recombinant interferon-? (IFN-?) in 28 oral carcinoma patients. The patients received daily intravenous injections of rIL-2 (1.2×105 U\\/m2) and rIFN-? (7.0×104 U\\/m2), and both natural killer (NK) and LAK activities were periodically examined. A significant increase in CD16+CD57+ and CD16+CD57- NK subsets was observed after the induction.

Kazunori Yoneda; Tetsuya Yamamoto; Eisaku Ueta; Tokio Osaki

1992-01-01

99

Potentiated lymphokine-activated killer cell activity generated by low-dose interleukin-2 and mismatched double-stranded RNA  

Microsoft Academic Search

Summary Lymphokine-activated killer (LAK) cell activity was measured in human peripheral blood mononuclear cells (PBMC) treated in vitro for 3 days with recombinant interleukin-2 (rIL-2) and mismatched double-stranded RNA (dsRNA). Lytic activity was measured utilizing K562 (NK-sensitive) and 786-0 (NK-resistant) target cells. PBMC cultured with rIL-2 (10–1000 BRMP U\\/ml) alone showed concentration-dependent lytic activity against the 786-0 target cells, while

Howard R. Hubbell; George D. Gibson; Robert D. Bigler

1992-01-01

100

Active immunization of metastatic melanoma patients with interleukin-2-transduced allogeneic melanoma cells: evaluation of efficacy and tolerability  

Microsoft Academic Search

From January 1994 to July 1996 we immunized metastatic melanoma patients with HLA-A2-compatible, interleukin-2 (IL-2)-secreting,\\u000a immunogenic melanoma lines in an attempt to induce a systemic reaction that might also affect distant melanoma lesions. Twelve\\u000a patients (6 male and 6 female) aged from 28 to 72 years, affected with visceral and\\/or subcutaneous (s.c.) melanoma metastases,\\u000a were treated. Two different HLA-A2+ melanoma

Filiberto Belli; Flavio Arienti; J. Sulé-Suso; C. Clemente; Luigi Mascheroni; Alessandro Cattelan; Cristina Santantonio; Gian Francesco Gallino; Cecilia Melani; Stefania Rao; Mario P. Colombo; Michele Maio; Natale Cascinelli; G. Parmiani

1997-01-01

101

Soluble interleukin-2 receptors, antineutrophil cytoplasmic antibodies, and other autoantibodies in patients with ulcerative colitis.  

PubMed Central

Ulcerative colitis (UC) is an inflammatory bowel disease of unknown aetiology. In this study, serum samples from 80 patients with UC were studied for the presence of various autoantibodies and soluble interleukin-2 receptor molecules (sIL-2Rs) in an attempt to determine the degree of activation of the immune system in this disease process. Autoantibodies detected included rheumatoid factors (in 5% of patients), antinuclear antibodies (in 51.3%), anti-Ro(SSA) (in 1.3%), anticardiolipin antibodies (IgG and/or IgM classes in 26.3%), anti-double stranded DNA (IgG or IgM classes in 45%), and antineutrophil cytoplasmic antibodies (ANCAs, in 30%). The ANCAs had a perinuclear pattern (p-ANCA) in 95.8%, without anti-myeloperoxidase activity, at least in an enzyme linked immunosorbent assay (ELISA) system. Raised concentrations of sIL-2R were found in 32.5% of patients (26/80, 18 with active and eight with inactive UC). The mean (SD) sIL-2R concentrations were significantly higher in patients with active UC (595 (219) u/ml v 406 (162) u/ml, p = 0.0001) and in patients with ANCAs (584 (177) u/ml in ANCA positive v 447 (212) u/ml in ANCA negative patients, p < 0.01). The sIL-2R concentrations were correlated with increased serum concentrations of C3c (r = 0.23, p < 0.05) or C4 (r = 0.4, p < 0.001) components of the complement system and erythrocyte sedimentation rate (ESR, r = 0.44, p = 0.0001). Platelets, ESR, and C3c were not associated with disease activity (p = 0.06, 0.33 and 0.86) whereas mean (SD) serum concentrations of C4 were higher in active disease (37.4 (11.9) mg/dl v 32.3 (10.3) mg/dl, p < 0.05). The sIL-2Rs had 53% sensitivity and 82.6% specificity for disease activity whereas platelet counts had 53% sensitivity and 58.7% specificity. To conclude, UC is accompanied by an autoimmune response that results in the production of several autoantibodies and cellular immune activation, as shown by the high sIL-2R concentration, is also present. The identification of the target antigen(s) of p-ANCA would possibly act as an indicator of disease activity if this distinct subset of ANCAs can be attributed to the pathogenesis of UC. The sIL-2R concentrations seem to be a useful laboratory marker for assessing activity of the disease. PMID:8504967

Dalekos, G N; Manoussakis, M N; Goussia, A C; Tsianos, E V; Moutsopoulos, H M

1993-01-01

102

Plasma nitrate plus nitrite changes during continuous intravenous infusion interleukin 2.  

PubMed Central

Nitric oxide (NO), a biologically active mediator generated in many cell types by the enzyme NO synthase, may play an important role in cardiovascular toxicity that is frequently observed in cancer patients during intravenous (i.v.) interleukin 2 (IL-2) therapy. The induction of NO synthase and the production of NO seem to be involved in the pathogenesis of the vascular leakage syndrome, as well as in the regulation of myocardial contractility. In the present study, we evaluated the pattern of plasmatic NO changes during multiple cycles of continuous i.v. infusion (CIVI) of IL-2 in ten advanced cancer patients (five males, five females, median age 59 years, range 33-67 years; eight affected by renal cell cancer and two affected by malignant melanoma). The patients received IL-2 at 18 MIU m-2 day-1 (14 cycles) or 9 MIU m-2 day-1 (seven cycles) for 96 h, repeated every 3 weeks. Interferon alpha (IFN alpha) was also administered subcutaneously (s.c) during the 3 week interval between IL-2 cycles. For each cycle, plasma samples were collected before treatment (t0), 24 h (t1), 48 h (t2), 72 h (t3) and 96 h (t4) after the start of IL-2 infusion, and 24 h after the end of the cycle. NO concentration was determined spectrophotometrically by measuring the accumulation of both nitrite and nitrate (after reduction to nitrite). The following observations may be drawn from data analysis: (1) plasma nitrate + nitrite significantly raised during treatment (P = 0.0226 for t0 vs t3), but statistical significance was retained only when cycles administered with IL-2 18 MIU m-2 day-1 are considered (P = 0.0329 for t0 vs t3; P = 0.0354 for t0 vs t2 vs t4) (dose-dependent pattern); (2) during subsequent cycles a significant trend toward a progressive increase of plasma nitrate + nitrite levels, with increasing cumulative dose of IL-2, was observed (linear regression coefficient r = 0.62, P = 0.0141 for t0; r = 0.80, P = 0.0003 for t1; r = 0.62, P = 0.013 for t2; r = 0.69, P = 0.045 for t3); (3) plasma nitrate + nitrite levels peaked earlier in subsequent cycles than in the first cycle; (4) all patients experienced hypotension. The mean of the systolic blood pressure values was significantly lower at the time of plasma nitrate + nitrite peak than at t0 (P = 0.0004); (5) the two cases of grade III hypotension occurred in patients with the higher mean and peak plasma nitrate + nitrite values. We conclude that determination of plasma nitrate + nitrite levels during CIVI IL-2 can usefully estimate, in a dose-dependent pattern, the degree of peripheral vascular relaxation and capillary leakage associated with cytokine action, clinically manifested as hypotension. However, isolated cardiac toxicity that continues to represent a relevant problem during IL-2 therapy, does not appear to correlate with plasma nitrate + nitrite levels; therefore, further studies are required to understand adequately the mechanisms underlying IL-2-induced cardiac toxicity. PMID:8883421

Citterio, G.; Pellegatta, F.; Lucca, G. D.; Fragasso, G.; Scaglietti, U.; Pini, D.; Fortis, C.; Tresoldi, M.; Rugarli, C.

1996-01-01

103

Theileria parva: reappearance of schizonts in infected lymphoblastoid cells treated with parvaquone is dependent on interleukin 2-like growth factors.  

PubMed

Schizont-infected cell lines derived by in vitro infection of bovine T cell clones with the Muguga isolate of Theileria parva were treated for 72 hr with the theileriacidal drug, parvaquone, at a concentration of 10 micrograms/ml. This treatment completely eliminated schizonts from the recovered cells, which failed to undergo further proliferation and died. However, treated cells cultured with either bovine T cell growth factor or human recombinant interleukin 2 remained viable, underwent proliferation, and in many instances, schizonts reappeared. When cultured in the presence of supernatant obtained from an actively growing T. parva-infected cell line, treated cells did not proliferate, but schizonts reappeared. The cells became transformed by the parasite and grew continuously in the absence of exogenous growth factor. The appearance of schizonts was preceded by the development of densely staining intracytoplasmic inclusions, visualized by light and electron microscopy. Electron-dense inclusions were shown to contain DNA. Hybridization of a T. parva-specific DNA probe to Southern blots of restriction enzyme-digested DNA prepared from parvaquone-treated cells which developed inclusions but not schizonts produced a pattern similar to that seen with DNA prepared from schizont-infected cells. We conclude that reorganization of schizonts can occur in T. parva-infected T lymphocytes cured of infection with parvaquone in the presence of interleukin 2 or growth factors produced by T. parva-infected cells. The implications of these results for the establishment of a carrier state following parvaquone therapy are discussed. PMID:2495228

Brown, W C; Shaw, M K; Conrad, P A; Dolan, T T

1989-04-01

104

Expression levels and genetic polymorphisms of interleukin-2 and interleukin-10 as biomarkers of Graves’ disease  

PubMed Central

The aim of the present study was to determine whether the expression levels of interleukin (IL)-2 and IL-10 may be used as biological markers in Graves’ disease (GD) patients. A total of 256 individuals, including 118 GD patients and 138 healthy individuals, were enrolled into the study. Blood samples were collected from each patient and healthy individual, which were then subjected to enzyme-linked immunosorbent assay (ELISA). Total RNA and total proteins were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, respectively. In addition, restriction fragment length polymorphism (RFLP) analysis was performed to detect the presence of genetic polymorphisms. The ELISA results indicated that the IL-2 and IL-10 serum levels in the GD patients were increased by ~5.2 and ~7-fold when compared with the levels in the healthy controls. The results of RT-qPCR indicated that the mRNA expression levels of IL-2 and IL-10 were upregulated in the GD patients when compared with the healthy controls. Furthermore, the western blot analysis results revealed that the protein expression levels of IL-2 and IL-10 were significantly increased in the GD patients. RFLP analysis indicated that the increased number of GG single nucleotide polymorphisms (SNPs) in the GD group were detected in the ?330 locus of the IL-2 promoter and the ?1082 locus of the IL-10 promoter. In addition, the results indicated that the relatively high rates of homozygous GG SNPs (IL-2 ?330T/G and IL-10 ?1082A/G polymorphisms) on the alleles may be associated with the incidence of GD. The serum, mRNA and protein expression levels of IL-2 and IL-10 were significantly increased in GD patients when compared with the levels in the healthy controls. In conclusion, the expression levels and genetic polymorphisms of IL-2 and IL-10 may be potential biomarkers for the incidence of Graves’ disease in the population studied.

LIANG, CUIGE; DU, WENHUA; DONG, QINGYU; LIU, XIAOMENG; LI, WENXIA; WANG, YUELI; GAO, GUANQI

2015-01-01

105

[Isolation, crystallization, and preliminary x-ray study of a Fab fragment of monoclonal antibody against human interleukin-2 and its complex with antigenic peptide].  

PubMed

Antigen-binding fragments (Fab) of mouse monoclonal antibodies to human interleukin-2 were obtained in preparative quantities by a modified procedure. These Fab-fragments were shown to be homogeneous according to the isoelectric focusing method. Various monocrystals of these free Fab-fragments and their complexes with the antigenic peptide corresponding to the 59-72 sequence of interleukin-2 were obtained. These were shown to be suitable for X-ray and were preliminarily studied by X-ray. PMID:10422589

Mikha?lova, I Iu; Mareeva, T Iu; Tsygannik, I N; Mikhaleva, I I; Onoprienko, L V; Vikhrov, A A; Markvicheva, E A; Panghorn, W; Duax, W; Nesmiianov, V A; Pletnev, V Z

1999-04-01

106

Lung-protective effects of the metalloporphyrinic peroxynitrite decomposition catalyst WW-85 in interleukin-2 induced toxicity.  

PubMed

Recombinant interleukin-2 (IL-2) therapy for malignancy is associated with a pulmonary vascular leakage syndrome (VLS) similar to that seen in sepsis. We investigated the possibility that the IL-2-induced VLS may be associated with the release of peroxynitrite (ONOO(-)), and used a model of IL-2-induced VLS in sheep to test the effects of the ONOO(-) decomposition catalyst WW-85. Eighteen sheep were chronically instrumented and randomly divided into three groups (n=6 per group): sham: lactated Ringer's solution, control: IL-2, and treatment: IL-2 and WW-85. Treatment with WW-85 significantly improved lung transvascular fluid flux, decreased lipid peroxidation, limited iNOS as well as PAR intensity, prevented tachycardia, and attenuated the increase in core body temperature resulting from IL-2 treatment. These findings suggest that ONOO(-) plays a pivotal role in the pathology of IL-2-induced pulmonary VLS, and that WW-85 may become a useful treatment option. PMID:18951875

Maybauer, Dirk M; Maybauer, Marc O; Szabó, Csaba; Westphal, Martin; Traber, Lillian D; Enkhbaatar, Perenlei; Murthy, Kanneganti G K; Nakano, Yoshimitsu; Salzman, Andrew L; Herndon, David N; Traber, Daniel L

2008-12-19

107

Inhibitory effect of 3'-azido-3'-deoxythymidine on proliferative responsiveness of CD8+ lymphocytes to interleukin-2.  

PubMed

Although several studies have shown that 3'-azido-3'-deoxythymidine (AZT) is not toxic for CD4+ lymphocytes, its effect on CD8+ cells has never been studied in a systematic way. We purified CD8+ cells from the peripheral blood mononuclear cells of both human immunodeficiency virus (HIV)-seronegative and HIV-infected individuals by means of magnetic beads that had been coated with monoclonal antibodies. We report that AZT, but not two other nucleosides tested, inhibited the interleukin-2-dependent proliferation of CD8+ lymphocytes in a dose-dependent manner. No such effect was observed with regard to CD4(+)-enriched populations. The AZT-mediated antiproliferative effect did not appear to be related to either the CD4+ count or to prior treatment with this drug in the case of HIV-seropositive subjects. PMID:8556489

Mercure, L; Lalonde, R; Phaneuf, D; Brenner, B; Wainberg, M A

1994-07-01

108

Recombinant interleukin-2 (rIL-2) with flavone acetic acid (FAA) in advanced malignant melanoma: immunological studies.  

PubMed Central

Natural killer (NK) cell activity and lymphokine activated killer (LAK) cell cytotoxicity were measured in patients receiving recombinant interleukin 2 (rIL-2) and flavone acetic acid (FAA) for treatment of progressing metastatic melanoma. NK activity was increased in 23 of 26 patients and LAK activity induced in 13 of 26 patients. However, levels of cytotoxicity in the present study were not significantly greater than a previous study using rIL-2 alone. LAK cell precursors demonstrated by in vitro incubation of pretreatment lymphocytes with IL-2 and subsequent cytotoxicity were no different in the patients compared to normal controls. Analysis of cell surface phenotypes failed to reveal any significant changes in the cell populations examined, including IL-2R and Leu 19. Although five patients had tumour response, one being complete, there was no correlation with the immunological parameters examined. PMID:2328217

Ghosh, A. K.; Mellor, M.; Prendiville, J.; Thatcher, N.

1990-01-01

109

Evaluation of the effect of continuous infusion recombinant interleukin-2 (bioleukin) on peripheral blood leucocytes of patients with terminal malignancy.  

PubMed Central

Seventeen patients with terminal malignancy have been entered into a sequential investigation of two doses of continuous infusion recombinant interleukin-2 (bioleukin) given in the setting of a general ward. After an initial experience of a dose of 300 micrograms m-2 in eight patients the remainder received 400 micrograms m-2. Temporary interruption of treatment at the first sign of any serious toxicity led to rapid resolution of side-effects. No patient needed intensive care support, although nine of 17 required temporary interruption of infusion, lasting on average 4 h. Median lymphocyte rebound on day 14 was 3.6 times the pre-treatment level. It remained above pre-treatment levels in four of five patients who had no shown disease progression at day 56 after more than 28 days off treatment. Minor responses occurred in five patients, lasting on average 4 months. PMID:2605101

Oliver, R. T.; Crosby, D.; Nouri, A.; Scott, E.; Galazka, A.

1989-01-01

110

Immunotherapy with recombinant human interleukin 2 in patients with hematological malignancies after bone marrow or peripheral blood stem cell transplantation.  

PubMed

High-dose chemotherapy in conjunction with bone marrow transplantation (BMT) or peripheral blood stem cell transplantation (PBSCT) is increasingly being used for treatment of patients with hematological malignancies. Residual tumor cells, resistant to high-dose chemoradiotherapy, are responsible for reccurence of the disease. Interleukin 2 (IL-2), a pleiotropic cytokine which plays a central role in immune response, has been introduced in several clinical trials in patients with hematological malignancies after BMT or PBSCT to increase immunocompetence of these patients and eradicate residual malignant cells. At present there is no general agreement on the optimum dosage or route of administration and clinical trials also gave conflicting results. Establishment of optimum dosage schedules and methods of administration should enable a better assessment of the place of IL-2 in the treatment of these patients. PMID:10483870

Frydecka, I; Kosmaczewska, A; Bo?ko, D; Ciszak, L; Kaczmarek, P

1999-01-01

111

Transcriptional repression of the interleukin-2 gene by vitamin D3: direct inhibition of NFATp/AP-1 complex formation by a nuclear hormone receptor.  

PubMed Central

T-lymphocyte proliferation is suppressed by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], the active metabolite of vitamin D3, and is associated with a decrease in interleukin 2 (IL-2), gamma interferon, and granulocyte-macrophage colony-stimulating factor mRNA levels. We report here that 1,25(OH)2D3-mediated repression in Jurkat cells is cycloheximide resistant, suggesting that it is a direct transcriptional repressive effect on IL-2 expression by the vitamin D3 receptor (VDR). We therefore examined vitamin D3-mediated repression of activated IL-2 expression by cotransfecting Jurkat cells with IL-2 promoter/reporter constructs and a VDR overexpression vector and by DNA binding. We delineated an element conferring both DNA binding by the receptor in vitro and 1,25(OH)2D3-mediated repression in vivo to a short 40-bp region encompassing an important positive regulatory element, NF-AT-1, which is bound by a T-cell-specific transcription factor, NFATp, as well as by AP-1. VDR DNA-binding mutants were unable to either bind to this element in vitro or repress in vivo; the VDR DNA-binding domain alone, however, bound the element but also could not repress IL-2 expression. These results indicate that DNA binding by VDR is necessary but not sufficient to mediate IL-2 repression. By combining partially purified proteins in vitro, we observed the loss of the bound NFATp/AP-1-DNA complex upon inclusion of VDR or VDR-retinoid X receptor. Order of addition and off-rate experiments indicate that the VDR-retinoid X receptor heterodimer blocks NFATp/AP-1 complex formation and then stably associates with the NF-AT-1 element. This direct inhibition by a nuclear hormone receptor of transcriptional activators of the IL-2 gene may provide a mechanistic explanation of how vitamin derivatives can act as potent immunosuppressive agents. PMID:7565732

Alroy, I; Towers, T L; Freedman, L P

1995-01-01

112

Interleukin-2-mediated inhibition of dendritic cell development correlates with decreased CD135 expression and increased monocyte/macrophage precursors.  

PubMed

We have previously shown that interleukin-2 (IL-2) inhibits dendritic cell (DC) development from mouse bone marrow (BM) precursors stimulated with the ligand for FMS-like tyrosine kinase 3 receptor (Flt3L), and have provided evidence that this inhibition occurs at the monocyte DC precursor stage of DC development. Here, we explored the mechanism of IL-2-mediated inhibition of DC development. First, we showed that these in vitro cultures accurately model DCs that develop in vivo by comparing gene and protein expression of the three main Flt3L-induced DC subsets from the BM, CD11b(+) and CD24(+) conventional DCs (cDCs) and plasmacytoid DCs (pDCs) with their respective ex vivo spleen DC subsets (CD11b(+), CD8(+) and pDCs). Next, gene expression changes were quantified in Flt3L DC subsets that developed in the presence of IL-2. These changes included increased expression of Bcl2l11, which encodes the apoptosis-inducing protein Bim, and decreased expression of Flt3 (CD135), the receptor that initiates DC development. Interleukin-2 also significantly reduced Flt3 protein expression on all three Flt3L DC subsets, and attenuated Flt3L-induced STAT3 phosphorylation in DCs. Based on these data, we hypothesized that decreased Flt3 signalling may divert BM precursors down monocyte and macrophage lineages. Indeed, addition of IL-2 led to increases in Flt3(-) cells, including cKit(+) Ly6C(+) CD11b(-) populations consistent with the recently identified committed monocyte/macrophage progenitor. Therefore, IL-2 can inhibit DC development via decreased signalling through Flt3 and increased monocyte/macrophage development. PMID:24954893

Guerrero, Alan D; Dong, Matthew B; Zhao, Yongge; Lau-Kilby, Annie; Tarbell, Kristin V

2014-12-01

113

Observations on the systemic administration of autologous lymphokine-activated killer cells and recombinant interleukin-2 to patients with metastatic cancer.  

PubMed

We describe here the preliminary results of the systemic administration of autologous lymphokine-activated killer (LAK) cells and the recombinant-derived lymphokine interleukin-2 to patients with advanced cancer. This regimen was based on animal models in which the systemic administration of LAK cells plus interleukin-2 mediated the regression of established pulmonary and hepatic metastases from a variety of murine tumors in several strains of mice. We treated 25 patients with metastatic cancer in whom standard therapy had failed. Patients received both 1.8 to 18.4 X 10(10) autologous LAK cells, generated from lymphocytes obtained through multiple leukaphereses, and up to 90 doses of interleukin-2. Objective regression of cancer (more than 50 per cent of volume) was observed in 11 of the 25 patients: complete tumor regression occurred in one patient with metastatic melanoma and has been sustained for up to 10 months after therapy, and partial responses occurred in nine patients with pulmonary or hepatic metastases from melanoma, colon cancer, or renal-cell cancer and in one patient with a primary unresectable lung adenocarcinoma. Severe fluid retention was the major side effect of therapy, although all side effects resolved after interleukin-2 administration was stopped. Further development of this approach and additional patient follow-up are required before conclusions about its therapeutic value can be drawn. PMID:3903508

Rosenberg, S A; Lotze, M T; Muul, L M; Leitman, S; Chang, A E; Ettinghausen, S E; Matory, Y L; Skibber, J M; Shiloni, E; Vetto, J T

1985-12-01

114

Laparoscopic cytoreductive nephrectomy as preparation for administration of systemic interleukin-2 in the treatment of metastatic renal cell carcinoma: a pilot study  

Microsoft Academic Search

Objectives. Cytoreductive nephrectomy is commonly performed in patients with metastatic renal cell carcinoma before systemic interleukin-2 (IL-2) therapy. Open nephrectomy is associated with prolonged recovery during which metastatic disease can progress. The feasibility of laparoscopic cytoreductive surgery in these patients with large renal tumors was examined. The role of tumor morcellation in reducing the recovery period and allowing earlier treatment

McClellan M Walther; J. Chris Lyne; Steven K Libutti; W. Marston Linehan

1999-01-01

115

mRNA  

NSDL National Science Digital Library

Template for protein synthesis. Each set of three bases, called codons, specifies a certain protein in the sequence of amino acids that comprise the protein. The sequence of a strand of mRNA is based on the sequence of a complementary strand of DNA.

Darryl Leja (National Human Genome Research Institute REV)

2005-04-04

116

Influence of tunicamycin, sialidase, and cholera toxin on gangliosides and T-lymphocyte responses to interleukin 2  

SciTech Connect

The authors have shown that gangliosides inhibit interleukin 2 (IL 2)-dependent proliferation of murine T cells. Tunicamycin (TM), sialidase, and cholera toxin-..beta.. subunit (..beta..-CT) are known modulators of cell surface glycoconjugates. To test the possible role of endogenous gangliosides in T cell responses to IL-2, the effect of these agents on ganglioside expression and cell proliferation was studied. Gangliosides were labelled for 24 hrs with /sup 3/H-glucosamine/galactose in the presence of IL-2 and purified sialidase, TM or ..beta..-CT. Gangliosides were isolated and the species separated by TLC. Alternatively, proliferation was assayed by /sup 3/H-thymidine uptake after 48 hrs culture. TM treatment at a concentration (10 ..mu..g/ml) that completely inhibited proliferation resulted in a 86% reduction of incorporation of saccharide precursors into gangliosides compared to a 50% reduction into proteins. Sialidase treatment (0.1 IU/ml) resulted in a 70% inhibition of proliferation and 30% reduction of radiolabel into gangliosides, of which 3 species were specifically reduced. ..beta..-CT, which binds to GM/sub 1/ and to a lesser extent GD/sub 1a/, caused a 50% reduction in proliferation response at 35 units/ml. The results support the hypothesis that gangliosides are involved in IL-2-dependent proliferation.

Semmes, O.J.; Bailey, J.M.; Merritt, W.D.

1986-05-01

117

4-Fluoro-3-nitrophenyl grafted gold electrode based platform for label free electrochemical detection of interleukin-2 protein.  

PubMed

A new platform based on 4-Fluoro-3-nitrophenyl (FNP) grafted gold disk electrode prepared via electrochemical reduction of 4-fluoro-3-nitrobenzene diazonium ion has been developed and utilized for biosensor fabrication. Anti-interleukin-2 (anti-IL2) antibody has been covalently immobilized onto FNP/Au surface and utilized for label free electrochemical impedance based detection of cytokine IL2. FNP acts as a bridge (cross-linker) between gold surface and anti-IL2, where fluoro group of FNP undergoes nucleophilic substitution by amino group of biomolecule and results in its covalent immobilization. The immobilization process and fabricated electrode have been characterized using contact angle (CA) measurements, cyclic voltammetry (CV) and electrochemical impedance (EIS) technique. CV studies show that FNP grafted surface provides conductive surface for anti-IL2 immobilization. The EIS response of studies as a function of IL2 concentrations exhibits a detection in linear range from 1 pg ml(-1) to 10 ng ml(-1) with minimum detectable concentration of 1 pg ml(-1). The electrode has been found to be selective against other cytokine molecules. PMID:24906083

Arya, Sunil K; Park, Mi Kyoung

2014-11-15

118

Proliferation induced by Plasmodium falciparum antigen and interleukin-2 production by lymphocytes isolated from malaria-immune individuals.  

PubMed Central

Affinity-purified Plasmodium falciparum soluble antigens (SPAg) isolated from in vitro cultures of the parasite were shown to be relatively free of nonspecific polyclonal activators. To determine the presence of lymphocytes with specificity against SPAg in the peripheral blood of malaria-immune individuals, the proliferative response and the interleukin-2 (IL-2) production of SPAg-activated mononuclear cells (MNCs) from individuals unexposed, sensitized, and immune to malaria were measured. It was found that MNC isolated from malaria-immune individuals proliferated in response to SPAg and that this activation resulted in measurable IL-2 production in 5 of 10 MNC cultures. MNC isolates from most unexposed individuals did not respond to SPAg. To establish which cells responded to SPAg, different subpopulations of MNCs were tested. Only T helper cells were found to respond, and they responded only when cocultured with monocytes. The finding of parasite-specific T helper cells in the blood of malaria-immune individuals and the fact that some of these cells were able to produce IL-2 in vitro support the hypothesis that in malaria the cellular part of the protective immune response is initiated by immune T cells. These cells may activate nonspecific effector cells (i.e., macrophages) that eliminate the parasite. PMID:2941375

Theander, T G; Bygbjerg, I C; Jepsen, S; Svenson, M; Kharazmi, A; Larsen, P B; Bendtzen, K

1986-01-01

119

Recombinant interleukin 2 and gamma-interferon act synergistically on distinct steps of in vitro terminal human B cell maturation.  

PubMed Central

The effects of recombinant interleukin 2 (IL-2) on the in vitro differentiation of human tonsillar B cells which were not preincubated with Staphylococcus aureus Cowan I or with anti-human IgM were investigated. IL-2 was shown to induce the generation of Ig-containing cells in a dose-dependent fashion from 2.5 to 2,500 U IL-2/ml. Conversely, the quantities of Ig secreted in the culture supernatant were found in the majority of experiments to peak at 25 U/ml. The possible presence, in cultures stimulated with IL-2, of cells that were capable of synthesizing Ig but that did not secrete the Ig they have produced was investigated. Among a number of factors tested, we found that gamma-interferon, which did not trigger in vitro B cell differentiation when used alone, can induce an increased secretion of Ig without noticeable change in the number of Ig-containing cells in cultures stimulated with IL-2. The possibility that gamma-interferon and IL-2 act on subsequent steps of in vitro B cell differentiation is discussed. PMID:3082936

Lê thi Bich-Thuy; Fauci, A S

1986-01-01

120

Antitumor activity of tumor-targeted RNA replicase-based plasmid that expresses interleukin-2 in a murine melanoma model  

PubMed Central

Double stranded RNA (dsRNA) has multiple antitumor mechanisms that may be used to control tumor growth. Previously we have shown that treatment of solid tumors with a plasmid that encodes Sindbis viral RNA replicase complex, pSIN-?, significantly inhibited the growth of tumors in mice. In the present study, we evaluated the feasibility of further improving the antitumor activity of the pSIN-? plasmid by incorporating interleukin-2 (IL2) gene into the plasmid. The resultant pSIN-IL2 plasmid was delivered to mouse melanoma cells that overexpress sigma receptor. Here we report that the pSIN-IL2 plasmid was more effective at controlling the growth of B16 melanoma in mice when complexed with sigma receptor-targeted liposomes than with the untargeted liposomes. Importantly, the pSIN-IL2 plasmid was more effective than pSIN-? plasmid at controlling the growth of B16 melanoma in mice, and B16 tumor-bearing mice that were treated with pSIN-IL2 had an elevated number of activated CD4+, CD8+, and natural killer cells, as compared to those treated with pSIN-?. The RNA replicase-based, IL2-expressing plasmid may have application in melanoma gene therapy. PMID:23641783

Rodriguez, B. Leticia; Blando, Jorge M.; Lansakara-P, Dharmika S. P.; Kiguchi, Yuriko; DiGiovanni, John; Cui, Zhengrong

2013-01-01

121

Reactivity of mitogen-induced autorosette-forming cells to interleukin 2 (T-cell growth factor).  

PubMed

The reactivity of mitogen-induced autologous rosette-forming cells (ARFC) to interleukin 2 (IL-2; T-cell growth factor) was studied in the present report. Both ARFC-enriched T cells and ARFC-depleted T cells, which were separated from concanavalin A (Con A)-activated T cells, were reactive to this factor. The IL-2 activity was absorbed by both ARFC-enriched and ARFC-depleted T cells, although ARFC-enriched T cells could absorb more IL-2 activity. Furthermore, ARFC were further inducible by IL-2 from non-ARFC. These results suggest the expression of the receptors for IL-2 on both ARFC and non-ARFC following mitogen stimulation. They further support the possibility that mitogen-induced ARFC, rather than being recruited only from such a minor T-cell subset as the spontaneous ARFC, are more likely the result of most T cells being responsive to mitogenic stimulation and expressing the receptors for autologous erythrocytes by the effects of IL-2 and mitogen. PMID:6600976

Ichikawa, Y; Daniels, J C

1983-02-15

122

Flavone acetic acid (FAA) with recombinant interleukin-2 (TIL-2) in advanced malignant melanoma. II: Induction of nitric oxide production.  

PubMed Central

Plasma samples were collected from 20 patients undergoing phase I clinical trial with flavone-8-acetic acid (FAA; 4.8 g m-2 per dose) in combination with recombinant human interleukin-2 (rhIL-2; 6-18 i.u. m-2 per day) for the treatment of metastatic melanoma. Samples were analysed for nitrate content as an indication of the oxidation of L-arginine to nitric oxide. Pretreatment plasma nitrate levels (53 +/- 4 microM) were significantly above those of healthy volunteers (19 +/- 4 microM). The maximum plasma nitrate concentration obtained after treatment, 190 +/- 29 microM (range 49 to 655 microM), was comparable to that of mice treated with FAA. Most of the increases occurred 3-5 days after initiation of a 5 day infusion of rhIL-2, but three of the increases occurred within 2 days of a 1 h infusion of FAA alone. The maximum plasma nitrate concentrations of the three patients which underwent remission (two complete, one partial) following treatment (368 +/- 143 microM) were significantly higher (P < 0.05) than those of patients with progressive disease. Hypotension was the major dose-limiting side effect, and there was no relationship between the degree of hypotension and the rise in plasma nitrate. The results provide evidence that treatment of patients with FAA and rhIL-2 induce the synthesis of nitric oxide, a physiological mediator and potential cytotoxic agent. PMID:1419615

Thomsen, L. L.; Baguley, B. C.; Rustin, G. J.; O'Reilly, S. M.

1992-01-01

123

The association of -330 interleukin-2 gene polymorphism with its plasma concentration in Iranian multiple sclerosis patients.  

PubMed

Multiple sclerosis (MS) is a chronic neuroinflammatory demyelinating disease of the central nervous system. The cytokine genes are involved in autoimmune diseases such as MS. In this study, we report the influence of -330 interleukin-2 (IL2) gene polymorphism on its plasma levels in a group of Iranian MS patients. In this study 100 MS patients and 100 ethnically, age, and sex matched healthy controls were selected from Medical Genetics Department of Sarem Women Hospital. Blood samples of all individuals were collected in EDTA tubes. The restriction fragment length polymorphism PCR (RFLP) method was applied to determine various alleles and genotypes in these individuals. Plasma concentration of IL2 was measured in all the samples using human IL2 kit. The frequency of -330 T/T IL2 genotype was higher in MS patients compared to normal individuals. Accordingly, the plasma levels of IL2 were significantly higher (P < 0.0001) in patients when compared to the control group. In conclusion, in case of MS patients the -330 T/T IL2 genotype is associated with higher plasma levels of IL2. PMID:24959373

Sayad, Arezou; Movafagh, Abolfazl

2014-01-01

124

The Association of ?330 Interleukin-2 Gene Polymorphism with Its Plasma Concentration in Iranian Multiple Sclerosis Patients  

PubMed Central

Multiple sclerosis (MS) is a chronic neuroinflammatory demyelinating disease of the central nervous system. The cytokine genes are involved in autoimmune diseases such as MS. In this study, we report the influence of ?330 interleukin-2 (IL2) gene polymorphism on its plasma levels in a group of Iranian MS patients. In this study 100 MS patients and 100 ethnically, age, and sex matched healthy controls were selected from Medical Genetics Department of Sarem Women Hospital. Blood samples of all individuals were collected in EDTA tubes. The restriction fragment length polymorphism PCR (RFLP) method was applied to determine various alleles and genotypes in these individuals. Plasma concentration of IL2 was measured in all the samples using human IL2 kit. The frequency of ?330 T/T IL2 genotype was higher in MS patients compared to normal individuals. Accordingly, the plasma levels of IL2 were significantly higher (P < 0.0001) in patients when compared to the control group. In conclusion, in case of MS patients the ?330 T/T IL2 genotype is associated with higher plasma levels of IL2. PMID:24959373

Sayad, Arezou; Movafagh, Abolfazl

2014-01-01

125

Two mutational hotspots in the interleukin-2 receptor gamma chain gene causing human X-linked severe combined immunodeficiency.  

PubMed Central

Human severe combined immunodeficiency (SCID), a syndrome of profoundly impaired cellular and humoral immunity, is most commonly caused by mutations in the X-linked gene for interleukin-2 (IL-2) receptor gamma chain (IL2RG). For mutational analysis of IL2RG in males with SCID, SSCP screening was followed by DNA sequencing. Of 40 IL2RG mutations found in unrelated SCID patients, 6 were point mutations at the CpG dinucleotide at cDNA 690-691, encoding amino acid R226. This residue lies in the extracellular domain of the protein in a region not previously recognized to be significantly conserved in the cytokine receptor gene family, 11 amino acids upstream from the highly conserved WSXWS motif. Three additional instances of mutation at another CpG dinucleotide at cDNA 879 produced a premature termination signal in the intracellular domain of IL2RG, resulting in loss of the SH2-homologous intracellular domain known to be essential for signaling from the IL-2 receptor complex. Mutations at these two hotspots constitute > 20% of the X-linked SCID mutations found by our group and a similar proportion of all reported IL2RG mutations. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7668284

Pepper, A E; Buckley, R H; Small, T N; Puck, J M

1995-01-01

126

Adoptive transfer of anti-cytomegalovirus effect of interleukin-2-activated bone marrow: potential application in transplantation.  

PubMed

This work is a continuation of our studies that showed that interleukin-2 (IL-2)-activated murine bone marrow (ABM) cells have potent cytotoxic potential against murine cytomegalovirus (MCMV)-infected targets in vitro, without loss of reconstitutive ability in vivo. Our data show that ABM cells lyse the MCMV-infected cells in vitro, at both acute and chronic stages of infection; this lysis is specific for the MCMV-infected cells. ABM cells supplemented with IL-2 therapy virtually eradicated the viral infection and prolonged the survival of MCMV-infected Balb/c mice, whether or not they were immunocompromised by irradiation (P less than .001 in both situations). Efficacy of ABM cells alone or IL-2 alone was less than the combination of ABM cells and IL-2. The efficacy of combination treatment with ABM cells and IL-2 in improving the survival of MCMV-infected mice was comparable, whether used in a preventive or a therapeutic setting. Therapy with ABM plus IL-2 also prevented the reactivation of chronic MCMV infection after irradiation. Preliminary findings indicate that Thy-1+ and asialo GM1+ cells limited the MCMV proliferation by approximately 30% and 80%, respectively, while BM macrophages limited the proliferation of MCMV by 100%. These results suggest that BM transplantation (BMT) with ABM cells followed by IL-2 therapy may constitute a novel strategy to improve the host resistance against cytomegalovirus infection after BMT. PMID:1650263

Agah, R; Charak, B S; Chen, V; Mazumder, A

1991-08-01

127

Influence of immunomodulatory drugs on the cytotoxicity induced by monoclonal antibody 17-1A and interleukin-2.  

PubMed

Patients treated with monoclonal antibodies and cytokines for cancer receive often co-medication, which may influence treatment efficacy. Therefore, we investigated with a flowcytometric cytotoxicity assay the effect of several immunomodulatory drugs on antibody dependent cellular cytotoxicity (ADCC), interleukin-2 (IL-2) induced cytotoxicity and IL-2-induced-ADCC. We found that dexamethasone markedly inhibited the IL-2 induced cytotoxicity and the IL-2-induced-ADCC. Ondansetron, a 5-HT-3 serotonin receptor antagonist augmented significantly ADCC. Clemastine, a histamine type-2 receptor antagonist augmented the IL-2-induced-ADCC. The TNF antagonist thalidomide suppressed ADCC whereas pentoxifylline proved to be ineffective. Other tested drugs namely ibuprofen and indomethacin, both prostaglandin E2 antagonists, cimetidine a histamine type-2 receptor antagonist, the opioid pethidine, prostaglandin E2 and histamine exerted minor effects or had no influence on the tested parameters. We conclude that glucocorticosteroids should be avoided with monoclonal antibody and cytokine treatment. According to our in vitro data the other drugs tested did not have a negative impact on cellular cytotoxicity and ADCC. PMID:17562330

Flieger, Dimitri; Varvenne, Michael; Kleinschmidt, Rolf; Schmidt-Wolf, Ingo G H

2007-03-01

128

In vivo cytokine production and recombinant interleukin 2 immunotherapy: an insight into the possible mechanisms underlying clinical responses.  

PubMed Central

Recombinant interleukin 2 (rIL-2), when given to patients with advanced malignant disease, induces a limited beneficial effect, with only 20-30% of patients with solid tumours responding. This present study has identified those patients with advanced colorectal cancer most likely to respond to rIL-2 therapy, by analysis of serum cytokine levels, prior to and during rIL-2 treatment, documented in responders and non-responders. Responders were found to have significantly lower pretreatment serum IL-6 and soluble IL-2 receptor levels (sIL-2R) than non-responders (P < 0.01 and P < 0.05 respectively). During rIL-2 infusion, responders developed high circulating levels of IL-6 and had low constant levels of prostaglandin E2 (PGE2). Non-responders failed to produce IL-6 and demonstrated elevated serum concentrations of PGE2, during infusions of rIL-2. Thus, an enhanced ongoing IL-6 and sIL-2R response, prior to therapy, was detrimental to subsequent treatment with rIL-2. Differential production and/or release of cytokines and prostaglandins, during therapy, further determined the likelihood of response to rIL-2. PMID:8198981

Deehan, D. J.; Heys, S. D.; Simpson, W. G.; Broom, J.; Franks, C.; Eremin, O.

1994-01-01

129

Treatment of metastatic renal cell carcinoma with subcutaneous interleukin 2: evidence for non-renal clearance of cytokines.  

PubMed Central

The circulating cytokine concentrations following administration of subcutaneous recombinant interleukin 2 (IL-2) in combination with interferon alpha and 5-fluorouracil used to treat advanced renal cancer were studied. One patient was anephric and on dialysis, and seven had normal biochemical renal function, although five had undergone single nephrectomy. The pharmacokinetics of IL-2 and changes in IL-6 and tumour necrosis factor (TNF)-alpha were essentially similar in all patients including the anephric patient, irrespective of the periods of dialysis, although at some time points, IL-2 concentrations were slightly higher in the anephric patient than in the others. These results show that for subcutaneous administration of low-dose IL-2, renal clearance of IL-2 is not important. This contrasts with high-dose, intravenous IL-2 where blood concentrations are higher and renal clearance seems to occur, perhaps because of saturation of the non-renal mechanisms of clearance. The subcutaneous route is certainly preferred if IL-2 is used in anephric patients and in those with impaired renal function, and it may be generally preferred for most purposes. PMID:9192992

Banks, R. E.; Forbes, M. A.; Hallam, S.; Jenkins, A.; Wadhwa, M.; Dilger, P.; Meager, A.; Thorpe, R.; Bowmer, C. J.; Joffe, J. K.; Patel, P.; Johnson, P. W.; Selby, P. J.

1997-01-01

130

Subcutaneous low-dose recombinant interleukin 2 and alpha-interferon in patients with metastatic renal cell carcinoma.  

PubMed Central

A double-institution phase II study was performed in patients with metastatic renal cell carcinoma treated subcutaneously (s.c.) with interleukin 2 (IL-2) and alpha-interferon (INF-alpha). Thirty-eight patients were treated over a course of 7 weeks. Initially (day 1 + 2) patients received s.c. IL-2 at 18 x 10(6) IU m-2. During the following 6 weeks, patients received s.c. IL-2 at 3.6 x 10(6) IU m-2 for 5 days per week and s.c. INF-alpha at 5 x 10(6) for 3 days per week. Thirty-eight patients were evaluated for response. An objective response was seen in seven patients (18.4 +/- 12.3%), with one complete response and six partial responses. Median duration of response was 6.7 months. Toxicity could be evaluated in 38 patients and was limited. Mild to moderate toxicity included fever (97%), fatigue or malaise (76%), nausea or vomiting (50%), anorexia (32%), hypotension (26%), neurological disturbances (26%) and hypercreatininaemia (39%). In addition, four grade IV haematological toxicities were noted. No cardiac side-effects were seen. IL-2 and INF-alpha given by this schedule can be safely administered in an outpatient setting. The objective response rate was similar to our previous treatments with high-dose IL-2 given as a continuous infusion. PMID:8198979

Ravaud, A.; Négrier, S.; Cany, L.; Merrouche, Y.; Le Guillou, M.; Blay, J. Y.; Clavel, M.; Gaston, R.; Oskam, R.; Philip, T.

1994-01-01

131

The clinical effects of prolonged treatment of patients with advanced cancer with low-dose subcutaneous interleukin-2 [corrected  

PubMed Central

Thirty-five patients with advanced malignant disease have been treated as outpatients with increasing doses (0.1-100 mcg) of interleukin 2 (IL2) by once daily self-administered subcutaneous (s.c.) injection, 5 days weekly for 8 weeks followed by a 4 week observation period. Systemic side effects were not experienced by patients at the 3 lower doses. Three patients required dose reduction from 100 mcg daily because of intolerance (fever, rash, lethargy, nausea and vomiting) and one patient was discontinued because of dyspnoea. We observed immunological effects at the 100 mcg dose (but not at the lower doses). These consisted of (a) a modest sustained lymphocytosis, (b) eosinophilia in six (out of nine) patients and (c) a significant rise in IL2-stimulated peripheral blood lymphocyte activated killer (LAK) cell activity in six (out of nine) patients to a mean of 2.0 times pretreatment levels (P less than 0.01). Two (out of nine) patients with renal cell carcinoma treated with 100 mcg daily had partial responses of duration 4 and 9 months respectively and a further three had disease stabilisation for at least 3 months. Low dose long-term s.c. IL2 is clinically and immunologically active, and in comparison to other IL2 regimens it has minor toxicity and is easy to administer. These characteristics make low dose s.c. IL2 suitable for study in the adjuvant setting. PMID:1997106

Stein, R. C.; Malkovska, V.; Morgan, S.; Galazka, A.; Aniszewski, C.; Roy, S. E.; Shearer, R. J.; Marsden, R. A.; Bevan, D.; Gordon-Smith, E. C.

1991-01-01

132

Effect of corticosteroid on the antitumor activity of lymphokine-activated killer cells and interleukin 2 in mice.  

PubMed

The adoptive transfer of lymphokine-activated killer (LAK) cells combined with low dose interleukin 2 (IL-2) mediates the regression of established pulmonary metastases in mice and has efficacy in the treatment of human cancer. Systemic administration of high dose IL-2 alone can mediate tumor regression. Cortisone acetate (CA), 25-75 mg/kg, was administered daily to mice receiving high dose IL-2 for 10 days. CA significantly reduced the toxicity induced by IL-2; 38 of 48 mice receiving CA survived compared to 0 of 30 controls (P less than 0.0001). In addition, CA administration caused a decrease in IL-2-induced 125I-labeled albumin leakage in mouse organs. However, CA abrogated the in vivo antitumor effect of high dose IL-2, and to a lesser extent the therapeutic effect of exogenous LAK cells plus lower dose IL-2. Mice treated with 100,000 units of IL-2 showed 98, 63, and 33% reductions of pulmonary metastases in Hanks' balanced salt solution, 25 mg Ca/kg, and 75 mg Ca/kg groups, respectively; treatment with LAK and 7,500 units of IL-2 resulted in reductions of 94, 77, and 57% in these same groups. CA treatment of animals did not affect LAK generation, although the absolute number of LAK precursors was greatly reduced. These results show that although CA can reduce the toxic effect(s) of IL-2, it can be detrimental to successful immunotherapy using this approach. PMID:3489525

Papa, M Z; Vetto, J T; Ettinghausen, S E; Mulé, J J; Rosenberg, S A

1986-11-01

133

Hematologic effects of immunotherapy with lymphokine-activated killer cells and recombinant interleukin-2 in cancer patients.  

