Subramaniyan, Sivakumar Allur; Kim, Sidong; Hwang, Inho
2016-10-01
The present study was carried out to understand the interaction between fibroblast and 3T3-L1 preadipocyte cells under H 2 O 2 -induced oxidative stress condition. H 2 O 2 (40 μM) was added in co-culture and monoculture of fibroblast and 3T3-L1 cell. The cells in the lower well were harvested for analysis and the process was carried out for both cells. The cell growth, oxidative stress markers, and antioxidant enzymes were analyzed. Additionally, the mRNA expressions of caspase-3 and caspase-7 were selected for analysis of apoptotic pathways and TNF-α and NF-κB were analyzed for inflammatory pathways. The adipogenic marker such as adiponectin and PPAR-γ and collagen synthesis markers such as LOX and BMP-1 were analyzed in the co-culture of fibroblast and 3T3-L1 cells. Cell viability and antioxidant enzymes were significantly increased in the co-culture compared to the monoculture under stress condition. The apoptotic, inflammatory, adipogenic, and collagen-synthesized markers were significantly altered in H 2 O 2 -induced co-culture of fibroblast and 3T3-L1 cells when compared with the monoculture of H 2 O 2 -induced fibroblast and 3T3-L1 cells. In addition, the confocal microscopical investigation indicated that the co-culture of H 2 O 2 -induced 3T3-L1 and fibroblast cells increases collagen type I and type III expression. From our results, we suggested that co-culture of fat cell (3T3-L1) and fibroblast cells may influence/regulate each other and made the cells able to withstand against oxidative stress and aging. It is conceivable that the same mechanism might have been occurring from cell to cell while animals are stressed by various environmental conditions.
FoxO1 regulates apoptosis induced by asbestos in the MT-2 human T-cell line.
Matsuzaki, Hidenori; Lee, Suni; Maeda, Megumi; Kumagai-Takei, Naoko; Nishimura, Yasumitsu; Otsuki, Takemi
2016-09-01
Asbestos is known to cause malignant mesothelioma and lung cancer. Recent studies implicate tumor immunity in the development of various tumors, including malignant mesothelioma. In order to establish an in vitro T-cell model to clarify the effects of long-term exposure of asbestos on tumor immunity, in this study, human T-cell line MT-2 cells were cultured with asbestos for longer than 8 months and the resultant cells (MT-2Rst) were assessed for the expression of forkhead transcription factor FoxO1. Gene expression analysis revealed that the amount of FoxO1 mRNA decreased after long-term exposure of the MT-2 cells to asbestos. In accordance with this reduction in FoxO1, pro-apoptotic Foxo1 target genes Puma, Fas ligand and Bim were also seen to be down-regulated in MT-2Rst cells. Furthermore, shRNA-mediated knock-down of FoxO1 reduced the number of apoptotic parental MT-2 cells after treatment with asbestos. On the other hand, over-expression of FoxO1 did not affect asbestos-induced apoptosis in MT-2Rst cells. These results suggested that FoxO1 played an important role in regulating asbestos-induced apoptosis and confirmed the presence of multiple pathways regulating resistance to asbestos in MT-2Rst cells.
Kim, Sang-Cheol; Kang, Jung-Il; Hyun, Jin-Won; Kang, Ji-Hoon; Koh, Young-Sang; Kim, Young-Heui; Kim, Ki-Ho; Ko, Ji-Hee; Yoo, Eun-Sook; Kang, Hee-Kyoung
2017-01-01
4-O-methylhonokiol, a neolignan compound from Magnolia Officinalis, has been reported to have various biological activities including hair growth promoting effect. However, although transforming growth factor-β (TGF-β) signal pathway has an essential role in the regression induction of hair growth, the effect of 4-O-methylhonokiol on the TGF-β signal pathway has not yet been elucidated. We thus examined the effect of 4-O-methylhonokiol on TGF-β-induced canonical and noncanonical pathways in HaCaT human keratinocytes. When HaCaT cells were pretreated with 4-O-methylhonokiol, TGF-β1-induced G1/G0 phase arrest and TGF-β1-induced p21 expression were decreased. Moreover, 4-O-methylhonokiol inhibited nuclear translocation of Smad2/3, Smad4 and Sp1 in TGF-β1-induced canonical pathway. We observed that ERK phosphorylation by TGF-β1 was significantly attenuated by treatment with 4-O-methylhonokiol. 4-O-methylhonokiol inhibited TGF-β1-induced reactive oxygen species (ROS) production and reduced the increase of NADPH oxidase 4 (NOX4) mRNA level in TGF-β1-induced noncanonical pathway. These results indicate that 4-O-methylhonokiol could inhibit TGF-β1-induced cell cycle arrest through inhibition of canonical and noncanonical pathways in human keratinocyte HaCaT cell and that 4-O-methylhonokiol might have protective action on TGF-β1-induced cell cycle arrest. PMID:28190316
Kim, Sang-Cheol; Kang, Jung-Il; Hyun, Jin-Won; Kang, Ji-Hoon; Koh, Young-Sang; Kim, Young-Heui; Kim, Ki-Ho; Ko, Ji-Hee; Yoo, Eun-Sook; Kang, Hee-Kyoung
2017-07-01
4- O -methylhonokiol, a neolignan compound from Magnolia Officinalis , has been reported to have various biological activities including hair growth promoting effect. However, although transforming growth factor-β (TGF-β) signal pathway has an essential role in the regression induction of hair growth, the effect of 4- O -methylhonokiol on the TGF-β signal pathway has not yet been elucidated. We thus examined the effect of 4- O -methylhonokiol on TGF-β-induced canonical and noncanonical pathways in HaCaT human keratinocytes. When HaCaT cells were pretreated with 4- O -methylhonokiol, TGF-β1-induced G1/G0 phase arrest and TGF-β1-induced p21 expression were decreased. Moreover, 4- O -methylhonokiol inhibited nuclear translocation of Smad2/3, Smad4 and Sp1 in TGF-β1-induced canonical pathway. We observed that ERK phosphorylation by TGF-β1 was significantly attenuated by treatment with 4- O -methylhonokiol. 4- O -methylhonokiol inhibited TGF-β1-induced reactive oxygen species (ROS) production and reduced the increase of NADPH oxidase 4 (NOX4) mRNA level in TGF-β1-induced noncanonical pathway. These results indicate that 4- O -methylhonokiol could inhibit TGF-β1-induced cell cycle arrest through inhibition of canonical and noncanonical pathways in human keratinocyte HaCaT cell and that 4- O -methylhonokiol might have protective action on TGF-β1-induced cell cycle arrest.
Possible role of PAPR-1 in protecting human HaCaT cells against cytotoxicity of SiO2 nanoparticles.
Gong, Chunmei; Yang, Linqing; Zhou, Jichang; Guo, Xiang; Zhuang, Zhixiong
2017-10-05
Nano-SiO 2 materials play a significant role in the engineered nanomaterials (ENMs) field. The ease of their production as well as their relatively low cost has promoted the wide use of these products in many fields. Nano-SiO 2 exposure is known to cause severe DNA damage; however, the underlying mechanisms remain poorly understood. In a previous study, we found that nano-SiO 2 exposure regulate the expression of the poly(ADP-ribose) polymerases-1 (PARP-1), a pivotal DNA repair gene, in human HaCaT cells. Here, we employed lentivirus-mediated RNA interference (RNAi) to knock down PAPR-1 expression in HaCaT cells and explored the potential role of PARP-1 in nano-SiO 2 induced cytotoxicity. We found that nano-SiO 2 treatment of HaCaT cells causes decreased cell viability, increased apoptosis and DNA damage. Nano-SiO 2 -treated HaCaT cells were also found to have slightly changed cell cycle distribution. Lentivirus-mediated PAPR-1 knockdown partially aggravated cytotoxicity and increased apoptosis induced by nano-SiO 2 treatment. Nano-SiO 2 had significant toxicity to human HaCaT cells and causes DNA damage. PAPR-1 knock-down cell line appears more sensitive to nano-SiO 2 than the control cells in DNA damage. The results suggest that PAPR-1 is involved in protecting cells from damage caused by nano-SiO 2 . Copyright © 2017 Elsevier B.V. All rights reserved.