PubMed

Immunotherapy with interleukin-2 (IL-2) and lymphokine-activated killer (LAK) cells generated from autologous lymphocytes has produced significant tumor regressions in patients with advanced cancer. In the current study, we reviewed the hematologic effects associated with this therapy in our initial 42 patients. Eighty-eight percent of the treated patients developed anemia that required greater than or equal to 4 units of red cell transfusions, and 43% received at least 8 units. Only a blood loss of 2 to 3 units could be attributed to repeated phlebotomy, cytophereses, and hemodilution. IL-2 administration also resulted in thrombocytopenia as well as lymphopenia and eosinophilia. Forty-three percent of patients developed platelet counts of less than or equal to 50,000/microL, and 36% of the total group required platelet transfusions. Mild neutropenia and a rebound lymphocytosis followed discontinuation of IL-2 treatment. To explore the possible mechanisms for these hematologic effects, standard hematopoietic colony assays were conducted on serial blood samples from five patients. IL-2 produced a significant decline in circulating erythroid (BFU-E) and granulocytic/macrophage (CFU-C) progenitors, which rebounded after the discontinuation of IL-2 therapy. Infusion of IL-2 also resulted in measurable serum levels of gamma-interferon. Some of the hematologic effects of immunotherapy with LAK cells and IL-2 may be the result of IL-2-mediated suppression of hematopoiesis. PMID:3495302

Ettinghausen, S E; Moore, J G; White, D E; Platanias, L; Young, N S; Rosenberg, S A

1987-06-01

134

Subcutaneous administration of interleukin 2 and interferon-alpha-2b in advanced renal cell carcinoma: a confirmatory study.  

PubMed Central

Recent clinical studies have suggested that the combination of subcutaneous recombinant human interleukin 2 (rIL-2) and interferon alpha (rIFN-alpha) is especially promising in advanced renal cell carcinoma. We assessed the safety, activity and toxicity of home therapy with these two agents in 50 patients. Each treatment cycle consisted of a 2 day pulse phase, with 9 x 10(6) IU m-2 of rIL-2 being given subcutaneously every 12 h, followed by a 6 week maintenance phase during which rIL-2 1.8 x 10(6) IU m-2 was administered subcutaneously every 12 h on days 1-5 and rIFN-alpha 2b 5 x 10(6) IU m-2 once a day on days 1, 3 and 5. Objective responses (CR+PR) occurred in 9/50 (18%) patients, six of whom (12%) achieved a complete response. Disease stabilisation was observed in 17 cases (34%) and 18 patients progressed during therapy. In the other six cases, treatment was interrupted early for toxicity or patient refusal. One patient died of myocardial infarction during the second cycle. The overall median survival was 12 months. Home therapy with subcutaneous rIL-2 + rIFN-alpha 2b proved to be active, feasible and moderately toxic, but serious adverse events can sometimes occur. PMID:8519672

Facendola, G.; Locatelli, M. C.; Pizzocaro, G.; Piva, L.; Pegoraro, C.; Pallavicini, E. B.; Signaroldi, A.; Meregalli, M.; Lombardi, F.; Beretta, G. D.

1995-01-01

135

Lymphokine-activated killer cell phenomenon. Lysis of natural killer-resistant fresh solid tumor cells by interleukin 2-activated autologous human peripheral blood lymphocytes  

Microsoft Academic Search

Activation in lectin-free interleukin 2 (IL-2) containing supernatants of peripheral blood mononuclear leukocytes (PBL) from cancer patients or normal individuals resulted in expression of cytotoxicity toward 20 of 21 natural killer (NK)-resistant fresh solid tumor cells tested. Fresh solid tumor cells were resistant to NK-mediated lysis in 10 autologous patients' PBL-tumor interactions, and from 17 normal individuals tested against 13

E. A. Grimm; A. Mazumder; H. Z. Zhang; S. A. Rosenberg

1982-01-01

136

The Immediate-Early Gene Product Egr1 Regulates the Human Interleukin2 Receptor b-Chain Promoter through Noncanonical Egr and Sp1 Binding Sites  

Microsoft Academic Search

The interleukin-2 IL-2 receptor b-chain (IL-2Rb) is an essential component of the receptors for IL-2 and IL-15. Although IL-2Rb is constitutively expressed by lymphocytes, its expression can be further induced by a number of stimuli, including phorbol 12-myristate 13-acetate (PMA). We have now characterized factors that bind to an enhancer region located between nucleotides 2170 and 2139 of the human

JIAN-XIN LIN; WARREN J. LEONARD

1997-01-01

137

Treatment of Endometriosis with Transvaginal Ultrasound-Guided Drainage and Recombinant Interleukin2 Left in the Cysts: A Third Clinical Trial  

Microsoft Academic Search

Background: To analyze the therapeutic results of recombinant interleukin-2 (rIL-2) left in the cysts after transvaginal ultrasound (US)-guided drainage of endometriomas as an alternative to surgery. Methods: Prospective and randomized clinical trial. A total of 25 consecutive patients were included. Two transvaginal US-guided punctures were performed, and 3 million IU of rIL-2 were left in the aspirated cysts once (group

Pedro Acién; Irene Velasco; Maribel Acién; Francisco Quereda

2010-01-01

138

Treatment of Endometriosis with Transvaginal Ultrasound-Guided Drainage under GnRH Analogues and Recombinant Interleukin2 Left in the Cysts  

Microsoft Academic Search

Background: To analyze the therapeutic results of one dose of 3 million IU of recombinant interleukin-2 (rIL-2) left intracyst (group I) versus two doses with a 1-month interval (group II) after transvaginal ultrasound (US)-guided drainage of endometriomas under the effect of GnRH analogues. Methods: Prospective and randomized clinical trial (helped by a random number table) at a University Hospital. Twenty-four

Pedro Acién; Gloria Pérez-Albert; Francisco J. Quereda; Marisa Sánchez-Ferrer; Ana García-Almela; Irene Velasco

2005-01-01

139

GnRH Analogues, Transvaginal Ultrasound-Guided Drainage and Intracystic Injection of Recombinant Interleukin2 in the Treatment of Endometriosis  

Microsoft Academic Search

We performed a double-blind, randomised controlled trial to evaluate the results of ultrasound-guided aspiration of endometriomas under the effect of GnRH analogues and a possible additional beneficial effect by leaving 600,000 IU of recombinant interleukin-2 (rIL-2) in the cysts. Twenty-four women with endometriosis-related symptoms, increased values of CA-125 and transvaginal ultrasonography showing endometriomas >3 cm who were initially sent to

Pedro Acién; Francisco J. Quereda; María-José Gómez-Torres; Rosa Bermejo; Mercedes Gutierrez

2003-01-01

140

Interleukin2-dependent long-term cultures of low-density lymphocytes allow the proliferation of lymphokine-activated killer cells with natural killer, Ti?\\/? or TNK phenotype  

Microsoft Academic Search

Summary We have developed a culture system for “longterm” growth of human lymphokine-activated killer (LAK) cells exhibiting an elevated, wide-spectrum antitumor cytotoxicity. The system allows the exponential growth of monocyte-depleted low-density lymphocytes in the presence of human serum and recombinant human interleukin-2 (103 U\\/ml), alone or in combination with interleukin-1 ? or ? (both at 10 U\\/ml). Eighteen cultures were

U. Testa; A. Care; E. Montesoro; C. Fossati; G. Giannella; R. Masciulli; M. Fagioli; D. Bulgarini; D. Habetswallner; G. Isacchi; P. G. Pelicci; C. Peschle

1990-01-01

141

Cytotoxic cell function and phenotypic analysis of peripheral blood mononuclear cells in cancer patients treated with low-dose interleukin-2 and mitomycin C  

Microsoft Academic Search

We previously found that the ability of peripheral blood mononuclear cells (PBM) of cancer patients to generate lymphokine-activated killer (LAK) cells became remarkably augmented after mitomycin C administration. On the basis of the clinical finding, we designed a treatment regimen comprised of 12 mg\\/m2 mitomycin C i. v. on day 1 and 700 U\\/m2 recombinant interleukin-2 (IL-2) i.v. every 12

Shinya Arinaga; Nobuya Karimine; Masashi Adachi; Hiroshi Inoue; Shigeru Nanbara; Tsukasa Asoh; Hiroaki Ueo; Tsuyoshi Akiyoshi

1993-01-01

142

Association of interleukin-2 and interferon-? production by blood mononuclear cells in infancy with parental allergy skin tests and with subsequent development of atopy  

Microsoft Academic Search

The mechanisms regulating the onset of atopic sensitization in human beings are not yet fully clarified. We assessed the capacity of mitogen-stimulated umbilical and peripheral blood mononuclear cells to produce interferon-? (IFN-?) and interleukin-2 (IL-2) at birth and at 9 months of age in 159 infants. Mononuclear cell production of both IFN-? and IL-2 at 9 months, but not at

Fernando D. Martinez; Debra A. Stern; Anne L. Wright; Catharine J. Holberg; Lynn M. Taussig; Marilyn Halonen

1995-01-01

143

Adoptive immunotherapy of advanced melanoma patients with interleukin-2 (IL2) and tumor-infiltrating lymphocytes selected in vitro with low doses of IL2  

Microsoft Academic Search

Freshly isolated tumor-infiltrating lymphocytes (TIL) from stage IV melanoma patients were cultured for 2 weeks with low doses of interleukin-2 (IL-2; 120 IU\\/ml), to select potentially for tumor-specific lymphocytes present in the neoplastic lesion, followed by high doses (6000 IU\\/ml) to achieve lymphocyte expansion. TIL were serially analyzed for their expansion, phenotype and cytotoxic activity against autologous and allogeneic tumor

Flavio Arientil; Filiberto Belli; Licia Rivoltinil; Carlo Gambacorti-Passerinil; Luigi Furlan; Luigi Mascheroni; Augusto Prada; Maurilia Rizzi; Edoardo Marchesil; Maurizio Vaglini; Giorgio Parmianil; Natale Cascinelli

1993-01-01

144

A dynamic assembly of diverse transcription factors integrates activation and cell-type information for interleukin 2 gene regulation.  

PubMed Central

The interleukin 2 (IL-2) gene is subject to two types of regulation: its expression is T-lymphocyte-specific and it is acutely dependent on specific activation signals. The IL-2 transcriptional apparatus integrates multiple types of biochemical information in determining whether or not the gene will be expressed, using multiple diverse transcription factors that are each optimally activated or inhibited by different signaling pathways. When activation of one or two of these factors is blocked IL-2 expression is completely inhibited. The inability of the other, unaffected factors to work is explained by the striking finding that none of the factors interacts stably with its target site in the IL-2 enhancer unless all the factors are present. Coordinate occupancy of all the sites in the minimal enhancer is apparently maintained by continuous assembly and disassembly cycles that respond to the instantaneous levels of each factor in the nuclear compartment. In addition, the minimal enhancer undergoes specific increases in DNase I accessibility, consistent with dramatic changes in chromatin structure upon activation. Still to be resolved is what interaction(s) conveys T-lineage specificity. In the absence of activating signals, the minimal IL-2 enhancer region in mature T cells is apparently unoccupied, exactly as in non-T lineage cells. However, in a conserved but poorly studied upstream region, we have now mapped several novel sites of DNase I hypersensitivity in vivo that constitutively distinguish IL-2 producer type T cells from cell types that cannot express IL-2. Thus a distinct domain of the IL-2 regulatory sequence may contain sites for competence- or lineage-marking protein contacts. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8790334

Rothenberg, E V; Ward, S B

1996-01-01

145

Cervical and Vulvar Cancer Risk in Relation to Joint Effects of Cigarette Smoking and Genetic Variation in Interleukin 2  

PubMed Central

Cigarette smoking is an established co-factor to human papillomavirus (HPV) in the development of cervical and vulvar squamous cell carcinoma (SCC), and may influence risk through an immunosuppressive pathway. Genetic variation in interleukin 2 (IL2), associated in some studies with inhibition of HPV-targeted immunity, may modify the effect of smoking on the risk of HPV-related anogenital cancers. We conducted a population-based case-only study to measure the departure from a multiplicative joint effect of cigarette smoking and IL2 variation on cervical and vulvar SCC. Genotyping of four IL2 tagSNPs (rs2069762, rs2069763, rs2069777, and rs2069778) was performed in 399 cervical and 486 vulvar SCC cases who had been interviewed regarding their smoking history. Compared to cases carrying the rs2069762 TT genotype, we observed significant departures from multiplicativity for smoking and carriership of the TG or GG genotypes in vulvar SCC risk (interaction odds ratio (IOR)=1.67, 95% confidence interval (CI): 1.16, 2.41). Carriership of one of three diplotypes together with cigarette smoking was associated with either a supra-multiplicative (TGCT/GGCC, IOR=2.09, 95% CI: 0.98, 4.46) or sub-multiplicative (TTCC/TGTC, IOR=0.37, 95% CI: 0.16, 0.85 or TGCT/TGCC, IOR=0.37, 95% CI: 0.15, 0.87) joint effect in vulvar cancer risk. For cervical SCC, departure from multiplicativity was observed for smokers homozygous for the rs2069763 variant allele (TT versus GG or GT genotypes) (IOR=1.87, 95% CI: 1.00, 3.48), and for carriership of the TTCC/TTCC diplotype, (IOR=2.08, 95% CI: 1.01, 4.30). These results suggest that cervical and vulvar SCC risk among cigarette smokers is modified by genetic variation in IL2. PMID:18628433

Hussain, Shehnaz K.; Madeleine, Margaret M.; Johnson, Lisa G.; Du, Qin; Malkki, Mari; Wilkerson, Hui-Wen; Farin, Federico M.; Carter, Joseph J.; Galloway, Denise A.; Daling, Janet R.; Petersdorf, Effie W.; Schwartz, Stephen M.

2008-01-01

146

Interactions between interleukin-2-activated lymphocytes and vascular endothelium: binding to and migration across specialized and non-specialized endothelia.  

PubMed Central

A prerequisite for the successful immunotherapy of solid tumours with interleukin-2 (IL-2)-activated lymphocytes is their ability to home to the tumour tissue. Lymphocyte homing is a complex process which is known to involve at least two independently regulated events: adhesion to the luminal surface of vascular endothelium and the subsequent transendothelial migration of lymphocytes. In this study we have used an in vitro model of lymphocyte homing which employs specialized high endothelium to ask whether IL-2-activated lymphocytes are able to migrate across vascular endothelium in order to leave the blood vessel. Both the adhesion of IL-2-activated cells and their migration across monolayers of cultured high endothelial cells (HEC) were increased in comparison with non-activated lymphocytes. The adhesion of IL-2-activated lymphocytes was mediated by lymphocyte function-associated antigen-1 (LFA-1) and a very late activation antigen-4 (VLA-4)-related pathway. LFA-1-dependent adhesion was mediated by ligands on HEC other than the intercellular adhesion molecule-1 (ICAM-1) and the VLA-4-related pathway was mediated by ligands other than the CS1 domain of fibronectin. HEC-adherent lymphocytes were enriched in natural killer (NK) cells and CD8+ T cells which are known to be the tumour-cytotoxic cells in IL-2-activated lymphocytes. However, there was no evidence of cytotoxicity towards the endothelial layer using a syngeneic model. The interaction of IL-2-activated lymphocytes and endothelial cells was not specific for high endothelium since equal numbers of activated lymphocytes bound to and migrated across aortic endothelium. The inability of IL-2-activated lymphocytes to discriminate between high endothelium and non-specialized 'flat' endothelium could be responsible for the widespread dissemination of the cells throughout the body following their adoptive transfer and the unwanted side-effects at non-involved sites. Images Figure 2 PMID:1398764

Pankonin, G; Reipert, B; Ager, A

1992-01-01

147

Interleukin 2 regulates Raf-1 kinase activity through a tyrosine phosphorylation-dependent mechanism in a T-cell line.  

PubMed Central

Previously we found that interleukin 2 (IL-2) induces tyrosine phosphorylation and activation of the serine/threonine-specific kinase encoded by the raf-1 protooncogene in a T-cell line, CTLL-2. Here we extended these findings by exploring the effects of selective removal of phosphate from tyrosines in p72-74-Raf-1 kinase that had been immunoprecipitated from IL-2-stimulated CTLL-2 cells. Treatment in vitro of IL-2-activated Raf-1 with the tyrosine-specific phosphatases CD45 and TCPTP (formerly called T-cell protein tyrosine phosphatase) reduced Raf kinase activity to nearly baseline levels. This effect was completely inhibited by the phosphatase inhibitor sodium orthovanadate. In contrast, treatment of Raf-1 with a serine/threonine-specific phosphatase, protein phosphatase 1 (PP-1), resulted in a more modest decrease in Raf in vitro kinase activity, and this effect was prevented by okadaic acid. Two-dimensional phosphoamino acid analysis confirmed the selective removal of phosphate from tyrosine by CD45 and from serine and threonine by PP-1. The immunoreactivity of p72-74-Raf-1 with anti-phosphotyrosine antibodies was also completely abolished by treatment with CD45 in the absence but not in the presence of sodium orthovanadate. These findings provide evidence that the IL-2-stimulated phosphorylation of Raf-1 on tyrosines plays an important role in upregulating the activity of this serine/threonine-specific kinase in CTLL-2 cells and, as such, provides a model system for studying the transfer of growth factor-initiated signals from protein tyrosine kinases to serine/threonine-specific kinases. Images Fig. 1 Fig. 2 Fig. 3 PMID:7685905

Turner, B C; Tonks, N K; Rapp, U R; Reed, J C

1993-01-01

148

Active immunization of metastatic melanoma patients with interleukin-2-transduced allogeneic melanoma cells: evaluation of efficacy and tolerability.  

PubMed

From January 1994 to July 1996 we immunized metastatic melanoma patients with HLA-A2-compatible, interleukin-2 (IL-2)-secreting, immunogenic melanoma lines in an attempt to induce a systemic reaction that might also affect distant melanoma lesions. Twelve patients (6 male and 6 female) aged from 28 to 72 years, affected with visceral and/or subcutaneous (s.c.) melanoma metastases, were treated. Two different HLA-A2+ melanoma lines were transduced with the human IL-2 gene (14932/IL-2 and 1B6/IL-2) and used as vaccine. Two groups of 4 patients each were injected s.c. with 5 x 10(7) and 15 x 10(7) irradiated 14932/IL-2 melanoma cells respectively, whereas a third group received 5 x 10(7) cells of the second line (1B6/IL-2). All patients received the vaccine on days 1, 13, 26; if no progression was evident, further immunizations were administered at monthly intervals. All patients were assessable for clinical response after at least three injections of the vaccine. In 4 cases a stabilization of disease lasting from 2 to 6 months was observed: in 2 of them a mixed type of response to treatment was noted with simultaneous evidence of regressing and non-responding lesions in the same patients. No signs of clinical response were found in the remaining patients. Nine patients died of disease between 3 and 24 months after the onset of therapy, whereas 3 were alive 3 months after the end of therapy. The local and systemic side-effects of treatment were mild. These results indicate that vaccination with cells bearing the appropriate antigens and releasing IL-2 locally can produce weak clinical responses, but also indicate that better results may be achieved through modifications of the vaccine, the schedule of immunization and/or a more appropriate selection of patients. PMID:9222277

Belli, F; Arienti, F; Sulé-Suso, J; Clemente, C; Mascheroni, L; Cattelan, A; Santantonio, C; Gallino, G F; Melani, C; Rao, S; Colombo, M P; Maio, M; Cascinelli, N; Parmiani, G; Sanatonio, C

1997-06-01

149

Limited antitumor T cell response in melanoma patients vaccinated with interleukin-2 gene-transduced allogeneic melanoma cells.  

PubMed

We have immunized advanced melanoma patients with a HLA-A2-compatible human melanoma line genetically modified to release interleukin-2 (IL-2), to elicit or increase a T cell-mediated anti-melanoma response that may affect distant lesions. Twelve stage-IV patients were injected subcutaneously at days 1, 13, 26, and 55 with IL-2 gene-transduced and irradiated melanoma cells at doses of 5 or 15 x 10(7) cells. Both local and systemic toxicities were mild, consisting of transient erythema at the vaccination site; fever occurred in a minority of patients. Three mixed responses were recorded. Seven patients were evaluable for immunological studies. Mixed tumor-lymphocyte cultures carried out with different allogeneic HLA-A2-matched melanoma lines as stimulators and targets revealed an increase in the MHC-unrestricted, but no changes in the MHC-restricted, cytotoxicity in peripheral blood lymphocytes (PBL) obtained after vaccination as compared with those obtained before vaccination. Increased recognition of the tyrosinase 368-376 peptide occurred in post-vaccination PBL of one patient, whereas a weak increase in recognition of the gp100 280-288 peptide was detectable in another patient; these 2 patients also recognized the gp100 457-466 peptide. After in vitro, stimulation with the only available autologous melanoma line, CD4+ cells with autologous tumor-specific cytotoxicity and ability to release interferon-gamma (IFN-gamma) were found in post- but not in pre-vaccination PBL. In the same patient, as well as in another patient, limiting dilution analysis showed that vaccination resulted in an increased frequency of melanoma-specific cytotoxic T lymphocyte (CTL) precursors. These results indicate that vaccination with cells releasing IL-2 locally can expand a T cell response against antigen(s) of autologous, untransduced tumor, although this response occurred in a minority of the melanoma patients studied. PMID:8930655

Arienti, F; Sulé-Suso, J; Belli, F; Mascheroni, L; Rivoltini, L; Melani, C; Maio, M; Cascinelli, N; Colombo, M P; Parmiani, G

1996-10-20

150

The 20th anniversary of interleukin-2 therapy: bimodal role explaining longstanding random induction of complete clinical responses  

PubMed Central

Background This year marks the twentieth anniversary of the approval by the US Food and Drug Administration of interleukin-2 (IL2) for use in cancer therapy, initially for renal cell carcinoma and later for melanoma. IL2 therapy for cancer has stood the test of time, with continued widespread use in Europe, parts of Asia, and the US. Clinical complete responses are variably reported at 5%–20% for advanced malignant melanoma and renal cell carcinoma, with strong durable responses and sustained long-term 5–10-year survival being typical if complete responses are generated. Methods The literature was reviewed for the actions and clinical effects of IL2 on subsets of T cells. The influence of IL2 on clinical efficacy was also sought. Results The review revealed that IL2 is capable of stimulating different populations of T cells in humans to induce either T effector or T regulatory responses. This apparent “functional paradox” has confounded a clear understanding of the mechanisms behind the clinical effects that are observed during and following administration of IL2 therapy. An average complete response rate of around 7% in small and large clinical trials using IL2 for advanced renal cell carcinoma and malignant melanoma has been shown from a recent review of the literature. Conclusion This review considers the published literature concerning the actions and emerging clinical effects of IL2 therapy, spanning its 20-year period in clinical use. It further details some of the recently described “bimodal” effects of IL2 to explain the apparent functional paradox, and how IL2 might be harnessed to emerge rapidly as a much more effective and predictable clinical agent in the near future. PMID:22904643

Coventry, Brendon J; Ashdown, Martin L

2012-01-01

151

Analysis of T cell function in autoimmune murine strains. Defects in production and responsiveness to interleukin 2  

PubMed Central

In the studies reported here, we have analyzed the production and consumption of T cell growth factor, more recently termed interleukin 2 (IL-2), as well as some cell-mediated immune functions, in murine strains [MRL, BXSB, NZB, and (NZB x NZWF1] manifesting systemic lupus erythematosus (SLE)-like syndromes. Young (4-6 wk) or old (4-8 mo) autoimmune or normal mice were studied and compared with regard to the following T cell functions in vitro after stimulation with concanavalin A (Con A): (a) mitogenic response; (b) IL-2 levels in culture supernates; and (c) the ability to respond to and adsorb IL-2. In addition, proliferative activity in the allogeneic mixed leukocyte culture and frequency of alloreactive cytotoxic T lymphocyte precursors (CTLp) were analyzed in some of these strains. Reduced Con A-induced mitogenic responses and IL-2 production appeared at 3-6 wk of age in the early, severe SLE developing strains MRL-Mp-lpr/lpr (MRL/l) and male BXSB and progressed thereafter. Similar defects appeared at a later stage in MRL/Mp-+/+ and (NZB x NZW)F1 hybrid mice, which develop late disease. Detailed analysis of cells from the enlarged lymph nodes and spleens of older MRL/l mice demonstrated that such cells: (a) responded poorly to Con A or allogeneic stimulator cells, even in the presence of exogenous IL-2; (b) did not suppress IL-2 production by normal spleen cells; (c) were relatively incapable of adsorbing or inactivating IL-2; and (d) had a markedly reduced anti-H-2b CTLp frequency in the mesenteric lymph nodes but a normal one in spleen. These results indicate that the proliferating Thy-1.2+, Lyt-1+ T cells in MRL/l mice are defective in their responses to mitogenic stimuli, in IL-2 production, and in expression of acceptor sites for IL-2. The relevance of these defects to the MRL/l disease as well as to the role of IL-2 in autoimmunity in general remains to be determined. PMID:6456321

1981-01-01

152

Combined effects of interferon alpha and interleukin 2 on the induction of a vascular leak syndrome in mice.  

PubMed

Immunotherapy with interleukin 2 (IL-2) alone or in combination with lymphokine-activated killer cells can mediate tumor regression in mice and in man. Further dose escalation of IL-2 along with lymphokine-activated killer cells has been prevented by the development of a vascular leak syndrome produced by IL-2. Because we have found that interferon alpha (IFN-alpha) or tumor necrosis factor (TNF-alpha) has synergistic antitumor effects when administered together with IL-2, we have tested the vascular leakage induced by these lymphokine combinations. We used a murine model to quantify vascular leakage by measuring the extravasation of 125I-albumin from the intravascular space as well as the wet and dry lung weights after treatment with different cytokines. Cytokines (or Hanks balanced salt solution) were administered to C57BL/6 mice and 4 h after the last injection the vascular leak was quantified. IFN-alpha alone did not cause extravasation of radiolabel or increase in wet lung weights, though when given in combination with IL-2, significantly greater extravasation (P less than 0.01) as well as increase in lung water weights (P less than 0.05) was observed compared to the response in mice treated with IL-2 alone. IFN-alpha in combination with IL-2 induced significant vascular leakage earlier than the response induced by IL-2 alone. For example treatment with IFN-alpha and IL-2 induced accumulation of 14,674 +/- 605 cpm in the lungs at day 1 while IL-2 alone induced 12,340 +/- 251 cpm. The degree of vascular leakage was highly related to the dose of IFN-alpha administered along with IL-2 and increased vascular leak syndrome was evident even at low doses (5000 units) of IFN-alpha. Immunosuppression of mice by pretreatment irradiation (500 rad) markedly decreased the development of vascular leak syndrome induced by IL-2 and IFN-alpha. Interestingly IFN-gamma and TNF-alpha did not induce vascular leakage in the lungs when given alone, and did not add or synergize with IL-2 in causing the syndrome. Thus the administration of IFN-alpha in combination with IL-2 produces a dose-limiting vascular leakage that is more severe than that caused by IL-2 alone, and may be mediated, directly or indirectly by host radiosensitive cells. PMID:2495179

Puri, R K; Rosenberg, S A

1989-01-01

153

Soluble interleukin-2 receptor is a thyroid hormone-dependent early-response marker in the treatment of thyrotoxicosis.  

PubMed Central

Thyrotoxic patients exhibit increased levels of immune activation molecules (soluble interleukin-2 receptor [sIL-2R], intercellular adhesion molecule-1 [ICAM-1], and endothelial-leukocyte adhesion molecule-1 [ELAM-1]) in serum, although the clinical significance of these measurements remains unclear. In a randomized 4-week study, we have recently shown that in the treatment of hyperthyroidism, the combination of cholestyramine and methimazole (MMI) resulted in faster lowering of serum thyroid-hormone levels than did MMI alone. Stored serial serum samples from patients participating in this randomized treatment trial were analyzed for sIL-2R, soluble ICAM-1 (sICAM-1), and soluble ELAM-1 (sELAM-1). The levels of all three molecules were elevated in patients with hyperthyroidism. Although the levels of sICAM-1 and sELAM-1 remained elevated through the 4-week follow-up period in both groups of patients, the sIL-2R levels (normal levels, 1.0 to 4.2 ng/ml) decreased significantly in the 10 patients who received cholestyramine in addition to MMI (week 0, 14.2 +/- 1.5 ng/ml; week 2, 10.8 +/- 1.2 ng/ml; week 4, 8.9 +/- 1.5 ng/ml). In eight patients who received MMI alone, sIL-2R decreased less rapidly (week 0, 12.3 +/- 1.4 ng/ml; week 2, 12.3 +/- 1.3 ng/ml; week 4, 10.9 +/- 1.3 ng/ml). sICAM-1 and sELAM-1 were elevated at baseline but did not decrease during therapy. In the former group, free thyroxine and free triiodothyronine decreased faster. These data show that levels of sIL-2R in serum, but not those of sICAM-1 and sELAM-1, may be of clinical use in the early follow-up evaluation of medically treated patients. PMID:9302209

Smallridge, R C; Tsokos, G C; Burman, K D; Porter, L; Cranston, T; Sfikakis, P P; Solomon, B L

1997-01-01

154

Adoptive immunotherapy of human pancreatic cancer with lymphokine-activated killer cells and interleukin-2 in a nude mouse model  

SciTech Connect

A pancreatic cancer cell line was grown in orthotopic and heterotopic positions in young Swiss/NIH nude mice, which were tested with adoptive immunotherapy. Mice were injected with 1 x 10(7) human cancer cells in the subcutaneous tissue and duodenal lobe of the pancreas. The mice were randomly divided into four groups: group IA (LAK + IL-2) (N = 25) received 2 X 10(7) human lymphokine-activated killer (LAK) cells from normal donors by tail vein injection followed by 10,000 units of human recombinant interleukin-2 (IL-2) given intraperitoneally every 12 hours for 28 days; group IB (IL-2) (N = 27) was given the same dose of IL-2 alone; group IC (RPMI-1640) (N = 18) received a placebo consisting of 1 ml of RPMI-1640 intraperitoneally every 12 hours; and group ID (LAK) (N = 14) received 2 X 10(7) LAK cells but no IL-2. Toxicity was significantly higher in group IB, with a mortality rate of 45.5% (10/22 animals) versus a 0% mortality (0/25) in group IA. None of the group IA or IB animals died of pancreatic cancer during the experiment. The animals that did not receive IL-2 died before 28 days in 14.2% of group IC and in 16.7% of group ID. The area under the growth curve of subcutaneous tumors during the course of treatment and the pancreatic tumor weight at the end of treatment were compared in each group. Subcutaneous tumors had a reduced rate of growth in group IA animals compared to all the other treatments. Pancreatic tumor growth was slowed in group IA. The animals treated with IL-2 alone (group IB) showed some slowing of tumor growth that was intermediate between group IA, group IC, and group ID. A similar experiment was done with irradiated (375 rad) mice. Nine nude mice with tumors were treated with LAK + IL-2 (group IIA), eight received IL-2 alone (group IIB), and seven received placebo (group IIC).

Marincola, F.M.; Da Pozzo, L.F.; Drucker, B.J.; Holder, W.D. Jr. (Stanford Univ. School of Medicine, CA (USA))

1990-11-01

155

Experience with the use of high-dose interleukin-2 in the treatment of 652 cancer patients.  

PubMed Central

We have administered 1039 courses of high-dose interleukin-2 (IL-2) to 652 cancer patients. Five hundred ninety-six patients had metastatic cancer that either had failed standard effective therapies or had disease for which no standard effective therapy existed, and 56 patients were treated in the absence of evaluable disease in the adjuvant setting. IL-2 was administered either alone (155 patients) or in conjunction with activated immune cells such as lymphokine activated killer (LAK) cells (214 patients) or tumor infiltrating lymphocytes (TIL) (66 patients), with other cytokines such as alpha interferon (a-IFN)(128 patients) or tumor necrosis factor (TNF)(38 patients), with monoclonal antibodies (32 patients), or with the chemotherapeutic agent cyclophosphamide (19 patients). Initial results with the treatment of high-dose IL-2 alone or in conjunction with LAK cells have indicated that objective regressions of cancer can be achieved in 20% to 35% of patients with selected advanced metastatic cancers. Although most responses have been seen in patients with metastatic renal cell cancer, melanoma, colorectal cancer, and non-Hodgkin's lymphoma, many histologic types of cancer have not been treated in significant numbers. These regressions can be durable; of 18 patients achieving a complete response, ten have not experienced recurrence at intervals from 18 to 52 months. Although combinations of IL-2 with TNF do not appear to result in increased responses, there is a suggestion in our initial phase I studies that the combination of a-IFN and IL-2 is more effective than the administration of cytokine alone and this combination deserves further study. Similarly the adoptive transfer of TIL in conjunction with IL-2 also appears to be more effective than the use of IL-2 alone. The toxic side effects in patients treated with high-dose IL-2 are presented and include malaise, nausea and vomiting, hypotension, fluid retention, and organ dysfunction. Treatment-related deaths were seen in 1% of all treatment courses and in 1.5% of patients. These studies demonstrate that a purely immunologic manipulation can mediate the regression of advanced cancers in selected patients and may provide a base for the development of practical, effective biologic treatments for some cancer patients. PMID:2679456

Rosenberg, S A; Lotze, M T; Yang, J C; Aebersold, P M; Linehan, W M; Seipp, C A; White, D E

1989-01-01

156

An Anti-Interleukin-2 Receptor Drug Attenuates T- Helper 1 Lymphocytes-Mediated Inflammation in an Acute Model of Endotoxin-Induced Uveitis  

PubMed Central

The aim of the present study was to evaluate the anti-inflammatory efficacy of Daclizumab, an anti-interleukin-2 receptor drug, in an experimental uveitis model upon a subcutaneous injection of lipopolysaccharide into Lewis rats, a valuable model for ocular acute inflammatory processes. The integrity of the blood-aqueous barrier was assessed 24 h after endotoxin-induced uveitis by evaluating two parameters: cell count and protein concentration in aqueous humors. The histopathology of all the ocular structures (cornea, lens, sclera, choroid, retina, uvea, and anterior and posterior chambers) was also considered. Enzyme-linked immunosorbent assays of the aqueous humor samples were performed to quantify the levels of the different chemokine and cytokine proteins. Similarly, a biochemical analysis of oxidative stress-related markers was also assessed. The inflammation observed in the anterior chamber of the eyes when Daclizumab was administered with endotoxin was largely prevented since the aqueous humor protein concentration substantially lowered concomitantly with a significant reduction in the uveal and vitreous histopathological grading. Th1 lymphocytes-related cytokines, such as Interleukin-2 and Interferon-?, also significantly reduced with related anti-oxidant systems recovery. Daclizumab treatment in endotoxin-induced uveitis reduced Th1 lymphocytes-related cytokines, such as Interleukin-2 and Interferon gamma, by about 60–70% and presented a preventive role in endotoxin-induced oxidative stress. This antioxidant protective effect of Daclizumab may be related to several of the observed Daclizumab effects in our study, including IL-6 cytokine regulatory properties and a substantial concomitant drop in INF?. Concurrently, Daclizumab treatment triggered a significant reduction in both the uveal histopathological grading and protein concentration in aqueous humors, but not in cellular infiltration. PMID:24595020

Navea, Amparo; Almansa, Inmaculada; Muriach, María; Bosch-Morell, Francisco

2014-01-01

157

Induction of myasthenia gravis, myositis, and insulin-dependent diabetes mellitus by high-dose interleukin-2 in a patient with renal cell cancer.  

PubMed

Interleukin-2 is an effective agent against renal cell carcinoma and melanoma, but it has been associated with autoimmune sequelae such as hypothyroidism and vitiligo. A 64-year-old man with non-insulin-dependent diabetes and metastatic renal cell carcinoma developed insulin-dependent diabetes after his first cycle of therapy with high-dose (HD) interleukin-2. After additional therapy with interleukin-2, the patient developed generalized myasthenia gravis (MG) and polymyositis, both of which responded to treatment with corticosteroids and plasmapheresis. To investigate the role of IL-2 in the development of these autoimmune complications, autoantibody titers were assayed from serum obtained before and after IL-2 treatment and after treatment with corticosteroids plus plasmapheresis. Before IL-2 treatment, the patient had antibodies directed against insulin, islet cell antigens, and striated muscle. Acetylcholine receptor antibody levels were normal before starting IL-2. After treatment with IL-2, the patient developed acetylcholine receptor binding antibodies and exhibited an increase in the striated muscle antibody titer from 1:40 to 1:160. Recovery from the MG and polymyositis was associated with substantial decreases in the acetylcholine receptor and striated muscle antibody titers. These findings suggest that HD IL-2 accelerated the progression of latent autoimmune diabetes and myositis in this patient whose tolerance to islet cell antigens and striated muscle had already been broken and precipitated a break in tolerance to the acetylcholine receptor resulting in the development of MG. This case demonstrates the importance of prompt recognition of IL-2-induced MG and shows how this complication can be successfully managed with aggressive therapy. PMID:12142560

Fraenkel, Paula G; Rutkove, Seward B; Matheson, Jean K; Fowkes, Mary; Cannon, Marie E; Patti, Mary-Elizabeth; Atkins, Michael B; Gollob, Jared A

2002-01-01

158

Effect of heavy-ion beam irradiation on the level of serum soluble interleukin-2 receptors in hamster cheek pouch carcinoma model.  

PubMed

Soluble interleukin-2 receptor (sIL-2R) is a glycoprotein derived from ? chain of interleukin 2 receptors of mononuclear as well as T-cell membranes. The aims of this study were to detect the changes of serum soluble interleukin-2 receptor (sIL-2R) levels following heavy-ion beam irradiation in the hamster model with cheek pouch carcinoma, as well as to examine the impact of immune status of the hamster cheek pouch carcinoma model using heavy-ion beam irradiation. sIL-2R serum levels were detected by radioimmunoassay (RIA) in 40 hamsters bearing cheek pouch carcinoma prior to and following exposure to heavy-ion beam irradiation, and 8 normal animals served as the control. The sIL-2R serum level in hamster cheek pouch carcinoma model was significantly increased as compared to the normal control group (P<0.05). Results showed that an increase in the irradiation dose led to a gradual decrease in the sIL-2R serum level. Additionally, a statistical significance was observed compared to the tumor group (P<0.05). In conclusion, alterations in serum sIL-2R expression have an effect on the hamsters cheek pouch carcinoma model subsequent to heavy-ion beam irradiation. An increase in the irradiation dose indicated a decreased tendency in serum sIL-2R content. Detection of serum level changes may lead to an improved understanding of heavy-ion irradiation in vivo immune status, which is crucial for clinical diagnosis and prognosis. It can also provide a sensitive indicator to help estimate the effects of heavy-ion cancer targets. PMID:24748984

An, Xiaoli; Li, Mingxin; Li, Na; Liu, Bin; Zhang, Hong; Wang, Jizeng

2014-05-01

159

Effect of heavy-ion beam irradiation on the level of serum soluble interleukin-2 receptors in hamster cheek pouch carcinoma model  

PubMed Central

Soluble interleukin-2 receptor (sIL-2R) is a glycoprotein derived from ? chain of interleukin 2 receptors of mononuclear as well as T-cell membranes. The aims of this study were to detect the changes of serum soluble interleukin-2 receptor (sIL-2R) levels following heavy-ion beam irradiation in the hamster model with cheek pouch carcinoma, as well as to examine the impact of immune status of the hamster cheek pouch carcinoma model using heavy-ion beam irradiation. sIL-2R serum levels were detected by radioimmunoassay (RIA) in 40 hamsters bearing cheek pouch carcinoma prior to and following exposure to heavy-ion beam irradiation, and 8 normal animals served as the control. The sIL-2R serum level in hamster cheek pouch carcinoma model was significantly increased as compared to the normal control group (P<0.05). Results showed that an increase in the irradiation dose led to a gradual decrease in the sIL-2R serum level. Additionally, a statistical significance was observed compared to the tumor group (P<0.05). In conclusion, alterations in serum sIL-2R expression have an effect on the hamsters cheek pouch carcinoma model subsequent to heavy-ion beam irradiation. An increase in the irradiation dose indicated a decreased tendency in serum sIL-2R content. Detection of serum level changes may lead to an improved understanding of heavy-ion irradiation in vivo immune status, which is crucial for clinical diagnosis and prognosis. It can also provide a sensitive indicator to help estimate the effects of heavy-ion cancer targets. PMID:24748984

AN, XIAOLI; LI, MINGXIN; LI, NA; LIU, BIN; ZHANG, HONG; WANG, JIZENG

2014-01-01

160

Synthesis and Optimization of the Labeling Procedure of 99m Tc-Hynic-Interleukin-2 for In vivo Imaging of Activated T lymphocytes  

Microsoft Academic Search

Introduction  We have previously described the labeling of interleukin-2 (IL2) with 123I and 99mTc-N3S. Both radiopharmaceuticals were successfully applied in humans to image several inflammatory lesions and autoimmune diseases\\u000a characterized by tissue infiltrating lymphocytes expressing the IL2 receptor (CD25). However, both radiopharmaceuticals had\\u000a some specific disadvantages, such as cost and time of synthesis.\\u000a \\u000a \\u000a \\u000a \\u000a Materials and methods  Here, we describe a new improved

Calogero D’Alessandria; Valentina di Gialleonardo; Marco Chianelli; Stephen J. Mather; Erik F. J. de Vries; Francesco Scopinaro; Rudi A. Dierck; Alberto Signore

2010-01-01

161

A modification of the local lymph node assay for contact allergenicity screening: measurement of interleukin-2 as an alternative to radioisotope-dependent proliferation assay.  

PubMed

The local lymph node assay is an effective prediction method for contact allergenicity, but employs radioisotopes. We investigated whether measurement of interleukin-2 (IL-2) production by lymph node cells could be used instead to predict contact allergenicity of chemicals. Test chemicals were applied for three consecutive days to both ears of Balb/c mice and the auricular lymph nodes were obtained on either the fourth or fifth day after the first application. Both IL-2 concentration in supernatant of the suspension and proliferative activity of lymph node cells were determined after 24-, 48-, 72-h cell culture in RPMI-1640 medium by ELISA and by measuring [3H]methylthymidine incorporation, respectively. These two methods detected allergenicity similarly except in the case of TNCB and oxazolone, which showed excessive proliferation-inducing capacity as compared to IL-2 release-increasing effect. Flow cytometry showed that these two chemicals also increased the percentage of Iad-positive cells in the lymph nodes, suggesting that these chemicals might induce not only cellular immunity but also humoral immunity. We conclude that interleukin-2 assay is a convenient and dependable method for screening strong contact allergens without using radioisotopes. PMID:7740543

Hatao, M; Hariya, T; Katsumura, Y; Kato, S

1995-04-12

162

Synthesis and Optimization of the Labeling Procedure of 99mTc-Hynic-Interleukin-2 for In vivo Imaging of Activated T lymphocytes  

PubMed Central

Introduction We have previously described the labeling of interleukin-2 (IL2) with 123I and 99mTc-N3S. Both radiopharmaceuticals were successfully applied in humans to image several inflammatory lesions and autoimmune diseases characterized by tissue infiltrating lymphocytes expressing the IL2 receptor (CD25). However, both radiopharmaceuticals had some specific disadvantages, such as cost and time of synthesis. Materials and methods Here, we describe a new improved method for labeling interleukin-2 with 99mTc using HYNIC-NHS and tricine as coligand. Several optimizations of reagent concentrations and labeling conditions were performed in order to standardize the procedure. After labeling, IL2 was purified by tC2 reverse-phase chromatography and tested in vitro and in vivo, in mice and in a normal volunteer. Results showed that this labeling procedure is cheap, fast, reliable, and reproducible, leading to a product with high specific activity (153 µCi/µg), high stability and capable of binding in vitro to CD25 positive cells. In vivo biodistribution in mice and human volunteer did not show any significant different from 99mTc-N3S-IL2. Conclusion In conclusion, the optimization of 99mTc-HYNIC-IL2 has a great advantage in terms of cost and time of production and a simple kit formulation can be considered for routine application to study patients affected by autoimmune diseases, graft rejection, or other chronic inflammatory disorders. PMID:19949980

D’Alessandria, Calogero; di Gialleonardo, Valentina; Chianelli, Marco; Mather, Stephen J.; de Vries, Erik F. J.; Scopinaro, Francesco; Dierck, Rudi A.

2009-01-01

163

mRNA transcript therapy.  

PubMed

mRNA is the central molecule of all forms of life. It is generally accepted that current life on Earth descended from an RNA world. mRNA, after its first therapeutic description in 1992, has recently come into increased focus as a method to deliver genetic information. The recent solution to the two main difficulties in using mRNA as a therapeutic, immune stimulation and potency, has provided the basis for a wide range of applications. While mRNA-based cancer immunotherapies have been in clinical trials for a few years, novel approaches; including, in vivo delivery of mRNA to replace or supplement proteins, mRNA-based generation of pluripotent stem cells, or genome engineering using mRNA-encoded meganucleases are beginning to be realized. This review presents the current state of mRNA drug technologies and potential applications, as well as discussing the challenges and prospects in mRNA development and drug discovery. PMID:25359562

Weissman, Drew

2015-02-01

164

Adoptive Cell Therapy for Patients with Melanoma, Using Tumor-Infiltrating Lymphocytes Genetically Engineered to Secrete Interleukin-2  

PubMed Central

Adoptive cell transfer of tumor-infiltrating lymphocytes (TILs) after lymphodepletion mediates regression in 50% of patients with metastatic melanoma. In vivo persistence and telomere length of the transferred cells correlate with antitumor response. In an attempt to prolong the in vivo survival of the transferred cells, TILs were genetically engineered to produce interleukin (IL)-2. In vitro, these transduced TILs secreted IL-2 while retaining tumor specificity and exhibited prolonged survival after IL-2 withdrawal. In a phase I/II clinical trial, seven evaluable patients received transduced TILs and one patient experienced a partial response associated with in vivo persistence of IL-2-transduced TILs in circulating lymphocytes. An additional five patients received transduced TILs in conjunction with IL-2 administration. Persistence of IL-2-transduced TILs was observed in three patients, including one partial responder. The transgene DNA as well as vector-derived IL-2 mRNA could be detected for 4 months in responding patients. The low response rate in this trial was possibly due to a reduction in telomere length in cells as a result of prolonged in vitro culture. In this study, insertion of the IL-2 gene into antitumor TILs increased their ability to survive after IL-2 withdrawal in vitro but did not increase their in vivo persistence or clinical effectiveness. PMID:18444786

HEEMSKERK, BIANCA; LIU, KE; DUDLEY, MARK. E.; JOHNSON, LAURA A.; KAISER, ANDREW; DOWNEY, STEPHANIE; ZHENG, ZHILI; SHELTON, THOMAS E.; MATSUDA, KANT; ROBBINS, PAUL F.; MORGAN, RICHARD A.; ROSENBERG, STEVEN A.

2008-01-01

165

Decreased expression of the interleukin 2 receptor on CD8 recipient lymphocytes in intestinal grafts rendered tolerant by liver transplantation in rats  

PubMed Central

Background—In a previous study, it was shown that a spontaneously tolerated DA (RT1a) liver allograft in a PVG (RT1c) recipient was able to induce tolerance of a DA small bowel graft performed 17 days later in spite of infiltration of the intestinal grafts by mononuclear cells. ?Aims—To compare the phenotype of graft infiltrating cells in rejecting and tolerated small bowel grafts in order to elucidate the mechanism(s) which block the graft infiltrating cells from mediating rejection. ?Methods—Multiparameter immunofluorescence was used to compare the phenotype and state of activation of donor and recipient cells isolated from intestinal grafts rejected or tolerated after liver transplantation. ?Results—Three differences were found. Firstly, there was a more rapid replacement of lamina propria (LP) cells by recipient lymphocytes in tolerated than in rejected grafts. Secondly, the proportion of LP recipient CD8??+ lymphocytes bearing the high affinity receptor for interleukin 2 was significantly less in tolerated grafts (1.1%, range 0-2%) than in rejected grafts (21.3%, range 9-26%). Finally, tolerated grafts contained significantly less NK lymphocytes (NKR-P1+) and macrophages than rejected intestinal allografts. ?Conclusions—These observations make it possible to delineate clear cut differences in the phenotype of cells infiltrating rejecting versus tolerated grafts. Furthermore, the data suggest that liver transplantation induces tolerance of intestinal grafts by hampering the activation of recipient TcR??+ CD8??+ T cells and subsequently the recruitment of non-specific effector cells. ?? Keywords: liver transplantation; small bowel transplantation; tolerance; intestinal T lymphocytes; interleukin 2 receptor; rat PMID:9824615

Sarnacki, S; Nakai, H; Calise, D; Azuma, T; Brousse, N; Revillon, Y; Cerf-Bensussan, N

1998-01-01

166

Prolonged survival of a patient affected by pancreatic adenocarcinoma with massive lymphocyte and dendritic cell infiltration after interleukin-2 immunotherapy. Report of a case.  

PubMed

Several studies have shown that there is a paucity of immune cells within the stroma of pancreatic adenocarcinoma, a very aggressive cancer with a median survival of about 18 months. A 65-year-old man presented with jaundice. Abdominal ultrasound revealed intra- and extrahepatic bile duct dilatation and a 45-mm diameter hypoechoic solid mass within the pancreatic head; a computed tomography scan excluded vascular infiltration and metastatic lesions. The patient received immunotherapy consisting of 6,000,000 IU human recombinant interleukin-2 administered subcutaneously twice a day for 3 consecutive days. Thirty-six hours after the last dose, he underwent a pylorus-preserving pancreatoduodenectomy. Because of the presence of high-grade dysplasia detected by intraoperative histological examination of a distal section, a spleen preserving total pancreatectomy was performed. The postoperative course was uneventful. The patient died 32 months after surgery because of local recurrence. Histopathology showed G3 pancreatic ductal adenocarcinoma infiltrating the anterior and posterior peripancreatic tissue, duodenal wall and intrapancreatic common bile duct, with sarcoma-like foci and a component of intraductal tumor involving the common bile duct. In the distal pancreas, widespread foci of pancreatic intraepithelial neoplasia (PanI2-3) were found. The Ki-67 proliferation index was 16%. TNM staging was pT3 pN1 R1. Sections were immunostained for the T-lymphocyte marker CD3 and for the dendritic cell marker CD1a. Intratumoral infiltration was high for CD1a+ cells and mild for CD3+ cells. Preoperative immunotherapy with interleukin-2 may contribute to massive stromal infiltration of immune cells in pancreatic adenocarcinoma. This may prolong the survival even in the presence of negative prognostic factors (age >65 years, tumor diameter >20 mm, R1, tumor grade G3). PMID:18705415

Nobili, Cinzia; Degrate, Luca; Caprotti, Roberto; Franciosi, Claudio; Leone, Biagio Eugenio; Trezzi, Rosangela; Romano, Fabrizio; Uggeri, Fabio; Uggeri, Franco

2008-01-01

167

Cytokine mRNA expression in lesions in cats with chronic gingivostomatitis.  

PubMed

Semiquantitative reverse transcription-PCR assays were developed to measure feline interleukin-2 (IL-2), IL-4, IL-5, IL-6, IL-10, and IL-12 (p35 & p40); gamma interferon (IFN-gamma); and glyceraldehyde-3-phosphate dehydrogenase mRNA concentrations in biopsies of feline oral mucosa. Biopsies were collected from 30 cats with chronic gingivostomatitis (diseased) prior to each cat receiving one of four treatments. In 23 cases replicate biopsies were collected 3 months after treatment commenced. Biopsies were also analyzed from 11 cats without clinical disease (nondiseased). Expression of IL-2, IL-10, IL-12 (p35 and p40), and IFN-gamma was detected in most nondiseased biopsies, while IL-6 was detected in a minority, and IL-4 and IL-5 were both undetectable. Compared to nondiseased cats, the diseased population showed a significant increase in the relative mRNA expression of IL-2, IL-4, IL-6, IL-10, IL-12 (p35 and p40), and IFN-gamma. In contrast, IL-5 mRNA expression was unchanged and was only detected in one case. No significant relationship was demonstrable between the change in relative expression of specific cytokine mRNA and the change in clinical severity of the local mucosal lesions over the treatment period. The results demonstrate that the normal feline oral mucosa is biased towards a predominantly (Th) type 1 profile of cytokine expression and that during the development of lesions seen in feline chronic gingivostomatitis there is a shift in the cytokine profile from a type 1 to a mixed type 1 and type 2 response. PMID:10391845

Harley, R; Helps, C R; Harbour, D A; Gruffydd-Jones, T J; Day, M J

1999-07-01

168

Mechanism of action of glucocorticosteroids. Inhibition of T cell proliferation and interleukin 2 production by hydrocortisone is reversed by leukotriene B4.  

PubMed Central

The mechanism whereby glucocorticosteroids are immunosuppressive is unknown. One potential mechanism of action of these compounds is inhibition of arachidonic acid metabolism. We found that the inhibition of lymphocyte proliferation by hydrocortisone or dexamethasone was mimicked by nonspecific lipoxygenase inhibitors and also by a specific 5-lipoxygenase inhibitor, but not by a specific cyclooxygenase inhibitor. Mitogen-stimulated cultures of T cells produce approximately 5 X 10(-9) M leukotriene B4 (LTB4) in 24 h. This production of LTB4 is completely inhibited by concentrations of hydrocortisone or lipoxygenase inhibitors that inhibit mitogen-induced [3H]thymidine incorporation. The inhibition of lymphocyte proliferation by either hydrocortisone or by the 5-lipoxygenase inhibitor was totally reversed by LTB4 but not by leukotriene C4 or leukotriene D4. LTB4 had no effect on the inhibition of lymphocyte proliferation by noncorticosteroids such as prostaglandin E2, histamine, or gamma-interferon. The inhibition of interleukin 2 (IL-2) production by hydrocortisone or dexamethasone was also completely reversed by exogenous LTB4. LTB4 alone did not cause IL-2 production or cell proliferation when added to resting lymphocytes. Thus, endogenous LTB4 production appears to be necessary but not sufficient for phytohemagglutinin-induced IL-2 production and lymphocyte proliferation. Glucocorticosteroids inhibit IL-2 production and lymphocyte proliferation by inhibiting endogenous LTB4 production. Images PMID:3007577

Goodwin, J S; Atluru, D; Sierakowski, S; Lianos, E A

1986-01-01

169

Recombinant human interleukin-2 reverses in vitro-deficient cell-mediated immune responses to tuberculin purified protein derivative by lymphocytes of tuberculous patients.  

PubMed Central

In vitro lymphocyte proliferative response to purified protein derivative of tuberculin (PPD) was investigated in patients with tuberculosis. Peripheral blood lymphocytes (PBL) from patients with advanced, refractory tuberculosis showed a significantly depressed response compared with the response of PBL from patients with newly diagnosed tuberculosis (P less than 0.01). A further characterization of this low responsiveness to PPD revealed that PBL from these advanced tuberculous patients failed to generate interleukin-2 (IL-2) in response to PPD stimulation. IL-2 receptor (Tac antigen) expression on the surface of T cells after PPD stimulation was also impaired, although to a lesser extent, in the patients with advanced, refractory tuberculosis. We attempted to overcome the depressed in vitro response observed in PBL from patients with advanced, refractory tuberculosis and found that the addition of exogenous, recombinant IL-2 returned the depressed PPD-induced PBL proliferation in these patients to the level of response observed in PBL from patients with newly diagnosed tuberculosis. The addition of recombinant IL-2 also had a restorative effect (up regulation) in vitro on the partly impaired PPD-induced IL-2 receptor expression by PBL from the patients with advanced, refractory tuberculosis. Our results suggest that recombinant IL-2 may offer a novel approach to the therapy of advanced, drug-resistant tuberculosis. PMID:3114146

Shiratsuchi, H; Okuda, Y; Tsuyuguchi, I

1987-01-01

170

Effects of medium quality on the expression of human interleukin-2 at high cell density in fermentor cultures of Escherichia coli K-12.  

PubMed Central

We examined the ability of transformed Escherichia coli cells in fermentor cultures to accumulate interleukin-2 (IL-2) intracellularly under temperature-regulated control of the phage lambda pL promoter. Induction of expression was undertaken at different culture optical densities, and specific IL-2 accumulation was found to decrease with increasing cell density at induction. Induction at higher culture optical densities was also accompanied by decreased growth during induction and increased acetate accumulation in the culture medium. Experiments were undertaken to study the effect of replacing spent medium by perfusion with fresh medium both before induction and during IL-2 expression at high cell density. Improved IL-2 expression was seen only when perfusion was continued past 1.6 h after the start of induction, and it was accompanied by a significant reduction in acetate buildup. Further improvements were not seen when perfusion was continued beyond hour 3 of induction. Replenishing medium components and decreasing the concentration of diffusible inhibitors before induction did not alleviate acetate buildup, growth limitation, or limitation of IL-2 synthesis. These results suggested that accumulation of diffusible inhibitors such as acetate during induction may be a significant factor limiting IL-2 expression in high-density cultures, but other factors intrinsic to the organism or the protein also played a major role. PMID:2180368

MacDonald, H L; Neway, J O

1990-01-01

171

Early Gamma Interferon and Interleukin-2 Responses to Vaccination Predict the Late Resting Memory in Malaria-Naïve and Malaria-Exposed Individuals?  

PubMed Central

Two different cell populations respond to potent T-cell-inducing vaccinations. The induction and loss of effector cells can be seen using an ex vivo enzyme-linked immunospot (ELISPOT) assay, but the more durable resting memory response is demonstrable by a cultured ELISPOT assay. The relationship of the early effector response to durable resting memory is incompletely understood. Effector phenotype is usually identified by gamma interferon (IFN-?) production, but interleukin-2 (IL-2) has been specifically linked to the differentiation of memory cells. Here, IFN-?- and IL-2-secreting effector cells were identified by an ex vivo ELISPOT assay 1 week after vaccination and compared with the resting memory responses detected by a cultured ELISPOT assay 3 months later. The different kinetics and induction of IL-2 by different vaccines and natural exposure are described. Furthermore, both early IFN-? and IL-2 production independently predicted subsequent memory responses at 3 months in malaria-naïve volunteers, but only IFN-? predicted memory in malaria-exposed volunteers. However, dual ELISPOT assays were also performed on malaria-exposed volunteers to identify cells producing both cytokines simultaneously. This demonstrated that double-cytokine-producing cells were highly predictive of memory. This assay may be useful in predicting vaccinations most likely to generate stable, long-term memory responses. PMID:16966412

Bejon, Philip; Keating, Sheila; Mwacharo, Jedidah; Kai, Oscar K.; Dunachie, Susanna; Walther, Michael; Berthoud, Tamara; Lang, Trudie; Epstein, Judy; Carucci, Daniel; Moris, Philippe; Cohen, Joe; Gilbert, Sarah C.; Peshu, Norbert; Marsh, Kevin; Hill, Adrian V. S.

2006-01-01

172

The additional methionine residue at the N-terminus of bacterially expressed human interleukin-2 affects the interaction between the N- and C-termini.  

PubMed

To gain insight into the origin of the difference in isoelectric point (pI) values for wild-type human interleukin-2 (IL-2) and IL-2 with an additional methionine residue at the N-terminus (Met-IL-2), conformational properties of the two molecular forms of IL-2 were compared by utilizing 1H NMR spectroscopy. Although overall conformations were conserved in the two forms, the presence of the additional methionine residue at the N-terminus induced chemical shift changes for residues Ala1 to Lys8 as well as for Thr133, which is located at the C-terminus. These observations indicate that the effect of the additional methionine residue is confined to the N- and C-terminal regions and unveil the existence of an interaction between the N- and C-terminal regions. The chemical shift change observed for Thr133 can be interpreted in terms of a change in pKa of the C-terminal carboxyl group, which interacts differently with the N-terminal amino group in the two forms of IL-2. It seems to be reasonable to conclude that the difference in pI values for the two forms of IL-2 is the consequence of the different interactions between the C- and N-terminal residues. PMID:11170412

Endo, S; Yamamoto, Y; Sugawara, T; Nishimura, O; Fujino, M

2001-01-30

173

65-kilodalton protein phosphorylated by interleukin 2 stimulation bears two putative actin-binding sites and two calcium-binding sites  

SciTech Connect

The authors have previously characterized a 65-kilodalton protein (p65) as an interleukin 2 stimulated phosphoprotein in human T cells and showed that three endopeptide sequences of p65 are present in the sequence of l-plastin. In this paper, they present the complete primary structure of p65 based on the cDNA isolated from a human T lymphocyte (KUT-2) cDNA library. Analysis of p65 sequences and the amino acid composition of cleaved p65 N-terminal peptide indicated that the deduced p65 amino acid sequence exactly coincides with that of l-plastin over the C-terminal 580 residues and has a 57-residue extension at the N-terminus to l-plastin. Computer-assisted structural analysis revealed that p65 is a multidomain molecule involving at least three intriguing functional domains: two putative calcium-binding sites along the N-terminal 80 amino acid residues; a putative calmodulin-binding site following the calcium-binding region; and two tandem repeats of putative actin-binding domains in its middle and C-terminal parts, each containing approximately 240 amino acid residues. These results suggest that p65 belongs to actin-binding proteins.

Zu, Youli; Shigesada, Katsuya; Hanaoka, Masao; Namba, Yuziro (Kyoto Univ. (Japan)); Nishida, Eisuke (Univ. of Tokyo (Japan)); Kubota, Ichiro (Suntory Bio-Pharma Tech Center, Gunma (Japan)); Kohno, Michiaki (Gifu Pharmaceutical Univ., (Japan))

1990-09-11

174

Interleukin 2 production, not the pattern of early T-cell antigen receptor-dependent tyrosine phosphorylation, controls anergy induction by both agonists and partial agonists.  

PubMed Central

Full activation of T cells requires signaling through the T-cell antigen receptor (TCR) and additional surface molecules interacting with ligands on the antigen-presenting cell. TCR recognition of agonist ligands in the absence of accessory signals frequently results in the induction of a state of unresponsiveness termed anergy. However, even in the presence of costimulation, anergy can be induced by TCR partial agonists. The unique pattern of early receptor-induced tyrosine phosphorylation events induced by partial agonists has led to the hypothesis that altered TCR signaling is directly responsible for the development of anergy. Here we show that anergy induction is neither correlated with nor irreversibly determined by the pattern of early TCR-induced phosphorylation. Rather, it appears to result from the absence of downstream events related to interleukin 2 receptor occupancy and/or cell division. This implies that the anergic state can be manipulated independently of the precise pattern of early biochemical changes following TCR occupancy, a finding with implications for understanding the induction of self-tolerance and the use of partial agonist ligands in the treatment of autoimmune diseases. Images Fig. 2 Fig. 4 PMID:8790400

Madrenas, J; Schwartz, R H; Germain, R N

1996-01-01

175

Ultra-low dose interleukin-2 promotes immune-modulating function of regulatory T cells and natural killer cells in healthy volunteers.  

PubMed

Low-dose interleukin-2 (IL-2) expands regulatory T cells (Tregs) and natural killer (NK) cells after stem cell transplantation (SCT) and may reduce graft-versus-host disease (GVHD). We hypothesized that ultra-low dose (ULD) IL-2 could serve as an immune-modulating agent for stem cell donors to prevent GVHD following SCT. However, the safety, dose level, and immune signatures of ULD IL-2 in immune-competent healthy subjects remain unknown. Here, we have characterized the phenotype and function of Tregs and NK cells as well as the gene expression and cytokine profiles of 21 healthy volunteers receiving 50,000 to 200,000 units/m(2)/day IL-2 for 5 days. ULD IL-2 was well tolerated and induced a significant increase in the frequency of Tregs with increased suppressive function. There was a marked expansion of CD56(bright) NK cells with enhanced interferon-? (IFN-?) production. Serum cytokine profiling demonstrated increase of IFN-? induced protein 10 (IP-10). Gene expression analysis revealed significant changes in a highly restricted set of genes, including FOXP3, IL-2RA, and CISH. This is the first study to evaluate global immune-modulating function of ULD IL-2 in healthy subjects and to support the future studies administrating ULD IL-2 to stem cell donors. PMID:24686272

Ito, Sawa; Bollard, Catherine M; Carlsten, Mattias; Melenhorst, Jan Joseph; Biancotto, Angélique; Wang, Ena; Chen, Jinguo; Kotliarov, Yuri; Cheung, Foo; Xie, Zhi; Marincola, Francesco; Tanimoto, Kazushi; Battiwalla, Minoo; Olnes, Matthew J; Perl, Shira; Schum, Paula; Hughes, Thomas E; Keyvanfar, Keyvan; Hensel, Nancy; Muranski, Pawel; Young, Neal S; Barrett, A John

2014-07-01

176

A phase I study of prolonged continuous infusion of low dose recombinant interleukin-2 in melanoma and renal cell cancer. Part II: Immunological aspects.  

PubMed Central

Previously we described the clinical aspects of a phase I study of prolonged continuous infusion of low-dose recombinant interleukin-2 (rIL-2). In the present paper we report several immunological effects in 13 patients with melanoma and renal cell cancer treated on an out-patient basis with rIL-2 for uninterrupted periods ranging from 5 to 18 weeks. Groups of three patients were treated at following dose levels 0.18, 0.6, 1.8 or 6 x 10(6) IU m-2 24 h-1 and one patient was treated with 3 x 10(6) IU m-2 24 h-1. Prolonged rIL-2 treatment resulted in a dose-dependent and sustained increase in the percentage and absolute number of (CD56+, CD8dim) natural killer cells. Within this population a preferential increase in the CD56bright cells with low expression of CD16 was observed. The CD27 antigen was also upregulated in the CD56bright CD16dim population. This increase of NK cells was accompanied by an enhancement of the cytotoxic capacity of the peripheral lymphocytes. No consistent signs of T cell activation or expansion were noted. PMID:8353046

Vlasveld, L. T.; Hekman, A.; Vyth-Dreese, F. A.; Rankin, E. M.; Scharenberg, J. G.; Voordouw, A. C.; Sein, J. J.; Dellemijn, T. A.; Rodenhuis, S.; Melief, C. J.

1993-01-01

177

Recombinant interleukin-2 (rIL-2) given intrasplenically and intravenously for advanced malignant melanoma. A phase I and II study.  

PubMed Central

Recombinant interleukin-2 (rIL-2) was used to treat 31 patients with progressing metastatic malignant melanoma. Only three patients had disease confined to non-visceral sites; the median number of organ sites involved was four. The first dose of rIL-2 was given intrasplenically (to stimulate cytotoxic cells in high concentration) via a femoral artery catheter, and four further i.v. doses were given over 6 days. A total of three courses at 21-day intervals was planned. Doses were escalated in 15 patients from 1 x 10(6) to 16.4 x 10(4) Cetus units m-2. The maximum tolerated dose (11.0 x 10(6) U m-2) was used in the other 16 patients. Of the 71 courses, severe but transient toxicity requiring interruption of rIL-2 or additional care occurred on three courses (dyspnoea) and 15 from hypotension, but the patients' performance status improved. Four patients had partial tumour responses although in only one patient did response occur in all sites of disease. However, responses occurred in visceral sites and six patients are alive at 9-16 months. IL-2 is of use in advanced melanoma and does not need complicated ICU facilities. PMID:2803954

Thatcher, N.; Dazzi, H.; Johnson, R. J.; Russell, S.; Ghosh, A. K.; Moore, M.; Chadwick, G.; Craig, R. D.

1989-01-01

178

Tumor necrosis factor. alpha. induces proteins that bind specifically to. kappa. B-like enhancer elements and regulate interleukin 2 receptor. alpha. -chain gene expression in primary human T lymphocytes  

Microsoft Academic Search

The authors have investigated the biochemical basis for the activation of interleukin 2 receptor α-subunit (IL-2Rα) gene expression in primary human T lymphocytes by a cytokine (tumor necrosis factor α), a T-cell mitogen (phorbol 12-myristate 13-acetate), and the transactivator protein (Tax) from the type I human T-cell leukemia virus. Using in vivo transfection techniques specifically designed for these primary T

J. W. Lowenthal; D. W. Ballard; E. Boehnlein; W. C. Greene

1989-01-01

179

Tumor Necrosis Factor alpha Induces Proteins that Bind Specifically to kappa B-Like Enhancer Elements and Regulate Interleukin 2 Receptor alpha Chain Gene Expression in Primary Human T Lymphocytes  

Microsoft Academic Search

We have investigated the biochemical basis for the activation of interleukin 2 receptor alpha -subunit (IL-2Ralpha ) gene expression in primary human T lymphocytes by a cytokine (tumor necrosis factor alpha ), a T-cell mitogen (phorbol 12-myristate 13-acetate), and the transactivator protein (Tax) from the type I human T-cell leukemia virus. Using in vivo transfection techniques specifically designed for these

John W. Lowenthal; Dean W. Ballard; Ernst Bohnlein; Warner C. Greene

1989-01-01

180

Intensive chemotherapy with idarubicin, ara-C, etoposide, and m-AMSA followed by immunotherapy with interleukin-2 for myelodysplastic syndromes and high-risk Acute Myeloid Leukemia (AML)  

Microsoft Academic Search

Intensive chemotherapy followed by treatment with interleukin-2 (IL-2) was evaluated in a prospective, randomized, multicenter\\u000a trial including 18 patients with refractory anemia with excess of blasts in transformation (RAEB-T), 86 patients with acute\\u000a myeloid leukemia (AML) evolving from myelodysplastic syndromes, and six patients with secondary AML after previous chemotherapy.\\u000a Median age was 58?years (range: 18–76?years). Forty-nine patients (45%) achieved a

A. Ganser; G. Heil; G. Seipelt; W. Hofmann; J. T. Fischer; W. Langer; W. Brockhaus; K. Kolbe; T. H. Ittel; N. Brack; H. G. Fuhr; P. Knuth; K. Höffken; L. Bergmann; D. Hoelzer

2000-01-01

181

Interleukin 2 induces both, growth and maturation of lectin reactive Lyt-2+ but not Lyt-2-precursor cells and regulates the cytolytic potential of effector cells.  

PubMed

This study investigated the requirements for lymphokines derived by recombinant (rec.) DNA technology for the induction of growth and maturation in highly purified lectin reactive T cell subsets. Nylon purified C57BL/6 lymph node T cells were treated with monoclonal anti-Lyt-2.2 or anti-L3T4 antibodies and fluorescence labeled (FITC) anti-immunoglobulin antibodies and were positively selected into Lyt-2+ (L3T4-) and Lyt-2- (L3T4+) lymphocyte subsets using a fluorescence-activated cell sorter. Sorted T lymphocytes, which were devoid of accessory cells were incubated either in bulk culture (2 X 10(2) - 3 X 10(4) cells/microculture) or under limiting dilution conditions (2.5-1,000 cells/well) with lectin (Concanavalin A, Leukoagglutinin) and rec. human Interleukin 2 (rec. hIL-2) and/or rec. mouse Interferon gamma (rec. mIFN-gamma). The data show that Lyt-2+ lymphocytes respond to lectin and rec. hIL-2 with growth and development of cytolytic activity in the absence of other exogenous factor(s) or accessory cells. The presence of monoclonal antibodies to the Interleukin 2 receptor during the sensitization phase ablated the induction of Con A reactive precursor cells of cytolytic lymphocytes (CTL-P) by either rec. hIL-2 or conventional IL-2 containing lymphokine sources, indicating the essential role of IL-2 during activation of Lyt-2+ T lymphocytes. In contrast, Lyt-2- lymphocytes could not be induced by lectin and rec. hIL-2 alone for proliferation and always required the presence of accessory cells for significant growth. Exogenous rec. m IFN gamma was unable to induce growth and cytolytic activity in Con A reactive Lyt-2+ cells and did not significantly effect their response to rec. hIL-2. Limiting dilution experiments revealed that 10-16% of the Lyt-2+ lymphocytes responded to Con A and rec. hIL-2 with growth (GTL-P). The frequencies of CTL-P, determined under similar conditions, were always lower compared to GTL-P. However the results suggest that the differences observed between both precursor populations is due to differential sensitivity of the detection system rather than to the recruitment of distinct T cell subsets. Furthermore, it was shown that at least 50% of lectin reactive CTL-P were induced by rec. hIL-2 to secrete IFN-gamma under optimal conditions. The finding that some of the conventional lymphokine sources were superior to rec. hIL-2 in the induction of growth and cytolytic activity suggests the existence of mediators distinct from IL-2 that regulate the expansion of CTL-P.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3086217

Hochgeschwender, U; Diamantstein, T; Prester, M; Nerz, G; Simon, M M

1986-04-01

182

Correlation between density of beta 2-adrenergic receptors on peripheral blood mononuclear cells and serum levels of soluble interleukin-2 receptors in patients with chronic inflammatory diseases.  

PubMed

In order to study the influence of the autonomic nervous system on the immune response, we evaluated the density and the equilibrium dissociation constants (KD) of beta 2-adrenergic receptors (beta 2R) on peripheral blood mononuclear cells (PBMC) in patients with rheumatoid arthritis (RA), Crohn's disease (CD), and in controls. Results were correlated with the serum concentrations of soluble interleukin-2 receptors (sIL-2R) as a marker for T-cell activation in vivo. The density of beta 2R was significantly decreased in patients with RA (P < 0.05) and CD (P < 0.05) as compared with controls. The number of beta 2R in patients with RA was significantly lower than in CD patients (P < 0.05). KD values of beta 2R were markedly but not significantly decreased in both patient groups as compared with control values. Serum concentrations of sIL-2R were significantly elevated in RA patients as compared with those in CD patients (P < 0.05) and controls (P < 0.05), while there was no difference between the latter two groups. In patients with RA, a significant negative correlation between beta 2R density and serum IL-2R levels (r = -0.66, P < 0.02) was observed. These results demonstrate the close correlation between the modulation of beta 2R on PBMC and the activation of the immune response. However, the role of beta 2R stimulation in the pathogenesis of immunologically mediated diseases remains to be clarified. PMID:1333967

Krause, A; Henrich, A; Beckh, K H; Von Wichert, P; Baerwald, C

1992-10-01

183

Retrospective analysis of the safety and efficacy of high-dose interleukin-2 after prior tyrosine kinase inhibitor therapy in patients with advanced renal cell carcinoma.  

PubMed

Although tyrosine kinase inhibitors (TKI) are the most common first-line therapy for metastatic renal cell carcinoma, high-dose interleukin-2 (HD-IL2) remains the only agent that provides durable complete responses. The optimal sequence of these agents remains uncertain. This retrospective multi-institutional study examined the safety and efficacy of HD-IL2 following TKI therapy. After IRB approval at 7 HD-IL2 centers, data relating to patient, disease, and treatment characteristics among 40 consecutive patients with metastatic renal cell carcinoma who were treated with HD-IL2 after at least 1 prior TKI therapy were retrospectively collected. The most common cardiac adverse events were grade 3 hypotension and vascular leak syndrome. Six patients (15%) experienced other grade ?3 cardiac adverse events. There were 2 treatment-related deaths due to congestive heart failure, occurring in 1 patient with short TKI to HD-IL2 interval and another patient with an abnormal baseline cardiac stress test. Best responses included 2 CRs (5%, duration 40+ and 62+ mo), 3 PRs (8%, duration 6, 11, and 24 mo), 13 SD (32%, median duration 12 mo), 20 PD (50%), and 2 not evaluable patients. Median overall survival was 22 months. Administration of HD-IL2 could be safe and effective after TKI therapy; however, careful selection of patients is critical. We recommend baseline cardiac risk factor assessment, screening with both cardiac stress test and echocardiogram, and allowing a TKI to HD-IL2 interval of at least 2 months. PMID:25075565

Lam, Elaine T; Wong, Michael K K; Agarwal, Neeraj; Redman, Bruce G; Logan, Theodore; Gao, Dexiang; Flaig, Thomas W; Lewis, Karl; Poust, Jamie; Monk, Paul; Jarkowski, Anthony; Sendilnathan, Arun; Bolden, Marcus; Kuzel, Timothy M; Olencki, Thomas

2014-09-01

184

Influence of Foreign DNA Introduction and Periplasmic Expression of Recombinant Human Interleukin-2 on Hydrogen Peroxide Quantity and Catalase Activity in Escherichia coli  

PubMed Central

Purpose: Oxidative stress is generated through imbalance between composing and decomposing of reactive oxygen species (ROS). This kind of stress was rarely discussed in connection with foreign protein production in Escherichia coli. Effect of cytoplasmic recombinant protein expression on Hydrogen peroxide concentration and catalase activity was previously reported. In comparison with cytoplasm, periplasmic space has different oxidative environment. Therefore, in present study we describe the effect of periplasmic expression of recombinant human interleukin-2 (hIL-2) on H2O2 concentration and catalase activity in Escherichia coli and their correlation with cell growth. Methods: Having constructed pET2hIL2 vector, periplasmic expression of hIL-2 was confirmed. Then, H2O2 concentration and catalase activity were determined at various ODs. Wild type and empty vector transformed cells were used as negative controls. Results: It was shown that H2O2 concentration in hIL-2 expressing cells was significantly higher than its concentration in wild type and empty vector transformed cells. Catalase activity and growth rate reduced significantly in hIL-2 expressing cells compared to empty vector transformed and wild type cells. Variation of H2O2 concentration and catalase activity is intensive in periplasmic hIL-2 expressing cells than empty vector containing cells. Correlation between H2O2 concentration elevation and catalase activity reduction with cell growth depletion are also demonstrated. Conclusion: Periplasmic expression of recombinant hIL-2 elevates the host cell’s hydrogen peroxide concentration possibly due to reduced catalase activity which has consequent suppressive effect on growth rate. PMID:24312866

Mahmoudi Azar, Lena; Mehdizadeh Aghdam, Elnaz; Karimi, Farrokh; Haghshenas, Babak; Barzegari, Abolfazl; Yaghmaei, Parichehr; Hejazi, Mohammad Saeid

2013-01-01

185

Retrospective Analysis of the Safety and Efficacy of High-dose Interleukin-2 After Prior Tyrosine Kinase Inhibitor Therapy in Patients With Advanced Renal Cell Carcinoma  

PubMed Central

Although tyrosine kinase inhibitors (TKI) are the most common first-line therapy for metastatic renal cell carcinoma, high-dose interleukin-2 (HD-IL2) remains the only agent that provides durable complete responses. The optimal sequence of these agents remains uncertain. This retrospective multi-institutional study examined the safety and efficacy of HD-IL2 following TKI therapy. After IRB approval at 7 HD-IL2 centers, data relating to patient, disease, and treatment characteristics among 40 consecutive patients with metastatic renal cell carcinoma who were treated with HD-IL2 after at least 1 prior TKI therapy were retrospectively collected. The most common cardiac adverse events were grade 3 hypotension and vascular leak syndrome. Six patients (15%) experienced other grade ?3 cardiac adverse events. There were 2 treatment-related deaths due to congestive heart failure, occurring in 1 patient with short TKI to HD-IL2 interval and another patient with an abnormal baseline cardiac stress test. Best responses included 2 CRs (5%, duration 40+ and 62+ mo), 3 PRs (8%, duration 6, 11, and 24 mo), 13 SD (32%, median duration 12 mo), 20 PD (50%), and 2 not evaluable patients. Median overall survival was 22 months. Administration of HD-IL2 could be safe and effective after TKI therapy; however, careful selection of patients is critical. We recommend baseline cardiac risk factor assessment, screening with both cardiac stress test and echocardiogram, and allowing a TKI to HD-IL2 interval of at least 2 months. PMID:25075565

Wong, Michael K. K.; Agarwal, Neeraj; Redman, Bruce G.; Logan, Theodore; Gao, Dexiang; Flaig, Thomas W.; Lewis, Karl; Poust, Jamie; Monk, Paul; Jarkowski, Anthony; Sendilnathan, Arun; Bolden, Marcus; Kuzel, Timothy M.; Olencki, Thomas

2014-01-01

186

Evidence for a structural motif in toxins and interleukin-2 that may be responsible for binding to endothelial cells and initiating vascular leak syndrome  

PubMed Central

The dose-limiting toxicity of interleukin-2 (IL-2) and immunotoxin (IT) therapy in humans is vascular leak syndrome (VLS). VLS has a complex etiology involving damage to vascular endothelial cells (ECs), extravasation of fluids and proteins, interstitial edema, and organ failure. IL-2 and ITs prepared with the catalytic A chain of the plant toxin, ricin (RTA), and other toxins, damage human ECs in vitro and in vivo. Damage to ECs may initiate VLS; if this damage could be avoided without losing the efficacy of ITs or IL-2, larger doses could be administered. In this paper, we provide evidence that a three amino acid sequence motif, (x)D(y), in toxins and IL-2 damages ECs. Thus, when peptides from RTA or IL-2 containing this sequence motif are coupled to mouse IgG, they bind to and damage ECs both in vitro and, in the case of RTA, in vivo. In contrast, the same peptides with a deleted or mutated sequence do not. Furthermore, the peptide from RTA attached to mouse IgG can block the binding of intact RTA to ECs in vitro and vice versa. In addition, RTA, a fragment of Pseudomonas exotoxin A (PE38-lys), and fibronectin also block the binding of the mouse IgG-RTA peptide to ECs, suggesting that an (x)D(y) motif is exposed on all three molecules. Our results suggest that deletions or mutations in this sequence or the use of nondamaging blocking peptides may increase the therapeutic index of both IL-2, as well as ITs prepared with a variety of plant or bacterial toxins. PMID:10097145

Baluna, Roxana; Rizo, Josep; Gordon, Brian E.; Ghetie, Victor; Vitetta, Ellen S.

1999-01-01

187

Oxazolone and ethanol induce colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2R?null mice engrafted with human peripheral blood mononuclear cells  

PubMed Central

Oxazolone-induced colitis in mice has become a recognized model to study the efficacy of therapeutics targeting the immunological response underlying the development of inflammatory bowel disease. However, this model cannot be used when therapeutics designed to address human targets do not interact with the respective murine counterpart. In this study, we examined the induction of oxazolone mediated colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2R?null (NOD-SCID IL2R?null) mice engrafted with human peripheral blood mononuclear cells (hPBMC) derived from patients suffering from ulcerative colitis (UC), atopic dermatitis (AD) and healthy volunteers. NOD-SCID IL2R? null mice were engrafted with hPBMC followed by challenge with oxazolone or ethanol vehicle. Mice developed the same symptoms as observed previously in immunocompetent mice. The clinical activity score increased and the colon architecture was characterized by the development of oedema, fibrosis, crypt loss and dense infiltration of predominantly T cells into the lamina propria. Fluorescence activated cell sorter (FACS) analysis of lymphocytes in the colon identified natural killer (NK) T cells as a major constituent. In contrast to studies with immunocompetent mice, we observed the same phenotype in the group challenged with ethanol vehicle. The phenotype was most pronounced in mice engrafted with PBMC derived from a patient suffering from UC, suggesting that the immunological history of the donors predisposes the engrafted mice to react to ethanol. The model described here has the potential to study the efficacy of therapeutics targeting human lymphocytes in a model which is more reflective of the human disease. In addition, it might be developed to elucidate molecular mechanisms underlying the disease. PMID:23574330

Nolte, T; Zadeh-Khorasani, M; Safarov, O; Rueff, F; Gülberg, V; Herbach, N; Wollenberg, A; Mueller, T; Siebeck, M; Wolf, E; Gropp, R

2013-01-01

188

Anti-L3T4 antibody treatment suppresses hepatic granuloma formation and abrogates antigen-induced interleukin-2 production in Schistosoma mansoni infection.  

PubMed Central

In murine schistosomiasis mansoni, granulomatous inflammation is an immune response that involves egg antigen presentation to T cells in the context of class II major histocompatibility complex determinants and subsequent inflammatory lymphokine production by delayed-hypersensitivity (TDH) lymphocytes. In the present study, monoclonal antibodies directed against L3T4, I-A, and Lyt-2 molecules were injected intraperitoneally into S. mansoni-infected mice to study the role of these membrane antigens in the process of granuloma formation. A dramatic suppression of the hepatic granuloma size and antigen-induced interleukin-2 (IL-2) production by spleen cells was seen in mice that received anti-L3T4 monoclonal antibody treatment. The total number of cells, especially the L3T4+ T cells, was greatly diminished in the spleens. Furthermore, histopathological study of the granulomas in stained liver sections demonstrated the paucity of eosinophils and macrophages, absence of epithelioid cells and multinucleated giant cells, and minimal collagen deposition within the lesions. Damaged hepatocytes were also seen surrounding these ill-formed granulomas. In contrast, anti-I-A monoclonal antibody treatment partially suppressed IL-2 production, although granuloma size and cellular composition remained the same. Mice that received anti-Lyt-2 monoclonal antibody did not show any changes in either IL-2 production or hepatic granulomatous inflammation. The data presented in this paper indicate a crucial role for L3T4 molecules present on a subset of class II major histocompatibility complex-restricted TDH cells in IL-2 production and the generation of the granulomatous response. Images PMID:3096893

Mathew, R C; Boros, D L

1986-01-01

189

Human Leukocyte Antigen and Interleukin 2, 10 and 12p40 Cytokine Responses to Measles: Is There Evidence of the HLA Effect?  

PubMed Central

HLA class I and class II associations were examined in relation to measles virus-specific cytokine responses in 339 healthy children who had received two doses of live attenuated measles vaccine. Multivariate linear regression modeling analysis revealed suggestions of associations between the expression of DPA1*0201 (p=0.03) and DPA1*0202 (p=0.09) alleles and interleukin-2 (IL-2) cytokine production (global p-value 0.06). Importantly, cytokine production and DQB1 allele associations (global p-value 0.04) revealed that the alleles with the strongest association with IL-10 secretion were DQB1*0302 (p=0.02), DQB1*0303 (p=0.07) and DQB1*0502 (p=0.06). Measles-specific IL-10 secretion associations approached significance with DRB1 and DQA1 loci (both global p-values 0.08). Specifically, suggestive associations were found between DRB1*0701 (p=0.07), DRB1*1103 (p=0.06), DRB1*1302 (p=0.08), DRB1*1303 (p=0.06), DQA1*0101 (p=0.08), and DQA1*0201 (p=0.04) alleles and measles-induced IL-10 secretion. Further, suggestive association was observed between specific DQA1*0505 (p=0.002) alleles and measles-specific IL-12p40 secretion (global p-value 0.09) indicating that cytokine responses to measles antigens are predominantly influenced by HLA class II genes. We found no associations between any of the alleles of HLA A, B, and Cw loci and cytokine secretion. These novel findings suggest that HLA class II genes may influence the level of cytokine production in the adaptive immune responses to measles vaccine. PMID:17234427

Ovsyannikova, Inna G.; Ryan, Jenna E.; Jacobson, Robert M.; Vierkant, Robert A.; Pankratz, V. Shane; Poland, Gregory A.

2007-01-01

190

[Significance of soluble interleukin-2 receptor alpha chain in the management of patients with malignant lymphoma: a multi-center study].  

PubMed

A multi-center series of 117 patients with malignant lymphoma were analyzed to evaluate the clinical significance of soluble interleukin-2 receptor alpha chain (sIL-2R alpha). The initial levels of sIL-2R alpha ranged from 277 U/ml to 22,800 U/ml with a mean level of 3,451 +/- 4,268 U/ml and a median level of 1,600 U/ml. The sIL-2R alpha levels of the diffuse lymphoma/intermediate-grade subtypes defined by the LSG classification/Working Formulation were higher than those of the follicular lymphoma/low-grade subtypes. There was a tendency for B-cell lymphomas to show higher sIL-2R alpha levels than T-cell lymphomas. The sIL-2R alpha level was correlated with the Ann Arbor clinical stage (I, II versus III, IV), presence or absence of B symptoms, and performance status (0, 1 versus 2, 3, 4) of the patients. The sIL-2R alpha levels were in good accordance with the four risk groups defined by the International Prognostic Indices. Of 21 patients whose tumor burden was serially measured, the coefficients of correlation between sIL-2R alpha and tumor mass were > 0.6 in 18 cases. Sixty-two patients achieved complete remission (CR) during the study; the initial and minimum sIL-2R alpha levels were lower than those of the non-CR patients. This study confirmed that sIL-2R alpha is a convenient and useful marker in the management of malignant lymphoma. PMID:11979748

Ohno, Hitoshi; Ishikawa, Takayuki; Kitajima, Hiroyuki; Nomura, Shosaku; Suzuki, Takayo; Konishi, Hiroshi; Ohno, Yoichiro; Onishi, Rie; Konaka, Yoshiteru; Arima, Nobuyoshi; Doi, Shoichi; Nasu, Kaori; Takahashi, Takayuki; Tsudo, Mitsuru; Fukuhara, Shirou; Uchiyama, Takashi

2002-03-01

191

Use of Antigen-Specific Interleukin-2 To Differentiate between Cattle Vaccinated with Mycobacterium bovis BCG and Cattle Infected with M. bovis  

PubMed Central

We describe here the application of a novel bovine interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) for the measurement of antigen-specific IL-2 in cattle naturally infected with Mycobacterium bovis and in cattle vaccinated with Mycobacterium bovis BCG and then experimentally challenged with pathogenic M. bovis. Supernatants from whole-blood cultures stimulated with mycobacterial antigen (bovine purified protein derivative [PPDB] or the peptide cocktail ESAT6-CFP10) were assessed using a sandwich ELISA consisting of a new recombinant monoclonal fragment capture antibody and a commercially available polyclonal anti-bovine-IL-2. The production of IL-2 was compared to the production of gamma interferon (IFN-?) in the same antigen-stimulated whole-blood supernatants. The data show that cattle infected with M. bovis produced quantifiable levels of antigen-specific IL-2, while IL-2 levels in cattle vaccinated with M. bovis BCG did not. Furthermore, cattle vaccinated with M. bovis BCG and then challenged with pathogenic M. bovis displayed a more rapid induction of IL-2 but ultimately had lower levels of infection-induced IL-2 than did unvaccinated challenge control cattle. These data suggest that IL-2 responses are not detectable post-BCG vaccination and that these responses may require infection with virulent M. bovis to develop. This may be useful to differentiate infected cattle from uninfected or BCG-vaccinated cattle, although the overall sensitivity is relatively low, particularly in single intradermal comparative cervical tuberculin (SICCT)-negative infected animals. Furthermore, the strength of the IL-2 response may correlate with pathology, which poses interesting questions on the immunobiology of bovine tuberculosis in contrast to human tuberculosis, which is discussed. PMID:24173026

McKinna, Lucy C.; Steinbach, Sabine; Dean, Gilly S.; Villarreal-Ramos, Bernardo; Whelan, Adam O.; Pirson, C.; Jones, Gareth J.; Clifford, Derek; Vordermeier, H. Martin

2014-01-01

192

Effects of in vivo administration of interleukin-2 (IL-2) and IL-4, alone and in combination, on ex vivo human basophil histamine release.  

PubMed

Human basophils possess receptors for interleukin-2 (IL-2) and IL-4. The effect of 3 days of intravenous administration of IL-2 and/or IL-4 on basophil histamine release was examined in three groups of patients receiving IL-2, IL-4, or the combination of agents as part of a protocol to treat malignant melanoma or renal cell carcinoma. Because all patients received ranitidine for control of side effects, a control group of patients receiving ranitidine for Zollinger-Ellison's syndrome was also studied. IL-4 significantly inhibited IgE-mediated histamine release, while there was a trend for enhancement of IgE-mediated histamine release by IL-2. Administration of the combination of IL-2 and IL-4 did not alter IgE-mediated basophil histamine release. Both IL-2 and IL-4, alone and in combination, enhanced basophil histamine release induced by histamine releasing factors in human nasal washings. The effect of IL-2 alone was significantly greater than that of IL-4 alone or the combination of IL-2 plus IL-4. Taken together, the data suggest that when coadministered, IL-4 may inhibit the effects of IL-2 on basophils. Neither cytokine exerted any effect on basophil histamine release induced by the calcium ionophore A23187, nor did ranitidine cause any effects on histamine release induced by any of the stimulants. Thus, human basophil reactivity can be affected by IL-2 and by IL-4. The role that these two cytokines play in basophil function in vivo is likely to be complex. PMID:1372188

White, M V; Igarashi, Y; Emery, B E; Lotze, M T; Kaliner, M A

1992-03-15

193

Phase I trial of biochemotherapy with cisplatin, temozolomide, and dose escalation of nab-paclitaxel combined with interleukin-2 and interferon-? in patients with metastatic melanoma  

PubMed Central

The primary objective of this study was to determine the safety, toxicity, and maximum tolerated dose of nanoparticle albumin-bound (nab)-paclitaxel as part of biochemotherapy for metastatic melanoma and to determine whether substituting nab-paclitaxel for less potent agents could increase response rates and duration. Treatment consisted of intravenous cisplatin (20 mg/m2) on days 1–4, oral temozolomide (250 mg/m2) on days 1–3, subcutaneous interferon-? (5 × 106 IU/m2) on days 1–5, and continuous intravenous interleukin-2 (9 × 106 IU/m2) for 96 h on days 1–4. A standard 3 + 3 dose escalation method was used; the nab-paclitaxel starting dose was 100 mg/m2 on day 1 and 70 mg/m2 on day 5. The treatment cycle was repeated every 3 weeks and toxicity was assessed weekly. Ten patients were enrolled. Dose-limiting toxicities included diarrhea, transaminasemia, and neutropenia. The maximum tolerated dose was not identified because the nab-paclitaxel dose on day 1 at the lowest planned dose (80 mg/m2) caused dose-limiting toxicity in two of five patients. Of the nine patients who were evaluable for response, five had a partial response. The median time to disease progression was 5.30 months and the median overall survival was 8.73 months. Six patients developed central nervous system metastasis at a median of 5.33 months after treatment initiation. Biochemotherapy including nab-paclitaxel according to the doses and schedule regimen used in the present study has significant toxicity. Substituting dacarbazine with temozolomide did not prevent central nervous system metastasis in patients with metastatic melanoma. PMID:24743052

Alrwas, Anas; Papadopoulos, Nicholas E.; Cain, Suzanne; Patel, Sapna P.; Kim, Kevin B.; Deburr, Tawania L.; Bassett, Roland; Hwu, Wen-Jen; Bedikian, Agop Y.; Davies, Michael A.; Woodman, Scott E.; Hwu, Patrick

2014-01-01

194

A Phase I Study of High-Dose Interleukin-2 With Sorafenib in Patients With Metastatic Renal Cell Carcinoma and Melanoma  

PubMed Central

This study was designed to evaluate the safety and feasibility of high-dose interleukin-2 (HD IL-2) followed by sorafenib in patients with metastatic melanoma (MM) and renal cell carcinoma (RCC). Biomarkers relevant to the antitumor effects of IL-2 that may be altered by sorafenib including the percentages of natural T-regulatory cells (Tregs), myeloid-derived suppressor cells (MDSC), and STAT5 phosphorylation (pSTAT5) in T cells were evaluated. We hypothesized that the proposed treatment schedule is feasible and safe and may lead to enhanced tumor response. A phase I dose escalation trial was conducted in patients with either metastatic RCC or MM. HD IL-2 (600,000 IU/kg IV q8h×8–12 doses) was administered on days 1–5 and 15–19, followed by sorafenib on days 29–82. The sorafenib dose was escalated. The percentage of Tregs, MDSC, and pSTAT5 in T cells were evaluated in peripheral blood by flow cytometry. Twelve of the 18 patients were evaluable for dose-limiting toxicity. No dose-limiting toxicity was observed. The treatment-related toxicity was predictable and did not seem to be additive with this schedule of administration. Partial responses were seen in 3 patients. No significant changes in the percentage of circulating Treg and MDSC were observed, whereas sorafenib did not adversely affect the ability of IL-2 to induce pSTAT5 in T cells. HD IL-2 followed by sorafenib was safe and feasible in patients with MM and RCC and did not adversely affect T-cell signaling through STAT5 in response to IL-2. PMID:24598448

Lam, Elaine; Mortazavi, Amir; Kendra, Kari; Lesinski, Gregory B.; Mace, Thomas A.; Geyer, Susan; Carson, William E.; Tahiri, Sanaa; Bhinder, Arvinder; Clinton, Steven K.; Olencki, Thomas

2014-01-01

195

A randomised dose escalation study of subcutaneous interleukin 2 with and without levamisole in patients with metastatic renal cell carcinoma or malignant melanoma.  