Aging Converts Innate B1a Cells into Potent CD8+ T Cell Inducers
Lee-Chang, Catalina; Bodogai, Monica; Moritoh, Kanako; Chen, Xin; Wersto, Robert; Sen, Ranjan; Young, Howard A.; Croft, Michael; Ferrucci, Luigi; Biragyn, Arya
2016-01-01
B-cell dysregulation in aging is thought to mostly occur in conventional B2 cells without affecting innate B1 cells. Elderly humans and mice also accumulate 4-1BBL+ MHC class-IHi CD86Hi B cells of unknown origin. Here we report that these cells, termed 4BL cells, are activated murine and possibly human B1a cells. The activation is mediated by aging human monocytes and murine peritoneal macrophages. The 4BL cells induce expression of 4-1BBL and IFNγR1 on B1a cells resulting in subsequent up regulation of membrane TNFα (mTNFα) and CD86. As a result, B1a cells induce expression of granzyme B in CD8+T cells by targeting TNFR2 via mTNFα while providing co-stimulation with CD86. Thus, for the first time, these results indicate that aging affects the function of B1a cells. Upon aging, these cells lose their tumor-supporting activity and become inducers of potentially antitumor and autoimmune CD8+T cells. PMID:26983789
Macaire, Héloïse; Riquet, Aurélien; Moncollin, Vincent; Biémont-Trescol, Marie-Claude; Duc Dodon, Madeleine; Hermine, Olivier; Debaud, Anne-Laure; Mahieux, Renaud; Mesnard, Jean-Michel; Pierre, Marlène; Gazzolo, Louis; Bonnefoy, Nathalie; Valentin, Hélène
2012-01-01
Human T lymphotropic virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia/lymphoma (ATLL). ATLL is a severe malignancy with no effective treatment. HTLV-1 regulatory proteins Tax and HTLV-1 basic leucine zipper factor (HBZ) play a major role in ATLL development, by interfering with cellular functions such as CD4+ T-cell survival. In this study, we observed that the expression of Bfl-1, an antiapoptotic protein of the Bcl-2 family, is restricted to HTLV-1-infected T-cell lines and to T-cells expressing both Tax and HBZ proteins. We showed that Tax-induced bfl-1 transcription through the canonical NF-κB pathway. Moreover, we demonstrated that Tax cooperated with c-Jun or JunD, but not JunB, transcription factors of the AP-1 family to stimulate bfl-1 gene activation. By contrast, HBZ inhibited c-Jun-induced bfl-1 gene activation, whereas it increased JunD-induced bfl-1 gene activation. We identified one NF-κB, targeted by RelA, c-Rel, RelB, p105/p50, and p100/p52, and two AP-1, targeted by both c-Jun and JunD, binding sites in the bfl-1 promoter of T-cells expressing both Tax and HBZ. Analyzing the potential role of antiapoptotic Bcl-2 proteins in HTLV-1-infected T-cell survival, we demonstrated that these cells are differentially sensitive to silencing of Bfl-1, Bcl-xL, and Bcl-2. Indeed, both Bfl-1 and Bcl-xL knockdowns decreased the survival of HTLV-1-infected T-cell lines, although no cell death was observed after Bcl-2 knockdown. Furthermore, we demonstrated that Bfl-1 knockdown sensitizes HTLV-1-infected T-cells to ABT-737 or etoposide treatment. Our results directly implicate Bfl-1 and Bcl-xL in HTLV-1-infected T-cell survival and suggest that both Bfl-1 and Bcl-xL represent potential therapeutic targets for ATLL treatment. PMID:22553204
HTLV-1 induces a Th1-like state in CD4+CCR4+ T cells
Araya, Natsumi; Sato, Tomoo; Ando, Hitoshi; Tomaru, Utano; Yoshida, Mari; Coler-Reilly, Ariella; Yagishita, Naoko; Yamauchi, Junji; Hasegawa, Atsuhiko; Kannagi, Mari; Hasegawa, Yasuhiro; Takahashi, Katsunori; Kunitomo, Yasuo; Tanaka, Yuetsu; Nakajima, Toshihiro; Nishioka, Kusuki; Utsunomiya, Atae; Jacobson, Steven; Yamano, Yoshihisa
2014-01-01
Human T-lymphotropic virus type 1 (HTLV-1) is linked to multiple diseases, including the neuroinflammatory disease HTLV-1–associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T cell leukemia/lymphoma. Evidence suggests that HTLV-1, via the viral protein Tax, exploits CD4+ T cell plasticity and induces transcriptional changes in infected T cells that cause suppressive CD4+CD25+CCR4+ Tregs to lose expression of the transcription factor FOXP3 and produce IFN-γ, thus promoting inflammation. We hypothesized that transformation of HTLV-1–infected CCR4+ T cells into Th1-like cells plays a key role in the pathogenesis of HAM/TSP. Here, using patient cells and cell lines, we demonstrated that Tax, in cooperation with specificity protein 1 (Sp1), boosts expression of the Th1 master regulator T box transcription factor (T-bet) and consequently promotes production of IFN-γ. Evaluation of CSF and spinal cord lesions of HAM/TSP patients revealed the presence of abundant CD4+CCR4+ T cells that coexpressed the Th1 marker CXCR3 and produced T-bet and IFN-γ. Finally, treatment of isolated PBMCs and CNS cells from HAM/TSP patients with an antibody that targets CCR4+ T cells and induces cytotoxicity in these cells reduced both viral load and IFN-γ production, which suggests that targeting CCR4+ T cells may be a viable treatment option for HAM/TSP. PMID:24960164
Cannabidiol (CBD) Induces Functional Tregs in Response to Low-Level T Cell Activation
Dhital, Saphala; Stokes, John V.; Park, Nogi; Seo, Keun-Seok; Kaplan, Barbara L.F.
2016-01-01
Many effects of the non-psychoactive cannabinoid, cannabidiol (CBD), have been described in immune responses induced by strong immunological stimuli. It has also been shown that CBD enhances IL-2 production in response to low-level T cell stimulation. Since IL-2, in combination with TGF-β1, are critical for Treg induction, we hypothesized that CBD would induce CD4+CD25+FOXP3+ Tregs in response to low-level stimulation. Low-level T cell stimulation conditions were established based on minimal CD25 expression in CD4+ cells using suboptimal PMA/Io (4 nM/0.05 μM, S/o), ultrasuboptimal PMA/Io (1 nM/0.0125 μM, Us/o) or soluble anti-CD3/28 (400-800 ng each, s3/28). CBD increased CD25+FOXP3+ cells from CD4+, CD4+CD25+, and CD4+CD25− T cells, as well as in CD4+ T cells derived from FOXP3-GFP mice. Most importantly, the Us/o + CBD-induced CD4+CD25+ Tregs robustly suppressed responder T cell proliferation, demonstrating that the mechanism by which CBD is immunosuppressive under low-level T cell stimulation involves induction of functional Tregs. PMID:27865421
Cannabidiol (CBD) induces functional Tregs in response to low-level T cell activation.
Dhital, Saphala; Stokes, John V; Park, Nogi; Seo, Keun Seok; Kaplan, Barbara L F
2017-02-01
Many effects of the non-psychoactive cannabinoid, cannabidiol (CBD), have been described in immune responses induced by strong immunological stimuli. It has also been shown that CBD enhances IL-2 production in response to low-level T cell stimulation. Since IL-2, in combination with TGF-β1, are critical for Treg induction, we hypothesized that CBD would induce CD4 + CD25 + FOXP3 + Tregs in response to low-level stimulation. Low-level T cell stimulation conditions were established based on minimal CD25 expression in CD4 + cells using suboptimal PMA/Io (4nM/0.05μM, S/o), ultrasuboptimal PMA/Io (1nM/0.0125μM, Us/o) or soluble anti-CD3/28 (400-800ng each, s3/28). CBD increased CD25 + FOXP3 + cells from CD4 + , CD4 + CD25 + , and CD4 + CD25 - T cells, as well as in CD4 + T cells derived from FOXP3-GFP mice. Most importantly, the Us/o+CBD-induced CD4 + CD25 + Tregs robustly suppressed responder T cell proliferation, demonstrating that the mechanism by which CBD is immunosuppressive under low-level T cell stimulation involves induction of functional Tregs. Copyright © 2016 Elsevier Inc. All rights reserved.