PubMed Central

We have examined the efficacy, toxicity and host immunological response of two different dose schedules of interleukin 2 (IL-2) given subcutaneously, daily for 3 months in patients with renal cell carcinoma (RCC) or metastatic melanoma (MM). We also examined the effect of adding the immune modulator levamisole to the two different schedules of IL-2. Thirty-nine patients were entered into two sequential phase I/II studies. Eighteen patients entered study 1 and were randomised to receive IL-2, 3 x 10(6) IU m-2 day-1, subcutaneously for 3 months with or without levamisole 50 mg t.d.s. p.o. on days 1-3 on alternate weeks. Twenty-one patients entered study 2 and were randomised to receive 5.4 x 10(6) IU m-2 day-1 subcutaneously for 3 months with or without levamisole 50 mg t.d.s. p.o. on days 1-3 on alternate weeks. Blood was taken for peripheral blood lymphocyte (PBL) phenotype analysis, and measurement of IL-2, soluble IL-2 receptor (sIL-2R) and neopterin concentration. Two patients with metastatic melanoma, one in each study, responded (11.8%); both received IL-2 alone. Observations of immunological parameters showed that treatment with subcutaneous IL-2 resulted in a significant rise in the percentage of PBLs bearing CD25, CD3/HLA-DR, CD56 and levels of IL-2 receptor and neopterin. The total white blood cell count (WBC) and total lymphocyte count rose significantly on day 18 compared with pretreatment levels. The addition of levamisole to either IL-2 schedule resulted in no significant changes in any immunological parameters. This study illustrates that prolonged subcutaneous IL-2 can be given safely in the outpatient setting. There was no evidence that levamisole acts as an immunomodulator in this study. PMID:8855983

Ahmed, F. Y.; Leonard, G. A.; A'Hern, R.; Taylor, A. E.; Lorentzos, A.; Atkinson, H.; Moore, J.; Nicolson, M. C.; Riches, P. G.; Gore, M. E.

1996-01-01

196

Changes in plasma levels of interleukin-2 receptor in relation to disease exacerbations and levels of anti-dsDNA and complement in systemic lupus erythematosus.  

PubMed Central

Interleukin-2 receptor (IL-2R) is expressed and released predominantly by activated T cells. In order to investigate whether disease exacerbations of systemic lupus erythematosus (SLE) are preceded by T cell activation, we prospectively measured levels of IL-2R once a month, from 6 months prior to exacerbations until 1 month afterwards. To assess the temporal relation between T cell activation and B cell activation, we measured, in addition, levels of anti-dsDNA, complement C3/C4, and total IgG. During a mean follow-up period of 23 months, 40 exacerbations occurred in 21 out of the 71 participating patients. For the present study one exacerbation per patient was evaluated. During exacerbation levels of IL-2R were increased in 18 out of the 21 cases and correlated with levels of anti-dsDNA (P less than 0.02), C3 (P less than 0.02), and C4 (P less than 0.01), but not with the score of the disease activity index. Levels of IL-2R rose prior to the exacerbation (P less than 0.02) and fell afterwards following treatment (P less than 0.05). Even in the absence of disease activity or during minor disease symptoms IL-2R levels were higher (P less than 0.01) than in healthy controls. Sixteen out of the 21 exacerbations (76%) were preceded by a significant increase in IL-2R. Changes in levels of anti-dsDNA and complement C3/C4 tended to precede changes in levels of IL-2R. We conclude that increased levels of IL-2R, compatible with T cell activation, are present in SLE already during inactive disease. These levels further increased prior to exacerbations of disease. As such, IL-2R is an indicator of disease activity in SLE. Serial measurement of IL-2R is a sensitive test for predicting disease exacerbations of SLE. PMID:2208793

ter Borg, E J; Horst, G; Limburg, P C; Kallenberg, C G

1990-01-01

197

In vivo administration of purified human interleukin 2. II. Half life, immunologic effects, and expansion of peripheral lymphoid cells in vivo with recombinant IL 2.  

PubMed

Purified recombinant human interleukin 2 (RIL 2) derived from E. coli containing the inserted gene encoding for IL 2 was administered to 20 patients with a variety of malignancies. Toxicity was dose related and included fever, chills, malaise, arthralgias, myalgias, and unexpectedly, weight gain related to marked fluid retention. All patients receiving more than 10(5) U/kg total cumulative dose developed evidence of fluid retention, and all patients requiring discontinuance of RIL 2 (11/20) received total doses of between 2.54 X 10(5) U/kg to 15.4 X 10(5) U/kg. The limiting dose with this preparation was 3000 U/kg/hr by continuous administration or 10(6) U/kg by bolus administration. IL 2 was rapidly cleared from the plasma, with a half life of 6.9 min, and a later delayed clearance was consistent with a two-compartment model, with slower release from the extravascular space back into the plasma compartment. A marked change in lymphoid cells in the periphery was noted with an early depletion of all lymphoid cells, followed by an expansion of such cells with continuous IL 2 administration. A twofold to 16-fold expansion of total lymphoid cells in the peripheral blood could be demonstrated. TAC+ cells representing up to 25% of the circulating peripheral blood mononuclear cells could be demonstrated with 3 wk of continuous RIL 2 administration. Interferon-gamma levels increased in patients treated with IL 2. Precursors of lymphokine-activated killer cells generated under standard conditions were depleted within 2 to 3 min after IL 2 administration, but repopulated the peripheral blood after 7 to 10 days of continuous IL 2 administration. No tumor regression was seen in any of the cancer patients treated with IL 2 alone. PMID:2993418

Lotze, M T; Matory, Y L; Ettinghausen, S E; Rayner, A A; Sharrow, S O; Seipp, C A; Custer, M C; Rosenberg, S A

1985-10-01

198

Phase I trial of biochemotherapy with cisplatin, temozolomide, and dose escalation of nab-paclitaxel combined with interleukin-2 and interferon-? in patients with metastatic melanoma.  

PubMed

The primary objective of this study was to determine the safety, toxicity, and maximum tolerated dose of nanoparticle albumin-bound (nab)-paclitaxel as part of biochemotherapy for metastatic melanoma and to determine whether substituting nab-paclitaxel for less potent agents could increase response rates and duration. Treatment consisted of intravenous cisplatin (20 mg/m) on days 1-4, oral temozolomide (250 mg/m) on days 1-3, subcutaneous interferon-? (5×10 IU/m) on days 1-5, and continuous intravenous interleukin-2 (9×10 IU/m) for 96 h on days 1-4. A standard 3+3 dose escalation method was used; the nab-paclitaxel starting dose was 100 mg/m on day 1 and 70 mg/m on day 5. The treatment cycle was repeated every 3 weeks and toxicity was assessed weekly. Ten patients were enrolled. Dose-limiting toxicities included diarrhea, transaminasemia, and neutropenia. The maximum tolerated dose was not identified because the nab-paclitaxel dose on day 1 at the lowest planned dose (80 mg/m) caused dose-limiting toxicity in two of five patients. Of the nine patients who were evaluable for response, five had a partial response. The median time to disease progression was 5.30 months and the median overall survival was 8.73 months. Six patients developed central nervous system metastasis at a median of 5.33 months after treatment initiation. Biochemotherapy including nab-paclitaxel according to the doses and schedule regimen used in the present study has significant toxicity. Substituting dacarbazine with temozolomide did not prevent central nervous system metastasis in patients with metastatic melanoma. PMID:24743052

Alrwas, Anas; Papadopoulos, Nicholas E; Cain, Suzanne; Patel, Sapna P; Kim, Kevin B; Deburr, Tawania L; Bassett, Roland; Hwu, Wen-Jen; Bedikian, Agop Y; Davies, Michael A; Woodman, Scott E; Hwu, Patrick

2014-08-01

199

Correlation of blood lymphocyte CTLA-4 (CD152) induction in Hodgkin's disease with proliferative activity, interleukin 2 and interferon-gamma production.  

PubMed

Expression of the downregulatory CTLA-4 molecule was determined on unstimulated and anti-CD3 + recombinant interleukin 2 (rIL-2)-stimulated peripheral blood T cells in Hodgkin's disease (HD) and correlated with the T-cells' proliferative activity, IL-2 and interferon (IFN)-gamma production. There was a negligible percentage of CTLA-4+/CD3+ cells before culture. The mean percentage of CTLA-4+/CD3+ lymphocytes increased gradually, peaked after 72 h of stimulation and returned to basal values after 96 h of stimulation. The mean proportion of CTLA-4+/CD3+ cells from untreated patients was significantly higher after 24, 48 and 72 h of stimulation compared with controls. The mean percentage of CTLA-4+/CD3+ cells from patients in clinical remission (CR) was lower than that of untreated patients, but remained significantly higher compared with controls. Lymphocytes from untreated HD patients showed impaired proliferative activity, IL-2 and IFN-gamma production compared with controls. The proliferative activity of the lymphocytes, IL-2 and IFN-gamma production remained significantly lower in CR compared with controls. The proportion of CTLA-4+/CD3+ cells negatively correlated with proliferative activity, IL-2 and IFN-gamma production in HD patients and controls. However, some untreated patients as well as patients in CR with normal mean fluorescence intensity values of CTLA-4 showed unimpaired T-cell function tests. Our study provides the first evidence of an increased expression of downregulatory CTLA-4 molecule on stimulated T-cells in HD, which could be one of the mechanisms of immune deficiency in this disease. PMID:12100149

Kosmaczewska, Agata; Frydecka, Irena; Bo?ko, Dorota; Ciszak, Lidia; Teodorowska, Renata

2002-07-01

200

Adoptive immunotherapy of advanced melanoma patients with interleukin-2 (IL-2) and tumor-infiltrating lymphocytes selected in vitro with low doses of IL-2.  

PubMed

Freshly isolated tumor-infiltrating lymphocytes (TIL) from stage IV melanoma patients were cultured for 2 weeks with low doses of interleukin-2 (IL-2; 120 IU/ml), to select potentially for tumor-specific lymphocytes present in the neoplastic lesion, followed by high doses (6000 IU/ml) to achieve lymphocyte expansion. TIL were serially analyzed for their expansion, phenotype and cytotoxic activity against autologous and allogenic tumor cells. A preferential lysis of autologous melanoma cells was obtained in long-term cultures of 7/13 cases (54%), while the remaining ones showed a major-histocompatibility-complex-unrestricted, lymphokine-activated-killer(LAK)-like activity at the time of in vivo injection. Sixteen patients with metastatic melanoma were infused with TIL (mean number: 6.8 x 10(9), range: 0.35 x 10(9)-20 x 10(9)) and IL-2 (mean dose: 130 x 10(6) IU, range: 28.8 x 10(6)-231 x 10(6) IU); 1 complete and 3 partial responses were observed in 12 evaluable patients (response rate 33%). In all responding patients, injected TIL showed an in vitro preferential lysis of autologous tumor cells, while in no cases were TIL with LAK-like activity associated with a clinical response. The mean autologous tumor cytotoxic activity of TIL at the time of in vivo injection was significantly higher in responding patients in comparison to nonresponding ones, suggesting that a marked and preferential cytolysis of autologous tumor cells is associated with the therapeutic efficacy of TIL. PMID:8477417

Arienti, F; Belli, F; Rivoltini, L; Gambacorti-Passerini, C; Furlan, L; Mascheroni, L; Prada, A; Rizzi, M; Marchesi, E; Vaglini, M

1993-05-01

201

Treatment of recurrent in transit metastases from cutaneous melanoma by isolation perfusion in extracorporeal circulation with interleukin-2 and lymphokine activated killer cells. A pilot study.  

PubMed

Chemoresistant melanoma cells are known to be susceptible in vitro to lymphokine activated killer (LAK) cells. To obtain a high LAK/tumour cell ratio in vivo and avoid systemic toxicity due to interleukin-2 (IL-2), we used IL-2 plus LAK cells in the treatment of in transit melanoma metastases of the limbs by isolation perfusion (IP). In vivo immunological modifications induced by this immunotherapeutic approach were also analysed. Six patients previously treated with IP in extracorporeal circulation with tumour cytotoxic drugs and presently relapsing or not responding, were submitted to locoregional adoptive therapy consisting of 5 days systemic administration of IL-2 (Proleukin, EuroCetus) (9-12 x 10(6) IU/m2/day c.i.). Autologous LAK cells were derived from leukapheresis and subsequent in vitro stimulation with IL-2; LAK cells were then given along with IL-2 (120-2400 IU/ml of perfusion priming) to the affected limb by IP. In addition, 7-16 x 10(9) LAK cells were administered by systemic infusion the day after together with IL-2 (9-12 x 10(6) IU/m2/day) by c.i. for 5 days. All patients concluded the treatment without major toxicity. The analysis of circulating lymphocytes obtained from extracorporeal circuit at different times revealed rapid disappearance of LAK cells, suggesting their extravasation and/or endothelial adhesion in perfused tissues. Clinical responses included four partial and one complete response; another patient had stable disease. All patients are presently alive. Follow-up after IP ranges from 8 to 22 months. PMID:1490114

Belli, F; Arienti, F; Rivoltini, L; Santinami, M; Mascheroni, L; Prada, A; Ammatuna, M; Marchesi, E; Parmiani, G; Cascinelli, N

1992-11-01

202

Evidence for a Structural Motif in Toxins and Interleukin-2 That May Be Responsible for Binding to Endothelial Cells and Initiating Vascular Leak Syndrome  

NASA Astrophysics Data System (ADS)

The dose-limiting toxicity of interleukin-2 (IL-2) and immunotoxin (IT) therapy in humans is vascular leak syndrome (VLS). VLS has a complex etiology involving damage to vascular endothelial cells (ECs), extravasation of fluids and proteins, interstitial edema, and organ failure. IL-2 and ITs prepared with the catalytic A chain of the plant toxin, ricin (RTA), and other toxins, damage human ECs in vitro and in vivo. Damage to ECs may initiate VLS; if this damage could be avoided without losing the efficacy of ITs or IL-2, larger doses could be administered. In this paper, we provide evidence that a three amino acid sequence motif, (x)D(y), in toxins and IL-2 damages ECs. Thus, when peptides from RTA or IL-2 containing this sequence motif are coupled to mouse IgG, they bind to and damage ECs both in vitro and, in the case of RTA, in vivo. In contrast, the same peptides with a deleted or mutated sequence do not. Furthermore, the peptide from RTA attached to mouse IgG can block the binding of intact RTA to ECs in vitro and vice versa. In addition, RTA, a fragment of Pseudomonas exotoxin A (PE38-lys), and fibronectin also block the binding of the mouse IgG-RTA peptide to ECs, suggesting that an (x)D(y) motif is exposed on all three molecules. Our results suggest that deletions or mutations in this sequence or the use of nondamaging blocking peptides may increase the therapeutic index of both IL-2, as well as ITs prepared with a variety of plant or bacterial toxins.

Baluna, Roxana; Rizo, Josep; Gordon, Brian E.; Ghetie, Victor; Vitetta, Ellen S.

1999-03-01

203

Structure of the interleukin-2 tyrosine kinase Src homology 2 domain; comparison between X-ray and NMR-derived structures.  

PubMed

The crystal structure of the interleukin-2 tyrosine kinase Src homology domain (Itk SH2) is described and it is found that unlike in studies of this domain using NMR spectroscopy, cis-trans-prolyl isomerization is not readily detected in the crystal structure. Based on similarities between the Itk SH2 crystal form and the cis form of the Itk SH2 NMR structure, it is concluded that it is likely that the prolyl imide bond at least in part adopts the cis conformation in the crystal form. However, the lack of high-resolution data and the dynamic nature of the proline-containing loop mean that the precise imide-bond conformation cannot be determined and prolyl cis-trans isomerization in the crystal cannot be ruled out. Given the preponderance of structures that have been solved by X-ray crystallography in the Protein Data Bank, this result supports the notion that prolyl isomerization in folded proteins has been underestimated among known structures. Interestingly, while the precise status of the proline residue is ambiguous, Itk SH2 crystallizes as a domain-swapped dimer. The domain-swapped structure of Itk SH2 is similar to the domain-swapped SH2 domains of Grb2 and Nck, with domain swapping occurring at the ?-meander region of all three SH2 domains. Thus, for Itk SH2 structural analysis by NMR spectroscopy and X-ray crystallography revealed very different structural features: proline isomerization versus domain-swapped dimerization, respectively. PMID:22297986

Joseph, Raji E; Ginder, Nathaniel D; Hoy, Julie A; Nix, Jay C; Fulton, D Bruce; Honzatko, Richard B; Andreotti, Amy H

2012-02-01

204

A pilot study of paclitaxel combined with gemcitabine followed by interleukin-2 and granulocyte macrophage colony-stimulating factor for patients with metastatic melanoma  

PubMed Central

It has been suggested that paclitaxel and gemcitabine modulate the immune system. This paper reports the safety and efficacy of paclitaxel plus gemcitabine followed by interleukin-2 (IL-2)and granulocyte macrophage colony-stimulating factor (GM-CSF), the PGIG chemobiotherapy, for patients with metastatic melanoma. All patients received 175 mg/m2 paclitaxel on day 1 and 800 mg/m2 gemcitabine on day two. IL-2 and GM-CSF were administered from day 4 to day 8 at a dosage of 2 MIU/m2 and 100 ?g, respectively. The PGIG chemobiotherapy was repeated every 21 d. Serum cytokine levels at baseline and at the end of the second cycle were measured via flow cytometry. Twenty-seven patients with metastatic melanoma accepted PGIG chemobiotherapy from August 2009 to March 2011. There were five patients that exhibited a partial response, 14 patients that exhibited a stable response and eight that displayed progressive disease. Therefore, the response rate was 18.5%, and the disease control rate was 70.4%. The median time to progression and median survival were 4 mo and 8 mo, respectively. The one-year and two-year survival rates were 25.9% and 18.5%, respectively. Frequent side effects included chills, fever, arthralgia, rash and pruritus. Among the 13 patients who experienced a rash and pruritus and the 14 patients who did not suffer from this side effect, the response rates and disease control rates were 30.8% vs 7.1% and 77% vs 64.2%, respectively. No relationship between serum IL-6 levels, clinical response, and either skin side effect was observed. The PGIG chemobiotherapy is safe and effective for the treatment of patients with advanced melanoma, but randomized trials are necessary to validate this effect. PMID:22954698

Peng, Rui-Qing; Ding, Ya; Zhang, Xing; Liao, Yuan; Zheng, Li-Min; Zhang, Xiao-Shi

2012-01-01

205

Displacement of SATB1-bound histone deacetylase 1 corepressor by the human immunodeficiency virus type 1 transactivator induces expression of interleukin-2 and its receptor in T cells.  

PubMed

One hallmark of human immunodeficiency virus type 1 (HIV-1) infection is the dysregulation of cytokine gene expression in T cells. Transfection of T cells with human T-cell leukemia type 1 or 2 transactivator results in the induction of the T-cell-restricted cytokine interleukin-2 (IL-2) and its receptor (IL-2Ralpha). However, no T-cell-specific factor(s) has been directly linked with the regulation of IL-2 and IL-2Ralpha transcription by influencing the promoter activity. Thymocytes from SATB1 (special AT-rich sequence binding protein 1) knockout mice have been shown to ectopically express IL-2Ralpha, suggesting involvement of SATB1 in its negative regulation. Here we show that SATB1, a T-cell-specific global gene regulator, binds to the promoters of human IL-2 and IL-2Ralpha and recruits histone deacetylase 1 (HDAC1) in vivo. SATB1 also interacts with Tat in HIV-1-infected T cells. The functional interaction between HIV-1 Tat and SATB1 requires its PDZ-like domain, and the binding of the HDAC1 corepressor occurs through the same. Furthermore, Tat competitively displaces HDAC1 that is bound to SATB1, leading to increased acetylation of the promoters in vivo. Transduction with SATB1 interaction-deficient soluble Tat (Tat 40-72) and reporter assays using a transactivation-negative mutant (C22G) of Tat unequivocally demonstrated that the displacement of HDAC1 itself is sufficient for derepression of these promoters in vivo. These results suggest a novel mechanism by which HIV-1 Tat might overcome SATB1-mediated repression in T cells. PMID:15713622

Kumar, P Pavan; Purbey, Prabhat Kumar; Ravi, Dyavar S; Mitra, Debashis; Galande, Sanjeev

2005-03-01

206

Displacement of SATB1-Bound Histone Deacetylase 1 Corepressor by the Human Immunodeficiency Virus Type 1 Transactivator Induces Expression of Interleukin-2 and Its Receptor in T Cells  

PubMed Central

One hallmark of human immunodeficiency virus type 1 (HIV-1) infection is the dysregulation of cytokine gene expression in T cells. Transfection of T cells with human T-cell leukemia type 1 or 2 transactivator results in the induction of the T-cell-restricted cytokine interleukin-2 (IL-2) and its receptor (IL-2R?). However, no T-cell-specific factor(s) has been directly linked with the regulation of IL-2 and IL-2R? transcription by influencing the promoter activity. Thymocytes from SATB1 (special AT-rich sequence binding protein 1) knockout mice have been shown to ectopically express IL-2R?, suggesting involvement of SATB1 in its negative regulation. Here we show that SATB1, a T-cell-specific global gene regulator, binds to the promoters of human IL-2 and IL-2R? and recruits histone deacetylase 1 (HDAC1) in vivo. SATB1 also interacts with Tat in HIV-1-infected T cells. The functional interaction between HIV-1 Tat and SATB1 requires its PDZ-like domain, and the binding of the HDAC1 corepressor occurs through the same. Furthermore, Tat competitively displaces HDAC1 that is bound to SATB1, leading to increased acetylation of the promoters in vivo. Transduction with SATB1 interaction-deficient soluble Tat (Tat 40-72) and reporter assays using a transactivation-negative mutant (C22G) of Tat unequivocally demonstrated that the displacement of HDAC1 itself is sufficient for derepression of these promoters in vivo. These results suggest a novel mechanism by which HIV-1 Tat might overcome SATB1-mediated repression in T cells. PMID:15713622

Kumar, P. Pavan; Purbey, Prabhat Kumar; Ravi, Dyavar S.; Mitra, Debashis; Galande, Sanjeev

2005-01-01

207

Interleukin2 and Cancer Therapy  

Microsoft Academic Search

IL-2 was originally isolated as a soluble factor with the property of enhancing T-lymphocyte proliferation in studies of the\\u000a human immunodeficiency virus (1). The earliest studies of its activity in the cellular therapy of cancer used partially-purified IL-2 from the Jurkat human\\u000a T-cell line. Subsequent studies used recombinant IL-2 produced in E. coli, an unlimited source of this valuable cytokine

Kim Margolin; Joseph Clark

208

Urinary-Cell mRNA Profile and Acute Cellular Rejection in Kidney Allografts  

PubMed Central

Background The standard test for the diagnosis of acute rejection in kidney transplants is the renal biopsy. Noninvasive tests would be preferable. Methods We prospectively collected 4300 urine specimens from 485 kidney-graft recipients from day 3 through month 12 after transplantation. Messenger RNA (mRNA) levels were measured in urinary cells and correlated with allograft-rejection status with the use of logistic regression. Results A three-gene signature of 18S ribosomal (rRNA)–normalized measures of CD3? mRNA and interferon-inducible protein 10 (IP-10) mRNA, and 18S rRNA discriminated between biopsy specimens showing acute cellular rejection and those not showing rejection (area under the curve [AUC], 0.85; 95% confidence interval [CI], 0.78 to 0.91; P<0.001 by receiver-operating-characteristic curve analysis). The cross-validation estimate of the AUC was 0.83 by bootstrap resampling, and the Hosmer–Lemeshow test indicated good fit (P = 0.77). In an external-validation data set, the AUC was 0.74 (95% CI, 0.61 to 0.86; P<0.001) and did not differ significantly from the AUC in our primary data set (P = 0.13). The signature distinguished acute cellular rejection from acute antibody-mediated rejection and borderline rejection (AUC, 0.78; 95% CI, 0.68 to 0.89; P<0.001). It also distinguished patients who received anti–interleukin-2 receptor antibodies from those who received T-cell–depleting antibodies (P<0.001) and was diagnostic of acute cellular rejection in both groups. Urinary tract infection did not affect the signature (P = 0.69). The average trajectory of the signature in repeated urine samples remained below the diagnostic threshold for acute cellular rejection in the group of patients with no rejection, but in the group with rejection, there was a sharp rise during the weeks before the biopsy showing rejection (P<0.001). Conclusions A molecular signature of CD3? mRNA, IP-10 mRNA, and 18S rRNA levels in urinary cells appears to be diagnostic and prognostic of acute cellular rejection in kidney allografts. (Funded by the National Institutes of Health and others.) PMID:23822777

Suthanthiran, Manikkam; Schwartz, Joseph E.; Ding, Ruchuang; Abecassis, Michael; Dadhania, Darshana; Samstein, Benjamin; Knechtle, Stuart J.; Friedewald, John; Becker, Yolanda T.; Sharma, Vijay K.; Williams, Nikki M.; Chang, Christina S.; Hoang, Christine; Muthukumar, Thangamani; August, Phyllis; Keslar, Karen S.; Fairchild, Robert L.; Hricik, Donald E.; Heeger, Peter S.; Han, Leiya; Liu, Jun; Riggs, Michael; Ikle, David N.; Bridges, Nancy D.; Shaked, Abraham

2013-01-01

209

Plasmodium falciparum evades mosquito immunity by disrupting JNK-mediated apoptosis of invaded midgut cells.  

PubMed

The malaria parasite, Plasmodium, must survive and develop in the mosquito vector to be successfully transmitted to a new host. The Plasmodium falciparum Pfs47 gene is critical for malaria transmission. Parasites that express Pfs47 (NF54 WT) evade mosquito immunity and survive, whereas Pfs47 knockouts (KO) are efficiently eliminated by the complement-like system. Two alternative approaches were used to investigate the mechanism of action of Pfs47 on immune evasion. First, we examined whether Pfs47 affected signal transduction pathways mediating mosquito immune responses, and show that the Jun-N-terminal kinase (JNK) pathway is a key mediator of Anopheles gambiae antiplasmodial responses to P. falciparum infection and that Pfs47 disrupts JNK signaling. Second, we used microarrays to compare the global transcriptional responses of A. gambiae midguts to infection with WT and KO parasites. The presence of Pfs47 results in broad and profound changes in gene expression in response to infection that are already evident 12 h postfeeding, but become most prominent at 26 h postfeeding, the time when ookinetes invade the mosquito midgut. Silencing of 15 differentially expressed candidate genes identified caspase-S2 as a key effector of Plasmodium elimination in parasites lacking Pfs47. We provide experimental evidence that JNK pathway regulates activation of caspases in Plasmodium-invaded midgut cells, and that caspase activation is required to trigger midgut epithelial nitration. Pfs47 alters the cell death pathway of invaded midgut cells by disrupting JNK signaling and prevents the activation of several caspases, resulting in an ineffective nitration response that makes the parasite undetectable by the mosquito complement-like system. PMID:25552553

Ramphul, Urvashi N; Garver, Lindsey S; Molina-Cruz, Alvaro; Canepa, Gaspar E; Barillas-Mury, Carolina

2015-02-01

210

Plasmodium falciparum evades mosquito immunity by disrupting JNK-mediated apoptosis of invaded midgut cells  

PubMed Central

The malaria parasite, Plasmodium, must survive and develop in the mosquito vector to be successfully transmitted to a new host. The Plasmodium falciparum Pfs47 gene is critical for malaria transmission. Parasites that express Pfs47 (NF54 WT) evade mosquito immunity and survive, whereas Pfs47 knockouts (KO) are efficiently eliminated by the complement-like system. Two alternative approaches were used to investigate the mechanism of action of Pfs47 on immune evasion. First, we examined whether Pfs47 affected signal transduction pathways mediating mosquito immune responses, and show that the Jun-N-terminal kinase (JNK) pathway is a key mediator of Anopheles gambiae antiplasmodial responses to P. falciparum infection and that Pfs47 disrupts JNK signaling. Second, we used microarrays to compare the global transcriptional responses of A. gambiae midguts to infection with WT and KO parasites. The presence of Pfs47 results in broad and profound changes in gene expression in response to infection that are already evident 12 h postfeeding, but become most prominent at 26 h postfeeding, the time when ookinetes invade the mosquito midgut. Silencing of 15 differentially expressed candidate genes identified caspase-S2 as a key effector of Plasmodium elimination in parasites lacking Pfs47. We provide experimental evidence that JNK pathway regulates activation of caspases in Plasmodium-invaded midgut cells, and that caspase activation is required to trigger midgut epithelial nitration. Pfs47 alters the cell death pathway of invaded midgut cells by disrupting JNK signaling and prevents the activation of several caspases, resulting in an ineffective nitration response that makes the parasite undetectable by the mosquito complement-like system. PMID:25552553

Ramphul, Urvashi N.; Garver, Lindsey S.; Molina-Cruz, Alvaro; Canepa, Gaspar E.; Barillas-Mury, Carolina

2015-01-01

211

JNK-mediated phosphorylation of DLK suppresses its ubiquitination to promote neuronal apoptosis  

PubMed Central

Neurons are highly polarized cells that often project axons a considerable distance. To respond to axonal damage, neurons must transmit a retrograde signal to the nucleus to enable a transcriptional stress response. Here we describe a mechanism by which this signal is propagated through injury-induced stabilization of dual leucine zipper-bearing kinase (DLK/MAP3K12). After neuronal insult, specific sites throughout the length of DLK underwent phosphorylation by c-Jun N-terminal kinases (JNKs), which have been shown to be downstream targets of DLK pathway activity. These phosphorylation events resulted in increased DLK abundance via reduction of DLK ubiquitination, which was mediated by the E3 ubiquitin ligase PHR1 and the de-ubiquitinating enzyme USP9X. Abundance of DLK in turn controlled the levels of downstream JNK signaling and apoptosis. Through this feedback mechanism, the ubiquitin–proteasome system is able to provide an additional layer of regulation of retrograde stress signaling to generate a global cellular response to localized external insults. PMID:23979718

Huntwork-Rodriguez, Sarah; Wang, Bei; Watkins, Trent; Ghosh, Arundhati Sengupta; Pozniak, Christine D.; Bustos, Daisy; Newton, Kim; Kirkpatrick, Donald S.

2013-01-01

212

JNK mediates UVB-induced apoptosis upstream lysosomal membrane permeabilization and Bcl-2 family proteins.  

PubMed

UVB irradiation induced phosphorylation of JNK and subsequent apoptosis in human melanocytes. Depletion of both JNK1 and JNK2 expression using siRNA transfection, protected against apoptosis, as detected by decreased nuclear fragmentation and caspase-3 activity, as well as reduced translocation of Bax to mitochondria. Moreover, release of cathepsin B and D from lysosomes to the cytosol was reduced when JNK expression was suppressed by siRNA, demonstrating a JNK dependent regulation of lysosomal membrane permeabilization. In unirradiated control melanocytes, coimmunoprecipitation showed that Bim was sequestered by Mcl-1, which had a pro-survival function. After UVB irradiation, a significant decrease in Mcl-1 protein level was found, which was prevented by addition of a proteasome inhibitor. The interaction between Bim and Mcl-1 was reduced in response to UVB irradiation and Bim was phosphorylated in a JNK dependent manner. In conclusion, these findings suggest JNK to have an important pro-apoptotic function following UVB irradiation in human melanocytes, by acting upstream of lysosomal membrane permeabilization and Bim phosphorylation. PMID:18651223

Bivik, Cecilia; Ollinger, Karin

2008-09-01

213

Increased excretion of soluble interleukin 2 receptors and free light chain immunoglobulins in the urine of patients with active lupus nephritis.  

PubMed Central

Samples of protein from the urine of 23 patients with lupus nephropathy and 15 patients with proteinuria who did not have systemic lupus erythematosus (SLE) were studied for the presence of cytokines, soluble interleukin 2 receptors (sIL-2R), and free light chain immunoglobulins. The patients with lupus nephropathy were divided into two groups with active (nephritis) and inactive inflammation (nephrosis) based on the results of the analysis of urine samples and renal histology. The crude urine proteins (5 mg/ml) after precipitation by 80% ammonium sulphate from 14 patients with lupus nephritis contained higher concentrations of sIL-2R (4.88 (SEM 1.27 ng/ml) than those from nine patients with nephrosis (1.11 (0.52) ng/ml) or 15 patients without SLE (1.31 (0.87) ng/ml). The concentration of sIL-2R in protein from urine samples was not correlated with the concentration in plasma and was inversely correlated with the excretion of protein in urine over 24 hours in patients with SLE. It is suggested that, in addition to leakage from the circulation, the local production of sIL-2R by inflamed kidneys is possible. The crude proteins in urine were further fractionated by gel filtration on Sephacryl S-200. Arbitrarily, four fractions could be obtained from urine from patients with SLE but only three fractions were found in the urine of patients without SLE. Fraction IV derived from patients with nephritis or nephrosis augmented the pokeweed mitogen induced [3H]thymidine uptake of mononuclear cells. In addition, the positive rates of free kappa (kappa) (35.7%) and lambda (lambda) (42.9%) chains in proteins in urine from nephritic patients were higher than those in the other two groups. These results suggest that the severity of inflammation in the kidneys of patients with lupus can be reflected by the increased excretion of sIL-2R, free light chain immunoglobulins, and cytokine-like molecules in urine. Images PMID:1550398

Tsai, C Y; Wu, T H; Sun, K H; Lin, W M; Yu, C L

1992-01-01

214

Long-term interleukin 2-dependent growth and cytotoxic activity of tumor-infiltrating lymphocytes from human squamous cell carcinomas of the head and neck.  

PubMed

Tumor-infiltrating lymphocytes (TIL) from 16 squamous cell carcinomas of head and neck (SCCH&N) and four nonsquamous cell carcinomas were studied. By immunoperoxidase staining in situ, the tumors studied were found to be infiltrated mainly by CD2+CD3+ cells, and 30-50% of the T-lymphocytes were HLA-DR positive and transferrin-receptor positive. They also contained scarce NKH1+ cells. When TIL as well as autologous peripheral blood lymphocytes (A-PBL) were cultured in 1,000 U/ml of recombinant interleukin 2 (rIL2), TIL proliferated in all but three cases, and A-PBL proliferated in all but two cases. Frequently, but not always, TIL expanded better than A-PBL. The median expansion for TIL was 100-fold and that for A-PBL was 31-fold in long-term cultures maintained for up to 88 days. TIL obtained from untreated primary SCCH&N were initially delayed for up to 20 days in their proliferative response to rIL2, but then grew well. In contrast, TIL and A-PBL from metastatic SCCH&N either did not proliferate or were delayed in their proliferative response for up to 40 or 50 days. A-PBL, when tested early (days 10-20 in culture), showed the highest cytotoxic activity against cultured and fresh tumor-cell targets, whereas TIL were most active later in culture (days 20-30). On a per culture basis, TIL achieved higher antitumor cytotoxicity than A-PBL. By day 80, lytic activities of most TIL cultures declined to undetectable levels. CD3+Leu19- T-lymphocytes were the major expanding cell population in most TIL cultures. However, these cells were poor mediators of antitumor cytotoxicity in TIL or A-PBL cultures as shown in cell sorting experiments. The antitumor effector cells expressed CD3-Leu19+ and/or CD3+Leu19+ phenotypes. On Giemsa-stained smears, these two types of IL2-expanded effector cells had the morphology of large granular lymphocytes. Our results indicate that TIL from human SCCH&N could be expanded and reach high levels of antitumor effector function in long-term cultures with rIL2. PMID:3315186

Heo, D S; Whiteside, T L; Johnson, J T; Chen, K N; Barnes, E L; Herberman, R B

1987-12-01

215

Durable responses and reversible toxicity of high-dose interleukin-2 treatment of melanoma and renal cancer in a Community Hospital Biotherapy Program  

PubMed Central

Background High-dose interleukin-2 (IL-2) has been FDA-approved for over 20 years, but it is offered only at a small number of centers with expertise in its administration. We analyzed the outcomes of patients receiving high-dose IL-2 in relation to the severity of toxicity to ascertain if response or survival were adversely affected. Methods A retrospective analysis of the outcomes of 500 patients with metastatic renal cell carcinoma (RCC) (n?=?186) or melanoma (n?=?314) treated with high-dose IL-2 between 1997 and 2012 at Providence Cancer Center was performed. IL-2 was administered at a dose of 600,000 international units per kg by IV bolus every 8 hours for up to 14 doses. A second cycle was administered 16 days after the first and patients with tumor regression could receive additional cycles. Survival and anti-tumor response were analyzed by diagnosis, severity of toxicity, number of IL-2 cycles and subsequent therapy. Results The objective response rate in melanoma was 28% (complete 12% and partial 16%), and in RCC was 24% (complete 7% and partial 17%). The 1-, 2- and 3-year survivals were 59%, 41% and 31%, for melanoma and 75%, 56% and 44%, for RCC, respectively. The proportion of patients with complete or partial response in both melanoma and RCC was higher in patients who a) required higher phenylephrine doses to treat hypotension (p?100,000 platelets; p?

2014-01-01

216

Monoclonal antibodies to interferon-gamma inhibit interleukin 2-dependent induction of growth and maturation in lectin/antigen-reactive cytolytic T lymphocyte precursors.  

PubMed

In this study we tested the effect of monoclonal antibodies (moAb) AN-18 to murine IFN-gamma on the generation of cytolytic T cells (CTL) from a homogeneous population of precursor cells (CTL-P). As responder cells, highly purified Lyt-2+ C57BL/6 lymph node T cells were used that had been positively selected by flow cytofluorometry on a cell sorter. Lyt-2+ cells were set up in bulk culture or in limiting dilution (LD) either with Con A or with P815 tumor cells as antigen and recombinant human interleukin 2 (rec.hIL 2) in the presence or absence of moAb AN-18 and tested for growth and development of CTL. The results show that moAb AN-18 but not the unrelated moAb AN-37 diminished or abrogated proliferative and cytolytic responses of Lyt-2+ lymphocytes to lectin and rec.hIL 2 in a dose-dependent manner. The inhibitory activity of the antibodies could be abolished by neutralizing moAb AN-18 with recombinant murine IFN-gamma (rec.mIFN-gamma) before their addition to culture. Kinetic analysis shows that the inhibitory effect of moAb AN-18 is only optimal when added at the beginning of culture or up to 48 hr after initiation. The frequencies of CTL-P responding either to Con A or to P815 tumor cells and rec.hIL 2 were reduced up to 10-fold in the presence of moAb AN-18. The inhibitory capacity of moAb AN-18 was also operative in cultures containing on the average one antigen-specific CTL-P. Together with the finding that activated CTL-P secrete IFN-gamma in response to rec.hIL 2 in a dose-dependent manner, the data suggest that endogenous IFN-gamma collaborates with exogenous IL 2 in the induction of CTL-P. The generation of CTL may therefore represent a case of autocrine growth regulation of normal lymphocytes, in which the same cell synthesizes and responds to its own factor. PMID:3082972

Simon, M M; Hochgeschwender, U; Brugger, U; Landolfo, S

1986-04-15

217

A phase I study of prolonged continuous infusion of low dose recombinant interleukin-2 in melanoma and renal cell cancer. Part I: Clinical aspects.  

PubMed Central

The optimal schedule for recombinant interleukin-2 (rIL-2) administration is unclear. Because the clinical and immunological effects of prolonged continuous exposure to rIL-2 are unknown, we have conducted a phase I study to assess the toxicity and feasibility of continuous low dose infusion of rIL-2 (EuroCetus) using central venous access with a portable infusion device on an out-patient basis. Twenty-two patients entered the study, 13 with melanoma and nine with renal cell cancer, age range 26-66 years (median 51), performance status less than or equal to 1. They were treated with one of the following doses per m2 per 24 h: 0.18 x 10(6) IU, 0.6 x 10(6) IU, 1.8 x 10(6) IU, 3 x 10(6) IU, 6 x 10(6) IU and 9 x 10(6) IU. Toxicity was evaluable in 20 patients receiving greater than or equal to 3 weeks treatment duration or in whom treatment was discontinued prematurely because of toxicity. Constitutional symptoms consisting of fatigue, malaise and fever up to 40 degrees C without significant organ dysfunction occurred with doses greater than or equal to 1.8 x 10(6) IU m-2. The maximum tolerated dose was 6 x 10(6) IU m-2 24 h-1. In all patients toxicity reached a peak at 3 weeks and resolved thereafter despite continued rIL-2 treatment. Peripheral blood eosinophilia (up to 66% of white blood cell count) followed the same pattern. An infection of the central venous access occurred in 55% of the patients but this was mostly asymptomatic. Thirteen patients were treated greater than or equal to 6 weeks and were evaluable for tumour response. A partial remission occurred in a patient with melanoma with a dose of 1.8 x 10(6) IU rIL-2 m-2 24 h-1. PMID:1586602

Vlasveld, L. T.; Rankin, E. M.; Hekman, A.; Rodenhuis, S.; Beijnen, J. H.; Hilton, A. M.; Dubbelman, A. C.; Vyth-Dreese, F. A.; Melief, C. J.

1992-01-01

218

NG-nitro-L-arginine methyl ester, an inhibitor of nitric oxide synthesis, ameliorates interleukin 2-induced capillary leakage and reduces tumour growth in adenocarcinoma-bearing mice.  

PubMed Central

We tested whether NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide (NO) synthesis, can prevent interleukin 2 (IL-2)-induced capillary leakage in tumour-bearing mice without compromising the therapeutic benefits of IL-2. C3H/HeJ female mice transplanted s.c. with 2.5 x 10(5) C3-L5 mammary carcinoma cells were treated with: nothing, IL-2 (ten injections of 15,000 Cetus units i.p. every 8 h), L-NAME (0.1, 0.5, or 1 mg ml-1 drinking water), IL-2 + L-NAME (0.1 or 0.5 or 1 mg ml-1 drinking water). Therapies were given in one round (IL-2, days 10-13; L-NAME, days 9-13) or in two rounds (IL-2, days 10-13 and 20-23; L-NAME, days 9-13 and days 19-23) after tumour transplantation. Capillary leakage was measured from the water contents of the pleural cavities, lungs, spleen and kidneys. Effects of the therapies on the primary tumour size and the number of spontaneous lung metastases were also recorded. NO production was measured as the nitrite + nitrate levels in the serum and in the pleural effusion. After the first round of therapies, addition of L-NAME significantly reduced IL-2-induced pulmonary oedema and water retention in the spleen in a dose-dependent manner. It also significantly reduced the IL-2-induced rise in NO levels in the serum and pleural fluid, but did not affect IL-2-induced pleural effusion or water retention in the kidney. At later stages of tumour growth (day 23), tumours themselves induced significant fluid retention in the lungs and the kidney, which was not aggravated further with the second round of IL-2 therapy. At this time, L-NAME therapy alone ameliorated tumour-induced pulmonary oedema. During both rounds of therapy different doses of L-NAME alone caused a reduction of primary tumour growth as well as spontaneous lung metastases, which improved further with the addition of IL-2. The combination therapy was at least as effective as IL-2 therapy. In summary, L-NAME had anti-tumour effects in vivo, reduced the severity of IL-2-induced capillary leakage in some organs and did not compromise anti-tumour efficacy of IL-2 therapy. Thus, L-NAME could be a valuable adjunct to IL-2-based cancer therapy. PMID:8546905

Orucevic, A.; Lala, P. K.

1996-01-01

219

Immunotherapy of murine sarcomas using lymphokine activated killer cells: optimization of the schedule and route of administration of recombinant interleukin-2.  

PubMed

Interleukin-2 (IL-2) at high doses or at low doses in concert with lymphokine-activated killer (LAK) cells can produce regression of established pulmonary and hepatic metastases from a variety of tumors in mice. IL-2 appears to mediate its antitumor effect through the generation of LAK cells in vivo from endogenous lymphocytes and by the stimulation of host and transferred LAK cell proliferation in tissues. In this paper we have investigated different strategies for IL-2 administration to determine which regimen produced maximal in vivo proliferation and optimal immunotherapeutic efficacy of LAK cells. Tissue expansion of lymphoid cells was assessed using an assay of in vivo labeling of dividing cells by the thymidine analogue, 5-[125I]iododeoxyuridine. The therapeutic effect of the different IL-2 administration protocols was determined by evaluating their efficacy in the treatment of established, 3-day pulmonary metastases from sarcomas in mice. The selection of IL-2 injection regimens for evaluation was based upon pharmacokinetic studies of IL-2 in mice. A single i.v. or i.p. dose yielded high peak IL-2 levels that could be measured for only a few hours after injection, while IL-2 given i.p. thrice daily produced titers that were detectable throughout the study periods (greater than or equal to 6 units/ml of serum after 100,000 units of IL-2 i.p. thrice daily). Using the proliferation and therapy models, we tested the same cumulative daily doses of IL-2 administered by i.v. or i.p. once daily, or i.p. thrice daily regimens. The i.p. thrice daily protocol stimulated greater lymphoid cell proliferation in the lungs, for example, than did the other regimens. Similarly, 300,000 units of IL-2 divided i.p. thrice daily were more successful in reducing metastases (n = 16) than was the entire dose given i.v. once daily (n = 190; P less than 0.05) or i.p. once daily (n = 71; P less than 0.05). When compared to the i.p. or i.v. once daily protocols, the i.p. thrice daily regimen for IL-2 also produced greater proliferation of exogenous LAK cells, as well as a more effective therapeutic outcome when IL-2 was combined with transferred LAK cells. Thus, sustained, lower levels of IL-2 were more effective than brief, high peak titers for stimulation of proliferation and antitumor activity. We then evaluated the effect of duration of IL-2 treatment as well as the number of LAK cell injections in the two models.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3486038

Ettinghausen, S E; Rosenberg, S A

1986-06-01

220

Antitumor efficacy of lymphokine-activated killer cells and recombinant interleukin-2 in vivo: survival benefit and mechanisms of tumor escape in mice undergoing immunotherapy.  

PubMed

The studies described in this paper showed that the combination of i.v.-transferred lymphokine-activated killer (LAK) cells and i.p. injections of recombinant interleukin-2 (RIL-2) was highly effective in vivo in reducing established pulmonary metastases of natural killer cell-resistant, MCA-105 sarcoma and B16 melanoma in mice. A 3-day in vitro incubation of normal C57BL/6 splenocytes in medium containing pure RIL-2 generated LAK cells that, when combined with RIL-2, reduced the mean number of established pulmonary micrometastases of the B16 melanoma and of the MCA-105 sarcoma from 179 and 140, respectively (in groups treated with Hanks' balanced salt solution alone), to 12 (P = 0.01) and 6 (P = 0.01), respectively. This combined immunotherapy also consistently resulted in significant prolongation of survival in mice with established, 3-day or 10-day pulmonary metastases of the MCA-105 sarcoma. Mice autopsied at time of death revealed a massive involvement of tumor in the lungs and liver in the group receiving Hanks' balanced salt solution alone compared to a small number of residual large lung or liver metastases in the group receiving LAK cells plus RIL-2. Experiments were designed to test whether variants existed in the original tumor cell inoculum that were resistant to killing by LAK cells and thus could account for the metastases that "escaped" the combined immunotherapy of LAK cells plus RIL-2 in vivo. Metastases of the MCA-105 sarcoma that escaped the combined therapy of LAK cells plus RIL-2 were dissected from the organs of mice upon autopsy and directly tested for susceptibility in vitro to lysis by LAK cells in 4-h and 18-h 51Cr release assays. Target cells derived from the metastases were lysed to an equivalent extent as those prepared from a fresh MCA-105 sarcoma that was growing s.c. In addition, successful reduction of pulmonary metastases established by the i.v. infusion of MCA-105 sarcoma cells obtained from metastases that escaped a prior round of therapy with LAK cells and RIL-2 could be achieved in vivo by the combined immunotherapy as well as by high doses of RIL-2 alone. Culture adapted, natural killer cell-resistant B16 melanoma cells surviving two successive treatments with LAK cells in vitro remained as susceptible to LAK cell lysis as untreated B16 melanoma cells in 18-h 51Cr release assays.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3484431

Mulé, J J; Ettinghausen, S E; Spiess, P J; Shu, S; Rosenberg, S A

1986-02-01

221

Increased vascular permeability in organs mediated by the systemic administration of lymphokine-activated killer cells and recombinant interleukin-2 in mice.  