Aging Converts Innate B1a Cells into Potent CD8+ T Cell Inducers.
Lee-Chang, Catalina; Bodogai, Monica; Moritoh, Kanako; Chen, Xin; Wersto, Robert; Sen, Ranjan; Young, Howard A; Croft, Michael; Ferrucci, Luigi; Biragyn, Arya
2016-04-15
B cell dysregulation in aging is thought to mostly occur in conventional B2 cells without affecting innate B1 cells. Elderly humans and mice also accumulate 4-1BBL(+)MHC class-I(Hi)CD86(Hi)B cells of unknown origin. In this article, we report that these cells, termed 4BL cells, are activated murine and possibly human B1a cells. The activation is mediated by aging human monocytes and murine peritoneal macrophages. They induce expression and activation of 4-1BBL and IFN-γR1 on B1a cells to subsequently upregulate membrane TNF-α and CD86. As a result, activated B1a/4BL cells induce expression of granzyme B in CD8(+)T cells by targeting TNFR2 via membrane TNF-α and providing costimulation with CD86. Thus, for the first time, to our knowledge, these results indicate that aging affects the function of B1a cells. Upon aging, these cells lose their tumor-supporting activity and become inducers of potentially antitumor and autoimmune CD8(+)T cells. Copyright © 2016 by The American Association of Immunologists, Inc.
Chen, Wen-Fang; Xu, Li; Yu, Chung-Him; Ho, Chui-Kwan; Wu, Ka; Leung, Gina CW; Wong, Man-Sau
2012-01-01
In the present study, we investigated the therapeutic effect of a classical TCM formula, Free Wanderer Powder (逍遙散 xiāo yáo sǎn), in a breast cancer mouse model induced with estrogen-insensitive breast cancer 4T1 cells. Ovariectomized Balb/c mice (6-8 weeks) or sham mice were injected into the fourth mammary fat pad with 4T1 cells in which tumors were palpable 7 days after injection. On the eighth day, the mice were divided into 4 groups and tubefed daily with vehicle, Free Wanderer Powder (逍遙散 xiāo yáo sǎn) formula or tamoxifen for 28 days. Tumor growth inhibition and the decrease of the average tumor mass were most evident in mice treated with Free Wanderer Powder (逍遙散 xiāo yáo sǎn). Free Wanderer Powder (逍遙散 xiāo yáo sǎn) treatment significantly reduced Bcl-2 and elevated Bax and p53 protein expressions in breast cancer tumor. These results were further confirmed by immunohistochemisty. Tamoxifen could decrease spleen mass and Bcl-2 protein expression, increase the Bax protein expression as well as exert uterotrophic effects by increasing uterus index and inducing the gene expressions in the uterus. Taken together, these results show that Free Wanderer Powder (逍遙散 xiāo yáo sǎn) treatment induced apoptosis at protein level and inhibited the tumor growth in 4T1-induced ovariectomized Balb/c female mice, indicating the possibility of its future use for treatment of estrogen-insensitive breast caner. PMID:24716117
David, Rachel; Ma, Liang; Ivetic, Aleksandar; Takesono, Aya; Ridley, Anne J.; Chai, Jian-Guo; Tybulewicz, Victor; Marelli-Berg, Federica M.
2016-01-01
Localization of primed T cells to antigenic tissue is essential for the development of effective immunity. Together with tissue-selective homing molecules, T-cell receptor (TCR)- and CD28-mediated signals have been shown to promote transendothelial migration of specific T cells into non-lymphoid antigen-rich tissue tissue. However, the cellular and molecular requirements for T-cell accumulation to target tissue following their recruitment are largely undefined. The guanine nucleotide exchange factor (GEF) Vav1 has an integral role in coupling TCR and CD28 to signalling pathways that regulate T cell activation and migration. Here, we have investigated the contribution of TCR- and CD28-induced Vav1 activity to the trafficking and localization of primed HY-specific CD4+ T cells to antigenic sites. Severe migratory defects displayed by Vav1-/- T cells in vitro were fully compensated by a combination of shear flow and chemokines, leading to normal recruitment of Vav1-/- T cells in vivo. In contrast, Vav1-/- T-cell retention into antigen-rich tissue was severely impaired, reflecting their inability to engage in sustained TCR- and CD28-mediated interactions with tissue-resident antigen-presenting cells (APCs). This novel function of APC-induced, TCR- and CD28-mediated Vav1 activity in the regulation of effector T-cell immunity highlights its potential as a therapeutic target in T-cell-mediated tissue damage. PMID:19060239
Acute Malaria Induces PD1+CTLA4+ Effector T Cells with Cell-Extrinsic Suppressor Function
Mackroth, Maria Sophia; Abel, Annemieke; Steeg, Christiane; Schulze zur Wiesch, Julian; Jacobs, Thomas
2016-01-01
In acute Plasmodium falciparum (P. falciparum) malaria, the pro- and anti-inflammatory immune pathways must be delicately balanced so that the parasitemia is controlled without inducing immunopathology. An important mechanism to fine-tune T cell responses in the periphery is the induction of coinhibitory receptors such as CTLA4 and PD1. However, their role in acute infections such as P. falciparum malaria remains poorly understood. To test whether coinhibitory receptors modulate CD4+ T cell functions in malaria, blood samples were obtained from patients with acute P. falciparum malaria treated in Germany. Flow cytometric analysis showed a more frequent expression of CTLA4 and PD1 on CD4+ T cells of malaria patients than of healthy control subjects. In vitro stimulation with P. falciparum-infected red blood cells revealed a distinct population of PD1+CTLA4+CD4+ T cells that simultaneously produced IFNγ and IL10. This antigen-specific cytokine production was enhanced by blocking PD1/PDL1 and CTLA4. PD1+CTLA4+CD4+ T cells were further isolated based on surface expression of PD1 and their inhibitory function investigated in-vitro. Isolated PD1+CTLA4+CD4+ T cells suppressed the proliferation of the total CD4+ population in response to anti-CD3/28 and plasmodial antigens in a cell-extrinsic manner. The response to other specific antigens was not suppressed. Thus, acute P. falciparum malaria induces P. falciparum-specific PD1+CTLA4+CD4+ Teffector cells that coproduce IFNγ and IL10, and inhibit other CD4+ T cells. Transient induction of regulatory Teffector cells may be an important mechanism that controls T cell responses and might prevent severe inflammation in patients with malaria and potentially other acute infections. PMID:27802341
Vaeth, Martin; Gogishvili, Tea; Bopp, Tobias; Klein, Matthias; Berberich-Siebelt, Friederike; Gattenloehner, Stefan; Avots, Andris; Sparwasser, Tim; Grebe, Nadine; Schmitt, Edgar; Hünig, Thomas; Serfling, Edgar; Bodor, Josef
2011-01-01
Inducible cAMP early repressor (ICER) is a transcriptional repressor, which, because of alternate promoter use, is generated from the 3′ region of the cAMP response modulator (Crem) gene. Its expression and nuclear occurrence are elevated by high cAMP levels in naturally occurring regulatory T cells (nTregs). Using two mouse models, we demonstrate that nTregs control the cellular localization of ICER/CREM, and thereby inhibit IL-2 synthesis in conventional CD4+ T cells. Ablation of nTregs in depletion of regulatory T-cell (DEREG) mice resulted in cytosolic localization of ICER/CREM and increased IL-2 synthesis upon stimulation. Direct contacts between nTregs and conventional CD4+ T cells led to nuclear accumulation of ICER/CREM and suppression of IL-2 synthesis on administration of CD28 superagonistic (CD28SA) Ab. In a similar way, nTregs communicated with B cells and induced the cAMP-driven nuclear localization of ICER/CREM. High levels of ICER suppressed the induction of nuclear factor of activated T cell c1 (Nfatc1) gene in T cells whose inducible Nfatc1 P1 promoter bears two highly conserved cAMP-responsive elements to which ICER/CREM can bind. These findings suggest that nTregs suppress T-cell responses by the cAMP-dependent nuclear accumulation of ICER/CREM and inhibition of NFATc1 and IL-2 induction. PMID:21262800
Vassbotn, F S; Skar, R; Holmsen, H; Lillehaug, J R
1992-09-01
The effect of platelet-derived growth factor (PDGF) on c-fos mRNA transcription was studied in the immortalized mouse embryo fibroblast C3H/10T1/2 Cl 8 (10T1/2) cells and the chemically transformed, tumorigenic subclone C3H/10T1/2 Cl 16 (Cl 16). In the 10T1/2 cells as well as the Cl 16 subclone, the dose-dependent PDGF stimulation of c-fos mRNA synthesis was similar in both logarithmically growing and confluent cultures. c-fos mRNA was induced severalfold by 12-O-tetradecanoylphorbol-13-acetate (TPA) in both 10T1/2 and Cl 16. Down-regulation of protein kinase C (PKC) activity by TPA pretreatment inhibited PDGF-stimulated c-fos mRNA expression in Cl 16 cells but did not affect this induction in the 10T1/2 cells. This inhibition was not a general phenomenon of 3-methylcholanthrene-mediated transformation of 10T1/2 cells since experiments with another transformed 10T1/2 cell clone, C3H/10T1/2 TPA 482, gave qualitatively the same results as the 10T1/2 cells. Receptor binding experiments showed that the nontransformed and transformed cells had a comparable number of PDGF receptors, 1.3 x 10(5) and 0.7 x 10(5) receptors per cell, respectively. Furthermore, cAMP-induced c-fos expression induced by forskolin is formerly shown to be independent of PKC down-regulation. In our experiments, forskolin induced c-fos expression in both clones. However, PKC down-regulation inhibited the forskolin-induced c-fos expression in Cl 16 cells. This apparently demonstrates cross talk between PKC and PKA in the c-fos induction pathway. The present results provide evidence for an impaired mechanism for activating c-fos expression through PKC-independent, PDGF-induced signal transduction in the chemically transformed Cl 16 fibroblasts compared to that in nontransformed 10T1/2 cells.