PubMed

Significant tumor regressions in mice with established carcinomas, sarcomas, and melanomas and in humans with advanced cancers have been observed following immunotherapy with lymphokine-activated killer (LAK) cells and recombinant interleukin-2 (IL-2). However, dose escalations of LAK cells and IL-2 have been prevented by the development of a vascular leak syndrome (VLS). Although IL-2 alone can produce this VLS, we investigated the role of transferred LAK cells, generated by the incubation of syngeneic splenocytes in IL-2, in mediating this phenomenon. We used a murine model to quantitate the vascular leak by measuring the extravasation of iv injected 125I-bovine serum albumin. A permeability index (PI) was calculated by dividing the mean cpm of tissues from treated mice by those from control animals. The systemic transfer of LAK cells and IL-2 produced a significantly greater extravasation of albumin in the lungs, liver, and kidneys than after Hanks' balanced salt solution, IL-2, or LAK cells alone (in the lungs, for example, PI = 4.7, 1.4, and 1.6 after LAK cells and IL-2, LAK cells alone, and IL-2 alone, respectively). To eliminate the contribution to the leak by host lymphocytes, we irradiated mice before cell transfer. As compared to controls, LAK cells and IL-2 resulted in higher extravasation in the lungs, liver, kidneys, and spleen. However, a similar vascular leak was not observed in the lungs, liver, and kidneys after treatment with IL-2 plus fresh or cultured (without IL-2) splenocytes. Moreover, the combination of IL-2 excipient and LAK cells or IL-2 and irradiated LAK cells did not produce a fluid leak. The development of the VLS by LAK cells was directly related to the dose of concurrently administered IL-2 and the number of injected cells. Depletion of Thy 1.2-positive lymphocytes using antibody and complement prior to generating the LAK cells used in adoptive transfer did not abrogate the VLS in any of the organs tested. Similarly, depletion of L3T4 and Lyt-2 positive cells in vivo using monoclonal antibodies prior to harvesting spleens for generation of LAK cells also had no impact on the VLS. In contrast, in vitro treatment of LAK precursor cells with antibody to asialo-GM-1 plus complement completely eliminated the VLS when the depleted cells were cultured in IL-2 and subsequently transferred.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3258039

Ettinghausen, S E; Puri, R K; Rosenberg, S A

1988-04-01

222

Neo-adjuvant chemo-(immuno-)therapy of advanced squamous-cell head and neck carcinoma: a multicenter, phase III, randomized study comparing cisplatin?+?5-fluorouracil (5FU) with cisplatin +?5FU?+?recombinant interleukin 2  

Microsoft Academic Search

We carried out an open, randomized, phase III, multicenter clinical trial to compare, in neo-adjuvant setting, the clinical\\u000a response and toxicity of the combination chemotherapy cisplatin?+?5-FU with the same combination plus s.c. recombinant interleukin-2\\u000a (rIL-2) in patients with advanced (stage III–IV) head and neck squamous-cell carcinoma (HNSCC). Regimen A was the classical\\u000a Al Sarraf treatment: 100?mg\\/m2 cisplatin i.v. on day

Giovanni Mantovani; Vittorio Gebbia; Mario Airoldi; Cesare Bumma; Paolo Contu; Alessandro Bianchi; Massimo Ghiani; Daniela Dessì; Elena Massa; Luigi Curreli; Biancarosa Lampis; Paola Lai; Carlo Mulas; Antonio Testa; Ernesto Proto; Gabrio Cadeddu; Giorgio Tore

1998-01-01

223

Soluble interleukin 2 receptor levels and cervical neoplasia: results from a population-based case-control study in Costa Rica.  

PubMed

Progression from infection with human papillomavirus (HPV) to cervical cancer in some women is thought to involve a permissive host environment, one in which immune response is mobilized in an inappropriate manner. In a previous study (A. Hildesheim et al., Cancer Epidemiol. Biomark. Prev., 6: 807-813, 1997), increasing levels of soluble interleukin 2 receptor (sIL-2R), a known proxy for general immune activation, was found to be positively associated with increasing levels of cervical neoplasia. We attempted to confirm this finding by conducting a nested case-control study of 478 women within a 10,000-woman population-based cohort in Costa Rica. We selected for the study all of the women diagnosed (at enrollment into the cohort) with: (a) low-grade squamous intraepithelial lesions (LSIL, n = 191); (b) high-grade squamous intraepithelial lesions (HSIL, n = 130); or (c) cancer (n = 37). Controls were 120 cytologically normal, HPV-negative women selected from a random sample of the entire cohort. A questionnaire was administered to participants to elicit information on cervical cancer risk factors. All of the women received a pelvic examination during which cervical cells were collected and used for HPV DNA testing by PCR. Blood samples were also collected. Plasma obtained from the blood samples was tested for sIL-2R levels by ELISA. Results indicated that sIL-2R levels increased with age. Among controls, we observed that 44.3% of women over the age of 50 had high levels of sIL-2R (defined as >735 units/ml) compared with 15.8% of women <30 years of age (P = 0.008). When women with cervical disease (LSIL+) were compared with controls, women in the upper quartile of the sIL-2R distribution had an age-adjusted odds ratio (OR) of 2.1 [95% confidence interval (CI), 1.1-4.1]. Comparing each advancing state of neoplasia with its precursor, we found that women with LSIL had higher sIL-2R levels than controls (OR for upper quartile of sIL-2R, 2.3; 95% CI, 1.1-5.2; comparing LSIL cases with controls); women diagnosed with HSIL were similar to the LSIL group (OR for upper quartile of sIL-2R, 1.1; 95% CI, 0.5-2.4; comparing HSIL cases with LSIL cases); and those with cancer had higher sIL-2R levels than subjects with an HSIL diagnosis (OR for upper quartile of sIL-2R = 1.8; 95% CI, 0.5-7.1; comparing cancer cases with HSIL cases). These data suggest that among our study subjects, sIL-2R levels most likely rise as a response to the events of infection and cancerous invasion, but that sIL-2R levels are unlikely to be predictive of disease progression among women with LSIL. PMID:10090303

Ung, A; Kramer, T R; Schiffman, M; Herrero, R; Bratti, M C; Burk, R D; Swanson, C A; Sherman, M E; Hutchinson, M L; Alfaro, M; Morales, J; Balmaceda, I; Hildesheim, A

1999-03-01

224

Analyzing mRNA Expression Using Single mRNA Resolution Fluorescent In Situ Hybridization  

PubMed Central

As the product of transcription and the blueprint for translation, mRNA is the main intermediate product of the gene expression pathway. The ability to accurately determine mRNA levels is, therefore, a major requirement when studying gene expression. mRNA is also a target of different regulatory steps, occurring in different subcellular compartments. To understand the different steps of gene expression regulation, it is therefore essential to analyze mRNA in the context of a single cell, maintaining spatial information. Here, we describe a stepwise protocol for fluorescent in situ hybridization (FISH) that allows detection of individual mRNAs in single yeast cells. This method allows quantitative analysis of mRNA expression in single cells, permitting “absolute” quantification by simply counting mRNAs. It further allows us to study many aspects of mRNA metabolism, from transcription to processing, localization, and mRNA degradation. PMID:20946829

Zenklusen, Daniel; Singer, Robert H.

2011-01-01

225

Translation drives mRNA quality control  

PubMed Central

There are three predominant forms of co-translational mRNA surveillance: nonsense-mediated decay (NMD), no-go decay (NGD) and non-stop decay (NSD). While discussion of these pathways often focuses on mRNA fate, there is growing consensus that there are other important outcomes of these processes that must be simultaneously considered. Here, we seek to highlight similarities between NMD, NGD and NSD and their likely origins on the ribosome during translation. PMID:22664987

Shoemaker, Christopher J.; Green, Rachel

2015-01-01

226

Self-amplifying mRNA vaccines.  

PubMed

This chapter provides a brief introduction to nucleic acid-based vaccines and recent research in developing self-amplifying mRNA vaccines. These vaccines promise the flexibility of plasmid DNA vaccines with enhanced immunogenicity and safety. The key to realizing the full potential of these vaccines is efficient delivery of nucleic acid to the cytoplasm of a cell, where it can amplify and express the encoded antigenic protein. The hydrophilicity and strong net negative charge of RNA impedes cellular uptake. To overcome this limitation, electrostatic complexation with cationic lipids or polymers and physical delivery using electroporation or ballistic particles to improve cellular uptake has been evaluated. This chapter highlights the rapid progress made in using nonviral delivery systems for RNA-based vaccines. Initial preclinical testing of self-amplifying mRNA vaccines has shown nonviral delivery to be capable of producing potent and robust innate and adaptive immune responses in small animals and nonhuman primates. Historically, the prospect of developing mRNA vaccines was uncertain due to concerns of mRNA instability and the feasibility of large-scale manufacturing. Today, these issues are no longer perceived as barriers in the widespread implementation of the technology. Currently, nonamplifying mRNA vaccines are under investigation in human clinical trials and can be produced at a sufficient quantity and quality to meet regulatory requirements. If the encouraging preclinical data with self-amplifying mRNA vaccines are matched by equivalently positive immunogenicity, potency, and tolerability in human trials, this platform could establish nucleic acid vaccines as a versatile new tool for human immunization. PMID:25620012

Brito, Luis A; Kommareddy, Sushma; Maione, Domenico; Uematsu, Yasushi; Giovani, Cinzia; Berlanda Scorza, Francesco; Otten, Gillis R; Yu, Dong; Mandl, Christian W; Mason, Peter W; Dormitzer, Philip R; Ulmer, Jeffrey B; Geall, Andrew J

2015-01-01

227

Expression of activation antigens CD69, HLA-DR, interleukin-2 receptor-alpha (IL-2R alpha) and IL-2R beta on T cells of human decidua at an early stage of pregnancy.  

PubMed Central

T cells of human decidua at an early stage of pregnancy were examined by flow cytometry for the expression of the T-cell-activation antigens CD69, HLA-DR, interleukin-2 receptor-alpha (IL-2R alpha) and IL-2R beta. The decidua contained a small number of T cells and both CD4+ and CD8+ subsets expressed CD69, HLA-DR, IL-2R alpha and IL-2R beta antigens significantly whereas, in peripheral blood, only a small number of T cells expressed these activation antigens. These findings indicate that T cells in the decidua in the first trimester of pregnancy are regionally activated. PMID:1592443

Saito, S; Nishikawa, K; Morii, T; Narita, N; Enomoto, M; Ichijo, M

1992-01-01

228

Inhibition of leukocyte function and interleukin-2 gene expression by 2-methylarachidonyl-(2'-fluoroethyl)amide, a stable congener of the endogenous cannabinoid receptor ligand anandamide  

SciTech Connect

Arachidonylethanolamide (anandamide, AEA) has been identified as an endogenous ligand for cannabinoid receptors CB1 and CB2. Characterization of the direct cannabimimetic actions of anandamide has been hampered by its short duration of action and rapid degradation in in vivo and in vitro systems to arachidonic acid, a precursor in the biosynthesis of a broad range of biologically active molecules. In the present studies, we utilized 2-methylarachidonyl-(2'-fluoroethyl)amide (F-Me-AEA), an analog of anandamide resistant to enzymatic degradation, to determine whether F-Me-AEA modulated T cell function similar to that of plant-derived cannabinoids. Indeed, F-Me-AEA at low micromolar concentrations exhibited a marked inhibition of phorbol ester plus calcium ionophore (PMA/Io)-induced IL-2 protein secretion and steady state mRNA expression. Likewise, a modest suppression of the mixed lymphocyte response was observed in the presence of F-Me-AEA indicating an alteration in T cell responsiveness to allogeneic MHC class II antigens. F-Me-AEA was also found to modestly inhibit forskolin-stimulated adenylate cyclase activity in thymocytes and splenocytes, a hallmark of cannabinoid receptor agonists. Further characterization of the influence of F-Me-AEA on the cAMP signaling cascade revealed an inhibition of CREB-1/ATF-1 phosphorylation and subsequently, an inhibition of CRE DNA binding activity. Characterization of nuclear binding proteins further revealed that NF-AT and, to a lesser extent, NF-{kappa}B DNA binding activities were also suppressed. These studies demonstrate that F-Me-AEA modulates T cell function in a similar manner to plant-derived and endogenous cannabinoids and therefore can be utilized as an amidase- and hydrolysis-resistant endogenous cannabinoid.

Kaplan, Barbara L.F. [Department of Pharmacology and Toxicology and Center for Integrative Toxicology, Michigan State University, East Lansing, MI 48824 (United States); Ouyang Yanli [Department of Pharmacology and Toxicology and Center for Integrative Toxicology, Michigan State University, East Lansing, MI 48824 (United States); Herring, Amy [Department of Pharmacology and Toxicology and Center for Integrative Toxicology, Michigan State University, East Lansing, MI 48824 (United States); Yea, Sung Su [Department of Biochemistry, College of Medicine, Inje University, Pusan 614-735 (Korea, Republic of); Razdan, Raj [Organix Inc., Woburn, MA 01801 (United States); Kaminski, Norbert E. [Department of Pharmacology and Toxicology and Center for Integrative Toxicology, Michigan State University, East Lansing, MI 48824 (United States)]. E-mail: kamins11@msu.edu

2005-06-01

229

Vaccination with Messenger RNA (mRNA)  

Microsoft Academic Search

Both DNA and mRNA can be used as vehicles for gene therapy. Because the immune system is naturally activated by foreign nucleic\\u000a acids thanks to the presence of Toll-like Receptors (TLR) in endosomes (TLR3, 7, and 8 detect exogenous RNA, while TLR9 can\\u000a detect exogenous DNA), the delivery of foreign nucleic acids usually induces an immune response directed against the

Steve Pascolo

230

Sensitivity of mRNA Translation  

E-print Network

Using the dynamic mean-field approximation of the totally asymmetric simple exclusion process (TASEP), we investigate the effect of small changes in the initiation, exit, and elongation rates along the mRNA strand on the steady state protein translation rate. We focus on two special cases where exact closed-form expressions for the translation rate sensitivity can be derived. We discuss the ramifications of our results in the context of functional genomics, molecular evolution, and synthetic biology.

Gilad Poker; Michael Margaliot; Tamir Tuller

2014-09-18

231

Regulation of yeast development by mRNA methylation  

E-print Network

The internal methylation of mRNA post-transcriptionally is an essential component of the mRNA editing machinery in virtually every eukaryotic system. Despite this ubiquity, little is known about the relevance, consequences ...

Agarwala, Sudeep D

2012-01-01

232

Understanding regulation of mRNA by RNA binding proteins  

E-print Network

Posttranscriptional regulation of mRNA by RNA-binding proteins plays key roles in regulating the transcriptome over the course of development, between tissues and in disease states. The specific interactions between mRNA ...

Robertson, Alexander De Jong

2014-01-01

233

Enhanced protective efficacy against Mycobacterium tuberculosis afforded by BCG prime-DNA boost regimen in an early challenge mouse model is associated with increased splenic interleukin-2-producing CD4 T-cell frequency post-vaccination.  

PubMed

The development of improved vaccines and vaccination strategies against Mycobacterium tuberculosis has been hindered by a limited understanding of the immune correlates of anti-tuberculosis protective immunity. Simple measurement of interferon-? frequency or production per se does not provide adequate prediction of immune protection. In this study, we examined the relationship between T-cell immune responses and protective efficacy conferred by the heterologous vaccination strategy, bacillus Calmette-Guérin (BCG) prime-Ag85A DNA boost (B/D), in an early challenge mouse model of pulmonary tuberculosis. The results demonstrated that mice vaccinated with the B/D regimen had a significantly reduced bacillary load compared with BCG-vaccinated mice, and the reduction in colony-forming units was associated with decreased pathology and lower levels of inflammatory cytokines in the infected lungs. Further analysis of immunogenicity showed that the superior protection afforded by the B/D regimen was associated with significantly increased frequency of splenic interleukin-2 (IL-2) -producing CD4 T cells and increased IL-2 production when measured as integrated mean fluorescence intensity post-vaccination as well. These data suggest that measurement of elevated frequency of IL-2-producing CD4 T cells or IL-2 production in the spleens of vaccinated mice can predict vaccine efficacy, at least in the B/D strategy, and add to the accumulating body of evidence suggesting that BCG prime-boost strategies may be a useful approach to the control of M. tuberculosis infection. PMID:24965530

Kang, Han; Yuan, Qin; Ma, Hui; Hu, Zhi-Dong; Han, De-Ping; Wu, Kang; Lowrie, Douglas B; Fan, Xiao-Yong

2014-12-01

234

Regulation of cell-type-specific interleukin-2 receptor alpha-chain gene expression: potential role of physical interactions between Elf-1, HMG-I(Y), and NF-kappa B family proteins.  

PubMed Central

The interleukin 2 receptor alpha-chain (IL-2R alpha) gene is rapidly and potently induced in T cells in response to mitogenic stimuli. Previously, an inducible enhancer between nucleotides -299 and -228 that contains NF-kappa B and CArG motifs was identified. We now report the characterization of a second essential positive regulatory element located between nucleotides -137 and -64 that binds Elf-1 and HMG-I(Y). This element had maximal activity in lymphoid cells, paralleling the cell type specificity of Elf-1 expression. Transcription from the IL-2R alpha promoter was inhibited when either the Elf-1 or the HMG-I(Y) binding site was mutated. Coexpression of both proteins activated transcription of the -137 to -64 element in COS-7 cells. Elf-1 physically associated with HMG-I and with NF-kappa B p50 and c-Rel in vitro, suggesting that protein-protein interactions might functionally coordinate the actions of the upstream and downstream positive regulatory elements. This is the first report of a physical interaction between an Ets family member and NF-kappa B family proteins. These findings provide significant new insights into the protein-protein and protein-DNA interactions that regulate cell-type-specific and inducible IL-2R alpha gene expression and also have implications for other genes regulated by Elf-1 and NF-kappa B family proteins. PMID:7862168

John, S; Reeves, R B; Lin, J X; Child, R; Leiden, J M; Thompson, C B; Leonard, W J

1995-01-01

235

Natural killer (NK) cell activity in human long-term bone marrow cultures (LTBMC): effects of interleukin-2 (IL-2) and granulocyte-macrophage colony-stimulating factor (GM-CSF) on the progenitor cells.  

PubMed Central

Human bone marrow-derived progenitor cells were studied in a long-term bone marrow culture system (LTBMC) dependent on an autologous stroma cell layer. The establishment of the stromal cell layer was facilitated by using marrow obtained from small pieces of sternum, which was cultured for 4 weeks without addition of exogenous growth factors. After this period, the response of LTBMC to two different cytokines [recombinant human interleukin-2 (rhIL-2) and recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF)] was investigated. Our results show proliferation in response to both cytokines and induction of differentiation of cells able to bind IL-2 and/or GM-CSF again. The two cytokines also generate cells responding to rhGM-CSF by colony formation. However, a difference with respect to morphology, phenotype and cytotoxic function of cells in the LTBMC, was noted between the two cytokines. Cells with large granular lymphocyte (LGL) morphology and cytotoxic activity against K562 and Daudi were generated only in the rhIL-2-supplemented LTBMC. This was compatible with a higher frequency of cells expressing the CD56+ phenotype in the IL-2-stimulated LTBMC as compared to the GM-CSF supplemented LTBMC. Our results also demonstrate the existence of a population of myeloid progenitor cells (CD33+) with ability to bind IL-2 in fresh bone marrow (BM). Images Figure 1 Figure 5 PMID:1634251

Sitnicka, E; Hansson, M

1992-01-01

236

mRNA localisation during development.  

PubMed

Although there are many differences, mRNA localisations in the Xenopus oocyte show some tantalizing similarities to those occurring in Drosophila development. As in Drosophila, transcripts localise to opposite poles of the oocyte, this localisation is hierarchical and occurs in a multistep process in which localisation is followed by anchoring at the cortex. This distinction between initial transport and long-term maintenance reflects the dynamic nature of the cytoskeleton: the microtubule tracks form and reform according to the needs of the cell so that stable localisation must be mediated by a more constant structure--the cortex. A possible exception is the localisation of gurken mRNA where it is unknown whether there are separate mechanisms for transport to and maintenance at the oocyte nucleus. However, gurken is responsible for the transmission of a transitory signal; once this has been received, and the fate of the recipient follicle cells determined, there is no further need for localisation. It is possible that the time scale over which the localisation machinery is stable is sufficient for transmission of this signal without the need for a separate maintenance phase. The existence of a nanos homologue, Xcat-2 (Mosquera et al., 1993), associated with the Xenopus germ plasm is particularly interesting because of the morphological and functional similarities between Drosophila polar granules, Caenorhabditis P-granules, and Xenopus germ plasm. These electron-dense protein-RNA complexes are maternally supplied and in each case segregate with the germ line. These granules may represent a fundamental conserved pathway to germ-cell specification and it is now at least a possibility that they are also involved in establishing the embryonic axis through translational repression. In the case of Drosophila, this occurs through localised nanos acting (via Pumilio) on nanos response elements in hunchback mRNA. No such regulatory pair has yet been demonstrated in C. elegans or X. laevis, but each contains a candidate for one half of the interaction: glp-1 could be a target for an unidentified nanos-like protein; Xcat-2 may control translation of an unknown NRE-containing mRNA. Another common feature of mRNA localisation is that in every case where the targeting signal has been determined, it has been mapped to a region of the 3' UTR capable of forming an extensive secondary structure (e.g., David and Ish-Horowicz, 1991; Dalby and Glover, 1992; Gavis and Lehmann, 1992; Kim-Ha et al., 1993; Kislauskis et al., 1993, 1994; Lantz and Schedl, 1994). In several cases, translational control and transcript stability signals have also been mapped to these regions (Jackson and Standart, 1990; Standart et al., 1990; Standart and Hunt, 1990; Davis and Ish-Horowicz, 1991; Wharton and Struhl, 1991; Dalby and Glover, 1993; Evans et al., 1994; Kim-Ha et al., 1995). The large secondary structures may provide a means for stably exposing sequence-specific regions of RNA to proteins. Due to the ease with which RNA forms base pairs, it is likely that rather than remaining single-stranded, RNA within the cell forms some sort of secondary structure. The geometry of purely double-stranded RNA does not permit sequence specific interactions between proteins and the bases because the major groove is inaccessible to amino acid side chains (Weeks and Crothers, 1993). However, the distortions to the dsRNA helix provided by bulges, pseudoknots, and the single-strand loop regions in stem-loop structures do present sequence information that can be "read" by proteins. The extensive 3'UTRs may produce a stable secondary structure which ensures that regulatory elements remain exposed in such regions rather than hidden in double-stranded stems. (ABSTRACT TRUNCATED) PMID:8612958

Micklem, D R

1995-12-01

237

Post-Transcriptional Control: mRNA Translation, Localization and Turnover 1531 Cytoplasmic deadenylation: regulation of mRNA  

E-print Network

Post-Transcriptional Control: mRNA Translation, Localization and Turnover 1531 Cytoplasmic of the protein, even after inactivation of transcription. On the other hand, shortening of the poly(A) tail (deadenylation) slows down translation of the mRNA, or prevents it entirely, by inducing mRNA decay. Thus

Passmore, Lori A.

238

Messenger RNA (mRNA) nanoparticle tumour vaccination  

NASA Astrophysics Data System (ADS)

Use of mRNA-based vaccines for tumour immunotherapy has gained increasing attention in recent years. A growing number of studies applying nanomedicine concepts to mRNA tumour vaccination show that the mRNA delivered in nanoparticle format can generate a more robust immune response. Advances in the past decade have deepened our understanding of gene delivery barriers, mRNA's biological stability and immunological properties, and support the notion for engineering innovations tailored towards a more efficient mRNA nanoparticle vaccine delivery system. In this review we will first examine the suitability of mRNA for engineering manipulations, followed by discussion of a model framework that highlights the barriers to a robust anti-tumour immunity mediated by mRNA encapsulated in nanoparticles. Finally, by consolidating existing literature on mRNA nanoparticle tumour vaccination within the context of this framework, we aim to identify bottlenecks that can be addressed by future nanoengineering research.

Phua, Kyle K. L.; Nair, Smita K.; Leong, Kam W.

2014-06-01

239

Evodiamine, a plant alkaloid, induces calcium/JNK-mediated autophagy and calcium/mitochondria-mediated apoptosis in human glioblastoma cells.  

PubMed

Glioblastomas, the most common primary gliomas, are characterized by increased invasion and difficult therapy. Major clinical medicines for treating gliomas merely extend the survival time for a number of months. Therefore, development of new agents against gliomas is important. Autophagy, a process for degrading damaged organelles and proteins, is an adaptive response to environmental stress. However, the role of autophagy in glioblastoma development still needs to be further investigated. Evodiamine, a major alkaloid isolated from Evodia rutaecarpa Bentham, has various pharmacological activities, such as inhibiting tumor growth and metastatic properties. However, the effects of evodiamine on glioblastomas and their detailed molecular mechanisms and autophagy formation are not well understood. In this study, we observed that evodiamine induced dose- and time-dependent apoptosis in glioma cells. Blockade of calcium channels in endoplasmic reticulum (ER) significantly reduced evodiamine-induced cytosolic calcium elevation, apoptosis, and mitochondrial depolarization, which suggests that evodiamine induces a calcium-mediated intrinsic apoptosis pathway. Interestingly, autophagy was also enhanced by evodiamine, and had reached a plateau by 24h. Pharmacological inhibition of autophagy resulted in increased apoptosis and reduced cell viability. Inhibition of ER calcium channel activation also significantly reduced evodiamine-induced autophagy. Inactivation of c-Jun N-terminal kinases (JNK) suppressed evodiamine-mediated autophagy accompanied by increased apoptosis. Furthermore, evodiamine-mediated JNK activation was abolished by BAPTA-AM, an intracellular calcium scavenger, suggesting that evodiamine mediates autophagy via a calcium-JNK signaling pathway. Collectively, these results suggest that evodiamine induces intracellular calcium/JNK signaling-mediated autophagy and calcium/mitochondria-mediated apoptosis in glioma cells. PMID:23774672

Liu, Ann-Jeng; Wang, Sheng-Hao; Chen, Ku-Chung; Kuei, Hsiu-Ping; Shih, Yung-Luen; Hou, Sz-Ying; Chiu, Wen-Ta; Hsiao, Sheng-Huang; Shih, Chwen-Ming

2013-09-01

240

Relative strength of the mitogenic and interleukin-2-production-inducing activities of staphylococcal exotoxins presumed to be causative exotoxins of toxic shock syndrome: toxic shock syndrome toxin-1 and enterotoxins A, B and C to murine and human T cells.  

PubMed Central

Several observations suggest that staphylococcal enterotoxins A, B and C (SEA, SEB and SEC, respectively), in addition to toxic shock syndrome toxin-1 (TSST-1), are causative exotoxins of toxic shock syndrome (TSS). Based on the view that polyclonal T cell activation with the causative exotoxins, resulting in over-production of lymphokines, is involved in the development of the pathological changes observed in TSS, we investigated the activities of these four exotoxins to induce proliferation and interleukin 2 production in murine and human lymphocytes by using in vitro culture systems. The results showed that all these exotoxins are strong polyclonal inducers of proliferation and interleukin 2 production in human T cells, whereas TSST-1 and SEA are strong and SEB and SEC are weak polyclonal inducers in murine T cells. These results suggest that SEA, SEB and SEC, in addition to TSST-1, are possibly involved as causative exotoxins in the development of the pathological changes observed in TSS. PMID:2784735

Uchiyama, T; Kamagata, Y; Yan, X J; Kawachi, A; Fujikawa, H; Igarashi, H; Okubo, M

1989-01-01

241

The Majority of Epidermal T Cells in Psoriasis Vulgaris Lesions can Produce Type 1 Cytokines, Interferon-?, Interleukin2, and Tumor Necrosis Factor-?, Defining TC1 (Cytotoxic T Lymphocyte) and TH1 Effector Populations:1 a Type 1 Differentiation Bias is also Measured in Circulating Blood T Cells in Psoriatic Patients  

Microsoft Academic Search

Psoriasis vulgaris is a skin disease potentially mediated by pro-inflammatory cytokines produced by type 1 lesional T cells. The capability of individual T cells to produce these cytokines in lesional skin is not known. In this study we measured the ability of lesional and peripheral blood T cells to produce intracellular interferon-?, tumor necrosis factor-?, interleukin-2, interleukin-4, and interleukin-10 proteins

Lisa M Austin; Maki Ozawa; Toyoko Kikuchi; Ian B Walters; James G Krueger

1999-01-01

242

Molecular cloning of gp42, a cell-surface molecule that is selectively induced on rat natural killer cells by interleukin 2: glycolipid membrane anchoring and capacity for transmembrane signaling  

PubMed Central

We have previously shown that in vitro culture of rat natural killer (NK) cells in high concentrations of recombinant interleukin 2 (rIL-2) leads to the expression of a surface glycoprotein with a molecular mass of approximately 42 kD. This glycoprotein, gp42, is not induced on other lymphocytes and thus provides a lineage-specific marker for rIL-2- activated NK cells. We here present the nucleotide sequence for gp42 cDNA. The open reading frame encodes 233 amino acids with three potential sites for N-linked glycosylation. The deduced amino acid sequence lacks an apparent transmembrane domain and instead contains a hydrophobic COOH terminus that is characteristic of glycosylphosphatidylinositol (GPI)-anchored surface proteins. Consistent with this, gp42 is cleaved from the NK-like cell line, RNK- 16, by phosphatidylinositol-specific phospholipase C (PI-PLC), as is gp42 expressed on CHO cells that have been transformed with gp42 cDNA. On rIL-2-activated NK cells, gp42 is resistant to PI-PLC, though our studies suggest that gp42 on these cells is still expressed as a GPI- anchored molecule. Antibody to gp42 stimulates in RNK-16 cells an increase in inositol phosphates and in intracellular calciu, signals that are associated with the activation of lymphocytes, including NK cells. rIL-2-activated NK cells, however, lack this response to gp42 as well as to other stimuli. Thus, gp42, the only NK-specific activation antigen, is a GPI-anchored surface molecule with the capacity to stimulate transmembrane signaling. PMID:1845873

1991-01-01

243

Interleukin-2 Engineering for improved therapeutic effectiveness  

E-print Network

(cont.) (K[d] [approximately] 10pM) for its private alpha receptor subunit, unlike wild-type IL-2 (K[d] [approximately] 10 nM). IL-2 mutants with picomolar affinity for IL-2R? stimulate T cell growth responses quantitatively ...

Rao, Balaji Madhav, 1978-

2004-01-01

244

Premature translation termination mediates triosephosphate isomerase mRNA degradation  

SciTech Connect

The authors characterized an anemia-inducing mutation in the human gene for triosephosphate isomerase (TPI) that resulted in the production of prematurely terminated protein and mRNA with a reduced cytoplasmic half-life. The mutation converted a CGA arginine codon to a TGA nonsense condon and generated a protein of 188 amino acids, instead of the usual 248 amino acids. To determine how mRNA primary structure and translation and influence mRNA stability, in vitro-mutagenized TPI alleles were introduced into cultured L cells and analyzed for their effect on TPI RNA metabolism. Results indicated that mRNA stability is decreased by all nonsense and frameshift mutations. To determine the relative contribution of the changes in mRNA structure and translation to the altered half-life, the effects of individual mutations were compared with the effects of second-site reversions that restored translation termination to normal. All mutations that resulted in premature translation termination reduced the mRNA half-life solely or mainly by altering the length of the mRNA that was translated. The only mutation that altered translation termination and that reduced the mRNA half-life mainly by affecting the mRNA structure was an insertion that shifted termination to a position downstream of the normal stop codon.

Daar, I.O.; Maquat, L.E.

1988-02-01

245

Premature translation termination mediates triosephosphate isomerase mRNA degradation.  

PubMed Central

We characterized an anemia-inducing mutation in the human gene for triosephosphate isomerase (TPI) that resulted in the production of prematurely terminated protein and mRNA with a reduced cytoplasmic half-life. The mutation converted a CGA arginine codon to a TGA nonsense codon and generated a protein of 188 amino acids, instead of the usual 248 amino acids. To determine how mRNA primary structure and translation influence mRNA stability, in vitro-mutagenized TPI alleles were introduced into cultured L cells and analyzed for their effect on TPI RNA metabolism. Results indicated that mRNA stability is decreased by all nonsense and frameshift mutations. To determine the relative contribution of the changes in mRNA structure and translation to the altered half-life, the effects of individual mutations were compared with the effects of second-site reversions that restored translation termination to normal. All mutations that resulted in premature translation termination reduced the mRNA half-life solely or mainly by altering the length of the mRNA that was translated. The only mutation that altered translation termination and that reduced the mRNA half-life mainly by affecting the mRNA structure was an insertion that shifted termination to a position downstream of the normal stop codon. Images PMID:2832737

Daar, I O; Maquat, L E

1988-01-01

246

Single mRNA Tracking in Live Cells  

PubMed Central

Asymmetric distribution of mRNA is a prevalent phenomenon observed in diverse cell types. The posttranscriptional movement and localization of mRNA provides an important mechanism to target certain proteins to specific cytoplasmic regions of their function. Recent technical advances have enabled real-time visualization of single mRNA molecules in living cells. Studies analyzing the motion of individual mRNAs have shed light on the complex RNA transport system. This chapter presents an overview of general approaches for single particle tracking and some methodologies that are used for single mRNA detection. PMID:20580973

Park, Hye Yoon; Buxbaum, Adina R.; Singer, Robert H.

2011-01-01

247

Regulation of mRNA Translation and Stability  

E-print Network

Regulation of mRNA Translation and Stability by microRNAs Marc Robert Fabian,1 Nahum Sonenberg,1, Montreal, Quebec, H3G 1Y6, Canada; email: marc.fabian@mail.mcgill.ca, nahum.sonenberg@mcgill.ca 2 Friedrich, plants, and protozoa. miRNAs function posttranscriptionally by usually base-pairing to the mRNA 3

Bedwell, David M.

248

MELEAGRIS GALLOPAVO PROGLUCAGON (GCG), CLASS A MRNA, COMPLETE CDS  

Technology Transfer Automated Retrieval System (TEKTRAN)

In mammals, the proglucagon gene produces a single identical mRNA transcript in pancreas and intestine. This transcript encodes glucagon and two glucagon-like peptide hormones (GLP-1 and GLP-2). Previously, we identified two distinct proglucagon mRNA transcripts in chickens, one that encodes glucag...

249

MELEAGRIS GALLOPAVO PROGLUCAGON (GCG), CLASS B MRNA, COMPLETE CDS  

Technology Transfer Automated Retrieval System (TEKTRAN)

In mammals, the proglucagon gene produces a single identical mRNA transcript in pancreas and intestine. This transcript encodes glucagon and two glucagon-like peptide hormones (GLP-1 and GLP-2). Previously, we identified two distinct proglucagon mRNA transcripts in chickens, one that encodes glucag...

250

Effects of gamma interferon, tumor necrosis factor alpha, and interleukin-2 on infection and proliferation of Theileria parva-infected bovine lymphoblasts and production of interferon by parasitized cells.  

PubMed Central

Theileria parva is a protozoan parasite that infects bovine B cells and alpha beta and gamma delta T cells and transforms them into continually proliferating cells. CD4+ T. parva-antigen-specific immune T cells have been shown to produce cytokines in response to stimulation with parasitized cells, and T. parva-infected lymphocytes produce and consume T-cell growth factors and interleukin-2 (IL-2). To ascertain the role of T-cell cytokines on T. parva infections, we evaluated recombinant gamma interferon (rIFN-gamma), rIL-2, and tumor necrosis factor alpha (rTNF-alpha) for their effects on establishment, proliferation, and survival of parasitized cells. The results indicate that neither rIFN-gamma nor rTNF-alpha had an enhancing or inhibitory effect on the growth of established T. parva-infected T-cell clones, whereas bovine rIL-2 increased the proliferation of infected B-cell and alpha beta T-cell clones but not that of gamma delta T-cell clones. To evaluate the effects of the cytokines on establishment of parasitized cell lines, peripheral blood mononuclear cells were cultured in their presence immediately following infection with T. parva sporozoites. Neither rIFN-gamma nor rIL-2 altered the proportion of cells initially developing schizonts, but both enhanced establishment of infected cell lines by about twofold. In contrast, rTNF-alpha resulted in about a 33% decrease in the proportion of schizont-infected cells. Inhibitory effects on establishment of parasitized cell lines by rTNF-alpha were no longer apparent by 12 days following infection. Tests conducted during this study indicated that T. parva-infected lymphocytes also spontaneously produce IFN that is neutralized by acidic pH treatment. In conclusion, we speculate that none of these T-cell cytokines are likely to have a profound inhibitory effect in vivo on T. parva infections. Instead, IFN-gamma and IL-2 may facilitate the establishment of infection by T. parva. PMID:1937812

DeMartini, J C; Baldwin, C L

1991-01-01

251

Selective Localization of Arc mRNA in Dendrites Involves Activity- and Translation-Dependent mRNA Degradation  

PubMed Central

Arc is an immediate early gene that is unique among neuronal mRNAs because its transcripts are transported into dendrites and accumulate near activated synapses, presumably to be translated locally. These qualities pose Arc as playing an important, yet not fully understood, role in the activity-dependent modifications of synapses that are thought to underlie memory storage. Here we show in vivo in rats that newly synthesized Arc mRNA accumulates at activated synapses and that synaptic activity simultaneously triggers mRNA decay that eliminates Arc mRNA from inactive dendritic domains. Arc mRNA degradation occurs throughout the dendrite and requires both NMDA receptor activation and active translation. Synaptic activation did not lead to decreases in another dendritic mRNA (?CaMKII), indicating that there is not a general activation of mRNA degradation in dendrites. These data reveal a novel mechanism for controlling mRNA distribution within dendrites and highlight activity-dependent mRNA degradation as a regulatory process involved in synaptic plasticity. PMID:24671994

Farris, Shannon; Lewandowski, Gail; Cox, Conor D.

2014-01-01

252

mRNA quantification via second harmonic super resolution microscopy  

NASA Astrophysics Data System (ADS)

Cell-specific information on quantity and localization of key mRNA transcripts in single-cell level are critical to the assessment of cancer risk, therapy efficacy, and effective prevention strategies. However, most available technologies for mRNA detection rely on cell extraction that inherently destroys the tissue context and provide only average expression levels from cell populations or whole tissues. In this paper, we proposed a novel super resolution concept, second harmonic generation (SHG) super-resolution microscopy (SHaSM), and apply that to detect single short mRNA transcript, Her2 mRNA, beyond the diffraction limit. Nano-sized SHG crystals, barium titanium oxide BaTiO3 (BTO), were functionalized with two complimentary strands of Her2 mRNA after the chemical surface-modification. Dimer schematic was used to improve the specificity of detection and quantification, where two BTO monomers bind to the Her2 mRNA to form a dimer and being visualized via the SHaSM. SHaSM is able to detect single BTO nanocrystal with ~20 nm spatial resolution, and differentiate BTO dimers (Her2 mRNA) from BTO monomers (non-specific bounded BTO nanocrystal) with high specificity.

Liu, Jing; Cho, Il-Hoon; Kadam, Ulhas; Irudayaraj, Joseph

2014-03-01

253

A bispecific single-chain antibody directed against EpCAM\\/CD3 in combination with the cytokines interferon ? and interleukin-2 efficiently retargets T and CD3 + CD56 + natural-killer-like T lymphocytes to EpCAM-expressing tumor cells  

Microsoft Academic Search

Cytokine-induced killer cells (CIK), generated in?vitro from peripheral blood mononuclear cells (PBMC) by addition of interferon\\u000a ? (IFN?), interleukin-2 (IL-2), IL-1 and a monoclonal antibody (mAb) against CD3, are highly efficient cytotoxic effector\\u000a cells with the CD3+CD56+ phenotype. In this study, we evaluated whether the cytotoxicity of these natural-killer-like T lymphocytes against the colorectal\\u000a tumor cell line HT29 can be

Dimitri Flieger; Peter Kufer; Imke Beier; Tilman Sauerbruch; Ingo G. H. Schmidt-Wolf

2000-01-01

254

Medium weight neurofilament mRNA in goldfish Mauthner axoplasm  

PubMed Central

Although axons are generally considered to lack the ability to synthesize proteins, the Mauthner axon (M-axon) of the goldfish has been reported to contain some of the basic components of the translational machinery, such as transfer RNA (tRNA), ribosomal RNA (rRNA), and ribosomes. To determine if the M-axon also contains mRNA, we isolated samples of M-axoplasm free of glial contamination as demonstrated by the absence of glial-specific mRNA and protein. Reverse transcription-polymerase chain reaction (RT-PCR) of M-axoplasmic cDNA in the presence of primers for the goldfish medium-weight neumfilament (NF-M) gene produced a single product of the expected length for RT-PCR amplification of goldfish NF-M mRNA. This mRNA might direct protein synthesis of NF-M within the M-axoplasm. PMID:8858614

Weiner, Orion D.; Zorn, Aaron M.; Krieg, Paul A.; Bittner, George D.

2010-01-01

255

5'Terminal structure and mRNA stability  

Microsoft Academic Search

Reovirus mRNAs with 5'-terminal m7GpppGm or GpppG are more stable than mRNA containing unblocked ppG 5'-ends when injected into Xenopus laevis oocytes or incubated in cell-free protein synthesising extracts of wheat germ and mouse L cells. The greater stability of mRNA with blocked 5' termini is not dependent upon translation but seems to result from protection against 5'-exonucleolytic degradation.

Yasuhiro Furuichi; Alba Lafiandra; Aaron J. Shatkin

1977-01-01

256

Regulation of Bone Sialoprotein mRNA by Steroid Hormones  

Microsoft Academic Search

In this report we demonstrate an increase in the steady-state level of bone sialoprotein (BSP) mRNA in rat calvaria and a rat osteosarcoma cell line (ROS 17\\/2.8) after treatment with the synthetic gluco- corticoid, dexamethasone. In contrast, 1.25- dthydroxyvitamin D3 reduced the amount of BSP mRNA in calvaria and inhibited the dexamethasone in- duction in ROS 17\\/2.8 cells. The increase

Bodil Jirskog-Hed; Sara Axelsson; Dick HeinegArd

2010-01-01

257

The highways and byways of mRNA decay  

Microsoft Academic Search

When considering the control of gene expression, the focus has traditionally been on transcriptional regulation. Recently, however, the large contribution made by mRNA decay has become difficult to ignore. Large-scale analyses indicate that as many as half of all changes in the amounts of mRNA in some responses can be attributed to altered rates of decay. In this article, we

Nicole L. Garneau; Jeffrey Wilusz; Carol J. Wilusz

2007-01-01

258

Erythropoietin Receptor mRNA Expression in Human Endothelial Cells  

Microsoft Academic Search

A previous report demonstrated that endothelial cells have erythropoietin receptors and respond to this hormone with enhanced proliferation. The present study demonstrates the existence of mRNA for erythropoietin receptor in human umbilical vein endothelial cells. We have reverse transcribed mRNA of endothelial cells and then used different PCR primers to amplify erythropoietin receptor target cDNA between exons 5 and 6

Athanasius Anagnostou; Ziyao Liu; Manfred Steiner; Kyung Chin; Eun S. Lee; Noubar Kessimian; Constance T. Noguchi

1994-01-01

259

Costimulation by B7-1 and LFA-3 targets distinct nuclear factors that bind to the interleukin-2 promoter: B7-1 negatively regulates LFA-3-induced NF-AT DNA binding.  

PubMed Central

We have characterized the regulation of nuclear factors involved in transcriptional control of the interleukin-2 (IL-2) promoter-enhancer activity in Jurkat T cells stimulated with superantigen presented on HLA-DR transfectants combined with the ligands LFA-3 (CD58) and B7-1 (CD80). Gel shift analyses showed that NF-AT was strongly induced in LFA-3-costimulated Jurkat T cells, suggesting that NF-AT is a key target nuclear factor for the CD2-LFA-3 pathway. Studies using HLA-DR-B7-1-LFA-3 triple transfectants showed that the LFA-3-induced NF-AT DNA binding activity was negatively regulated by B7-1 costimulation. In contrast, induction of a CD28 response complex containing only c-Rel proteins was seen after B7-1 costimulation. Both LFA-3 costimulation and B7-1 costimulation induced the AP-1 and NF-kappaB nuclear factors. Distinct compositions of the NF-AT complexes were seen in B7-1- and LFA-3-costimulated cells. LFA-3 induced primarily Jun-D, Fra-1, and Fra-2, while B7-1 induced June-D-Fos complexes. In contrast, AP-1 and NF-kappaB complexes induced in B7-1- and LFA-3-costimulated T cells showed similar contents. Transient transfection of Jurkat T cells with a construct encoding the IL-2 enhancer-promoter region (position -500 to +60) linked to a luciferase reporter gene revealed that B7-1 costimulation was required to induce strong transcriptional activity. Combined B7-1-LFA-3 costimulation resulted in a synergistic increase in IL-2 transcriptional activity. Multimers of the AP-1, NF-AT, NF-kappaB, and CD28 response elements showed distinct kinetics and activity after LFA-3 and B7-1 costimulation and revealed that B7-1 and LFA-3 converge to superinduce transcriptional activity of the AP-1, NF-AT, and CD28 response elements. Transcriptional studies with an IL-2 enhancer-promoter carrying a mutation in the CD28 response element site revealed that the activity was reduced by 80% after B7-1 and B7-1-LFA-3 costimulation whereas the transcriptional activity induced by LFA-3 was unaffected. Our data strongly suggest a selectivity in induction of nuclear factors by the CD2-LFA-3 and CD28-B7-1 pathways. This selectivity may contribute to regulation of the levels of IL-2 induced by LFA-3 and B7-1 costimulation and favor autocrine and paracrine T-cell responses, respectively. PMID:9032258

Parra, E; Varga, M; Hedlund, G; Kalland, T; Dohlsten, M

1997-01-01

260

Polyadenylation accelerates degradation of chloroplast mRNA.  

PubMed Central

The expression of chloroplast genes is regulated by several mechanisms, one of which is the modulation of RNA stability. To understand how this regulatory step is controlled during chloroplast development, we have begun to define the mechanism of plastid mRNA degradation. We show here that the degradation petD mRNA involves endonucleolytic cleavage at specific sites upstream of the 3' stem-loop structure. The endonucleolytic petD cleavage products can be polyadenylated in vitro, and similar polyadenylated RNA products are detectable in vivo. PCR analysis of the psbA and psaA-psaB-rps14 operons revealed other polyadenylated endonucleolytic cleavage products, indicating that poly(A) addition appears to be an integral modification during chloroplast mRNA degradation. Polyadenylation promotes efficient degradation of the cleaved petD RNAs by a 3'-5' exoribonuclease. Furthermore, polyadenylation also plays an important role in the degradation of the petD mRNA 3' end. Although the 3' end stem-loop is usually resistant to nucleases, adenylation renders the secondary structure susceptible to the 3'-5' exoribonuclease. Analysis of 3' ends confirms that polyadenylation occurs in vivo, and reveals that the extent of adenylation increases during the degradation of plastid mRNA in the dark. Based on these results, we propose a novel mechanism for polyadenylation in the regulation of plastid mRNA degradation. Images PMID:9003789

Kudla, J; Hayes, R; Gruissem, W

1996-01-01

261

HDAC3 regulates stability of estrogen receptor ? mRNA  

SciTech Connect

Highlights: ? HDAC inhibitors decrease the stability of ER? mRNA in MCF-7 cells. ? HDAC3 is involved in maintaining ER? mRNA stability in MCF-7 cells. ? ER? mRNA instability by knockdown of HDAC3 reduces the estrogen-dependent proliferation of ER?-positive MCF-7 cells. ? HDAC3 specific inhibitor will be one of new drugs for ER?-positive breast cancers. -- Abstract: Estrogen receptor alpha (ER?) expression is a risk factor for breast cancer. HDAC inhibitors have been demonstrated to down-regulate ER? expression in ER?-positive breast cancer cell lines, but the molecular mechanisms are poorly understood. Here, we showed that HDAC inhibitors decrease the stability of ER? mRNA, and that knockdown of HDAC3 decreases the stability of ER? mRNA and suppresses estrogen-dependent proliferation of ER?-positive MCF-7 breast cancer cells. In the Oncomine database, expression levels of HDAC3 in ER?-positive tumors are higher than those in ER?-negative tumors, thus suggesting that HDAC3 is necessary for ER? mRNA stability, and is involved in the estrogen-dependent proliferation of ER?-positive tumors.