Selleri, Silvia; Dieng, Mame Massar; Nicoletti, Simon; Louis, Isabelle; Beausejour, Christian; Le Deist, Françoise
2013-01-01
The mechanisms by which mesenchymal stromal cells (MSCs) induce immunomodulation are still poorly understood. In the current work, we show by a combination of polymerase chain reaction (PCR) array, flow cytometry, and multiplex cytokine data analysis that during the inhibition of an alloantigen-driven CD4+ T-cell response, MSCs induce a fraction of CD4+ T-cells to coexpress interferon-γ (IFNγ) and interleukin-10 (IL-10). This CD4+ IFNγ+ IL-10+ cell population shares properties with recently described T-cells originating from switched Th1 cells that start producing IL-10 and acquire a regulatory function. Here we report that IL-10-producing Th1 cells accumulated with time during T-cell stimulation in the presence of MSCs. Moreover, MSCs caused stimulated T-cells to downregulate the IFNγ receptor (IFNγR) without affecting IL-10 receptor expression. Further, the inhibitory effect of MSCs could be reversed by an anti-IFNγR-blocking antibody, indicating that IFNγ is one of the major players in MSC-induced T-cell suppression. Stimulated (and, to a lesser extent, resting) CD4+ T-cells treated with MSCs were able to inhibit the proliferation of autologous CD4+ T-cells, demonstrating their acquired regulatory properties. Altogether, our results suggest that the generation of IL-10-producing Th1 cells is one of the mechanisms by which MSCs can downmodulate an immune response. PMID:23167734
Stephen-Victor, Emmanuel; Karnam, Anupama; Fontaine, Thierry; Beauvais, Anne; Das, Mrinmoy; Hegde, Pushpa; Prakhar, Praveen; Holla, Sahana; Balaji, Kithiganahalli N; Kaveri, Srini V; Latgé, Jean-Paul; Aimanianda, Vishukumar; Bayry, Jagadeesh
2017-12-05
Human dendritic cell (DC) response to α-(1,3)-glucan polysaccharide of Aspergillus fumigatus and ensuing CD4+ T-cell polarization are poorly characterized. α-(1,3)-Glucan was isolated from A. fumigatus conidia and mycelia cell wall. For the analysis of polarization, DCs and autologous naive CD4+ T cells were cocultured. Phenotype of immune cells was analyzed by flow cytometry, and cytokines by enzyme-linked immunosorbent assay (ELISA). Blocking antibodies were used to dissect the role of Toll-like receptor 2 (TLR2) and programmed death-ligand 1 (PD-L1) in regulating α-(1,3)-glucan-mediated DC activation and T-cell responses. DCs from TLR2-deficient mice were additionally used to consolidate the findings. α-(1,3)-Glucan induced the maturation of DCs and was dependent in part on TLR2. "α-(1,3)-Glucan-educated" DCs stimulated the activation of naive T cells and polarized a subset of these cells into CD4+CD25+FoxP3+ regulatory T cells (Tregs). Mechanistically, Treg stimulation by α-(1,3)-glucan was dependent on the PD-L1 pathway that negatively regulated interferon-gamma (IFN-γ) secretion. Short α-(1,3)-oligosaccharides lacked the capacity to induce maturation of DCs but significantly blocked α-(1,3)-glucan-induced Treg polarization. PD-L1 dictates the balance between Treg and IFN-γ responses induced by α-(1,3)-glucan. Our data provide a rationale for the exploitation of immunotherapeutic approaches that target PD-1-PD-L1 to enhance protective immune responses to A. fumigatus infections. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.
HTLV-1-induced leukotriene B4 secretion by T cells promotes T cell recruitment and virus propagation
Percher, Florent; Curis, Céline; Pérès, Eléonore; Artesi, Maria; Rosewick, Nicolas; Jeannin, Patricia; Gessain, Antoine; Gout, Olivier; Mahieux, Renaud; Ceccaldi, Pierre-Emmanuel; Van den Broeke, Anne; Duc Dodon, Madeleine; Afonso, Philippe V.
2017-01-01
The human T-lymphotropic virus type 1 (HTLV-1) is efficiently transmitted through cellular contacts. While the molecular mechanisms of viral cell-to-cell propagation have been extensively studied in vitro, those facilitating the encounter between infected and target cells remain unknown. In this study, we demonstrate that HTLV-1-infected CD4 T cells secrete a potent chemoattractant, leukotriene B4 (LTB4). LTB4 secretion is dependent on Tax-induced transactivation of the pla2g4c gene, which encodes the cytosolic phospholipase A2 gamma. Inhibition of LTB4 secretion or LTB4 receptor knockdown on target cells reduces T-cell recruitment, cellular contact formation and virus propagation in vitro. Finally, blocking the synthesis of LTB4 in a humanized mouse model of HTLV-1 infection significantly reduces proviral load. This results from a decrease in the number of infected clones while their expansion is not impaired. This study shows the critical role of LTB4 secretion in HTLV-1 transmission both in vitro and in vivo. PMID:28639618
Suppression of T cell-induced osteoclast formation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Karieb, Sahar; Fox, Simon W., E-mail: Simon.fox@plymouth.ac.uk
2013-07-12
Highlights: •Genistein and coumestrol prevent activated T cell induced osteoclast formation. •Anti-TNF neutralising antibodies prevent the pro-osteoclastic effect of activated T cells. •Phytoestrogens inhibit T cell derived TNF alpha and inflammatory cytokine production. •Phytoestrogens have a broader range of anti-osteoclastic actions than other anti-resorptives. -- Abstract: Inhibition of T cell derived cytokine production could help suppress osteoclast differentiation in inflammatory skeletal disorders. Bisphosphonates are typically prescribed to prevent inflammatory bone loss but are not tolerated by all patients and are associated with an increased risk of osteonecrosis of the jaw. In light of this other anti-resorptives such as phytoestrogens aremore » being considered. However the effect of phytoestrogens on T cell-induced osteoclast formation is unclear. The effect of genistein and coumestrol on activated T cell-induced osteoclastogenesis and cytokine production was therefore examined. Concentrations of genistein and coumestrol (10{sup −7} M) previously shown to directly inhibit osteoclast formation also suppressed the formation of TRAP positive osteoclast induced by con A activated T cells, which was dependent on inhibition of T cell derived TNF-α. While both reduced osteoclast formation their mechanism of action differed. The anti-osteoclastic effect of coumestrol was associated with a dual effect on con A induced T cell proliferation and activation; 10{sup −7} M coumestrol significantly reducing T cell number (0.36) and TNF-α (0.47), IL-1β (0.23) and IL-6 (0.35) expression, whereas genistein (10{sup −7} M) had no effect on T cell number but a more pronounced effect on T cell differentiation reducing expression of TNF-α (0.49), IL-1β (0.52), IL-6 (0.71) and RANKL (0.71). Phytoestrogens therefore prevent the pro-osteoclastic action of T cells suggesting they may have a role in the control of inflammatory bone loss.« less
Jia, Dan; Zheng, Weijiang; Jiang, Honglin
2018-06-01
The C3H10T1/2 cells are considered mesenchymal stem cells (MSCs) because they can be induced to become the progenitor cells for myocytes, adipocytes, osteoblasts, and chondrocytes by the DNA methyltransferase inhibitor 5'-azacytidine. In this study, we determined the effect of growth hormone (GH) on the myogenic and adipogenic lineage commitment in C3H10T1/2 cells. The C3H10T1/2 cells were treated with recombinant bovine GH in the presence or absence of 5'-azacytidine for 4 days. The myogenic commitment in C3H10T1/2 cells was assessed by immunostaining them for MyoD, the marker for myoblasts, and by determining their capacity to differentiate into the multinucleated myotubes. The adipogenic commitment in C3H10T1/2 cells was assessed by determining their ability to differentiate into adipocytes. Myotubes and adipocyteswere identified by immunocytochemistry and Oil Red O staining, respectively. C3H10T1/2 cells treated with 5'-azacytidine and GH for 4 days contained a greater percentage of MyoD-positive cells than those treated with 5'-axacytidine alone (P < 0.05). The former generated more myotubes than the latter upon induced myoblast differentiation (P < 0.05). However, C3H10T1/2 cells treated with GH alone did not form any myotubes. C3H10T1/2 cells treated with 5'-azacytidine formed adipocytes upon adipocyte differentiation induction, whereas C3H10T1/2 cells treated with GH alone did not form any adipocytes. C3H10T1/2 cells treated with both 5'-azacytidine and GH formed fewer adipocytes than those treated with 5'-azacytidine alone (P < 0.05). Both GHR and IGF-I mRNA expression in C3H10T1/2 cells were increased by 5'-azacytidine (P < 0.05), but neither was affected by GH. Overall, this study showed that GH enhanced 5'-azacytidine-induced commitment in C3H10T1/2 cells to myoblasts but inhibited 5'-azacytidine-induced commitment to preadipocytes. These results support the possibility that GH stimulates skeletal muscle growth and inhibits adipose
Pellerin, Laurence; Jenks, Jennifer Anne; Chinthrajah, Sharon; Dominguez, Tina; Block, Whitney; Zhou, Xiaoying; Noshirvan, Arram; Gregori, Silvia; Roncarolo, Maria Grazia; Nadeau, Kari Christine; Bacchetta, Rosa
2018-01-01
Peanut allergy (PA) is a life-threatening condition that lacks regulator-approved treatment. Regulatory T type 1 (T R 1) cells are potent suppressors of immune responses and can be induced in vivo upon repeated antigen exposure or in vitro by using tolerogenic dendritic cells. Whether oral immunotherapy (OIT) leads to antigen-specific T R 1 cell induction has not been established. We sought to determine whether peanut-specific T R 1 cells can be generated in vitro from peripheral blood of patients with PA at baseline or during OIT and whether they are functional compared with peanut-specific T R 1 cells induced from healthy control (HC) subjects. Tolerogenic dendritic cells were differentiated in the presence of IL-10 from PBMCs of patients with PA and HC subjects pulsed with the main peanut allergens of Arachis hypogaea, Ara h 1 and 2, and used as antigen-presenting cells for autologous CD4 + T cells (CD4 + T cells coincubated with tolerogenic dendritic cells pulsed with the main peanut allergens [pea-T10 cells]). Pea-T10 cells were characterized by the presence of CD49b + lymphocyte-activation gene 3 (LAG3) + T R 1 cells, antigen-specific proliferative responses, and cytokine production. CD49b + LAG3 + T R 1 cells were induced in pea-T10 cells at comparable percentages from HC subjects and patients with PA. Despite their antigen specificity, pea-T10 cells of patients with PA with or without OIT, as compared with those of HC subjects, were not anergic and had high T H 2 cytokine production upon peanut-specific restimulation. Peanut-specific T R 1 cells can be induced from HC subjects and patients with PA, but those from patients with PA are functionally defective independent of OIT. The unfavorable T R 1/T H 2 ratio is discussed as a possible cause of PA T R 1 cell impairment. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.
Pannexin1 channels act downstream of P2X7 receptors in ATP-induced murine T-cell death
Shoji, Kenji F; Sáez, Pablo J; Harcha, Paloma A; Aguila, Hector L; Sáez, Juan C
2014-01-01
Death of murine T cells induced by extracellular ATP is mainly triggered by activation of purinergic P2X7 receptors (P2X7Rs). However, a link between P2X7Rs and pannexin1 (Panx1) channels, which are non-selective, has been recently demonstrated in other cell types. In this work, we characterized the expression and cellular distribution of pannexin family members (Panxs 1, 2 and 3) in isolated T cells. Panx1 was the main pannexin family member clearly detected in both helper (CD4+) and cytotoxic (CD8+) T cells, whereas low levels of Panx2 were found in both T-cell subsets. Using pharmacological and genetic approaches, Panx1 channels were found to mediate most ATP-induced ethidium uptake since this was drastically reduced by Panx1 channel blockers (10Panx1, Probenecid and low carbenoxolone concentration) and absent in T cells derived from Panx1−/− mice. Moreover, electrophysiological measurements in wild-type CD4+ cells treated with ATP unitary current events and pharmacological sensitivity compatible with Panx1 channels were found. In addition, ATP release from T cells treated with 4Br-A23187, a calcium ionophore, was completely blocked with inhibitors of both connexin hemichannels and Panx1 channels. Panx1 channel blockers drastically reduced the ATP-induced T-cell mortality, indicating that Panx1 channels mediate the ATP-induced T-cell death. However, mortality was not reduced in T cells of Panx1−/− mice, in which levels of P2X7Rs and ATP-induced intracellular free Ca2+ responses were enhanced suggesting that P2X7Rs take over Panx1 channels lose-function in mediating the onset of cell death induced by extracellular ATP. PMID:24590064
Immunomodulation and T Helper TH1/TH2 Response Polarization by CeO2 and TiO2 Nanoparticles
Schanen, Brian C.; Das, Soumen; Reilly, Christopher M.; Warren, William L.; Self, William T.; Seal, Sudipta; Drake, Donald R.