Oie, Shohei; Matsuzaki, Kazuya; Yokoyama, Wataru; Murayama, Akiko; Yanagisawa, Junn, E-mail: junny@agbi.tsukuba.ac.jp

2013-03-08

262

Neurochemical and Behavioral Changes Induced by Interleukin2 and Soluble Interleukin2 Receptors  

Microsoft Academic Search

It has become abundantly clear that in addition to regulating immune function, interleukin (IL)-2 potently modulates brain\\u000a neurochemical activity and behavior. Central targets of IL-2 include mesolimbic and mesostriatal dopaminergic processes, hippocampal\\u000a cholinergic activity, and monoaminergic and neuroendocrine activity in hypothalamus, and brainstem. Of further significance,\\u000a IL-2 influences related behaviors and responses, including (but not limited to) exploratory activity, stereotypic

Steven S. Zalcman; Randall T. Woodruff; Ruchika Mohla; Allan Siegel

263

mRNA surveillance by the Caenorhabditis elegans smg genes.  

PubMed

mRNAs that contain premature stop codons are unstable in most eukaryotes, but the mechanism of their degradation is largely unknown. We demonstrate that functions of the six C. elegans smg genes are necessary for rapid turnover of nonsense mutant mRNAs of the unc-54 myosin heavy chain gene. Nonsense alleles of unc-54 express mRNAs that are unstable in smg(+) genetic backgrounds but have normal or near normal stability in smg(-) backgrounds. smg mutations also stabilize mRNA of unc-54(r293), a small deletion that removes the unc-54 polyadenylation site and expresses an aberrant mRNA. Most unc-54 nonsense mutations are recessive in both smg(+) and smg(-) genetic backgrounds. However, four specific alleles are recessive when smg(+) and dominant when smg(-). These smg-dependent dominant alleles express nonsense mutant polypeptides that disrupt thick filament and/or sarcomere assembly. All four alleles are predicted to express nonsense fragment polypeptides that contain most of the myosin globular head domain without an attached rod segment. By degrading messages that contain premature stop codons, the smg genes eliminate mRNAs that encode potentially toxic protein fragments. We propose that this system of mRNA turnover protects cells from their own errors of transcription, mRNA processing, or mRNA transport. PMID:8104846

Pulak, R; Anderson, P

1993-10-01

264

Lipid-based mRNA vaccine delivery systems.  

PubMed

Synthetic mRNAs can become biopharmaceutics allowing vaccination against cancer, bacterial and virus infections. Clinical trials with direct administration of synthetic mRNAs encoding tumor antigens demonstrated safety and induction of tumor-specific immune responses. Although immune responses are generated by naked mRNAs, their formulations with chemical carriers are expected to provide more specificity and internalization in dendritic cells (DCs) for better immune responses and dose reduction. This review reports lipid-based formulations (LBFs) that have proved preclinical efficacy. The selective delivery of mRNA LBFs to favor intracellular accumulation in DCs and reduction of the effective doses is discussed, notably to decorate LBFs with carbohydrates or glycomimetics allowing endocytosis in DCs. We also report how smart intracellular delivery is achieved using pH-sensitive lipids or polymers for an efficient mRNA escape from endosomes and limitations regarding cytosolic mRNA location for translation. PMID:25540984

Midoux, Patrick; Pichon, Chantal

2015-02-01

265

Interplay between viruses and host mRNA degradation  

PubMed Central

Messenger RNA degradation is a fundamental cellular process that plays a critical role in regulating gene expression by controlling both the quality and the abundance of mRNAs in cells. Naturally, viruses must successfully interface with the robust cellular RNA degradation machinery to achieve an optimal balance between viral and cellular gene expression and establish a productive infection in the host. In the past several years, studies have discovered many elegant strategies that viruses have evolved to circumvent the cellular RNA degradation machinery, ranging from disarming the RNA decay pathways and co-opting the factors governing cellular mRNA stability to promoting host mRNA degradation that facilitates selective viral gene expression and alters the dynamics of host-pathogen interaction. This review summarizes the current knowledge of the multifaceted interaction between viruses and cellular mRNA degradation machinery to provide an insight into the regulatory mechanisms that influence gene expression in viral infections. PMID:23274304

Narayanan, Krishna; Makino, Shinji

2013-01-01

266

Reovirus progeny subviral particles synthesize uncapped mRNA.  

PubMed Central

Reovirus progeny subviral particles were isolated from L-cells at late times postinfection. It has been shown (D. Skup and S. Millward, J. Virol. 34: 490--496, 1980) that these progeny subviral particles have masked capping enzymes, indicating that mRNA synthesized by these particles should be uncapped. When progeny subviral particles were used for mRNA synthesis in vitro, they failed to incorporate the beta-phosphate of [beta-32P]GTP into the 5' terminal. Direct analysis of reovirus mRNA synthesized by progeny subviral particles in the presence of either [alpha-32P]GTP or [alpha-32P]CTP indicated that the 5' terminal was uncapped, having the structure pGpC... The implications of this finding to the reovirus replicative cycle are discussed. Images PMID:7373718

Zarbl, H; Skup, D; Millward, S

1980-01-01

267

mRNA surveillance mitigates genetic dominance in Caenorhabditis elegans  

Microsoft Academic Search

Nonsense mutant mRNAs are unstable in all eucaryotes tested, a phenomenon termed nonsense-mediated mRNA decay (NMD) or mRNA\\u000a surveillance. Functions of the seven smg genes are required for mRNA surveillance in Caenorhabditis elegans. In Smg(+) genetic backgrounds, nonsense-mutant mRNAs are unstable, while in Smg(?) backgrounds such mRNAs are stable. Previous\\u000a work has demonstrated that the elevated level of nonsense-mutant mRNAs

B. M. Cali; P. Anderson

1998-01-01

268

Regulation of mRNA trafficking by nuclear pore complexes.  

PubMed

Over the last two decades, multiple studies have explored the mechanisms governing mRNA export out of the nucleus, a crucial step in eukaryotic gene expression. During transcription and processing, mRNAs are assembled into messenger ribonucleoparticles (mRNPs). mRNPs are then exported through nuclear pore complexes (NPCs), which are large multiprotein assemblies made of several copies of a limited number of nucleoporins. A considerable effort has been put into the dissection of mRNA export through NPCs at both cellular and molecular levels, revealing the conserved contributions of a subset of nucleoporins in this process, from yeast to vertebrates. Several reports have also demonstrated the ability of NPCs to sort out properly-processed mRNPs for entry into the nuclear export pathway. Importantly, changes in mRNA export have been associated with post-translational modifications of nucleoporins or changes in NPC composition, depending on cell cycle progression, development or exposure to stress. How NPC modifications also impact on cellular mRNA export in disease situations, notably upon viral infection, is discussed. PMID:25184662

Bonnet, Amandine; Palancade, Benoit

2014-01-01

269

BIOMARKERS OF ENDOCRINE DISRUPTION AT THE MRNA LEVEL  

EPA Science Inventory

Denslow, Nancy D., Christopher J. Bowman, Gillian Robinson, H. Stephen Lee, Ronald J. Ferguson, Michael J. Hemmer and Leroy C. Folmar. 1999. Biomarkers of Endocrine Disruption at the mRNA Level. In: Environmental Toxicology and Risk Assessment: Standardization of Biomarkers for ...

270

Analysis of mRNA recognition by human thymidylate synthase.  

PubMed

Expression of hTS (human thymidylate synthase), a key enzyme in thymidine biosynthesis, is regulated on the translational level through a feedback mechanism that is rarely found in eukaryotes. At low substrate concentrations, the ligand-free enzyme binds to its own mRNA and stabilizes a hairpin structure that sequesters the start codon. When in complex with dUMP (2'-deoxyuridine-5'-monophosphate) and a THF (tetrahydrofolate) cofactor, the enzyme adopts a conformation that is unable to bind and repress expression of mRNA. Here, we have used a combination of X-ray crystallography, RNA mutagenesis and site-specific cross-linking studies to investigate the molecular recognition of TS mRNA by the hTS enzyme. The interacting mRNA region was narrowed to the start codon and immediately flanking sequences. In the hTS enzyme, a helix-loop-helix domain on the protein surface was identified as the putative RNA-binding site. PMID:25423174

Brunn, Nicholas D; Dibrov, Sergey M; Kao, Melody B; Ghassemian, Majid; Hermann, Thomas

2014-01-01

271

Regulation of mRNA Trafficking by Nuclear Pore Complexes  

PubMed Central

Over the last two decades, multiple studies have explored the mechanisms governing mRNA export out of the nucleus, a crucial step in eukaryotic gene expression. During transcription and processing, mRNAs are assembled into messenger ribonucleoparticles (mRNPs). mRNPs are then exported through nuclear pore complexes (NPCs), which are large multiprotein assemblies made of several copies of a limited number of nucleoporins. A considerable effort has been put into the dissection of mRNA export through NPCs at both cellular and molecular levels, revealing the conserved contributions of a subset of nucleoporins in this process, from yeast to vertebrates. Several reports have also demonstrated the ability of NPCs to sort out properly-processed mRNPs for entry into the nuclear export pathway. Importantly, changes in mRNA export have been associated with post-translational modifications of nucleoporins or changes in NPC composition, depending on cell cycle progression, development or exposure to stress. How NPC modifications also impact on cellular mRNA export in disease situations, notably upon viral infection, is discussed. PMID:25184662

Bonnet, Amandine; Palancade, Benoit

2014-01-01

272

Analysis of mRNA recognition by human thymidylate synthase  

PubMed Central

Expression of hTS (human thymidylate synthase), a key enzyme in thymidine biosynthesis, is regulated on the translational level through a feedback mechanism that is rarely found in eukaryotes. At low substrate concentrations, the ligand-free enzyme binds to its own mRNA and stabilizes a hairpin structure that sequesters the start codon. When in complex with dUMP (2?-deoxyuridine-5?-monophosphate) and a THF (tetrahydrofolate) cofactor, the enzyme adopts a conformation that is unable to bind and repress expression of mRNA. Here, we have used a combination of X-ray crystallography, RNA mutagenesis and site-specific cross-linking studies to investigate the molecular recognition of TS mRNA by the hTS enzyme. The interacting mRNA region was narrowed to the start codon and immediately flanking sequences. In the hTS enzyme, a helix–loop–helix domain on the protein surface was identified as the putative RNA-binding site. PMID:25423174

Brunn, Nicholas D.; Dibrov, Sergey M.; Kao, Melody B.; Ghassemian, Majid; Hermann, Thomas

2014-01-01

273

Transcription Expression and Clinical Significance of Dishevelled-3 mRNA and ?-Catenin mRNA in Pleural Effusions from Patients with Lung Cancer  

PubMed Central

Objective. To evaluate diagnostic utility of Dishevelled-3 (DVL-3) mRNA and ?-catenin mRNA expression in pleural effusions of patients with lung cancer. Methods. DVL-3 mRNA and ?-catenin mRNA levels were assessed by performing RT-PCR on pleural effusion specimens from patients with lung cancer (n = 75) and with lung benign disease (n = 51). Results. The expressions of DVL-3 mRNA and ?-catenin mRNA were significantly higher in malignant than in benign lung disease (P < 0.01) and were obviously higher than cytology in adenocarcinoma (P < 0.01). In single use, DVL-3 mRNA had the highest specificity (94.1%) and PPV (95.7%), whereas ?-catenin mRNA had the highest sensitivity (92.0%) and NPV (88.5%). When combinations of markers were evaluated together, DVL-3 mRNA and ?-catenin mRNA gave a high-diagnostic performance: sensitivity of 100.0%, NPV of 100.0%, and accuracy of 96.0%, respectively. Conclusion. As molecular markers of detecting pleural micrometastasis, DVL-3 mRNA and ?-catenin mRNA are helpful to diagnose the cancer cells in pleural effusions of patients with lung cancer. PMID:22461838

Li, Xiao-Yan; Liu, Shu-Li; Cha, Na; Zhao, Yu-Jie; Wang, Shao-Cheng; Li, Wei-Nan; Wang, En-Hua; Wu, Guang-Ping

2012-01-01

274

Differential protein occupancy profiling of the mRNA transcriptome  

PubMed Central

Background RNA-binding proteins (RBPs) mediate mRNA biogenesis, translation and decay. We recently developed an approach to profile transcriptome-wide RBP contacts on polyadenylated transcripts by next-generation sequencing. A comparison of such profiles from different biological conditions has the power to unravel dynamic changes in protein-contacted cis-regulatory mRNA regions without a priori knowledge of the regulatory protein component. Results We compared protein occupancy profiles of polyadenylated transcripts in MCF7 and HEK293 cells. Briefly, we developed a bioinformatics workflow to identify differential crosslinking sites in cDNA reads of 4-thiouridine crosslinked polyadenylated RNA samples. We identified 30,000 differential crosslinking sites between MCF7 and HEK293 cells at an estimated false discovery rate of 10%. 73% of all reported differential protein-RNA contact sites cannot be explained by local changes in exon usage as indicated by complementary RNA-seq data. The majority of differentially crosslinked positions are located in 3? UTRs, show distinct secondary-structure characteristics and overlap with binding sites of known RBPs, such as ELAVL1. Importantly, mRNA transcripts with the most significant occupancy changes show elongated mRNA half-lives in MCF7 cells. Conclusions We present a global comparison of protein occupancy profiles from different cell types, and provide evidence for altered mRNA metabolism as a result of differential protein-RNA contacts. Additionally, we introduce POPPI, a bioinformatics workflow for the analysis of protein occupancy profiling experiments. Our work demonstrates the value of protein occupancy profiling for assessing cis-regulatory RNA sequence space and its dynamics in growth, development and disease. PMID:24417896

2014-01-01

275

Imaging individual mRNA molecules using multiple singly labeled probes.  

PubMed

We describe a method for imaging individual mRNA molecules in fixed cells by probing each mRNA species with 48 or more short, singly labeled oligonucleotide probes. This makes each mRNA molecule visible as a computationally identifiable fluorescent spot by fluorescence microscopy. We demonstrate simultaneous detection of three mRNA species in single cells and mRNA detection in yeast, nematodes, fruit fly wing discs, and mammalian cell lines and neurons. PMID:18806792

Raj, Arjun; van den Bogaard, Patrick; Rifkin, Scott A; van Oudenaarden, Alexander; Tyagi, Sanjay

2008-10-01

276

The majority of epidermal T cells in Psoriasis vulgaris lesions can produce type 1 cytokines, interferon-gamma, interleukin-2, and tumor necrosis factor-alpha, defining TC1 (cytotoxic T lymphocyte) and TH1 effector populations: a type 1 differentiation bias is also measured in circulating blood T cells in psoriatic patients.  

PubMed

Psoriasis vulgaris is a skin disease potentially mediated by pro-inflammatory cytokines produced by type 1 lesional T cells. The capability of individual T cells to produce these cytokines in lesional skin is not known. In this study we measured the ability of lesional and peripheral blood T cells to produce intracellular interferon-gamma, tumor necrosis factor-alpha, interleukin-2, interleukin-4, and interleukin-10 proteins as detected by flow cytometric analysis. Cytokine synthesis was induced by activation with ionomycin/phorbol myristate acetate (in the presence of Brefeldin A, which inhibits the exocytosis of these cytokines). After stimulation, we found relatively high percentages of epidermal CD8 and CD4 T cells capable of producing interferon-gamma, tumor necrosis factor-alpha, and interleukin-2, whereas few T cells, < 11%, expressed interleukin-4 or interleukin-10. Hence both CD8+ and CD4+ T cells are capable of type 1 effector functions (TC1 and TH1, respectively). This activation scheme was repeated on peripheral blood T cells from psoriatic patients versus healthy controls, where we also found a type 1 bias. In order to evaluate quantitatively the type 1 cytokine bias, we compared the frequency of type 2 interleukin-4 producing versus type 1 interferon-gamma producing T cells in our assay and found a shift towards type 1 producing cells. This shift reveals a type 1 differentiation bias in both lesional areas and in the peripheral blood, which may indicate an imbalance within the T cell population, which is contributing to the chronic or sustained immunologic activation of T cells found in this disease. PMID:10571730

Austin, L M; Ozawa, M; Kikuchi, T; Walters, I B; Krueger, J G

1999-11-01

277

Regulation of mRNA Translation by Signaling Pathways  

PubMed Central

mRNA translation is the most energy consuming process in the cell. In addition, it plays a pivotal role in the control of gene expression and is therefore tightly regulated. In response to various extracellular stimuli and intracellular cues, signaling pathways induce quantitative and qualitative changes in mRNA translation by modulating the phosphorylation status and thus the activity of components of the translational machinery. In this work we focus on the phosphoinositide 3-kinase (PI3K)/AKT and the mitogen-activated protein kinase (MAPK) pathways, as they are strongly implicated in the regulation of translation in homeostasis, whereas their malfunction has been linked to aberrant translation in human diseases, including cancer. PMID:22888049

Roux, Philippe P.; Topisirovic, Ivan

2012-01-01

278

The Current Status of Vertebrate Cellular mRNA IRESs  

PubMed Central

Internal ribosome entry sites/segments (IRESs) were first discovered over 20 years ago in picornaviruses, followed by the discovery of two other types of IRES in hepatitis C virus (HCV), and the dicistroviruses, which infect invertebrates. In the meantime, reports of IRESs in eukaryotic cellular mRNAs started to appear, and the list of such putative IRESs continues to grow to the point in which it now stands at ?100, 80% of them in vertebrate mRNAs. Despite initial skepticism from some quarters, there now seems universal agreement that there is genuine internal ribosome entry on the viral IRESs. However, the same cannot be said for cellular mRNA IRESs, which continue to be shrouded in controversy. The aim of this article is to explain why vertebrate mRNA IRESs remain controversial, and to discuss ways in which these controversies might be resolved. PMID:23378589

Jackson, Richard J.

2013-01-01

279

Early nonsense: mRNA decay solves a translational problem  

Microsoft Academic Search

Gene expression is highly accurate and rarely generates defective proteins. Several mechanisms ensure this fidelity, including specialized surveillance pathways that rid the cell of mRNAs that are incompletely processed or that lack complete open reading frames. One such mechanism, nonsense-mediated mRNA decay, is triggered when ribosomes encounter a premature translation-termination — or nonsense — codon. New evidence indicates that the

Nadia Amrani; Matthew S. Sachs; Allan Jacobson

2006-01-01

280

Compilation of E. coli mRNA promoter sequences.  

PubMed Central

An updated compilation of 300 E. coli mRNA promoter sequences is presented. For each sequence the most recent relevant paper was checked, to verify the location of the transcriptional start position as identified experimentally. We comment on the reliability of the sequence databanks and analyze the conservation of known promoter features in the current compilation. This database is available by E-mail. PMID:8479900

Lisser, S; Margalit, H

1993-01-01

281

Cytokine mRNA quantification by real-time PCR  

Microsoft Academic Search

Real-time PCR represents a new methodology that accurately quantifies nucleic acids. This has been made possible by the use of fluorogenic probes, which are presented in two forms, namely hydrolysis probes (also called TaqMan probes) and hybridisation probes. We decided to apply this methodology to cytokine mRNA quantification and this led us to the development of a protocol that provides

Patrick Stordeur; Lionel F. Poulin; Ligia Craciun; Ling Zhou; Liliane Schandené; Aurore de Lavareille; Stanislas Goriely; Michel Goldman

2002-01-01

282

Oligonucleotide switches and nanomaterials for intracellular mRNA sensing  

NASA Astrophysics Data System (ADS)

We describe here the conjugation of polymethylmethacrylate nanoparticles to particular oligonucleotide switches, termed molecular beacons (MBs), as potential intracellular nanosensors. Survivin mRNA targeting MBs have been used with Atto647N and Blackberry 650 as fluorophore/quencher pair. The nanosensors have been characterized in vitro by investigating the analytical performances of the chosen molecular beacon and its functionalities after conjugation to the nanoparticles.

Tombelli, S.; Ballestri, M.; Giambastiani, G.; Giannetti, A.; Guerrini, A.; Sotgiu, G.; Trono, C.; Tuci, G.; Varchi, G.; Baldini, F.

2013-06-01

283

Vibrational force alters mRNA expression in osteoblasts  

NASA Technical Reports Server (NTRS)

Serum-deprived mouse osteoblastic (MC3T3E1) cells were subjected to a vibrational force modeled by NASA to simulate a space shuttle launch (7.83 G rms). The mRNA levels for eight genes were investigated to determine the effect of vibrational force on mRNA expression. The mRNA levels of two growth-related protooncogenes, c-fos and c-myc, were up-regulated significantly within 30 min after vibration, whereas those of osteocalcin as well as transforming growth factor-beta1 were decreased significantly within 3 h after vibration. No changes were detected in the levels of beta-actin, histone H4, or cytoplasmic phospholipase A2 after vibration. No basal levels of cyclooxygenase-2 expression were detected. In addition, the extracellular concentrations of prostaglandin E2 (PGE2), a potent autocrine/paracrine growth factor in bone, were not significantly altered after vibration most likely due to the serum deprivation state of the osteoblasts. In comparison with the gravitational launch profile, vibrational-induced changes in gene expression were greater both in magnitude and number of genes activated. Taken together, these data suggest that the changes in mRNA expression are due to a direct mechanical effect of the vibrational force on the osteoblast cells and not to changes in the local PGE2 concentrations. The finding that launch forces induce gene expression is of utmost importance since many of the biological experiments do not dampen vibrational loads on experimental samples. This lack of dampening of vibrational forces may partially explain why 1-G onboard controls sometimes do not reflect 1-G ground controls. These data may also suggest that scientists use extra ground controls that are exposed to launch forces, have these forces dampened on launched samples, or use facilities such as Biorack that provide an onboard 1-G centrufuge in order to control for space shuttle launch forces.

Tjandrawinata, R. R.; Vincent, V. L.; Hughes-Fulford, M.

1997-01-01

284

The utility of protein and mRNA correlation.  

PubMed

Transcriptomic, proteomic, and metabolomic measurements are revolutionizing the way we model and predict cellular behavior, and multi-omic comparisons are being published with increased regularity. Some have expected a trivial and predictable correlation between mRNA and protein; however, the manifest complexity of biological regulation suggests a more nuanced relationship. Indeed, observing this lack of strict correlation provides clues for new research topics, and has the potential for transformative biological insight. PMID:25467744

Payne, Samuel H

2015-01-01

285

Decreased albumin mRNA in immunodeficient wasted' mice  

SciTech Connect

Mice bearing the autosomal recessive gene wst (wst/wst) develop a wasting syndrome' that leads to death by 28-32 days of age. These mice have faulty repair of damage induced by ionizing radiation, immunodeficiency at secretory sites, and neurologic abnormalities. In addition to a progressively more apparent wasted phenotype, wst/wst mice show other features of failure to thrive and malnutrition. Daily body weights of the animals revealed a loss in weight between 25 and 30 days of age, a time during which normal littermates were progressively and rapidly gaining weight. Albumin mRNA levels were measured by dilution dot blot hybridizations of liver-derived RNA preparations from wasted mice, littermates, and parental controls. In all wasted mice, albumin mRNA levels were reduced 5 to 10 fold compared to controls. Northern blots revealed that the albumin mRNA present in wasted mice was normal in length though reduced in amount. These results suggest there may be a relationship between low albumin synthesis and the wasting syndrome of the wst/wst mouse.

Libertin, C.R.; Buczek, N.; Weaver, P.; Mobarhan, S.; Woloschak, G.E. (Loyola Univ. of Chicago, Maywood, IL (United States) Argonne National Lab., IL (United States))

1991-03-15

286

Identification of phloem-mobile mRNA.  

PubMed

Signaling between cells, tissues and organs is essential for multicellular organisms to coordinate and adapt their development and growth to internal and environmental changes. Plants have evolved a plant-specific symplasmic pathway, called plasmodesmata, for efficient intercellular communication, in addition to the receptor-ligand-based apoplasmic pathway. Long-distance signaling between distant organs is enabled via the phloem tube system, where plasmodesmata contribute to phloem loading and unloading for photosynthate allocation. In addition to signaling by small molecules such as metabolites and phytohormones, the transport of proteins, small RNAs and mRNAs is also considered an important mechanism to achieve long-distance signaling in plants. Recent studies on phloem-mobile proteins and small RNAs have revealed their role in crucial physiological processes including flowering, systemic silencing and nutrient allocation. However, the biological role of mRNAs found in the phloem tube is not yet clear, though their mobility over long-distances has been well evidenced. To gain this knowledge, it is important to collect further information on mRNA profiles in the phloem translocation stream. In this review, I summarize the current approaches to identifying the mRNA population in the phloem translocation system, and discuss the possible role of short- and long-distance mRNA transport. PMID:25516498

Notaguchi, Michitaka

2015-01-01

287

Nonsense-mediated mRNA decay factors act in concert to regulate common mRNA targets  

Microsoft Academic Search

Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that degrades mRNAs containing nonsense codons, and regulates the expression of naturally occurring transcripts. While NMD is not essential in yeast or nematodes, UPF1, a key NMD effector, is essential in mice. Here we show that NMD components are required for cell proliferation in Drosophila. This raises the question of whether NMD

JAN REHWINKEL; IVICA LETUNIC; JEROEN RAES; PEER BORK; ELISA IZAURRALDE

2005-01-01

288

Enhancing mRNA stability through the addition of stabilizing untranslated regions  

E-print Network

Messenger-RNA (mRNA) therapy, in which mRNA is introduced to cells or tissues to cause a transient expression of specific genes, has applications ranging from tissue engineering to neural regeneration. This transient nature ...

Tran, Thi (Thi Thi Kim)

2011-01-01

289

Characterization of a b-Actin mRNA Zipcode-Binding Protein  

Microsoft Academic Search

Localization of b-actin mRNA to the leading edge offibroblasts requires the presence of conserved elements in the 3* untranslated region of the mRNA, including a 54-nucleotide element which has been termed the \\

ANTHONY F. ROSS; YURI OLEYNIKOV; EDWARD H. KISLAUSKIS; KRISHAN L. TANEJA; ANDROBERT H. SINGER

1997-01-01

290

Identification of mRNA binding proteins that regulate the stability of LDL receptor mRNA through AU-rich elements  

Microsoft Academic Search

The 3'untranslated region (UTR) of human LDL receptor (LDLR) mRNA contains three AU-rich elements (AREs) responsible for rapid mRNA turnover and mediates the stabilization induced by berberine (BBR). However, the identities of the specific RNA binding proteins involved in the regulation of LDLR mRNA stability at the steady state level or upon BBR treatment are unknown. By conducting small interfering

Hai Li; Wei Chen; Yue Zhou; Parveen Abidi; Orr Sharpe; William H. Robinson; Fredric B. Kraemer; Jingwen Liu

2009-01-01

291

Urinary mRNA in systemic lupus erythematosus.  

PubMed

Systemic lupus erythematosus (SLE) is a relapsing autoimmune disease with clinical manifestations that affect multiple organ systems. Lupus nephritis is recognized as one of the most severe organ involvements in SLE and affects half of the lupus patients. Notably, lupus nephritis is characterized by intrarenal lymphocyte activation and inflammation. Since most of the cytokines exert their effects in a paracrine fashion, measuring their expression at the site of pathology should be of biological relevance. Although kidney biopsy is widely used to determine the histology and severity of lupus nephritis, this invasive procedure has its own risk and is not practical for serial monitoring. In the past decade, extraction and quantification of messenger RNA (mRNA) from urinary sediment has emerged as a robust laboratory technique. Quantification of mRNA expression in urinary sediment has been tested as a noninvasive means to assess the disease activity of SLE patients. Available published evidence, however, is limited to small-scale studies. Based on the result of these studies, a number of cytokine and transcript factor genes have been found to have potential for the differentiation between active and inactive SLE, between proliferative and membranous types of lupus nephritis, assessment of the systemic lupus activity or histological activity of kidney biopsy specimen, monitoring of treatment response in active lupus nephritis, or detection of lupus disease flare in clinically quiescent patients. Being a simple and noninvasive method, urinary mRNA level deserves further studies to validate its role in risk stratification and monitoring of therapeutic response in patients with lupus nephritis. PMID:24772668

Szeto, Cheuk-Chun; Kwan, Bonnie Ching-Ha; Tam, Lai-Shan

2013-01-01

292

mRNA Regulation by Puf Domain Proteins  

NSDL National Science Digital Library

Puf domain proteins bind specific sequences in mRNAs to regulate their translation or stability, or both. Neither the mechanism of their action nor the identities of targeted mRNAs have been well defined. Recent work suggests that Puf proteins generally act by recruiting Pop2, a deadenylation enzyme that is part of a large complex. Recent work from a separate group defines a subset of the Drosophila transcriptome that is bound by the fly Puf protein, Pumilio. Together, these papers substantially increase our understanding of the biology of the Puf family of mRNA regulators.

Robin P. Wharton (NC;Howard Hughes Medical Institute and Duke University Medical School Durham REV); Aneel K. Aggarwal (NY;Mount Sinai School of Medicine, New York REV)

2006-09-26

293

[Age-dependent changes in mRNA transport (nucleus-cytoplasm)].  

PubMed

Transport of mRNA from nucleus to cytoplasm is an ATP-dependent process which occurs strictly vectorially. Because the mRNA is structurally bound during transport, mRNA transport is a "solid-state" process consisting of i) mRNA release from the nuclear matrix, ii) mRNA translocation through the nuclear pore, and iii) cytoskeletal binding. We identified and purified the following components involved in the translocation step: i) the nuclear envelope (NE) nucleoside triphosphatase (NTPase) which is stimulated by the 3'poly(A) tail of mRNA, ii) the poly(A)-recognizing mRNA carrier, iii) the NE protein kinase, and iv) the NE phosphatase. In addition, we found that an RNA helicase activity is present in NE, which also may be involved in RNA transport. Our results show that, besides poly(A), also double-stranded RNA structures may modulate RNA export. The amount of mRNA released from nuclei markedly decreases with age. Evidence is presented that this age-dependent change is caused by an impairment of polyadenylation of mRNA, hnRNA processing, release of mRNA from nuclear matrix, and translocations of mRNA from nuclear to cytoplasmic compartment (decrease in activities of NE NTPase, protein kinase, and phosphatase; decrease in poly(A)-binding affinity of mRNA carrier). PMID:8212790

Müller, W E; Agutter, P S; Prochnow, D J; Fasold, H; Sève, A P; Tsiapalis, C M; Schröder, H C

1993-01-01

294

PKA isoforms coordinate mRNA fate during nutrient starvation  

PubMed Central

Summary A variety of stress conditions induce mRNA and protein aggregation into mRNA silencing foci, but the signalling pathways mediating these responses are still elusive. Previously we demonstrated that PKA catalytic isoforms Tpk2 and Tpk3 localise with processing and stress bodies in Saccharomyces cerevisiae. Here, we show that Tpk2 and Tpk3 are associated with translation initiation factors Pab1 and Rps3 in exponentially growing cells. Glucose starvation promotes the loss of interaction between Tpk and initiation factors followed by their accumulation into processing bodies. Analysis of mutants of the individual PKA isoform genes has revealed that the TPK3 or TPK2 deletion affects the capacity of the cells to form granules and arrest translation properly in response to glucose starvation or stationary phase. Moreover, we demonstrate that PKA controls Rpg1 and eIF4G1 protein abundance, possibly controlling cap-dependent translation. Taken together, our data suggest that the PKA pathway coordinates multiple stages in the fate of mRNAs in association with nutritional environment and growth status of the cell. PMID:22899713

Tudisca, Vanesa; Simpson, Clare; Castelli, Lydia; Lui, Jennifer; Hoyle, Nathaniel; Moreno, Silvia; Ashe, Mark; Portela, Paula

2012-01-01

295

PKA isoforms coordinate mRNA fate during nutrient starvation.  

PubMed

A variety of stress conditions induce mRNA and protein aggregation into mRNA silencing foci, but the signalling pathways mediating these responses are still elusive. Previously we demonstrated that PKA catalytic isoforms Tpk2 and Tpk3 localise with processing and stress bodies in Saccharomyces cerevisiae. Here, we show that Tpk2 and Tpk3 are associated with translation initiation factors Pab1 and Rps3 in exponentially growing cells. Glucose starvation promotes the loss of interaction between Tpk and initiation factors followed by their accumulation into processing bodies. Analysis of mutants of the individual PKA isoform genes has revealed that the TPK3 or TPK2 deletion affects the capacity of the cells to form granules and arrest translation properly in response to glucose starvation or stationary phase. Moreover, we demonstrate that PKA controls Rpg1 and eIF4G(1) protein abundance, possibly controlling cap-dependent translation. Taken together, our data suggest that the PKA pathway coordinates multiple stages in the fate of mRNAs in association with nutritional environment and growth status of the cell. PMID:22899713

Tudisca, Vanesa; Simpson, Clare; Castelli, Lydia; Lui, Jennifer; Hoyle, Nathaniel; Moreno, Silvia; Ashe, Mark; Portela, Paula

2012-11-01

296

mRNA quality control at the 5? end*  

PubMed Central

All eukaryotic mRNAs are capped at their 5? end. Capping of mRNAs takes place co-transcriptionally and involves three steps. The intermediates of the capping process, as well as the uncapped 5? tri-phosphate RNA, are resistant to decapping and degradation by known factors, leading to the assumption that the capping process always proceeds to completion. This view was recently drastically changed. A novel family of enzymes, including the yeast proteins Rai1, Dxo1/Ydr370C, and the mammalian protein DXO/Dom3Z, has been identified. These enzymes catalyze the conversion of the improperly capped mRNAs to 5? mono-phosphate RNA, allowing them to be degraded by 5?-3? exoribonucleases. Several of these enzymes also possess 5?-3? exoribonuclease activities themselves, and can single-handedly clear the improperly capped mRNAs. Studying of these enzymes has led to the realization that mRNA capping does not always proceed to completion, and the identification of an mRNA capping quality control mechanism in eukaryotes. In this paper, we briefly review recent advances in this area. PMID:24793761

Zhai, Li-ting; Xiang, Song

2014-01-01

297

The Arabidopsis translatome cell-specific mRNA atlas  

PubMed Central

Plants consist of distinct cell types distinguished by position, morphological features and metabolic activities. We recently developed a method to extract cell-type specific mRNA populations by immunopurification of ribosome-associated mRNAs. Microarray profiles of 21 cell-specific mRNA populations from seedling roots and shoots comprise the Arabidopsis Translatome dataset. This gene expression atlas provides a new tool for the study of cell-specific processes. Here we provide an example of how genes involved in a pathway limited to one or few cell-types can be further characterized and new candidate genes can be predicted. Cells of the root endodermis produce suberin as an inner barrier between the cortex and stele, whereas the shoot epidermal cells form cutin as a barrier to the external environment. Both polymers consist of fatty acid derivates, and share biosynthetic origins. We use the Arabidopsis Translatome dataset to demonstrate the significant cell-specific expression patterns of genes involved in those biosynthetic processes and suggest new candidate genes in the biosynthesis of suberin and cutin. PMID:20220312

Mustroph, Angelika

2010-01-01

298

Retaining MKP1 expression and attenuating JNK-mediated apoptosis by RIP1 for cisplatin resistance through miR-940 inhibition  

PubMed Central

The elucidation of chemoresistance mechanisms is important to improve cancer patient survival. In this report, we investigated the role and mechanism through which receptor-interacting protein 1 (RIP1), a mediator in cell survival and death signaling, participates in cancer's response to chemotherapy. In lung cancer cells, knockdown of RIP1 substantially increased cisplatin-induced apoptotic cytotoxicity, which was associated with robust JNK activation. The expression of the JNK inactivating phosphatase, MKP1, was substantially reduced in RIP1 knockdown cells. Although MKP1 protein stability was not altered by RIP1 suppression, the synthesis rate of MKP1 was dramatically reduced in RIP1-suppressed cells. Furthermore, we found that the expression of miR-940 was substantially increased in RIP1 knockdown cells. Knockdown of miR-940 restored MKP1 expression and attenuated cisplatin-induced JNK activation and cytotoxicity. Importantly, ectopic expression of MKP1 effectively attenuated cisplatin-induced JNK activation and cytotoxicity. In addition, activation of the JNK upstream signaling kinase, MKK4, was also potentiated in RIP1 knockdown cells. Altogether, our results suggest that RIP1 contributes to cisplatin resistance by suppressing JNK activation that involves releasing miR-940-mediated inhibition of MKP1 and suppressing activation of MKK4. Intervention targeting the RIP1/miR-940/MKP1/JNK pathway may be used to sensitize platinum-based chemotherapy. PMID:24675421

He, Weiyang; Padilla, Mabel T.; Zhang, Lin; Wang, Xia; Zhang, Bin; Lin, Yong

2014-01-01

299

Corrections for mRNA extraction and sample normalization errors find increased mRNA levels may compensate for cancer haplo-insufficiency  

PubMed Central

The relative mRNA levels of differentially expressed (DE) and housekeeping (HK) genes of six aneuploid cancer lines with large-scale genomic changes identified by SNP/SKY analysis were compared with similar genes in diploid cells. The aneuploid cancer lines had heterogeneous genomic landscapes with subdiploid, diploid, and supradiploid regions and higher overall gene copy numbers compared with diploid cells. The mRNA levels of the haploid, diploid, and triploid HK genes were found to be higher after correction of easily identifiable mRNA measurement errors. Surprisingly, diploid and aneuploid HK gene mRNA levels were the same by standard expression array analyses, despite the higher copy numbers of the cancer cell HK genes. This paradoxical result proved to be due to inaccurate inputs of true intra-cellular mRNAs for analysis. These errors were corrected by analyzing the expression intensities of DE and HK genes in mRNAs extracted from equal cell numbers (50:50) of intact cancer cell and lymphocyte mixtures. Correction for both mRNA extraction/sample normalization errors and total gene copy numbers found the SUIT-2 and PC-3 cell lines' cancer genes both had ?50% higher mRNA levels per single allele than lymphocyte gene alleles. These increased mRNA levels for single transcribed cancer alleles may restore functional mRNA levels to cancer genes rendered haplo-insufficient by the genetic instability of cancer. PMID:24327546

Weaver, David A; Nestor-Kalinoski, Andrea L; Craig, Kristen; Gorris, Matthew; Parikh, Tejal; Mabry, Helen; Allison, David C

2014-01-01

300

Identification of Secondary Structure in the 5' Untranslated Region of the Human Adrenomedullin mRNA with Implications for the Regulation of mRNA Translation  

E-print Network

RNA with Implications for the Regulation of mRNA Translation Fabienne Brenet1,# , Nadège Dussault1,# , Christine Delfino' UTR, Translational control, Stem-loop structure, RNA-binding proteins. Abbreviated Title : Regulatory role of the stem-loop in translation of AM mRNA. Number of pages : 30 Number of figures : 6 Number

Paris-Sud XI, Université de

301

Biomarkers of endocrine disruption at the mRNA level  

SciTech Connect

A large number of estrogen-mimicking, anthropogenic chemicals capable of disrupting normal reproductive function have been identified. The ubiquitous distribution of these compounds, many as components of complex industrial or municipal waste, has spurred an effort to develop methods to screen for chemicals which disrupt normal endocrine regulation of reproduction. The authors have developed assays that both allow exposure of animals in vivo and measure the response at the level of gene activation. The authors have developed a probe for measuring the induction of vitellogenin mRNA by Northern Blot in livers of sheepshead minnows treated with 17-{beta}-estradiol. The authors have also developed a strategy for using Differential Display Polymerase Chain Reaction for determining gene induction profiles following exposure to estradiol. These methods should be adaptable to a variety of structurally diverse estrogen mimics.

Denslow, N.D.; Bowman, C.J.; Robinson, G.; Lee, H.S.; Ferguson, R.J.; Hemmmer, M.J.; Folmar, L.C.

1999-07-01

302

Picornavirus Modification of a Host mRNA Decay Protein  

PubMed Central

ABSTRACT Due to the limited coding capacity of picornavirus genomic RNAs, host RNA binding proteins play essential roles during viral translation and RNA replication. Here we describe experiments suggesting that AUF1, a host RNA binding protein involved in mRNA decay, plays a role in the infectious cycle of picornaviruses such as poliovirus and human rhinovirus. We observed cleavage of AUF1 during poliovirus or human rhinovirus infection, as well as interaction of this protein with the 5? noncoding regions of these viral genomes. Additionally, the picornavirus proteinase 3CD, encoded by poliovirus or human rhinovirus genomic RNAs, was shown to cleave all four isoforms of recombinant AUF1 at a specific N-terminal site in vitro. Finally, endogenous AUF1 was found to relocalize from the nucleus to the cytoplasm in poliovirus-infected HeLa cells to sites adjacent to (but distinct from) putative viral RNA replication complexes. PMID:23131833

Rozovics, Janet M.; Chase, Amanda J.; Cathcart, Andrea L.; Chou, Wayne; Gershon, Paul D.; Palusa, Saiprasad; Wilusz, Jeffrey; Semler, Bert L.

2012-01-01

303

PROLYL CARBOXYPEPTIDASE mRNA EXPRESSION IN THE MOUSE BRAIN  

PubMed Central

Prolyl carboxypeptidase (PRCP), a serine protease, is widely expressed in the body including liver, lung, kidney and brain, with a variety of known substrates such as plasma prekallikrein, bradykinin, angiotensins II and III, and -MSH, suggesting its role in the processing of tissue-specific substrates. In the brain, PRCP has been shown to inactivate hypothalamic -MSH, thus modulating melanocortin signaling in the control of energy metabolism. While its expression pattern has been reported in the hypothalamus, little is known on the distribution of PRCP throughout the mouse brain. This study was undertaken to determine PRCP expression in the mouse brain. Radioactive in situ hybridization was performed to determine endogenous PRCP mRNA expression. In addition, using a gene-trap mouse model for PRCP deletion, X-gal staining was performed to further determine PRCP distribution. Results from both approaches showed that PRCP gene is broadly expressed in the brain. PMID:24161824

Jeong, Jin Kwon; Diano, Sabrina

2013-01-01

304

Prolyl carboxypeptidase mRNA expression in the mouse brain.  

PubMed

Prolyl carboxypeptidase (PRCP), a serine protease, is widely expressed in the body including liver, lung, kidney and brain, with a variety of known substrates such as plasma prekallikrein, bradykinin, angiotensins II and III, and ?-MSH, suggesting its role in the processing of tissue-specific substrates. In the brain, PRCP has been shown to inactivate hypothalamic ?-MSH, thus modulating melanocortin signaling in the control of energy metabolism. While its expression pattern has been reported in the hypothalamus, little is known on the distribution of PRCP throughout the mouse brain. This study was undertaken to determine PRCP expression in the mouse brain. Radioactive in situ hybridization was performed to determine endogenous PRCP mRNA expression. In addition, using a gene-trap mouse model for PRCP deletion, X-gal staining was performed to further determine PRCP distribution. Results from both approaches showed that PRCP gene is broadly expressed in the brain. PMID:24161824

Jeong, Jin Kwon; Diano, Sabrina

2014-01-13

305

Clustered bottlenecks in mRNA translation and protein synthesis  

NASA Astrophysics Data System (ADS)

We construct an algorithm that generates large, band-diagonal transition matrices for a totally asymmetric exclusion process (TASEP) with local hopping rate inhomogeneities. The matrices are diagonalized numerically to find steady-state currents of TASEPs with local variations in hopping rate. The results are then used to investigate clustering of slow codons along mRNA. Ribosome density profiles near neighboring clusters of slow codons interact, enhancing suppression of ribosome throughput when such bottlenecks are closely spaced. Increasing the slow codon cluster size, beyond ˜ 3-4, does not significantly reduce ribosome current. Our results are verified by extensive Monte-Carlo simulations and provide a biologically-motivated explanation for the experimentally-observed clustering of low-usage codons.

Chou, Tom; Lakatos, Greg

2004-03-01

306

Clustered bottlenecks in mRNA translation and protein synthesis  

E-print Network

We construct an algorithm that generates large, band-diagonal transition matrices for a totally asymmetric exclusion process (TASEP) with local hopping rate inhomogeneities. The matrices are diagonalized numerically to find steady-state currents of TASEPs with local variations in hopping rate. The results are then used to investigate clustering of slow codons along mRNA. Ribosome density profiles near neighboring clusters of slow codons interact, enhancing suppression of ribosome throughput when such bottlenecks are closely spaced. Increasing the slow codon cluster size, beyond $\\approx 3-4$, does not significantly reduce ribosome current. Our results are verified by extensive Monte-Carlo simulations and provide a biologically-motivated explanation for the experimentally-observed clustering of low-usage codons.

Tom Chou; Greg Lakatos

2003-10-29

307

Increased apoB100 mRNA in inbred strains of mice by estrogen is caused by decreased RNA editing protein mRNA.  

PubMed

Estrogen administration to rats diminishes all apoproteins and lipoproteins from plasma. In contrast, some inbred strains of mice raise their plasma apoB and LDL levels by more than 2-fold (Srivastava et al, 1993, Eur. J. Biochem. 216, 527-538). Further studies with 13 inbred strains of mice given 3 micrograms beta-estradiol/g body weight/day for 5 consecutive days suggest that some mouse strains increased their apoB and LDL levels and some did not. To examine the mechanism of influence of genetic factors on apoB regulation, two strains, C57L and C57BL, that increased their VLDL- and LDL-cholesterol, and 2 strains, BALB and C3H, that did not, were chosen. Estrogen increased plasma apoB levels selectively in the strains C57L and C57BL, termed as 'responders,' but did not change in BALB and C3H, termed as 'non-responders.' One of the mechanisms for increased plasma apoB levels could be through increased production of apoB-containing particles. This possibility was investigated. ApoB and REPR mRNA were quantified by RNase protection assay, and apoB-100 mRNA by apoB mRNA editing assay. Hepatic apoB mRNA increased by 30% in 'non-responders,' but decreased by 20% in the 'responders.' However, apoB-100 mRNA increased relative to apoB-48 mRNA in all the 4 strains by 50%. The mRNA for RNA editing protein (REPR) decreased in all strains, suggesting that apoB-100 mRNA increased as a result of decreased apoB mRNA editing activity. These results suggest that:(a) modulation of apoB mRNA by estrogen was strain-specific;(b) increased apoB100 mRNA in inbred strains of mice were caused by decreased apoB mRNA editing activity; and (c) the differences in the plasma apoB levels among 'responder' and 'nonresponder' strains of mice occur through mechanisms other than the apoB mRNA editing. PMID:7626051

Srivastava, R A

1995-07-17

308

Transfection Efficiency and Transgene Expression Kinetics of mRNA Delivered in Naked and Nanoparticle Format  

PubMed Central

Transfection efficiencies and transgene expression kinetics of messenger RNA (mRNA), an emerging class of nucleic acid-based therapeutics, have been poorly characterized. In this study, we evaluated transfection efficiencies of mRNA delivered in naked and nanoparticle format in vitro and in vivo using GFP and luciferase as reporters. While mRNA nanoparticles transfect primary human and mouse dendritic cells (DCs) efficiently in vitro, naked mRNA could not produce any detectable gene product. Protein expression of nanoparticle-mediated transfection in vitro peaks rapidly within 5-7 hours and decays in a biphasic manner. In vivo, naked mRNA is more efficient than mRNA nanoparticles when administered subcutaneously. In contrast, mRNA nanoparticle performs better when administered intranasally and intravenously. Gene expression is most transient when delivered intravenously in nanoparticle format with an apparent half-life of 1.4 hours and lasts less than 24 hours, and most sustained when delivered in the naked format subcutaneously at the base of tail with an apparent half-life of 18 hours and persists for at least 6 days. Notably, exponential decreases in protein expression are consistently observed post-delivery of mRNA in vivo regardless of the mode of delivery (naked or nanoparticle) or the site of administration. This study elucidates the performance of mRNA transfection and suggests a niche for mRNA therapeutics when predictable in vivo transgene expression kinetics is imperative. PMID:23306021

Phua, Kyle K.L; Leong, Kam W.; Nair, Smita K.