2013-01-01
Immunomodulation by nanoparticles, especially as related to the biochemical properties of these unique materials, has scarcely been explored. In an in vitro model of human immunity, we demonstrate two catalytic nanoparticles, TiO2 (oxidant) and CeO2 (antioxidant), have nearly opposite effects on human dendritic cells and T helper (TH) cells. For example, whereas TiO2 nanoparticles potentiated DC maturation that led towards TH1-biased responses, treatment with antioxidant CeO2 nanoparticles induced APCs to secrete the anti-inflammatory cytokine, IL-10, and induce a TH2-dominated T cell profile. In subsequent studies, we demonstrate these results are likely explained by the disparate capacities of the nanoparticles to modulate ROS, since TiO2, but not CeO2 NPs, induced inflammatory responses through an ROS/inflammasome/IL-1β pathway. This novel capacity of metallic NPs to regulate innate and adaptive immunity in profoundly different directions via their ability to modulate dendritic cell function has strong implications for human health since unintentional exposure to these materials is common in modern societies. PMID:23667525
Cancer cell-derived IL-1α induces CCL22 and the recruitment of regulatory T cells
Wiedemann, Gabriela Maria; Knott, Max Martin Ludwig; Vetter, Viola Katharina; Rapp, Moritz; Haubner, Sascha; Fesseler, Julia; Kühnemuth, Benjamin; Layritz, Patrick; Thaler, Raffael; Kruger, Stephan; Ormanns, Steffen; Mayr, Doris; Endres, Stefan; Anz, David
2016-01-01
ABSTRACT In cancer patients, immunosuppression through regulatory T cells (Treg) is a crucial component of tumor immune evasion and contributes to disease progression. Tumor-infiltrating Treg in particular suppress local effector T cell responses and are associated with poor prognosis in tumors such as human pancreatic cancer or hepatocellular carcinoma (HCC). The chemokine CCL22 is known to recruit Treg into the tumor tissue and many types of human tumors are known to express high levels of CCL22. The mechanisms leading to intratumoral secretion of CCL22 are so far unknown. We demonstrate here that intratumoral CCL22 is induced in tumor-infiltrating immune cells through cancer cell-derived interleukin-1 (IL-1α). In pancreatic cancer and HCC, CCL22 is produced by intratumoral dendritic cells, while the cancer cells themselves do not secrete CCL22 in vitro and in vivo. Incubation of human peripheral blood mononuclear cells (PBMC) or murine splenocytes with tumor cells or tumor cell supernatants strongly induced CCL22 secretion in vitro. Tumor cell supernatants contained IL-1 and CCL22 induction in PBMC could be specifically prevented by the IL-1 receptor antagonist anakinra or by transfection of tumor cell lines with IL-1 siRNA, leading to a suppression of Treg migration. In conclusion, we identify here tumor cell-derived IL-1α as a major inducer of the Treg attracting chemokine CCL22 in human cancer cells. Therapeutic blockade of the IL-1 pathway could represent a promising strategy to inhibit tumor-induced immunosuppression. PMID:27757295
6-gingerol inhibits rosiglitazone-induced adipogenesis in 3T3-L1 adipocytes.
Tzeng, Thing-Fong; Chang, Chia Ju; Liu, I-Min
2014-02-01
We investigated the effects of 6-gingerol ((S)-5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-3-decanone) on the inhibition of rosiglitazone (RGZ)-induced adipogenesis in 3T3-L1 cells. The morphological changes were photographed based on staining lipid accumulation by Oil-Red O in RGZ (1 µmol/l)-treated 3T3-L1 cells without or with various concentrations of 6-gingerol on differentiation day 8. Quantitation of triglycerides content was performed in cells on day 8 after differentiation induction. Differentiated cells were lysed to detect mRNA and protein levels of adipocyte-specific transcription factors by real-time reverse transcription-polymerase chain reaction and Western blot analysis, respectively. 6-gingerol (50 µmol/l) effectively suppressed oil droplet accumulation and reduced the sizes of the droplets in RGZ-induced adipocyte differentiation in 3T3-L1 cells. The triglyceride accumulation induced by RGZ in differentiated 3T3-L1 cells was also reduced by 6-gingerol (50 µmol/l). Treatment of differentiated 3T3-L1 cells with 6-gingerol (50 µmol/l) antagonized RGZ-induced gene expression of peroxisome proliferator-activated receptor (PPAR)γ and CCAAT/enhancer-binding protein α. Additionally, the increased levels of mRNA and protein in adipocyte-specific fatty acid binding protein 4 and fatty acid synthase induced by RGZ in 3T3-L1 cells were decreased upon treatment with 6-gingerol. Our data suggests that 6-gingerol may be beneficial in obesity, by reducing adipogenesis partly through the down-regulating PPARγ activity. Copyright © 2013 John Wiley & Sons, Ltd.
Lysophosphatidic acid induces chemotaxis in MC3T3-E1 osteoblastic cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Masiello, Lisa M.; Fotos, Joseph S.; Galileo, Deni S.
2006-07-01
Lysophosphatidic acid (LPA) is a bioactive lipid that has pleiotropic effects on a variety of cell types and enhances the migration of endothelial and cancer cells, but it is not known if this lipid can alter osteoblast motility. We performed transwell migration assays using MC3T3-E1 osteoblastic cells and found LPA to be a potent chemotactic agent. Quantitative time-lapse video analysis of osteoblast migration after wounds were introduced into cell monolayers indicated that LPA stimulated both migration velocity and the average migration distance per cell. LPA also elicited substantial changes in cell shape and actin cytoskeletal structure; lipid-treated cells contained fewermore » stress fibers and displayed long membrane processes that were enriched in F-actin. Quantitative RT-PCR analysis showed that MC3T3-E1 cells express all four known LPA-specific G protein-coupled receptors (LPA1-LPA4) with a relative mRNA abundance of LPA1 > LPA4 > LPA2 >> LPA3. LPA-induced changes in osteoblast motility and morphology were antagonized by both pertussis toxin and Ki16425, a subtype-specific blocker of LPA1 and LPA3 receptor function. Cell migration in many cell types is linked to changes in intracellular Ca2+. Ki16425 also inhibited LPA-induced Ca2+ signaling in a dose-dependent manner, suggesting a link between LPA-induced Ca2+ transients and osteoblast chemotaxis. Our data show that LPA stimulates MC3T3-E1 osteoblast motility via a mechanism that is linked primarily to the G protein-coupled receptor LPA1.« less
Kasimsetty, Sashi G; Shigeoka, Alana A; Scheinok, Andrew A; Gavin, Amanda L; Ulevitch, Richard J; McKay, Dianne B
2017-08-01
Nucleotide-binding oligomerization domain (Nod)-containing proteins Nod1 and Nod2 play important roles in the innate immune response to pathogenic microbes, but mounting data suggest these pattern recognition receptors might also play key roles in adaptive immune responses. Targeting Nod1 and Nod2 signaling pathways in T cells is likely to provide a new strategy to modify inflammation in a variety of disease states, particularly those that depend on Ag-induced T cell activation. To better understand how Nod1 and Nod2 proteins contribute to adaptive immunity, this study investigated their role in alloantigen-induced T cell activation and asked whether their absence might impact in vivo alloresponses using a severe acute graft versus host disease model. The study provided several important observations. We found that the simultaneous absence of Nod1 and Nod2 primed T cells for activation-induced cell death. T cells from Nod1 × 2 -/- mice rapidly underwent cell death upon exposure to alloantigen. The Nod1 × 2 -/- T cells had sustained p53 expression that was associated with downregulation of its negative regulator MDM2. In vivo, mice transplanted with an inoculum containing Nod1 × 2 -/- T cells were protected from severe graft versus host disease. The results show that the simultaneous absence of Nod1 and Nod2 is associated with accelerated T cell death upon alloantigen encounter, suggesting these proteins might provide new targets to ameliorate T cell responses in a variety of inflammatory states, including those associated with bone marrow or solid organ transplantation. Copyright © 2017 by The American Association of Immunologists, Inc.
Tournoy, K G; Hove, C; Grooten, J; Moerloose, K; Brusselle, G G; Joos, G F
2006-01-01
Non-specific anti-inflammatory medication is actually the treatment of choice for controlling the T-helper type 2 (Th-2) cell-driven airway inflammation in asthma. The induction of counterbalancing Th-1 cell clones, long considered a promising approach for immunotherapy, has failed to fulfil its promise because of potentially detrimental side-effects. This is therefore probably not a valid option for the treatment of asthma. With the increasing awareness that active immune mechanisms exist to control inflammatory responses, interest rises to investigate whether these can be exploited to control allergen-induced airway disease. The induction of antigen-specific T cells with suppressive characteristics (regulatory T cells) is therefore a potentially interesting approach. These regulatory T cells mediate tolerance in healthy, non-atopic individuals and have the potential of becoming an effective means of preventing allergen-induced airway inflammation and possibly of suppressing ongoing allergic immune responses. Here we review the available knowledge about allergen-induced suppressive immunity obtained from animal models taking into account the different developmental stages of allergic airway disease.
Wan, Jiangbo; Huang, Fang; Hao, Siguo; Hu, Weiwei; Liu, Chuanxu; Zhang, Wenhao; Deng, Xiaohui; Chen, Linjun; Ma, Liyuan; Tao, Rong
2017-01-01
Tr1 cells can induce peripheral tolerance to self- and foreign antigens, and have been developed as a therapeutic tool for the induction of tolerance to transplanted tissue. We explored the feasibility of generating Tr1 cells by using IL-10 gene-modified recipient DCs (DCLV-IL-10) to stimulate donor naive CD4+ T cells. We also investigated some biological properties of Tr1 cells. DCLV-IL-10 were generated through DCs transduced with a lentivirus vector carrying the IL-10 gene, and Tr1 cells were produced by using DCLV-IL-10 to stimulate naive CD4+ T cells. The effects of Tr1 cells on T-cell proliferation and the occurrence of graft versus host disease (GVHD) following allogeneic stem-cell transplantation (allo-HSCT) were investigated. The DCLV-IL-10-induced Tr1 cells co-expressed LAG-3 and CD49b. Moreover, they also expressed CD4, CD25, and IL-10, but not Foxp3, and secreted significantly higher levels of IL-10 (1,729.36 ± 185.79 pg/mL; P < 0.001) and INF-γ (1,524.48 ± 168.65 pg/mL; P < 0.01) than the control T cells upon the stimulation by allogeneic DCs. Tr1 cells markedly suppressed T-lymphocyte proliferation and the mixed lymphocytic response (MLR) in vitro. The mice used in the allo-HSCT model had longer survival times and lower clinical and pathological GVHD scores than the control mice. IL-10 gene-modified DC-induced Tr1 cells may be used as a potent cellular therapy for the prevention of GVHD after allo-HSCT. © 2017 The Author(s). Published by S. Karger AG, Basel.
GAD-specific T cells are induced by GAD-alum treatment in Type-1 diabetes patients.
Pihl, Mikael; Barcenilla, Hugo; Axelsson, Stina; Chéramy, Mikael; Åkerman, Linda; Johansson, Ingela; Ludvigsson, Johnny; Casas, Rosaura
2017-03-01
Administration of Glutamic Acid Decarboxylase (GAD) 65 formulated in aluminium hydroxide preserved insulin secretion in a phase II trial in recent onset Type 1 Diabetes. A subsequent European phase III trial was closed at 15months after failing to reach primary endpoint, but the majority of the Swedish patients completed the 21months follow-up. We studied the frequencies and phenotype of T cells, suppressive capacity of Tregs, GAD 65 -induced proliferation, and frequencies of T cells with a GAD 65 -specific TCR in Swedes participating in the trial. Stimulation with GAD 65 induced activated T cells and also cells with a suppressive phenotype. Activated GAD 65 -specific effector T cells were detected by tetramer staining while the frequency of GAD 65 -specific Treg was not affected by the treatment. Additional doses of GAD-alum increased frequencies of CD25 + CD127 + , but had no effect on CD25 hi CD127 lo . Our findings indicate that GAD-alum treatment primarily induced activated T cells. GAD 65 -specific cells were mainly of activated phenotype. Copyright © 2017 Elsevier Inc. All rights reserved.
Gross, Christine; Wiesmann, Veit; Millen, Sebastian; Kalmer, Martina; Wittenberg, Thomas; Gettemans, Jan; Thoma-Kress, Andrea K
2016-10-01
The delta-retrovirus Human T-cell leukemia virus type 1 (HTLV-1) preferentially infects CD4+ T-cells via cell-to-cell transmission. Viruses are transmitted by polarized budding and by transfer of viral biofilms at the virological synapse (VS). Formation of the VS requires the viral Tax protein and polarization of the host cytoskeleton, however, molecular mechanisms of HTLV-1 cell-to-cell transmission remain incompletely understood. Recently, we could show Tax-dependent upregulation of the actin-bundling protein Fascin (FSCN-1) in HTLV-1-infected T-cells. Here, we report that Fascin contributes to HTLV-1 transmission. Using single-cycle replication-dependent HTLV-1 reporter vectors, we found that repression of endogenous Fascin by short hairpin RNAs and by Fascin-specific nanobodies impaired gag p19 release and cell-to-cell transmission in 293T cells. In Jurkat T-cells, Tax-induced Fascin expression enhanced virus release and Fascin-dependently augmented cell-to-cell transmission to Raji/CD4+ B-cells. Repression of Fascin in HTLV-1-infected T-cells diminished virus release and gag p19 transfer to co-cultured T-cells. Spotting the mechanism, flow cytometry and automatic image analysis showed that Tax-induced T-cell conjugate formation occurred Fascin-independently. However, adhesion of HTLV-1-infected MT-2 cells in co-culture with Jurkat T-cells was reduced upon knockdown of Fascin, suggesting that Fascin contributes to dissemination of infected T-cells. Imaging of chronically infected MS-9 T-cells in co-culture with Jurkat T-cells revealed that Fascin's localization at tight cell-cell contacts is accompanied by gag polarization suggesting that Fascin directly affects the distribution of gag to budding sites, and therefore, indirectly viral transmission. In detail, we found gag clusters that are interspersed with Fascin clusters, suggesting that Fascin makes room for gag in viral biofilms. Moreover, we observed short, Fascin-containing membrane extensions surrounding
Wiesmann, Veit; Millen, Sebastian; Wittenberg, Thomas; Gettemans, Jan; Thoma-Kress, Andrea K.
2016-01-01
The delta-retrovirus Human T-cell leukemia virus type 1 (HTLV-1) preferentially infects CD4+ T-cells via cell-to-cell transmission. Viruses are transmitted by polarized budding and by transfer of viral biofilms at the virological synapse (VS). Formation of the VS requires the viral Tax protein and polarization of the host cytoskeleton, however, molecular mechanisms of HTLV-1 cell-to-cell transmission remain incompletely understood. Recently, we could show Tax-dependent upregulation of the actin-bundling protein Fascin (FSCN-1) in HTLV-1-infected T-cells. Here, we report that Fascin contributes to HTLV-1 transmission. Using single-cycle replication-dependent HTLV-1 reporter vectors, we found that repression of endogenous Fascin by short hairpin RNAs and by Fascin-specific nanobodies impaired gag p19 release and cell-to-cell transmission in 293T cells. In Jurkat T-cells, Tax-induced Fascin expression enhanced virus release and Fascin-dependently augmented cell-to-cell transmission to Raji/CD4+ B-cells. Repression of Fascin in HTLV-1-infected T-cells diminished virus release and gag p19 transfer to co-cultured T-cells. Spotting the mechanism, flow cytometry and automatic image analysis showed that Tax-induced T-cell conjugate formation occurred Fascin-independently. However, adhesion of HTLV-1-infected MT-2 cells in co-culture with Jurkat T-cells was reduced upon knockdown of Fascin, suggesting that Fascin contributes to dissemination of infected T-cells. Imaging of chronically infected MS-9 T-cells in co-culture with Jurkat T-cells revealed that Fascin’s localization at tight cell-cell contacts is accompanied by gag polarization suggesting that Fascin directly affects the distribution of gag to budding sites, and therefore, indirectly viral transmission. In detail, we found gag clusters that are interspersed with Fascin clusters, suggesting that Fascin makes room for gag in viral biofilms. Moreover, we observed short, Fascin-containing membrane extensions
DOE Office of Scientific and Technical Information (OSTI.GOV)
Li, Qiang; Kamemura, Kazuo, E-mail: k_kamemura@nagahama-i-bio.ac.jp
2014-07-18
Highlights: • The majority of EWS localizes stably in the cytosol in 3T3-L1 preadipocytes. • Adipogenic stimuli induce the nuclear localization of EWS. • Adipogenesis promotes O-GlcNAcylation of EWS. • O-GlcNAcylation stimulates the recruitment of EWS to the nuclear periphery. - Abstract: Although the Ewing sarcoma (EWS) proto-oncoprotein is found in the nucleus and cytosol and is associated with the cell membrane, the regulatory mechanisms of its subcellular localization are still unclear. Here we found that adipogenic stimuli induce the nuclear localization of EWS in 3T3-L1 cells. Tyrosine phosphorylation in the C-terminal PY-nuclear localization signal of EWS was negative throughoutmore » adipogenesis. Instead, an adipogenesis-dependent increase in O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation of EWS was observed. Pharmacological inactivation of O-GlcNAcase in preadipocytes promoted perinuclear localization of EWS. Our findings suggest that the nuclear localization of EWS is partly regulated by the glycosylation.« less
Curcumin inhibits adipogenesis induced by benzyl butyl phthalate in 3T3-L1 cells.