2013-01-01

309

Functional Link between the Mammalian Exosome and mRNA Decapping  

Microsoft Academic Search

Mechanistic understanding of mammalian mRNA turnover remains incomplete. We demonstrate that the 3? to 5? exoribonuclease decay pathway is a major contributor to mRNA decay both in cells and in cell extract. An exoribonuclease-dependent scavenger decapping activity was identified that follows decay of the mRNA and hydrolyzes the residual cap. The decapping activity is associated with a subset of the

Zuoren Wang; Megerditch Kiledjian

2001-01-01

310

New Study Shows Protein and mRNA Levels Are Not Correlated | Physical Sciences in Oncology  

Cancer.gov

Ever since molecular biologists sorted out the general mechanisms that cells use to turn DNA into messenger RNA (mRNA) and then into protein, researchers have largely assumed that the amount of mRNA a cell makes during gene transcription will correlate with the subsequent amount of protein the cell produces from that mRNA. Indeed, aggregate measurements made from large numbers of cells have long supported this assumption.

311

Coupling mRNA processing with transcription in time and space  

PubMed Central

Maturation of mRNA precursors often occurs simultaneously with their synthesis by RNA polymerase II (Pol II). The co-transcriptional nature of mRNA processing has permitted the evolution of coupling mechanisms that coordinate transcription with mRNA capping, splicing, editing and 3? end formation. Recent experiments using sophisticated new methods for analysis of nascent RNA have provided important insights into the relative amount of co-transcriptional and post-transcriptional processing, the relationship between mRNA elongation and processing, and the role of the Pol II carboxy-terminal domain (CTD) in regulating these processes. PMID:24514444

Bentley, David L.

2015-01-01

312

Eukaryotic mRNA capping enzyme-guanylate covalent intermediate.  

PubMed

Incubation of HeLa cell mRNA guanylyltransferase (GTP:mRNA guanylyltransferase, EC 2.7.7.50) with [alpha-32P]GTP and a divalent cation in the absence of an RNA acceptor results in the formation of a covalent enzyme-guanylate complex. The complex, after purification by phosphocellulose chromatography, can transfer its bound GMP moiety to pyrophosphate, regenerating GTP, or to the 5'-diphosphate end of poly(A), forming a cap structure G(5')pppA(pA)n. The GMP-polypeptide has a molecular weight of 65,000 and is stable to heating in the presence of sodium dodecyl sulfate. On the basis of the alkali-stable and acid-labile nature of the bond and its susceptibility to nucleophilic attack by hydroxylamine at low pH, the GMP-polypeptide linkage appears to be a phosphoamine bond. After digestion with trypsin, a single GMP-peptide was resolved by two dimensional electrophoresis and chromatography. PMID:6281766

Venkatesan, S; Moss, B

1982-01-01

313

Eukaryotic mRNA capping enzyme-guanylate covalent intermediate.  

PubMed Central

Incubation of HeLa cell mRNA guanylyltransferase (GTP:mRNA guanylyltransferase, EC 2.7.7.50) with [alpha-32P]GTP and a divalent cation in the absence of an RNA acceptor results in the formation of a covalent enzyme-guanylate complex. The complex, after purification by phosphocellulose chromatography, can transfer its bound GMP moiety to pyrophosphate, regenerating GTP, or to the 5'-diphosphate end of poly(A), forming a cap structure G(5')pppA(pA)n. The GMP-polypeptide has a molecular weight of 65,000 and is stable to heating in the presence of sodium dodecyl sulfate. On the basis of the alkali-stable and acid-labile nature of the bond and its susceptibility to nucleophilic attack by hydroxylamine at low pH, the GMP-polypeptide linkage appears to be a phosphoamine bond. After digestion with trypsin, a single GMP-peptide was resolved by two dimensional electrophoresis and chromatography. Images PMID:6281766

Venkatesan, S; Moss, B

1982-01-01

314

Polyamines cause elevation of steroid 5?-reductase mRNA levels by suppressing mRNA degradation in C6 glioma cells.  

PubMed

Polyamines are widely distributed in living organisms, and considered to play a potential role in various cellular processes. The effects of polyamines on gene expression as well as cell proliferation have been suggested to be closely associated with the physiological and pathological functions. However, it seems necessary to investigate their potential roles in the regulation of cellular metabolism and functions. Previously, glial cells have been suggested to be involved in the protection and preservation of neuronal functions, probably through the production of neurotrophic factors in the brain. On the other hand, neuroactive 5?-reduced steroids promote glial cell differentiation, resulting in enhancement of their ability to produce brain-derived neurotrophic factor (BDNF). Based on these findings, polyamines are assumed to stimulate the expression of the gene encoding steroid 5?-reductase (5?-R), which can induce the production of neuroactive 5?-reduced steroids in glial cells. The effects of polyamines on 5?-R mRNA levels in C6 glioma cells were examined as a model experiment. In consequence, spermine (SPM) and spermidine (SPD), but not putrescine (PUT), have been shown to elevate 5?-R mRNA levels without activating the 5?-R promoter. Furthermore, SPM increased 5?-R mRNA levels under the conditions in which the mRNA biosynthesis was inhibited. Therefore, it can be speculated that polyamines increase 5?-R mRNA levels as a consequence of suppressing the degradation of mRNA. PMID:24800957

Morita, Kyoji; Lee, Mi-Sook; Her, Song; Nishibori, Naoyoshi

2014-10-01

315

Quantifying mRNA targeting to P-bodies in living human cells reveals their dual role in mRNA decay and storage.  

PubMed

The 5'-to-3' mRNA degradation machinery localizes to cytoplasmic processing bodies (P-bodies), which are non-membranous structures found in all eukaryotes. Although P-body function has been intensively studied in yeast, less is known about their role in mammalian cells, such as whether P-body enzymes are actively engaged in mRNA degradation or whether P-bodies serve as mRNA storage depots, particularly during cellular stress. We examined the fate of mammalian mRNAs in P-bodies during translational stress, and show that mRNAs accumulate within P-bodies during amino acid starvation. The 5' and 3' ends of the transcripts residing in P-bodies could be identified, but poly(A) tails were not detected. Using the MS2 mRNA-tagging system for mRNA visualization in living cells, we found that a stationary mRNA population formed in P-bodies during translational stress, which cleared gradually after the stress was relieved. Dcp2-knockdown experiments showed that there is constant degradation of part of the P-body-associated mRNA population. This analysis demonstrates the dual role of P-bodies as decay sites and storage areas under regular and stress conditions. PMID:25128566

Aizer, Adva; Kalo, Alon; Kafri, Pinhas; Shraga, Amit; Ben-Yishay, Rakefet; Jacob, Avi; Kinor, Noa; Shav-Tal, Yaron

2014-10-15

316

Interrelations between translation and general mRNA degradation in yeast  

PubMed Central

Messenger RNA (mRNA) degradation is an important element of gene expression that can be modulated by alterations in translation, such as reductions in initiation or elongation rates. Reducing translation initiation strongly affects mRNA degradation by driving mRNA toward the assembly of a decapping complex, leading to decapping. While mRNA stability decreases as a consequence of translational inhibition, in apparent contradiction several external stresses both inhibit translation initiation and stabilize mRNA. A key difference in these processes is that stresses induce multiple responses, one of which stabilizes mRNAs at the initial and rate-limiting step of general mRNA decay. Because this increase in mRNA stability is directly induced by stress, it is independent of the translational effects of stress, which provide the cell with an opportunity to assess its response to changing environmental conditions. After assessment, the cell can store mRNAs, reinitiate their translation or, alternatively, embark on a program of enhanced mRNA decay en masse. Finally, recent results suggest that mRNA decay is not limited to non-translating messages and can occur when ribosomes are not initiating but are still elongating on mRNA. This review will discuss the models for the mechanisms of these processes and recent developments in understanding the relationship between translation and general mRNA degradation, with a focus on yeast as a model system. How to cite this article: WIREs RNA 2014, 5:747–763. doi: 10.1002/wrna.1244 PMID:24944158

Huch, Susanne; Nissan, Tracy

2014-01-01

317

Development of an interleukin 2 receptor targeted gene therapy vehicle  

E-print Network

, holds great promise as a potential tool in the treatment of T cell cancers, autoimmune disease, and organ transplantation. Cancers of the T cells such as leukemia and lymphoma affect 100,000 people annually (The Leukemia & Lymphoma Society, 2004...). Furthermore, it is estimated that approximately 70.4 million patients in the United States suffer from autoimmune related disease (Arthritis Foundation, 2002; National Multiple Sclerosis Society, 2003-2004). Current nonspecific therapies involving the use...

Wattanakaroon, Wanida

2006-08-16

318

Lymphocyte Subsets and Interleukin-2 Receptors in Autistic Children.  

ERIC Educational Resources Information Center

Blood samples were obtained from 10 male autistic children, ages 7-15 years, and 10 controls. The children with autism had a lower percentage of helper-inducer cells and a lower helper:suppressor ratio, with both measures inversely related to the severity of autistic symptoms. (Author/DB)

Denney, Douglas R.; And Others

1996-01-01

319

Rituximab Plus Interleukin-2 in Treating Patients With Hematologic Cancer  

ClinicalTrials.gov

B-cell Adult Acute Lymphoblastic Leukemia; Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Nodal Marginal Zone B-cell Lymphoma; Noncontiguous Stage II Adult Burkitt Lymphoma; Noncontiguous Stage II Adult Diffuse Large Cell Lymphoma; Noncontiguous Stage II Adult Diffuse Mixed Cell Lymphoma; Noncontiguous Stage II Adult Diffuse Small Cleaved Cell Lymphoma; Noncontiguous Stage II Adult Immunoblastic Large Cell Lymphoma; Noncontiguous Stage II Adult Lymphoblastic Lymphoma; Noncontiguous Stage II Grade 1 Follicular Lymphoma; Noncontiguous Stage II Grade 2 Follicular Lymphoma; Noncontiguous Stage II Grade 3 Follicular Lymphoma; Noncontiguous Stage II Mantle Cell Lymphoma; Noncontiguous Stage II Marginal Zone Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Diffuse Small Cleaved Cell Lymphoma; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Splenic Marginal Zone Lymphoma; Stage III Adult Burkitt Lymphoma; Stage III Adult Diffuse Large Cell Lymphoma; Stage III Adult Diffuse Mixed Cell Lymphoma; Stage III Adult Diffuse Small Cleaved Cell Lymphoma; Stage III Adult Immunoblastic Large Cell Lymphoma; Stage III Adult Lymphoblastic Lymphoma; Stage III Grade 1 Follicular Lymphoma; Stage III Grade 2 Follicular Lymphoma; Stage III Grade 3 Follicular Lymphoma; Stage III Mantle Cell Lymphoma; Stage III Marginal Zone Lymphoma; Stage III Small Lymphocytic Lymphoma; Stage IV Adult Burkitt Lymphoma; Stage IV Adult Diffuse Large Cell Lymphoma; Stage IV Adult Diffuse Mixed Cell Lymphoma; Stage IV Adult Diffuse Small Cleaved Cell Lymphoma; Stage IV Adult Immunoblastic Large Cell Lymphoma; Stage IV Adult Lymphoblastic Lymphoma; Stage IV Grade 1 Follicular Lymphoma; Stage IV Grade 2 Follicular Lymphoma; Stage IV Grade 3 Follicular Lymphoma; Stage IV Mantle Cell Lymphoma; Stage IV Marginal Zone Lymphoma; Stage IV Small Lymphocytic Lymphoma

2013-06-05

320

Differential targeting of VDAC3 mRNA isoforms influences mitochondria morphology  

PubMed Central

Intracellular targeting of mRNAs has recently emerged as a prevalent mechanism to control protein localization. For mitochondria, a cotranslational model of protein import is now proposed in parallel to the conventional posttranslational model, and mitochondrial targeting of mRNAs has been demonstrated in various organisms. Voltage-dependent anion channels (VDACs) are the most abundant proteins in the outer mitochondrial membrane and the major transport pathway for numerous metabolites. Four nucleus-encoded VDACs have been identified in Arabidopsis thaliana. Alternative cleavage and polyadenylation generate two VDAC3 mRNA isoforms differing by their 3? UTR. By using quantitative RT-PCR and in vivo mRNA visualization approaches, the two mRNA variants were shown differentially associated with mitochondria. The longest mRNA presents a 3? extension named alternative UTR (aUTR) that is necessary and sufficient to target VDAC3 mRNA to the mitochondrial surface. Moreover, aUTR is sufficient for the mitochondrial targeting of a reporter transcript, and can be used as a tool to target an unrelated mRNA to the mitochondrial surface. Finally, VDAC3–aUTR mRNA variant impacts mitochondria morphology and size, demonstrating the role of mRNA targeting in mitochondria biogenesis. PMID:24889622

Michaud, Morgane; Ubrig, Elodie; Filleur, Sophie; Erhardt, Mathieu; Ephritikhine, Geneviève; Maréchal-Drouard, Laurence; Duchêne, Anne-Marie

2014-01-01

321

mRNA quality control: Marking the message for life or death  

Microsoft Academic Search

A protein complex deposited upstream of exon–exon junctions after pre-mRNA splicing may serve a dual role in mRNA quality control by directing mRNA nuclear export and, possibly, serving as a downstream ‘mark’ for nonsense-mediated decay.

Jens Lykke-Andersen

2001-01-01

322

PROLONGED FASTING AND CORTISOL REDUCE MYOSTATIN MRNA LEVELS IN TILAPIA LARVAE, SHORT-TERM FASTING ELEVATES  

Technology Transfer Automated Retrieval System (TEKTRAN)

Myostatin negatively regulates muscle growth and development and has recently been characterized in several fishes. We measured fasting myostatin mRNA levels in adult tilapia skeletal muscle and in whole larvae. Although fasting reduced some growth indices in adults, skeletal muscle myostatin mRNA...

323

Differentiation equations III: mRNA transcription-protein translation model  

NSDL National Science Digital Library

The first video segment presents a canonical mathematical example from quantitative biology, in which mRNA is transcribed from a gene sequence, and protein is translated from mRNA. The second segment uses eigenvector-eigenvalue analysis to sketch the trajectories of the system in a phase portrait. Finally, the third segment generalizes the linear stability analysis used to study this example.

Liao, David

324

GALLUS GALLUS PREPROGLUCAGON (GCG), CLASS A MRNA, TRANSCRIPT VARIANT 1, COMPLETE CDS  

Technology Transfer Automated Retrieval System (TEKTRAN)

In mammals, the proglucagon gene produces a single identical mRNA transcript in pancreas and intestine. This transcript encodes glucagon and two glucagon-like peptide hormones (GLP-1 and GLP-2). Previously, two distinct proglucagon mRNA transcripts have been identified in chickens, one in pancreas ...

325

GALLUS GALLUS PREPROGLUCAGON (GCG), CLASS B MRNA, TRANSCRIPT VARIANT 1, COMPLETE EDS  

Technology Transfer Automated Retrieval System (TEKTRAN)

The proglucagon gene in mammals produces a single identical mRNA transcript in all tissues where it is expressed. This transcript encodes glucagon and two glucagon-like peptide hormones (GLP-1 and GLP-2). Previously, two distinct proglucagon mRNA transcripts have been identified in chickens, one in...

326

Regulation of the mRNA half-life in breast cancer  

PubMed Central

The control of the half-life of mRNA plays a central role in normal development and in disease progression. Several pathological conditions, such as breast cancer, correlate with deregulation of the half-life of mRNA encoding growth factors, oncogenes, cell cycle regulators and inflammatory cytokines that participate in cancer. Substantial stability means that a mRNA will be available for translation for a longer time, resulting in high levels of protein gene products, which may lead to prolonged responses that subsequently result in over-production of cellular mediators that participate in cancer. The stability of these mRNA is regulated at the 3’UTR level by different mechanisms involving mRNA binding proteins, micro-RNA, long non-coding RNA and alternative polyadenylation. All these events are tightly inter-connected to each other and lead to steady state levels of target mRNAs. Compelling evidence also suggests that both mRNA binding proteins and regulatory RNAs which participate to mRNA half-life regulation may be useful prognostic markers in breast cancers, pointing to a potential therapeutic approach to treatment of patients with these tumors. In this review, we summarize the main mechanisms involved in the regulation of mRNA decay and discuss the possibility of its implication in breast cancer aggressiveness and the efficacy of targeted therapy. PMID:25114848

Griseri, Paola; Pagès, Gilles

2014-01-01

327

Physiological activation of brown adipose tissue destabilizes thermogenin mRNA.  

PubMed

The amount of mRNA coding for the brown fat specific uncoupling protein thermogenin was followed in the brown adipose tissue of adult mice. As expected, cold exposure or norepinephrine injection caused an increase in the amount of thermogenin mRNA. However, contrary to expectation, the half-life of thermogenin mRNA was dramatically reduced, from about 18 h to about 3 h, when the mice were cold exposed. This destabilization of thermogenin mRNA was not related to the activity of protein synthesis. It was concluded that in brown adipose tissue an unusual mechanism operates which leads to a destabilization of thermogenin mRNA under the same physiological conditions which increase thermogenin gene expression. PMID:3121388

Jacobsson, A; Cannon, B; Nedergaard, J

1987-11-30

328

CPSF30 and Wdr33 directly bind to AAUAAA in mammalian mRNA 3' processing.  

PubMed

AAUAAA is the most highly conserved motif in eukaryotic mRNA polyadenylation sites and, in mammals, is specifically recognized by the multisubunit CPSF (cleavage and polyadenylation specificity factor) complex. Despite its critical functions in mRNA 3' end formation, the molecular basis for CPSF-AAUAAA interaction remains poorly defined. The CPSF subunit CPSF160 has been implicated in AAUAAA recognition, but direct evidence has been lacking. Using in vitro and in vivo assays, we unexpectedly found that CPSF subunits CPSF30 and Wdr33 directly contact AAUAAA. Importantly, the CPSF30-RNA interaction is essential for mRNA 3' processing and is primarily mediated by its zinc fingers 2 and 3, which are specifically targeted by the influenza protein NS1A to suppress host mRNA 3' processing. Our data suggest that AAUAAA recognition in mammalian mRNA 3' processing is more complex than previously thought and involves multiple protein-RNA interactions. PMID:25301780

Chan, Serena L; Huppertz, Ina; Yao, Chengguo; Weng, Lingjie; Moresco, James J; Yates, John R; Ule, Jernej; Manley, James L; Shi, Yongsheng

2014-11-01

329

In the right place at the right time: visualizing and understanding mRNA localization.  

PubMed

The spatial regulation of protein translation is an efficient way to create functional and structural asymmetries in cells. Recent research has furthered our understanding of how individual cells spatially organize protein synthesis, by applying innovative technology to characterize the relationship between mRNAs and their regulatory proteins, single-mRNA trafficking dynamics, physiological effects of abrogating mRNA localization in vivo and for endogenous mRNA labelling. The implementation of new imaging technologies has yielded valuable information on mRNA localization, for example, by observing single molecules in tissues. The emerging movements and localization patterns of mRNAs in morphologically distinct unicellular organisms and in neurons have illuminated shared and specialized mechanisms of mRNA localization, and this information is complemented by transgenic and biochemical techniques that reveal the biological consequences of mRNA mislocalization. PMID:25549890

Buxbaum, Adina R; Haimovich, Gal; Singer, Robert H

2015-02-01

330

An Unbiased Analysis Method to Quantify mRNA Localization Reveals Its Correlation with Cell Motility  

PubMed Central

SUMMARY Localization of mRNA is a critical mechanism used by a large fraction of transcripts to restrict its translation to specific cellular regions. Although current high- resolution imaging techniques provide ample information, the analysis methods for localization have either been qualitative or employed quantification in non-randomly selected regions of interest. Here, we describe an analytical method for objective quantification of mRNA localization using a combination of two characteristics of its molecular distribution, polarization and dispersion. The validity of the method is demonstrated using single-molecule FISH images of budding yeast and fibroblasts. Live-cell analysis of endogenous ?-actin mRNA in mouse fibroblasts reveals that mRNA polarization has a half- life of ~16 min and is cross-correlated with directed cell migration. This novel approach provides insights into the dynamic regulation of mRNA localization and its physiological roles. PMID:22832165

Park, Hye Yoon; Trcek, Tatjana; Wells, Amber L.; Chao, Jeffrey A.; Singer, Robert H.

2014-01-01

331

Challenges and advances towards the rational design of mRNA vaccines.  

PubMed

In recent years, mRNA vaccines have emerged as a safe and potent approach for the induction of cellular immune responses. Whereas initial studies were limited to the ex vivo loading of dendritic cells (DCs) with antigen-encoding mRNA, recent progress has led to the development of improved mRNA vaccines that enable direct in vivo targeting of DCs. Although preclinical studies demonstrated their potency in inducing antitumor immunity, several bottlenecks hinder the broader application of mRNA vaccines. In this review, we discuss the challenges associated with mRNA-based vaccination strategies, the technological advances that have been made to overcome these limitations, and the hurdles that remain to be tackled for the development of an optimal mRNA vaccine. PMID:24138818

Pollard, Charlotte; De Koker, Stefaan; Saelens, Xavier; Vanham, Guido; Grooten, Johan

2013-12-01

332

Treatment of neurological disorders by introducing mRNA in vivo using polyplex nanomicelles.  

PubMed

Sensory nerve disorders are difficult to cure completely considering poor nerve regeneration capacity and difficulties in accurately targeting neural tissues. Administering mRNA is a promising approach for treating neurological disorders because mRNA can provide proteins and peptides in their native forms for mature non-dividing neural cells, without the need of entering their nuclei. However, direct mRNA administration into neural tissues in vivo has been challenging due to too unstable manner of mRNA and its strong immunogenicity. Thus, using a suitable carrier is essential for effective mRNA administration. For this purpose, we established a novel carrier based on the self-assembly of polyethylene glycol (PEG)-polyamino acid block copolymer, i.e. polyplex nanomicelles. To investigate the feasibility and efficacy of mRNA administration for the treatment of sensory nerve disorders, we used a mouse model of experimentally induced olfactory dysfunction. Intranasal administration of mRNA-loaded nanomicelles provided an efficient and sustained protein expression for nearly two days in nasal tissues, particularly in the lamina propria which contains olfactory nerve fibers, with effectively regulating the immunogenicity of mRNA. Consequently, once-daily intranasal administration of brain-derived neurotrophic factor (BDNF)-expressing mRNA using polyplex nanomicelles remarkably enhanced the neurological recovery of olfactory function along with repairing the olfactory epithelium to a nearly normal architecture. To the best of our knowledge, this is the first study to show the therapeutic potential of introducing exogenous mRNA for the treatment of neurological disorders. These results indicate the feasibility and safety of using mRNA, and provide a novel strategy of mRNA-based therapy. PMID:25599855

Baba, Miyuki; Itaka, Keiji; Kondo, Kenji; Yamasoba, Tatsuya; Kataoka, Kazunori

2015-03-10

333

Quantitative analysis of survivin mRNA expression in bladder transitional cell carcinomas.  

PubMed

We assessed the expression levels of survivin mRNA in bladder transitional cell carcinoma (TCC) to provide additional information regarding its malignant potential. The real-time PCR method was used to detect the survivin mRNA level for 21 bladder tumor specimens, and for urinary exfoliated cells from 12 newly diagnosed bladder tumor patients. All bladder tumor specimens and 7 of 12 voided urine specimens expressed survivin mRNA. In tumor specimens, high grade, high stage tumors had the tendency to express more survivin mRNA. Of 12 superficial bladder tumor patients who had transurethral resection of bladder tumor (TURB), 3 showed high survivin mRNA expression and intravesical recurrence after the surgery. However, for the patients who had total cystectomy due to invasive tumor, no relations were observed between the survivin mRNA expression level and development of local recurrence and/or distant metastasis. Our results suggested that the quantitative analysis of the survivin mRNA may indicate local malignant potential, which contribute to the possibility of an intravesical recurrence. PMID:18323166

Kitsukawa, Shin-ichi; Aoyagi, Teiichiro; Noda, Kenjiro; Ito, Takaaki; Yamamoto, Yutaka; Hosoda, Satoru; Otsuru, Norihiko; Matsumoto, Tetsuo

2008-02-01

334

Reduced beta 2-microglobulin mRNA levels in transgenic mice expressing a designed hammerhead ribozyme.  

PubMed Central

We have generated three artificial hammerhead ribozymes, denoted 'Rz-b', 'Rz-c' and 'Rz-d', with different specificities for exon II of the mouse beta-2-microglobulin (beta 2M) mRNA. In this study we tested for ribozyme mediated reduction of beta 2M mRNA in a cell line and in transgenic mice. Transfections of either of the Rz-b, Rz-c or Rz-d plasmids into a mouse cell-line (NIH/3T3) revealed reductions of beta 2M mRNA substrate in each case. Ribozyme expression in individual transfected clones was accompanied with an up to 80% reduction of beta 2M mRNA levels. Rz-c was selected for a transgenic study. Seven Rz-c transgenic founder animals were identified from which three ribozyme expressing families were established and analysed. Expression of the ribozyme transgene was tested for and detected in lung, kidney and spleen. Expression was accompanied with reduction of the beta 2M mRNA levels of heterozygous (Rz+/-) animals compared to non-transgenic litter mates. The effect was most pronounced in lung with more than 90% beta 2M mRNA reduction in individual mice. In summary, expression of our ribozymes in a cell free system, in a cell-line and in transgenic mice were all accompanied with reductions of beta 2M mRNA levels. Images PMID:8036151

Larsson, S; Hotchkiss, G; Andäng, M; Nyholm, T; Inzunza, J; Jansson, I; Ahrlund-Richter, L

1994-01-01

335

Half-life of Escherichia coli polyadenylated lipoprotein mRNA.  

PubMed

In the light of recent evidence that the half-life of bacterial mRNA may be modulated by polyadenylation at the 3' end, we determined the half-life of polyadenylated lpp mRNA, which is an abundant and comparatively stable message encoding a major lipoprotein of the outer membrane. Messenger RNAs were pulse-labeled with [3H] adenosine and poly(A) RNA was isolated at various times after pulse-labeling by affinity chromatography on oligo(dT) cellulose. The amount of lpp mRNA remaining was quantitated by hybridization with the antisense strand of the lpp gene cloned in M13mp18. The observed half-life for polyadenylated lpp mRNA was 12 min. This represents the first half-life measurement for a specific polyadenylated mRNA in E. coli, and is slightly longer than the half-life of total lpp RNA reported earlier. It coincides with the functional half-life for lpp mRNA determined by Inouye and coworkers by measuring the rate of lipoprotein synthesis at various times after rifampicin addition. This suggest that polyadenylated lpp RNA is the predominant and translationally active form of lpp mRNA within the cell. PMID:9192102

Taljanidisz, J; Shen, P; Sarkar, N

1997-06-01

336

Transcription Control Pathways Decode Patterned Synaptic Inputs into Diverse mRNA Expression Profiles  

PubMed Central

Synaptic plasticity requires transcription and translation to establish long-term changes that form the basis for long term memory. Diverse stimuli, such as synaptic activity and growth factors, trigger synthesis of mRNA to regulate changes at the synapse. The palette of possible mRNAs is vast, and a key question is how the cell selects which mRNAs to synthesize. To address this molecular decision-making, we have developed a biochemically detailed model of synaptic-activity triggered mRNA synthesis. We find that there are distinct time-courses and amplitudes of different branches of the mRNA regulatory signaling pathways, which carry out pattern-selective combinatorial decoding of stimulus patterns into distinct mRNA subtypes. Distinct, simultaneously arriving input patterns that impinge on the transcriptional control network interact nonlinearly to generate novel mRNA combinations. Our model combines major regulatory pathways and their interactions connecting synaptic input to mRNA synthesis. We parameterized and validated the model by incorporating data from multiple published experiments. The model replicates outcomes of knockout experiments. We suggest that the pattern-selectivity mechanisms analyzed in this model may act in many cell types to confer the capability to decode temporal patterns into combinatorial mRNA expression. PMID:24787753

Jain, Pragati; Bhalla, Upinder S.

2014-01-01

337

Intranasal mRNA nanoparticle vaccination induces prophylactic and therapeutic anti-tumor immunity  

PubMed Central

Direct in vivo administration of messenger RNA (mRNA) delivered in both naked and nanoparticle formats are actively investigated because the use of dendritic cells transfected ex vivo with mRNA for cancer therapy is expensive and needs significant infrastructure. Notably, intravenous and subcutaneous injections are the only routes of administration tested for mRNA nanoparticle tumor vaccination. In this report, we demonstrate that tumor immunity can be achieved via nasal administration of mRNA. Mice nasally immunized with mRNA delivered in nanoparticle format demonstrate delayed tumor progression in both prophylactic and therapeutic immunization models. The observed tumor immunity correlates with splenic antigen-specific CD8+ T cells and is achieved only when mRNA is delivered in nanoparticle but not in naked format. In conclusion, we demonstrate, as a proof-of-concept, a non-invasive approach to mRNA tumor vaccination, increasing its potential as a broadly applicable and off-the-shelf therapy for cancer treatment. PMID:24894817

Phua, Kyle K. L.; Staats, Herman F.; Leong, Kam W.; Nair, Smita K.

2014-01-01

338

NONOates regulate KCl cotransporter-1 and -3 mRNA expression in vascular smooth muscle cells.  

PubMed

Nitric oxide (NO) donors regulate KCl cotransport (KCC) activity and cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in sheep erythrocytes and in primary cultures of rat vascular smooth muscle cells (VSMCs), respectively. In this study, we used NONOates as rapid and slow NO releasers to provide direct evidence implicating NO as a regulator of KCC3 gene expression at the mRNA level. In addition, we used the expression of KCC3 mRNA to further investigate the mechanism of action of these NO donors at the cellular level. Treatment of VSMCs with rapid NO releasers, like NOC-5 and NOC-9, as well as with the direct NO-independent soluble guanylyl cyclase (sGC) stimulator YC-1, acutely increased KCC3 mRNA expression in a concentration- and time-dependent manner. The slow NO releaser NOC-18 had no effect on KCC3 gene expression. A specific NO scavenger completely prevented the NONOate-induced KCC3 mRNA expression. Inhibition of sGC with LY-83583 blocked the NONOate- and YC-1-induced KCC3 mRNA expression. This study shows that in primary cultures of rat VSMCs, the fast NO releasers NOC-9 and NOC-5, but not the slow NO releaser NOC-18, acutely upregulate KCC3 mRNA expression in a NO/sGC-dependent manner. PMID:12521940

Di Fulvio, Mauricio; Lauf, Peter K; Shah, Shalin; Adragna, Norma C

2003-05-01

339

Single-molecule modeling of mRNA degradation by miRNA: Lessons from data  

E-print Network

Recent experimental results on the effect of miRNA on the decay of its target mRNA have been analyzed against a previously hypothesized single molecule degradation pathway. According to that hypothesis, the silencing complex (miRISC) first interacts with its target mRNA and then recruits the protein complexes associated with NOT1 and PAN3 to trigger deadenylation (and subsequent degradation) of the target mRNA. Our analysis of the experimental decay patterns allowed us to refine the structure of the degradation pathways at the single molecule level. Surprisingly, we found that if the previously hypothesized network was correct, only about 7% of the target mRNA would be regulated by the miRNA mechanism, which is inconsistent with the available knowledge. Based on systematic data analysis, we propose the alternative hypothesis that NOT1 interacts with miRISC before binding to the target mRNA. Moreover, we show that when miRISC binds alone to the target mRNA, the mRNA is degraded more slowly, probably through a deadenylation-independent pathway. The new biochemical pathway we propose both fits the data and paves the way for new experimental work to identify new interactions.

Celine Sin; Davide Chiarugi; Angelo Valleriani

2014-10-20

340

The Formation of the Bicoid Morphogen Gradient Requires Protein Movement from Anteriorly Localized mRNA  

PubMed Central

The Bicoid morphogen gradient directs the patterning of cell fates along the anterior-posterior axis of the syncytial Drosophila embryo and serves as a paradigm of morphogen-mediated patterning. The simplest models of gradient formation rely on constant protein synthesis and diffusion from anteriorly localized source mRNA, coupled with uniform protein degradation. However, currently such models cannot account for all known gradient characteristics. Recent work has proposed that bicoid mRNA spatial distribution is sufficient to produce the observed protein gradient, minimizing the role of protein transport. Here, we adapt a novel method of fluorescent in situ hybridization to quantify the global spatio-temporal dynamics of bicoid mRNA particles. We determine that >90% of all bicoid mRNA is continuously present within the anterior 20% of the embryo. bicoid mRNA distribution along the body axis remains nearly unchanged despite dynamic mRNA translocation from the embryo core to the cortex. To evaluate the impact of mRNA distribution on protein gradient dynamics, we provide detailed quantitative measurements of nuclear Bicoid levels during the formation of the protein gradient. We find that gradient establishment begins 45 minutes after fertilization and that the gradient requires about 50 minutes to reach peak levels. In numerical simulations of gradient formation, we find that incorporating the actual bicoid mRNA distribution yields a closer prediction of the observed protein dynamics compared to modeling protein production from a point source at the anterior pole. We conclude that the spatial distribution of bicoid mRNA contributes to, but cannot account for, protein gradient formation, and therefore that protein movement, either active or passive, is required for gradient formation. PMID:21390295

Kneeland, Thomas B.; Wieschaus, Eric F.; Gregor, Thomas

2011-01-01

341

Expression and stability of c-sis mRNA in human glioblastoma cells  

SciTech Connect

The production of platelet-derived growth factor like (PDGF-like) material by glioblastomas may be involved in the conversion of normal cells to tumor cells. In an investigation of this problem, the authors have examined some of the properties of the platelet-derived growth factor B-chain mRNA (c-sis mRNA) by a sensitive and quantitative RNA-RNA solution hybridization method. In 5 out of 8 human glioblastoma cell lines, c-sis mRNA was present, and in the line with the highest level, there were approximately 4-10 molecules per cell. The half-lives of the c-sis mRNA in two glioblastoma cell lines were 2.6 and 3.4 h, while in human umbilical vein endothelial (HUVE) and bladder carcinoma (T24) cells they were 1.6 and 2.5 h, respectively. Inhibiting protein synthesis produced no significant alteration of the c-sis mRNA half-lives in the glioblastoma or HUVE cells. The A-U-rich sequence at the 3' end of the c-sis mRNA therefore does not appear to affect the mRNA stability in the presence of cycloheximide as it does in other transcripts. The similarity of the c-sis mRNA half-lives in normal and tumor cells suggests that regulation of stability of c-sis mRNA is not a major factor in tumorigenesis in the glioblastoma cell lines examined.

Press, R.D.; Samols, D.; Goldthwait, D.A.

1988-07-26

342

Location of an inducible nitric oxide synthase mRNA in the normal kidney.  

PubMed

An inducible nitric oxide synthase (iNOS) mRNA was found primarily in the outer medulla of normal rat kidney. Identification of the mRNA was based upon the specificity of the oligonucleotide primers used for PCR amplification, PCR-Southern blot analysis and the nucleic acid sequence of the cloned PCR product. In addition to the outer medulla, glomeruli prepared from normal rat kidney contained significant amounts of an iNOS mRNA. These results suggest that there may be tonic influences in the outer medulla of the normal rat kidney resulting in the "steady-state" presence of an iNOS mRNA. Cortical tubules and the inner medulla were found to contain detectable but lesser amounts of the iNOS mRNA. The outer medulla was microdissected into proximal straight tubule (PST), medullary thick ascending limb (MTAL), medullary collecting duct (MCD) and vasa recta bundle (VRB). The iNOS mRNA was found primarily in the MTAL with minor amounts in the MCD and VRB of normal rat kidney. Animals were injected with lipopolysaccharide (LPS) and sacrificed 24 hours later. Treatment with LPS caused at least a 20-fold increase in the amount of iNOS mRNA in the liver or in macrophages isolated from the peritoneum. Endotoxin treatment led to over a 10-fold increase in iNOS mRNA content in glomeruli and the inner medulla. The iNOS mRNA level of the outer medulla was increased two- to threefold due to LPS treatment. PMID:7516453

Morrissey, J J; McCracken, R; Kaneto, H; Vehaskari, M; Montani, D; Klahr, S

1994-04-01

343

Strong association between mRNA folding strength and protein abundance in S. cerevisiae  

PubMed Central

One of the open questions in regulatory genomics is how the efficiency of gene translation is encoded in the coding sequence. Here we analyse recently generated measurements of folding energy in Saccharomyces cerevisiae, showing that genes with high protein abundance tend to have strong mRNA folding (mF; R=0.68). mF strength also strongly correlates with ribosomal density and mRNA levels, suggesting that this relation at least partially pertains to the efficiency of translation elongation, presumably by preventing aggregation of mRNA molecules. PMID:22249164

Zur, Hadas; Tuller, Tamir

2012-01-01

344

Spatial distribution and temporal accumulation of mRNA encoding diamine oxidase during lentil ( Lens culinaris Medicus) seedling development  

Microsoft Academic Search

During the first phases of lentil epicotyl development (up to 72 h), diamine oxidase (DAO) mRNA accumulation and the time course of enzyme activity are correlated. In the following days total DAO activity remained unchanged but the mRNA level decreased. Both enzyme activity and mRNA accumulation showed a spatial distribution along the epicotyl that was also developmentally regulated. DAO mRNA

Riccardo Angelini; Giuseppina Rea; Rodolfo Federico; Renato D'Ovidio

1996-01-01

345

Visual detection of Akt mRNA in living cell using gold nanoparticle beacon  

NASA Astrophysics Data System (ADS)

PI3K-Akt signaling pathway plays the key role in cell apoptosis and survival, and the components of PI3K /Akt signaling pathway are often abnormally expressed in human tumors. Therefore, determination of the Akt (protein kinase B, PKB) messenger ribonucleic acid (mRNA) expression is significantly important in understanding the mechanism of tumor progression. In this study, we designed a special hairpin deoxyribonucleic acid (DNA) functionalized with gold nanoparticles and fluorescein isothiocyanate(FITC) as a beacon for detecting human Akt mRNA. Spectrofluorometer was used to detect the fluorescence quenching and recovery of the beacons, and laser confocal scanning microscopy was adopted to image Akt mRNA in cells. The results showed that this beacon could sensitively and quantitatively measure the Akt mRNA in living cells . This strategy is potentially useful for the cellular imaging of RNA or protein expression in living cells.

Ma, Yi; Tian, Caiping; Li, Siwen; Wang, Zhaohui; Gu, Yueqing

2014-09-01

346

Polyadenylylation destabilizes the rpsO mRNA of Escherichia coli.  

PubMed Central

The rpsO mRNA, encoding ribosomal protein S15, is only partly stabilized when the three ribonucleases implicated in its degradation--RNase E, polynucleotide phosphorylase, and RNase II--are inactivated. In the strain deficient for RNase E and 3'-to-5' exoribonucleases, degradation of this mRNA is correlated with the appearance of posttranscriptionally elongated molecules. We report that these elongated mRNAs harbor poly(A) tails, most of which are fused downstream of the 3'-terminal hairpin at the site where transcription terminates. Poly(A) tails are shorter in strains containing 3'-to-5' exoribonucleases. Inactivation of poly(A) polymerase I (pcnB) prevents polyadenylylation and stabilizes the rpsO mRNA if RNase E is inactive. In contrast polyadenylylation does not significantly modify the stability of rpsO mRNA undergoing RNase E-mediated degradation. Images Fig. 2 Fig. 3 Fig. 4 PMID:7732015

Hajnsdorf, E; Braun, F; Haugel-Nielsen, J; Régnier, P

1995-01-01

347

Characterization of polydnavirus-encoded mRNA in parasitized armyworm larvae.  

PubMed

We have isolated five cDNA clones encoding Cotesia kariyai polydnavirus (CkPDV) mRNAs transcribed in parasitized host larvae of Pseudaletia separata. One of the cDNAs encoding the longest 2.0 kb CkPDV mRNA was sequenced and characterized. Southern hybridization analyses using the cloned cDNA as a probe showed that CkPDV mRNA was homologous to one of CkPDV DNA segments, 5.6 kbp DNA segment A. The 2.0-kb mRNA was demonstrated as being expressed in the parasitized host larvae by Northern-blot analyses. When specific host tissues were examined, the 2.0-kb mRNA was detected mainly in haemocytes. This RNA increased in relative abundance after 2 and 4 h post-parasitization when the immune response of host haemocytes appeared compromised. PMID:8969465

Yamanaka, A; Hayakawa, Y; Noda, H; Nakashima, N; Watanabe, H

1996-06-01

348

Eucaryotic RNA polymerase conditional mutant that rapidly ceases mRNA synthesis.  

PubMed Central

We have isolated a yeast conditional mutant which rapidly ceases synthesis of mRNA when subjected to the nonpermissive temperature. This mutant (rpb1-1) was constructed by replacing the wild-type chromosomal copy of the gene encoding the largest subunit of RNA polymerase II with one mutagenized in vitro. The rapid cessation of mRNA synthesis in vivo and the lack of RNA polymerase II activity in crude extracts indicate that the mutant possesses a functionally defective, rather than an assembly-defective, RNA polymerase II. The shutdown in mRNA synthesis in the rpb1-1 mutant has pleiotropic effects on the synthesis of other RNAs and on the heat shock response. This mutant provides direct evidence that the RPB1 protein has a functional role in mRNA synthesis. Images PMID:3299050

Nonet, M; Scafe, C; Sexton, J; Young, R

1987-01-01

349

Inefficient SRP Interaction with a Nascent Chain Triggers a mRNA Quality Control Pathway  

PubMed Central

SUMMARY Misfolded proteins are often cytotoxic, unless cellular systems prevent their accumulation. Data presented here uncover a novel mechanism by which defects in secretory proteins lead to a dramatic reduction in their mRNAs and protein expression. When mutant signal sequences fail to bind to the signal recognition particle (SRP) at the ribosome exit site, the nascent chain instead contacts Argonaute2 (Ago2) and the mutant mRNAs are specifically degraded. Severity of signal sequence mutations correlated with increased proximity of Ago2 to nascent chain, and mRNA degradation. Ago2 knockdown inhibited degradation of the mutant mRNA, while overexpression of Ago2 or knockdown of SRP54 promoted degradation of secretory protein mRNA. The results reveal a previously unappreciated general mechanism of translational quality control, in which specific mRNA degradation preemptively regulates aberrant protein production (RAPP). PMID:24439374

Karamyshev, Andrey L.; Patrick, Anna E.; Karamysheva, Zemfira N.; Griesemer, Dustin S.; Hudson, Henry; Tjon-Kon-Sang, Sandra; Nilsson, IngMarie; Otto, Hendrik; Liu, Qinghua; Rospert, Sabine; von Heijne, Gunnar; Johnson, Arthur E.; Thomas, Philip J.

2014-01-01

350

Screening of Different Organs of Rats for HCA2 Receptor mRNA.  

PubMed

Interest in hydroxy - carboxylic acid 2 (HCA2) receptor has been raised since it is the target of antidyslipidemic drug nicotinic acid. The present study aimed to evaluate the presence of mRNA of this receptor in different organs of laboratory rat. Twenty two different organs of rats including mesenteric fat, epididymis (head, body and tail), testis, ovary, xiphoid process, liver, adrenal gland, femoral head, proximal epiphyseal and metaphyseal bone marrow of femur, esophagus, glandular stomach, forestomach, intestines, colons, heart, spleen, kidney, trachea, lung, skeletal muscle (quadriceps), cerebrum and cerebellum were removed and examined for HCA2 mRNA by RT- PCR method. The mRNA for HCA2 receptor was detected in all analyzed tissues. In conclusion, the different organs of rat express HCA2 receptor mRNA which makes a proper animal model for future studies on the physiological and pharmacological roles of this receptor in vivo. PMID:25035863

Shomali, Tahoora; Mosleh, Najmeh; Kamalpour, Mohammad

2014-01-01

351

Regulation of gadd153 mRNA expression by hypoxia in pulmonary artery smooth muscle cells.  

PubMed

Hypoxia causes pulmonary hypertension and induces oxygen radicals in pulmonary artery smooth muscle cells (PASMCs). Since oxidative stress regulates gaddl53 expression, we examined gaddl53 mRNA in PASMCs cultured in a hypoxic environment. Gadd153 mRNA content was increased in PASMCs cultured for 24 hours in 1% oxygen. This increase was not abrogated by inhibition of protein synthesis. To explore the signaling pathways mediating hypoxic regulation of gaddl53 mRNA, the impact of calcium channel blockade by verapamil, G protein inhibition by pertussis toxin, and protein kinase C (PKC) down-regulation, was examined. Although none of these interventions reduced basal expression of gaddl53 mRNA in PASMCs, all of them suppressed the induction by hypoxia. In contrast, antioxidants had no effect. These observations indicate hypoxia induces gaddl53 expression in PASMCs through common signaling pathways. PMID:11758972

Chen, R; Harrod, K S; Olson, J W; Gillespie, M N

2000-01-01

352

Exogenous mRNA delivery and bioavailability in gene transfer mediated by piggyBac transposition  

PubMed Central

Background Up to now, the different uptake pathways and the subsequent intracellular trafficking of plasmid DNA have been largely explored. By contrast, the mode of internalization and the intracellular routing of an exogenous mRNA in transfected cells are poorly investigated and remain to be elucidated. The bioavailability of internalized mRNA depends on its intracellular routing and its potential accumulation in dynamic sorting sites for storage: stress granules and processing bodies. This question is of particular significance when a secure transposon-based system able to integrate a therapeutic transgene into the genome is used. Transposon vectors usually require two components: a plasmid DNA, carrying the gene of interest, and a source of transposase allowing the integration of the transgene. The principal drawback is the lasting presence of the transposase, which could remobilize the transgene once it has been inserted. Our study focused on the pharmacokinetics of the transposition process mediated by the piggyBac transposase mRNA transfection. Exogenous mRNA internalization and trafficking were investigated towards a better apprehension and fine control of the piggyBac transposase bioavailability. Results The mRNA prototype designed in this study provides a very narrow expression window of transposase, which allows high efficiency transposition with no cytotoxicity. Our data reveal that exogenous transposase mRNA enters cells by clathrin and caveolae-mediated endocytosis, before finishing in late endosomes 3 h after transfection. At this point, the mRNA is dissociated from its carrier and localized in stress granules, but not in cytoplasmic processing bodies. Some weaker signals have been observed in stress granules at 18 h and 48 h without causing prolonged production of the transposase. So, we designed an mRNA that is efficiently translated with a peak of transposase production 18 h post-transfection without additional release of the molecule. This confines the integration of the transgene in a very small time window. Conclusion Our results shed light on processes of exogenous mRNA trafficking, which are crucial to estimate the mRNA bioavailability, and increase the biosafety of transgene integration mediated by transposition. This approach provides a new way for limiting the transgene copy in the genome and their remobilization by mRNA engineering and trafficking. PMID:24070093

2013-01-01

353

Post-Transcriptional Regulation of 5-Lipoxygenase mRNA Expression via Alternative Splicing and Nonsense-Mediated mRNA Decay  

PubMed Central

5-Lipoxygenase (5-LO) catalyzes the two initial steps in the biosynthesis of leukotrienes (LT), a group of inflammatory lipid mediators derived from arachidonic acid. Here, we investigated the regulation of 5-LO mRNA expression by alternative splicing and nonsense-mediated mRNA decay (NMD). In the present study, we report the identification of 2 truncated transcripts and 4 novel 5-LO splice variants containing premature termination codons (PTC). The characterization of one of the splice variants, 5-LO?3, revealed that it is a target for NMD since knockdown of the NMD factors UPF1, UPF2 and UPF3b in the human monocytic cell line Mono Mac 6 (MM6) altered the expression of 5-LO?3 mRNA up to 2-fold in a cell differentiation-dependent manner suggesting that cell differentiation alters the composition or function of the NMD complex. In contrast, the mature 5-LO mRNA transcript was not affected by UPF knockdown. Thus, the data suggest that the coupling of alternative splicing and NMD is involved in the regulation of 5-LO gene expression. PMID:22363630

Ochs, Meike J.; Sorg, Bernd L.; Pufahl, Laura; Grez, Manuel; Suess, Beatrix; Steinhilber, Dieter

2012-01-01

354

Cloning of cDNA to rat mammary-gland fatty acid synthase mRNA. Evidence for the expression of two mRNA species during lactation.  

PubMed Central

A cDNA library was constructed in the expression vector lambda gt11, by synthesizing cDNA from size-selected poly(A) RNA from lactating rat mammary gland, using random hexanucleotide primers. Using this library we identified two recombinants which, on addition of a lac z inducer, produced proteins recognized by affinity-purified anti-fatty-acid synthase antibody, and which, therefore, contained fatty acid synthase coding sequences. The inserts were subcloned, were shown to be between 500 and 600 base pairs in size, and to cross-hybridize. The cloned DNA was then used in Northern hybridizations with mRNA isolated at various stages throughout lactation. Two mRNA species were identified of approx. 9.7 and 10.4 kilobases, which increased and decreased in parallel during lactation, reaching a peak at 12-13 days. Both mRNA species disappeared rapidly if the pups were removed prematurely. This study provides evidence that, during hormonal induction in lactation, regulation of the level of fatty acid synthase protein can be accounted for by variation in the level of mRNA. Images Fig. 1. Fig. 2. Fig. 3. PMID:3342031

Braddock, M; Hardie, D G

1988-01-01

355

The myosin motor, Myo4p, binds Ash1 mRNA via the adapter protein, She3p  

E-print Network

supported by the important observations that green fluorescent protein (GFP)-tagged Ash1 mRNA moves fromThe myosin motor, Myo4p, binds Ash1 mRNA via the adapter protein, She3p Peter A. Takizawa) In Saccharomyces cerevisiae, mRNA encoding the cell-fate deter- minant Ash1p is localized to the distal tip

Vale, Ronald D.

356

Quality control of mRNA 3'-end processing is linked to the nuclear exosome  

Microsoft Academic Search

An emerging theme in messenger RNA metabolism is the coupling of nuclear pre-mRNA processing events, which contributes to mRNA quality control. Most eukaryotic mRNAs acquire a poly(A) tail during 3'-end processing within the nucleus, and this is coupled to efficient export of mRNAs to the cytoplasm. In the yeast Saccharomyces cerevisiae, a common consequence of defective nuclear export of mRNA

Patricia Hilleren; Terri McCarthy; Michael Rosbash; Roy Parker; Torben Heick Jensen

2001-01-01

357

Effects of Ginseng and Echinacea on Cytokine mRNA Expression in Rats  

PubMed Central

The aim of the study was to determine the effect of ginseng and echinacea on the mRNA expression of IL-10, TNF-?, and TGF-?1 in healthy rats. Six-week-old male Fischer 344 rats (n = 48) were used. The animals were divided into three equal groups, as follows: control (C); ginseng (G); echinacea (E). While the C group was fed a standard rat diet (Purina) ad libitum for a period of 40 days, the G and E groups animals received the same diet containing 0.5?g/kg of Panax ginseng root powder and 0.75?g/kg of Echinacea purpurea root powder, respectively. Blood samples were obtained from 8 rats in each group after 20 and 40 days of treatment, and the mRNA expression of IL-10, TNF-?, and TGF-?1 was determined. After 20 days of treatment, the expression of IL-10 mRNA in the G group was different from the C group (P < 0.05); however, after 40 days of treatment, there was no difference between the groups. There was no difference after 20 and 40 days of treatment between the groups with respect to the expression of TGF-?1 mRNA. After 20 days of treatment, the expression of TNF-? mRNA in the E group was higher (P < 0.05) than the C group. After 40 days of treatment, the expression of TNF-? mRNA was similar in all of the groups. Based on the current study, the increase in expression of IL-10 mRNA in the G group and the increase in expression of TNF-? mRNA in the E group support the use of these plants for purposes of modulating the immune system. However, a more detailed study regarding the effects of ginseng and echinacea on these cytokines and other cytokines is needed. PMID:22666172

Ulu???k, Deniz; Keskin, Ercan

2012-01-01

358

T-Lymphocyte cytokine mRNA expression in cystic echinococcosis  

Microsoft Academic Search

In the present study we investigated cytokine mRNA expression by peripheral blood mononuclear cells (PBMC) from patients with cystic echinococcosis (CE) after stimulation with different antigens. By using reverse transcriptase polymerase chain reaction (RT-PCR) we could demonstrate that restimulation with crude Echinococcus granulosus antigen (Eg-Ag) induced or enhanced Th2 cytokine mRNA expression, especially IL-5 (by using antigen from sheep cyst

Susanne Fauser; Peter Kern

1997-01-01

359

Postmortem quantitative mRNA analyses of death investigation in forensic pathology: An overview and prospects  

Microsoft Academic Search

To analyze pathophysiological dynamics of the death process using mRNA quantification, previous studies investigated pulmonary surfactant-associated protein (SP-A), as well as hypoxia-inducible factor 1 (HIF-1) and its downstream factors. Quantitative assays of these mRNA transcripts were established using TaqMan real-time RT-PCR. Experimental studies showed that most of these factors in forensic autopsy materials gradually degraded in patterns similar to those

Dong Zhao; Takaki Ishikawa; Li Quan; Tomomi Michiue; Bao-Li Zhu; Hitoshi Maeda

2009-01-01

360

Factors Altering Ribozyme-Mediated Cleavage of Tumor Necrosis Factor-? mRNA in Vitro  

Microsoft Academic Search

Hammerhead ribozymes are capable of cleaving RNA in a sequence specific mannerin vitro.However, the complex environment of the cell differs dramatically from the conditionsin vitro.Therefore, we explored cleavage of full-length target RNA with two ribozymes targeted against the murine tumor necrosis factor-alpha (TNF-?) mRNA. These ribozymes cleaved TNF-? mRNA within a pool of total cellular RNAin vitro,but less efficiently than

Kevin O. Kisich; Stephen J. Freedland; Kent L. Erickson

1997-01-01

361

Analysis of mRNA expression for interleukin-1 genes on human testicular cells  

Microsoft Academic Search

We have investigated mRNA expression for IL-1? and IL-1? gene on fractionated human testicular cells. Using RT-PCR and Northern blot hybridization technique we detected the presence of IL-1? transcripts, predominantly in the intratubular compartment of the testis, comprising gametogenic and Sertoli cells. We were also able to detect mRNA for IL-1? on the testicular interstitium, but at significantly lower levels.

Micha? Janitz; Dorota Fiszer; Andrzej ?ukaszyk; Witold Skorupski; Maciej Kurpisz

1995-01-01

362

D1 and D2 dopamine receptor mRNA in rat brain.  

PubMed Central

Physiological and pharmacological criteria have divided dopamine receptors into D1 and D2 subtypes, and genes encoding these subtypes have recently been cloned. Based on the sequences of the cloned receptors, we prepared oligodeoxynucleotide probes to map the cellular expression of the corresponding mRNAs in rat brain by in situ hybridization histochemistry. These mRNAs showed largely overlapping yet distinct patterns of expression. The highest levels of expression for both mRNAs were observed in the caudate-putamen, nucleus accumbens, and olfactory tubercle. Within the caudate-putamen, 47 +/- 6% and 46 +/- 5% of the medium-sized neurons (10-15 microns) expressed the D1 and D2 mRNAs, respectively, and only the D2 mRNA was observed in the larger neurons (greater than 20 microns). The D1 and D2 mRNAs were expressed in most cortical regions, with the highest levels in the prefrontal and entorhinal cortices. Within neocortex, D1 mRNA was observed primarily in layer 6 and D2 mRNA in layers 4-5. Within the amygdala, D1 mRNA was observed in the intercalated nuclei, and D2 mRNA in the central nucleus. Within the hypothalamus, D1 mRNA was observed in the suprachiasmatic nucleus and D2 mRNA in many of the dopaminergic cell groups. Within the septum, globus pallidus, superior and inferior colliculi, mammillary bodies, and substantia nigra only D2 mRNA was detected. These data provide insight into the neuroanatomical basis of the differential effects of drugs that act on D1 or D2 receptors. Images PMID:1825729

Weiner, D M; Levey, A I; Sunahara, R K; Niznik, H B; O'Dowd, B F; Seeman, P; Brann, M R

1991-01-01

363

Nerve growth factor (NGF) and NGF mRNA change in rat uterus during pregnancy  

Microsoft Academic Search

During pregnancy, the uterus undergoes a profound sympathetic denervation. To explore whether this is associated with changes in neurotrophic factors, we assayed nerve growth factor (NGF) and NGF mRNA in the uterus of non-pregnant and pregnant rats. In the uterine horn, the concentration of NGF and its mRNA decreased during middle and late pregnancy. However, when values were corrected for

Fusun G Varol; Anne-Marie Duchemin; Norton H Neff; Maria Hadjiconstantinou

2000-01-01

364

Sleep deprivation differentially alters the mRNA and protein levels of neurogranin in rat brain  

Microsoft Academic Search

The mRNA level of the 17-kDa protein neurogranin (NG), a postsynaptic substrate of the protein kinase C, has previously been found to be decreased in rat forebrain after 24-h sleep deprivation (SD). To investigate the functional significance of this finding in various forebrain regions, the effect of 24-h SD on the mRNA level and the protein level of NG was

Martin Neuner-Jehle; Thomas A. Rhyner; Alexander A. Borbély

1995-01-01

365

Expression pattern of cytosolic glutathione peroxidase (cGPx) mRNA during mouse embryogenesis.  

PubMed

The selenoprotein cytosolic glutathione peroxidase (cGPx) is ubiquitously distributed in a variety of organs, and its primary function is to protect oxidative damage. To investigate the spatial and temporal expression pattern of cGPx mRNA in embryogenesis, as this has not been studied before, reverse transcription-polymerase chain reaction (RT-PCR) was carried out in a thermal cycler using mouse-specific cGPx primers, and in situ hybridization was performed in whole embryos or embryonic tissues using digoxigenin-labeled mouse cGPx riboprobes. Expression of cGPx mRNA was detected in all the embryos retrieved from embryonic days (EDs) 7.5 to 18.5. On EDs 10.5-12.5, cGPx mRNA was highly expressed in the margin of forelimb and hindlimb buds and dorsally in the cranial neural tube, including the telencephalon, diencephalon, and hindbrain neural tube. On ED 13.5, cGPx mRNA was accumulated especially in vibrissae, forelimb and hindlimb plates, tail, and spinal cord. On EDs 14.5-16.5, cGPx mRNA was found in the developing brain, Rathke's pouch, thymus, lung, and liver. On ED 17.5, the expression of cGPx mRNA was apparent in various tissues such as brain, submandibular gland, vibrissae, heart, lung, liver, stomach, intestine, pancreas, skin, and kidney. In particular, cGPx mRNA was greatly expressed in epithelial linings and metabolically active sites such as whisker follicles, alveolar epithelium of lung, surface epithelium and glandular region of stomach, skin epithelium, and cortex and tubules of kidney. Overall results indicate that cGPx mRNA is expressed in developing embryos, cell-specifically and tissue-specifically, suggesting that cGPx may function to protect the embryo against reactive oxygen species and/or hydroperoxides massively produced by the intracellular or extracellular environment. PMID:15789223

Baek, In-Jeoung; Yon, Jung-Min; Lee, Beom Jun; Yun, Young Won; Yu, Wook-Joon; Hong, Jin Tae; Ahn, Byeongwoo; Kim, Yun-Bae; Kim, Dae Joong; Kang, Jong-Koo; Nam, Sang-Yoon

2005-04-01

366

Ascorbic acid 2-phosphate enhances albumin mRNA expression and secretion of porcine hepatocytes  

Microsoft Academic Search

The effect of ascorbic acid 2-phosphate (Asc2P) was tested on porcine and rat mature hepatocytes in vitro. Asc2P enhanced the porcine albumin mRNA expression. The albumin secretion released into the culture medium from the porcine hepatocytes was also increased by Asc2P. However, Asc2P did not give any effects to rat hepatocytes on albumin mRNA expression and albumin secretion. This result

Dawei Yang; Toshie Koyama; Ai Okamura; Yoshiaki Shiba; Takayuki Akimoto; Makoto Kodama

2004-01-01

367

Relating mRNA and protein biomarker levels in a Dehalococcoides and Methanospirillum-containing community.  

PubMed

To better understand the quantitative relationships between messenger RNA (mRNA) and protein biomarkers relevant to bioremediation, we quantified and compared respiration-associated gene products in an anaerobic syntrophic community. Respiration biomarkers for Dehalococcoides, an organohalide reducer, and Methanospirillum, a hydrogenotrophic methanogen, were quantified via qRT-PCR for mRNA and multiple reaction monitoring (MRM) of proteotypic peptides for protein. mRNA transcripts of the Dehalococcoides reductive dehalogenases PceA, TceA, and DMC1545, and hydrogenase HupL, as well as the Methanospirillum oxidoreductases MvrD and FrcA were shown to be similarly regulated with respect to their temporal responses to substrate addition. However, MvrD was two orders of magnitude lower in mRNA abundance. Per cell, Dehalococcoides protein biomarkers quantified were more abundant than Methanospirillum proteins. Comparing mRNA with protein abundance, poor correlations were observed between mRNA transcript levels and the net protein produced. For example, Dehalococcoides HupL and TceA transcripts were similarly abundant though TceA was far more abundant at the protein level (167?±?121 vs. 1095?±?337 proteins per cell, respectively). In Methanospirillum, MvrD maintained comparable per-cell protein abundance to FrcA (42?±?14 vs. 60?±?1 proteins per cell, respectively) despite the significantly lower transcript levels. Though no variability in protein decay rates was observed, the mRNA translation rate quantified for TceA was greater than the other Dehalococcoides targets monitored. These data suggest that there is considerable variation in the relationship between mRNA abundance and protein production both across transcripts within an organism and across organisms. This highlights the importance of empirically based studies for interpreting biomarker levels in environmentally relevant organisms. PMID:25467924

Rowe, Annette R; Mansfeldt, Cresten B; Heavner, Gretchen L; Richardson, Ruth E

2014-12-01

368

Changes in mRNA expression levels of solute carrier transporters in inflammatory bowel disease patients.  

PubMed

Inflammatory bowel disease (IBD) is an inflammatory condition that affects the gastrointestinal tract. The solute carrier (SLC) superfamily of transporters comprise proteins involved in the uptake of drugs, hormones, and other biologically active compounds. The purpose of this study was to determine the mRNA expression levels of 15 solute carrier transporters in two regions of the intestine in IBD patients. Endoscopic biopsy specimens were taken from two locations (terminal ileum and colon) for histological examination and RNA extraction. We quantitatively measured the mRNA expression of 15 SLC transporters in 107 IBD patients (53 with Crohn's disease and 54 with ulcerative colitis) and 23 control subjects. mRNA expression was evaluated using the quantitative reverse transcription-polymerase chain reaction technique. We observed that in the ileum of IBD patients, mRNA levels for serotonin transporter, equilibrative nucleoside transporter (ENT) 1, ENT2, and organic anion-transporting polypeptide (OATP) 2B1 were significantly elevated, whereas levels for apical sodium-dependent bile acid transporter (ASBT) and organic zwitterion/cation transporter (OCTN) 2 were significantly lower. In colon, mRNA levels for ENT1, ENT2, concentrative nucleoside transporter (CNT) 2, OATP2B1, and OATP4A1 were significantly higher, whereas mRNA levels for OCTN2 were significantly decreased. In inflamed colon of IBD patients the mRNA expression levels of ENT1, ENT2, CNT2, OATP2B1, OATP4A1, and peptide transporter 1 were significantly higher. We conclude that intestinal SLC mRNA levels are dysregulated in IBD patients, which may be linked to the inflammation of the tissue and provides an indication about the role of inflammatory signaling in regulation of SLC expression. PMID:19487253

Wojtal, Kacper A; Eloranta, Jyrki J; Hruz, Petr; Gutmann, Heike; Drewe, Jürgen; Staumann, Alex; Beglinger, Christoph; Fried, Michael; Kullak-Ublick, Gerd A; Vavricka, Stephan R

2009-09-01

369

mRNA Expression Analysis of Tissue Sections and Single Cells  

Microsoft Academic Search

Thirty years of literature have shown that changes in mRNA abundance provide insight into cellular functioning. Indeed, mRNA abundance measurements have been used to determine how effective particular drugs are in eliciting cellular responses, in monitoring behavioral responses, and in several other areas of neurobiology. Although investigation of individual mRNAs was informative, there are many thousands of mRNAs expressed in

James Eberwine; Janet Estee Kacharmina; Christine Andrews; Kevin Miyashiro; Tracy McIntosh; Kevin Becker; Tanya Barrett; Dave Hinkle; Gersham Dent; Paolo Marciano

370

RNA-seq approaches for determining mRNA abundance in Leishmania.  

PubMed

High-throughput sequencing of cDNA copies of mRNA (RNA-seq) provides a digital read-out of mRNA levels over several orders of magnitude, as well as mapping the transcripts to the nucleotide level. Here we describe an RNA-seq approach that exploits the 39-nucleotide mini-exon or spliced leader (SL) sequence found at the 5' end of all Leishmania (and other trypanosomatid) mRNAs. PMID:25388116

Haydock, Andrew; Terrao, Monica; Sekar, Aarthi; Ramasamy, Gowthaman; Baugh, Loren; Myler, Peter J

2015-01-01

371

Phosphorylation is a regulatory mechanism in apolipoprotein B mRNA editing.  

PubMed Central

The editing of apolipoprotein B (apoB) mRNA is under tissue-specific, developmental and metabolic regulation. We found that multiple protein kinase inhibitors or activators increased apoB mRNA editing up to 2.5-fold in Caco-2 cells and 3-8-fold in McA7777 and FAO rat cells respectively. The phosphorylation-agent-induced modulation is independent of the apolipoprotein B editing catalytic subunit 1 (APOBEC-1) and of apoB mRNA expression levels, indicating the involvement of a protein modification, such as phosphorylation, regulating the cellular editing of apoB mRNA. Transient expression of protein kinase C-? more than doubled apoB mRNA editing in FAO cells. Chronic exposure to ethanol, a treatment known to increase the expression of protein kinases and to change protein phosphorylation status, increased apoB mRNA editing in FAO cells up to 2.5-fold without increasing the mRNA abundance of APOBEC-1. The elimination of potential phosphorylation sites 47 and 72 of human APOBEC-1 decreased its activity to approx. one-eighth of control levels by a Ser(47)-->Ala mutation, but more than doubled the activity by a Ser(72)-->Ala mutation. The activity modulation was reversed by a Ser-->Asp mutation at sites 47 and 72, which introduced a phosphorylation-like carbonic acid group. Both human APOBEC-1 dephosphorylated by alkaline phosphase and the Ser(47,72)-to-alanine double mutant protein demonstrated a shifted isoelectric focusing pattern compared with the wild type, indicating phosphorylation at these sites. Taken together, these results suggest that phosphorylation might be an important mechanism in the regulation of apoB mRNA editing. PMID:11463337

Chen, Z; Eggerman, T L; Patterson, A P

2001-01-01

372

Assays of Adenylate Uridylate-Rich Element-Mediated mRNA Decay in Cells  

PubMed Central

The abundance of a cytoplasmic mRNA in eukaryotes often determines the level of the encoded protein product. The rates at which an mRNA is synthesized, exported, and degraded collectively contribute to its abundance in all cell types. Numerous mRNAs, particularly those encoding structural proteins, are very stable, with half-lives in the order of many hours. In contrast, mRNAs encoding regulatory proteins, including oncoproteins, cytokines, and signaling proteins, are relatively unstable with half-lives of an hour or less. As a result, modest changes in their decay rates affect their levels over a relatively short time period. This is particularly important to ensure rapid responses to extracellular signaling events. Messenger RNAs often harbor sequence elements that dictate their degradation rates. Adenylate uridylate (A+U)-rich elements (AREs), first identified in 1986, are perhaps the best characterized sequences that promote rapid mRNA degradation. These elements, localized within 3?-untranslated regions, sometimes contain AUUUA pentamers within an overall U-rich sequence, but this does not always define a bona fide ARE. Thus, experimental validation is essential before bestowing upon a suspected A+U-rich sequence the title of “ARE.” This chapter describes a reporter gene system that permits quantitative assessment of the effects of candidate A+U-rich sequences on mRNA half-life. This system employs tetracycline-controlled transcriptional silencing of the reporter gene, isolation of total-cell RNA at selected time points, quantitative reverse transcriptase polymerase chain reaction analysis of reporter mRNA levels, and nonlinear regression analysis of mRNA level as a function of time to quantitatively define parameters describing mRNA decay kinetics. Finally, this chapter describes more specialized assays to characterize ARE-mediated mRNA decay pathways, including deadenylation, and discusses decapping. PMID:19215753

Ysla, Riza M.; Wilson, Gerald M.; Brewer, Gary

2013-01-01

373

Estrogen induction and overexpression of fibulin-1C mRNA in ovarian cancer cells  

Microsoft Academic Search

Fibulin-1 is an extracellular matrix protein induced by estradiol in estrogen receptor (ER) positive ovarian cancer cell lines. Alternative splicing of fibulin-1 mRNA results in four different variants named A, B, C and D that may have distinct biological functions. We studied the relative expression of fibulin-1 mRNA variants and their estrogen regulation in human ovarian cancer cells. In ovarian

Frederic Moll; Dionyssios Katsaros; Gwendal Lazennec; Nicolas Hellio; Pascal Roger; Pierre-Ludovic Giacalone; Dany Chalbos; Thierry Maudelonde; Henri Rochefort; Pascal Pujol

2002-01-01

374

Mechanisms of RNAi: mRNA cleavage fragments may indicate stalled RISC  

Microsoft Academic Search

The molecular mechanism of RNA interference (RNAi) is under intense investigation. We previously demon- strated the existence of inactive siRNAs and also of mRNA cleavage in vivo in human cells. Here it is shown that some siRNAs with low activity leave mRNA cleavage fragments while an siRNA with higher activity does not. The pattern is consistent with both short-term (4-24

Torgeir Holen

2005-01-01

375

Molecular Characterization of Fetal Alcohol Syndrome Using mRNA Differential Display  

Microsoft Academic Search

The molecular pathogenesis of fetal alcohol syndrome (FAS) has not been well elucidated. The technique of mRNA differential display was used to characterize the etiology and to identify potential markers for FAS. Out of approximately 1,080 mRNA transcripts in mouse embryos that were analyzed, the levels of three mRNAs were altered by ethanol. Two of these mRNAs (one novel and

Insong James Lee; Yunjo Soh; Byoung Joon Song

1997-01-01

376

iBioSeminar: The Life of Eukaryotic mRNA  

NSDL National Science Digital Library

The control of mRNA production and function is a key aspect of the regulation of gene expression. Eukaryotic cells, the control of mRNA localization, translation and degradation in the cytoplasm allow for the proper regulation of the amount, duration, and location of protein production. The basic mechanisms of these processes are understood and reveal that the mechanisms of localization, translation, and degradation are interconnected.

Roy Parker (University of Arizona and Howard Hughes Medical Institute;)

2010-10-07

377

A stem-loop structure directs oskar mRNA to microtubule minus ends.  

PubMed

mRNA transport coupled with translational control underlies the intracellular localization of many proteins in eukaryotic cells. This is exemplified in Drosophila, where oskar mRNA transport and translation at the posterior pole of the oocyte direct posterior patterning of the embryo. oskar localization is a multistep process. Within the oocyte, a spliced oskar localization element (SOLE) targets oskar mRNA for plus end-directed transport by kinesin-1 to the posterior pole. However, the signals mediating the initial minus end-directed, dynein-dependent transport of the mRNA from nurse cells into the oocyte have remained unknown. Here, we show that a 67-nt stem-loop in the oskar 3' UTR promotes oskar mRNA delivery to the developing oocyte and that it shares functional features with the fs(1)K10 oocyte localization signal. Thus, two independent cis-acting signals, the oocyte entry signal (OES) and the SOLE, mediate sequential dynein- and kinesin-dependent phases of oskar mRNA transport during oogenesis. The OES also promotes apical localization of injected RNAs in blastoderm stage embryos, another dynein-mediated process. Similarly, when ectopically expressed in polarized cells of the follicular epithelium or salivary glands, reporter RNAs bearing the oskar OES are apically enriched, demonstrating that this element promotes mRNA localization independently of cell type. Our work sheds new light on how oskar mRNA is trafficked during oogenesis and the RNA features that mediate minus end-directed transport. PMID:24572808

Jambor, Helena; Mueller, Sandra; Bullock, Simon L; Ephrussi, Anne

2014-04-01

378

Regulation of cyclin A mRNA in leech embryonic stem cells  

Microsoft Academic Search

Leech embryos undergo invariant patterns of cleavage to yield identifiable cells that have characteristic timings of cell\\u000a division. To elucidate how these cell-specific differences in cell-cycle timing are regulated, we have isolated a leech cyclin A cDNA clone and determined the patterns of cyclin A mRNA localization in identified cells of Helobdella leech embryos. The intensity of cyclin A mRNA

Yongmei Chen; S. T. Bissen

1997-01-01

379

Isoeugenol destabilizes IL-8 mRNA expression in THP-1 cells through induction of the negative regulator of mRNA stability tristetraprolin.  

PubMed

We previously demonstrated in the human promyelocytic cell line THP-1 that all allergens tested, with the exception of the prohapten isoeugenol, induced a dose-related release of interleukin-8 (IL-8). In the present study, we investigated whether this abnormal behavior was regulated by the AU-rich element-binding proteins HuR and tristetraprolin (TTP) or by the downstream molecule suppressor of cytokine signaling (SOCS)-3. The contact allergens isoeugenol, diethylmaleate (DEM), and 2,4-dinitrochlorobenzene (DNCB), and the irritant salicylic acid were used as reference compounds. Chemicals were used at concentrations that induced a 20% decrease in cell viability as assessed by propidium iodide staining, namely 100 ?g/ml (0.61 mM) for isoeugenol, 100 ?g/ml (0.58 mM) for DEM, 3 ?g/ml (14.8 ?M) for DNCB, and 250 ?g/ml (1.81 mM) for salicylic acid. Time course experiments of IL-8 mRNA expression and assessment of IL-8 mRNA half-life, indicated a decreased IL-8 mRNA stability in isoeugenol-treated cells. We could demonstrate that a combination and regulation of HuR and TTP following exposure to contact allergens resulted in a different modulation of IL-8 mRNA half-life and release. The increased expression of TTP in THP-1 cells treated with isoeugenol results in destabilization of the IL-8 mRNA, which can account for the lack of IL-8 release. In contrast, the strong allergen DNCB failing to up-regulate TTP, while inducing HuR, resulted in longer IL-8 mRNA half-life and protein release. SOCS-3 was induced only in isoeugenol-treated cells; however, its modulation did not rescue the lack of IL-8 release, indicating that it is unlikely to be involved in the lack of IL-8 production. Finally, the destabilization effect of isoeugenol on IL-8 mRNA expression together with SOCS-3 expression resulted in an anti-inflammatory effect, as demonstrated by the ability of isoeugenol to modulate LPS or ionomycin-induced cytokine release. PMID:21969073

Galbiati, Valentina; Carne, Alice; Mitjans, Montserrat; Galli, Corrado Lodovico; Marinovich, Marina; Corsini, Emanuela

2012-02-01

380

CRISPR-mediated targeted mRNA degradation in the archaeon Sulfolobus solfataricus  

PubMed Central

The recently discovered clustered regularly interspaced short palindromic repeat (CRISPR)-mediated virus defense represents an adaptive immune system in many bacteria and archaea. Small CRISPR RNAs cause cleavage of complementary invading nucleic acids in conjunction with an associated protein or a protein complex. Here, we show CRISPR-mediated cleavage of mRNA from an invading virus in the hyperthermophilic archaeon Sulfolobus solfataricus. More than 40% of the targeted mRNA could be cleaved, as demonstrated by quantitative polymerase chain reaction. Cleavage of the mRNA was visualized by northern analyses and cleavage sites were mapped. In vitro, the same substrates were cleaved by the purified CRISPR-associated CMR complex from Sulfolobus solfataricus. The in vivo system was also re-programmed to knock down mRNA of a selected chromosomal gene (?-galactosidase) using an artificial miniCRISPR locus. With a single complementary spacer, ?50% reduction of the targeted mRNA and of corresponding intracellular protein activity was achieved. Our results demonstrate in vivo cleavage of mRNA in a prokaryote mediated by small RNAs (i.e. analogous to RNA interference in eukaryotes) and the re-programming of the system to silence specific genes of interest. PMID:24603867

Zebec, Ziga; Manica, Andrea; Zhang, Jing; White, Malcolm F.; Schleper, Christa

2014-01-01

381

Dynactin suppresses the retrograde movement of apically localized mRNA in Drosophila blastoderm embryos.  

PubMed

Motor dependent transport of mRNA is a key mechanism in axis specification during development. Apical transport and anchoring of wingless and pair-rule transcripts in the Drosophila syncytial blastoderm embryo is mediated by cytoplasmic Dynein, the major minus end directed microtubule dependent molecular motor. Here, we show that, despite apical transport of mRNA being highly directional, mRNA particles often pause and move backward toward the plus ends of microtubules. We suggest that this retrograde movement helps overcome cellular obstructions. We show that the plus end movement of apical mRNA is independent of the major plus end microtubule motors Kinesin-1 and Kinesin-2. In contrast, Dynactin, a Dynein processivity factor, is required to suppress retrograde mRNA movements, as well as for efficient minus end motility. We propose that Dynein itself, rather than the activity of a plus end motor, is responsible for the plus end movements of the mRNA and that Dynactin is involved in preventing short reverse movements of the Dynein motor, known to occur in vitro. PMID:17901156

Vendra, Georgia; Hamilton, Russell S; Davis, Ilan

2007-11-01

382

Expression of neuropsin mRNA in the mouse embryo and the pregnant uterus.  

PubMed

Neuropsin is a novel serine protease whose mRNA is expressed in the mouse central nervous system. We examined the expression of neuropsin mRNA during embryonic development using Northern and in situ hybridization in non-neural tissues. The pregnant uterus showed strong expression of neuropsin mRNA, whereas the nonpregnant uterus did not express this mRNA. Expression was first detected in the primary decidual zone at 5.5 days post coitum and was maximized at 10 days post coitum, decreasing remarkably thereafter. During mouse organogenesis, neuropsin expression was observed in the developing heart, lung, thymus, pituitary, choroid plexus, and epithelial linings of the skin, oral cavity, tongue, esophagus, and forestomach. In adult mouse organs, neuropsin mRNA was expressed in epithelial tissues covered by keratinocytes with moderate density, whereas low expression was observed in lung, thymus, and spleen. Neuropsin mRNA expression in developing organs and adult keratinocytes suggests that neuropsin is associated with extracellular matrix modifications and cell migrations. PMID:9487112

Chen, Z L; Momota, Y; Kato, K; Taniguchi, M; Inoue, N; Shiosaka, S; Yoshida, S

1998-03-01

383

Three-Dimensional Mapping of mRNA Export through the Nuclear Pore Complex  

PubMed Central

The locations of transcription and translation of mRNA in eukaryotic cells are spatially separated by the nuclear envelope (NE). Plenty of nuclear pore complexes (NPCs) embedded in the NE function as the major gateway for the export of transcribed mRNAs from the nucleus to the cytoplasm. Whereas the NPC, perhaps one of the largest protein complexes, provides a relatively large channel for macromolecules to selectively pass through it in inherently three-dimensional (3D) movements, this channel is nonetheless below the diffraction limit of conventional light microscopy. A full understanding of the mRNA export mechanism urgently requires real-time mapping of the 3D dynamics of mRNA in the NPC of live cells with innovative imaging techniques breaking the diffraction limit of conventional light microscopy. Recently, super-resolution fluorescence microscopy and single-particle tracking (SPT) techniques have been applied to the study of nuclear export of mRNA in live cells. In this review, we emphasize the necessity of 3D mapping techniques in the study of mRNA export, briefly summarize the feasibility of current 3D imaging approaches, and highlight the new features of mRNA nuclear export elucidated with a newly developed 3D imaging approach combining SPT-based super-resolution imaging and 2D-to-3D deconvolution algorithms. PMID:25393401

Schnell, Steven J.; Ma, Jiong; Yang, Weidong

2014-01-01

384

Dynactin suppresses the retrograde movement of apically localized mRNA in Drosophila blastoderm embryos  

PubMed Central

Motor dependent transport of mRNA is a key mechanism in axis specification during development. Apical transport and anchoring of wingless and pair-rule transcripts in the Drosophila syncytial blastoderm embryo is mediated by cytoplasmic Dynein, the major minus end directed microtubule dependent molecular motor. Here, we show that, despite apical transport of mRNA being highly directional, mRNA particles often pause and move backward toward the plus ends of microtubules. We suggest that this retrograde movement helps overcome cellular obstructions. We show that the plus end movement of apical mRNA is independent of the major plus end microtubule motors Kinesin-1 and Kinesin-2. In contrast, Dynactin, a Dynein processivity factor, is required to suppress retrograde mRNA movements, as well as for efficient minus end motility. We propose that Dynein itself, rather than the activity of a plus end motor, is responsible for the plus end movements of the mRNA and that Dynactin is involved in preventing short reverse movements of the Dynein motor, known to occur in vitro. PMID:17901156

Vendra, Georgia; Hamilton, Russell S.; Davis, Ilan

2007-01-01

385

Cytokine mRNA expression in intestine from normal and inflammatory bowel disease patients.  

PubMed

Cytokines are involved in the regulation of normal immune events and may be important in the development or perpetuation of immune events in inflammatory bowel disease. We have previously shown that normal human mononuclear cells from tonsil, spleen, and peripheral blood exhibit tissue and stimulus-specific patterns of cytokine mRNA expression. The aim of this study was to determine if disease-dependent differences of cytokine mRNA expression could be found in the intestine. Total RNA was isolated from intestinal mucosa and lamina propria mononuclear cells from inflammatory bowel disease patients and controls. cDNA probes specific for interleukins (IL)-1, -4, -5, and -6 and transforming growth factor-beta were used. IL-1 beta mRNA and TGF-beta mRNA steady state expressions were higher in inflammatory bowel disease specimens than in normal intestine. In addition, mononuclear cell specimens had stronger cytokine mRNA expression than mucosal specimens. The steady state mRNA expression of proinflammatory cytokines is higher in inflammatory bowel disease, consistent with the ongoing inflammation seen. PMID:8440073

McCabe, R P; Secrist, H; Botney, M; Egan, M; Peters, M G

1993-01-01

386

Increased muscle ubiquitin mRNA levels in gastric cancer patients.  

PubMed

The intramuscular ATP-dependent ubiquitin (Ub)-proteasome proteolytic system is hyperactivated in experimental cancer cachexia. The present study aimed at verifying whether the expression of the muscle Ub mRNA is altered in patients with cancer. Total muscle RNA was extracted using the guanidinium isothiocyanate/phenol/chloroform method from rectus abdominis biopsies obtained intraoperatively from 20 gastric cancer (GC) patients and 10 subjects with benign abdominal diseases (CON) undergoing surgery. Ub mRNA levels were measured by northern blot analysis. Serum soluble tumor necrosis factor receptor (sTNFR) was measured by ELISA. Ub mRNA levels (arbitrary units, means +/- SD) were 2,345 +/- 195 in GC and 1,162 +/- 132 in CON (P = 0.0005). Ub mRNA levels directly correlated with disease stage (r = 0.608, P = 0.005), being 1,945 +/- 786 in stages I and II, 2,480 +/- 650 in stage III, and 3,799 +/- 66 in stage IV. Ub mRNA and sTNFR did not correlate with age and nutritional parameters. This study confirms experimental data indicating an overexpression of muscle Ub mRNA in cancer cachexia. Lack of correlation with nutritional status suggests that Ub activation in human cancer is an early feature that precedes any clinical sign of cachexia. PMID:11294777

Bossola, M; Muscaritoli, M; Costelli, P; Bellantone, R; Pacelli, F; Busquets, S; Argilès, J; Lopez-Soriano, F J; Civello, I M; Baccino, F M; Rossi Fanelli, F; Doglietto, G B

2001-05-01

387

Viscum album-Mediated COX-2 Inhibition Implicates Destabilization of COX-2 mRNA  

PubMed Central

Extensive use of Viscum album (VA) preparations in the complementary therapy of cancer and in several other human pathologies has led to an increasing number of cellular and molecular approaches to explore the mechanisms of action of VA. We have recently demonstrated that, VA preparations exert a potent anti-inflammatory effect by selectively down-regulating the COX-2-mediated cytokine-induced secretion of prostaglandin E2 (PGE2), one of the important molecular signatures of inflammatory reactions. In this study, we observed a significant down-regulation of COX-2 protein expression in VA-treated A549 cells however COX-2 mRNA levels were unaltered. Therefore, we hypothesized that VA induces destabilisation of COX-2 mRNA, thereby depleting the available functional COX-2 mRNA for the protein synthesis and for the subsequent secretion of PGE2. To address this question, we analyzed the molecular degradation of COX-2 protein and its corresponding mRNA in A549 cell line. Using cyclohexamide pulse chase experiment, we demonstrate that, COX-2 protein degradation is not affected by the treatment with VA whereas experiments on transcriptional blockade with actinomycin D, revealed a marked reduction in the half life of COX-2 mRNA due to its rapid degradation in the cells treated with VA compared to that in IL-1?-stimulated cells. These results thus demonstrate that VA-mediated inhibition of PGE2 implicates destabilization of COX-2 mRNA. PMID:25664986

Saha, Chaitrali; Hegde, Pushpa; Friboulet, Alain; Bayry, Jagadeesh; Kaveri, Srinivas V.

2015-01-01

388

mRNA Transfection of Mouse and Human Neural Stem Cell Cultures  

PubMed Central

The use of synthetic mRNA as an alternative gene delivery vector to traditional DNA-based constructs provides an effective method for inducing transient gene expression in cell cultures without genetic modification. Delivery of mRNA has been proposed as a safer alternative to viral vectors in the induction of pluripotent cells for regenerative therapies. Although mRNA transfection of fibroblasts, dendritic and embryonic stem cells has been described, mRNA delivery to neurosphere cultures has not been previously reported. Here we sought to establish an efficient method for delivering mRNA to primary neurosphere cultures. Neurospheres derived from the subventricular zone of adult mice or from human embryonic stem cells were transfected with EGFP mRNA by lipofection and electroporation. Transfection efficiency and expression levels were monitored by flow cytometry. Cell survival following transfection was examined using live cell counting and the MTT assay. Both lipofection and electroporation provided high efficiency transfection of neurospheres. In comparison with lipofection, electroporation resulted in increased transfection efficiencies, but lower expression per cell and shorter durations of expression. Additional rounds of lipofection renewed EGFP expression in neurospheres, suggesting this method may be suitable for reprogramming applications. In summary, we have developed a protocol for achieving high efficiency transfection rates in mouse and human neurosphere cell culture that can be applied for future studies of gene function studies in neural stem cells, such as defining efficient differentiation protocols for glial and neuronal linages. PMID:24386231

McLenachan, Samuel; Zhang, Dan; Palomo, Ana Belén Alvarez; Edel, Michael J.; Chen, Fred K.

2013-01-01

389

Expression of RANKL and OPG mRNA in periodontal disease: possible involvement in bone destruction.  

PubMed

Periodontitis is a complex, multifactorial process affected by bacterial plaque-components and host defense mechanisms. Inflammation of the periodontitium may lead the destruction of the underlying ligament and alveolar bone. Receptor activator of NF-kappaB ligand (RANKL), a novel TNF receptor-related protein is an important factor for osteoclast differentiation and activation. Given osteolysis by osteoclast has been demonstrated in periodontitis, we hypothesized that RANKL expression may be associated with bone destruction in periodontitis. We used semi-quantitative RT-PCR to compare the gene expression of RANKL and osteoprogerin (OPG), a decoy receptor of RANKL, between moderate and advanced periodontitis, and healthy subjects. The level of RANKL mRNA was highest in advanced periodontitis. In contrast, the level of OPG mRNA in both advanced and moderate periodontitis was lower than that in the healthy group. It appears that the ratio of RANKL to OPG mRNA in periodontitis has increased. To determine the localization of RANKL gene transcripts in gingival tissue at the cellular level, in situ hybridization was performed using digoxigenin-labeled specific riboprobes. RANKL mRNA was expressed in inflammatory cells, mainly lymphocyte and macrophages. In addition, proliferating epithelium in the vicinity of inflammatory cells expressed high levels of RANKL mRNA. In short, our data suggest that up regulation of RANKL mRNA in both inflammatory cells and epithelium may be associated with the activation of osteoclastic bone destruction in periodontitis. PMID:12469211

Liu, D; Xu, J K; Figliomeni, L; Huang, L; Pavlos, N J; Rogers, M; Tan, A; Price, P; Zheng, M H

2003-01-01

390

Reduced FMR1 mRNA translation efficiency in fragile X patients with premutations.  

PubMed Central

The Fragile X mental retardation gene (FMR1) contains a polymorphic trinucleotide CGG repeat in the 5' untranslated region (UTR) of the FMR1 messenger. We have characterized three lymphoblastoid cell lines derived from unrelated male carriers of a premutation that overexpress FMR1 mRNA and show reduced FMRP level compared to normal cells. The analysis of polysomes/mRNPs distribution of mRNA in the cell lines with a premutation shows that the polysomal association of FMR1 mRNA, which is high in normal cells, becomes progressively lower with increasing CGG repeat expansion. In addition, we could detect a very low level of FMR1 mRNA in a lymphoblastoid cell line from a patient with a full mutation. In this case, FMR1 mRNA is not at all associated with polysomes, in agreement with the complete absence of FMRP. The impairment of FMR1 mRNA translation in patients with the Fragile X syndrome with FMR1 premutation is the cause of the lower FMRP levels that leads to the clinical involvement. PMID:12515381

Primerano, Beatrice; Tassone, Flora; Hagerman, Randi J; Hagerman, Paul; Amaldi, Francesco; Bagni, Claudia

2002-01-01

391

The iron-sensing aconitase B binds its own mRNA to prevent sRNA-induced mRNA cleavage.  

PubMed

Aconitase is an iron-sulfur protein and a major enzyme of the TCA cycle that catalyzes the conversion of citrate to isocitrate under iron-rich conditions. In Escherichia coli, aconitase B (AcnB) is a typical moonlighting protein that can switch to its apo form (apo-AcnB) which favors binding its own mRNA 3'UTR and stabilize it when intracellular iron become scarce. The small regulatory RNA (sRNA) RyhB has previously been shown to promote RNase E-dependent degradation of acnB mRNA when it was expressed from an ectopic arabinose-dependent promoter, independently of intracellular iron levels. In marked contrast, we report here that expression of RyhB under low-iron conditions did not result in acnB mRNA degradation even when RyhB was bound to acnB ribosome binding site (RBS). Genetic and biochemical evidence suggested that, under low-iron conditions, apo-AcnB bound to acnB 3'UTR close to a RNase E cleavage site that is essential for RyhB-induced acnB mRNA degradation. Whereas RyhB can block acnB translation initiation, RNase E-dependent degradation of acnB was prevented by apo-AcnB binding close to the cleavage site. This previously uncharacterized regulation suggests an intricate post-transcriptional mechanism that represses protein expression while insuring mRNA stability. PMID:25092924

Benjamin, Julie-Anna M; Massé, Eric

2014-01-01

392

The iron-sensing aconitase B binds its own mRNA to prevent sRNA-induced mRNA cleavage  

PubMed Central

Aconitase is an iron–sulfur protein and a major enzyme of the TCA cycle that catalyzes the conversion of citrate to isocitrate under iron-rich conditions. In Escherichia coli, aconitase B (AcnB) is a typical moonlighting protein that can switch to its apo form (apo-AcnB) which favors binding its own mRNA 3?UTR and stabilize it when intracellular iron become scarce. The small regulatory RNA (sRNA) RyhB has previously been shown to promote RNase E-dependent degradation of acnB mRNA when it was expressed from an ectopic arabinose-dependent promoter, independently of intracellular iron levels. In marked contrast, we report here that expression of RyhB under low-iron conditions did not result in acnB mRNA degradation even when RyhB was bound to acnB ribosome binding site (RBS). Genetic and biochemical evidence suggested that, under low-iron conditions, apo-AcnB bound to acnB 3?UTR close to a RNase E cleavage site that is essential for RyhB-induced acnB mRNA degradation. Whereas RyhB can block acnB translation initiation, RNase E-dependent degradation of acnB was prevented by apo-AcnB binding close to the cleavage site. This previously uncharacterized regulation suggests an intricate post-transcriptional mechanism that represses protein expression while insuring mRNA stability. PMID:25092924

Benjamin, Julie-Anna M.; Massé, Eric

2014-01-01

393

Oestradiol reduces Liver Receptor Homolog-1 mRNA transcript stability in breast cancer cell lines  

SciTech Connect

Highlights: •LRH-1 is an orphan nuclear receptor that regulates tumor proliferation. •In breast cancer, high mRNA expression is associated with ER+ status. •In ER?ve cells, despite very low mRNA, we found abundant LRH-1 protein. •Our data show distinctly different LRH-1 protein isoforms in ER? and ER+ breast cancer cells. •This is due to differences in LRH-1 mRNA and protein stability rates. -- Abstract: The expression of orphan nuclear receptor Liver Receptor Homolog-1 (LRH-1) is elevated in breast cancer and promotes proliferation, migration and invasion in vitro. LRH-1 expression is regulated by oestrogen (E{sub 2}), with LRH-1 mRNA transcript levels higher in oestrogen receptor ? (ER?) positive (ER+) breast cancer cells compared to ER? cells. However, the presence of LRH-1 protein in ER? cells suggests discordance between mRNA transcript levels and protein expression. To understand this, we investigated the impact of mRNA and protein stability in determining LRH-1 protein levels in breast cancer cells. LRH-1 transcript levels were significantly higher in ER+ versus ER? breast cancer cells lines; however LRH-1 protein was expressed at similar levels. We found LRH-1 mRNA and protein was more stable in ER? compared to ER+ cell lines. The tumor-specific LRH-1 variant isoform, LRH-1v4, which is highly responsive to E{sub 2}, showed increased mRNA stability in ER? versus ER+ cells. In addition, in MCF-7 and T47-D cell lines, LRH-1 total mRNA stability was reduced with E{sub 2} treatment, this effect mediated by ER?. Our data demonstrates that in ER? cells, increased mRNA and protein stability contribute to the abundant protein expression levels. Expression and immunolocalisation of LRH-1 in ER? cells as well as ER? tumors suggests a possible role in the development of ER? tumors. The modulation of LRH-1 bioactivity may therefore be beneficial as a treatment option in both ER? and ER+ breast cancer.

Lazarus, Kyren A. [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia) [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia); Environmental and Biotechnology Centre, Swinburne University, Hawthorn, Victoria 3122 (Australia); Zhao, Zhe; Knower, Kevin C. [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia)] [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia); To, Sarah Q. [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia) [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia); Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria 3168 (Australia); Chand, Ashwini L. [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia)] [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia); Clyne, Colin D., E-mail: Colin.clyne@princehenrys.org [Cancer Drug Discovery Laboratory, Prince Henry’s Institute of Medical Research, Clayton, Victoria 3168 (Australia); Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria 3168 (Australia)

2013-08-30

394

172 RELATIONSHIPS AMONG GRANULOSA CELL FGF9 mRNA, FOLLICLE SIZE, AND APOPTOSIS IN CATTLE.  

PubMed

Fibroblast growth factor 9 (FGF9) has been suggested to act as a dedifferentiation factor during bovine folliculogenesis, reducing steroidogenesis and increasing cell proliferation in granulosa (GC) and theca (TC) cells, but whether endogenous GC production of FGF9 change during bovine folliculogenesis and atresia/apoptosis is unknown. The objective of these studies was to investigate th