Sakuma, Satoru; Sumida, Maki; Endoh, Yukiko; Kurita, Ayaka; Yamaguchi, Ayana; Watanabe, Tomoki; Kohda, Tetsuya; Tsukiyama, Yui; Fujimoto, Yohko
2017-08-15
Phthalates are a group of endocrine disrupting chemicals and may have contributed to the recent global obesity health crisis. Increased adipogenesis via the peroxisome proliferator-activated receptor γ (PPARγ)-CCAAT-enhancer binding protein α (C/EBPα) pathway could be one critical mechanism responsible for phthalate-induced weight gain. On the other hand, curcumin has been shown to inhibit adipogenesis in cells and animal models. The present study was undertaken to evaluate, for the first time, whether curcumin could reduce adipogenesis induced by benzyl butyl phthalate (BBP) via downregulation of the PPARγ-C/EBPα pathway. 3T3-L1 preadipocytes were differentiated by treating them with insulin, dexamethasone, and 3-isobutyl-1-methylxanthine in the presence of BBP, with or without curcumin. Cells that were grown in the presence of BBP alone showed a significant increase in triacylglycerol (TG) levels. In addition, the number of Oil Red O-stained cells and the mRNA expression levels of PPARγ, C/EBPα, adiponectin, and tumor necrosis factor-α (TNFα) were significantly increased. However, treatment with BBP in combination with curcumin resulted in major reductions in TG levels, the numbers of Oil Red O-stained cells, and the mRNA expression levels of the four proteins. These results suggest that curcumin might be an inhibitor of BBP-induced weight gain and inflammation via stimulation of adipocyte differentiation and TNFα generation. Curcumin may, therefore, be a potential medication for preventing the harmful effects of phthalates. Copyright © 2017 Elsevier Inc. All rights reserved.
RasGRP1 regulates antigen-induced developmental programming by naive CD8 T cells.
Priatel, John J; Chen, Xiaoxi; Huang, Yu-Hsuan; Chow, Michael T; Zenewicz, Lauren A; Coughlin, Jason J; Shen, Hao; Stone, James C; Tan, Rusung; Teh, Hung Sia
2010-01-15
Ag encounter by naive CD8 T cells initiates a developmental program consisting of cellular proliferation, changes in gene expression, and the formation of effector and memory T cells. The strength and duration of TCR signaling are known to be important parameters regulating the differentiation of naive CD8 T cells, although the molecular signals arbitrating these processes remain poorly defined. The Ras-guanyl nucleotide exchange factor RasGRP1 has been shown to transduce TCR-mediated signals critically required for the maturation of developing thymocytes. To elucidate the role of RasGRP1 in CD8 T cell differentiation, in vitro and in vivo experiments were performed with 2C TCR transgenic CD8 T cells lacking RasGRP1. In this study, we report that RasGRP1 regulates the threshold of T cell activation and Ag-induced expansion, at least in part, through the regulation of IL-2 production. Moreover, RasGRP1(-/-) 2C CD8 T cells exhibit an anergic phenotype in response to cognate Ag stimulation that is partially reversible upon the addition of exogenous IL-2. By contrast, the capacity of IL-2/IL-2R interactions to mediate Ras activation and CD8 T cell expansion and differentiation appears to be largely RasGRP1-independent. Collectively, our results demonstrate that RasGRP1 plays a selective role in T cell signaling, controlling the initiation and duration of CD8 T cell immune responses.
Liu, Zuqiang; Falo, Louis D; You, Zhaoyang
2011-07-01
Although high mobility group box 1 (HMGB1) in tumor cells is involved in many aspects of tumor progression, its role in tumor immune suppression remains elusive. Host cell-derived IL-10 suppressed a naturally acquired CD8 T cell-dependent antitumor response. The suppressive activity of tumor-associated Foxp3(+)CD4(+)CD25(+) regulatory T cells (Treg) was IL-10 dependent. Neutralizing HMGB1 impaired tumor cell-promoted IL-10 production by Treg. Short hairpin RNA-mediated knockdown of HMGB1 (HMGB1 KD) in tumor cells did not affect tumor cell growth but uncovered naturally acquired long-lasting tumor-specific IFN-γ- or TNF-α-producing CD8 T cell responses and attenuated their ability to induce Treg, leading to naturally acquired CD8 T cell- or IFN-γ-dependent tumor rejection. The data suggest that tumor cell-derived HMGB1 may suppress naturally acquired CD8 T cell-dependent antitumor immunity via enhancing Treg to produce IL-10, which is necessary for Treg-mediated immune suppression.
Joshi, Nikhil S.; Cui, Weiguo; Dominguez, Claudia; Chen, Jonathan H.; Hand, Timothy W.; Kaech, Susan M.
2011-01-01
Memory CD8 T cells acquire TEM properties following reinfection, and may reach terminally differentiated, senescent states (“Hayflick limit”) after multiple infections. The signals controlling this process are not well understood, but we found that the degree of 2o effector and memory CD8 T cell differentiation was intimately linked to the amount of T-bet expressed upon reactivation and pre-existing memory CD8 T cell number (i.e., 1o memory CD8 T cell precursor frequency) present during secondary infection. Compared to naïve cells, memory CD8 T cells were predisposed towards terminal effector (TE) cell differentiation because they could immediately respond to IL-12 and induce T-bet, even in the absence of antigen. TE cell formation following 2o or 3o infections was dependent on increased T-bet expression because T-bet+/− cells were resistant to these phenotypic changes. Larger numbers of pre-existing memory CD8 T cells limited the duration of 2o infection and the amount of IL-12 produced, and consequently, this reduced T-bet expression and the proportion of 2o TE CD8 T cells that formed. Together, these data show that, over repeated infections, memory CD8 T cell quality and proliferative fitness is not strictly determined by the number of serial encounters with antigen or cell divisions, but is a function of the CD8 T cell differentiation state, which is genetically controlled in a T-bet-dependent manner. This differentiation state can be modulated by pre-existing memory CD8 T cell number and the intensity of inflammation during reinfection. These results have important implications for vaccinations involving prime-boost strategies. PMID:21930973
IL-10 Producing B Cells Ability to Induce Regulatory T Cells Is Maintained in Rheumatoid Arthritis
Mielle, Julie; Audo, Rachel; Hahne, Michael; Macia, Laurence; Combe, Bernard; Morel, Jacques; Daien, Claire
2018-01-01
Despite growing evidence highlighting the relevance of increasing IL-10-producing B cells (B10+cells) in autoimmune diseases, their functions in patients are still unknown. The aim of this study was to evaluate the functions of CpG-induced B10+ cells isolated from healthy controls (HC) and rheumatoid arthritis (RA) patients, on naïve T cell differentiation. We demonstrated that CpG-induced B10+ cells from HC drove naïve T cell differentiation toward regulatory T cells (Treg cells) and IL-10-producing T cells (Tr1) through IL-10 secretion and cellular contacts. B10+ cells from HC did not decrease T helper 1 (Th1) nor and tumor necrosis factor α producing T cell (TNFα+ T cell) differentiation. We showed that in RA, B10+ cells could also induce Treg cells and Tr1 from naïve T cells. Contrary to HC, B10+ cells from RA patients increased naïve T cell conversion into Th1. Interestingly, PD-L2, a programmed death-1 (PD-1) ligand that inhibits PD-L1 and promotes Th1 differentiation, was overexpressed on RA B10+ cells compared to HC B10+ cells. Together, our findings showed that CpG-induced B10+ cells may be used to increase Treg cells in patients with RA. However, CpG may not be the most adequate stimuli as CpG-induced B10+ cells also increased inflammatory T cells in those patients. PMID:29774031
Kimura, Ryuichiro; Senba, Masachika; Cutler, Samuel J; Ralph, Stephen J; Xiao, Gutian; Mori, Naoki
2013-01-01
Human T cell leukemia virus type I (HTLV-I) is the etiologic agent of adult T cell leukemia (ATL) and various inflammatory disorders including HTLV-I-associated myelopathy/tropical spastic paraparesis. HTLV-I oncoprotein Tax is known to cause permanent activation of many cellular transcription factors including nuclear factor-κB (NF-κB), cyclic adenosine 3′,5′-monophosphate response element-binding protein, and activator protein 1 (AP-1). Here, we show that NF-κB-binding cofactor inhibitor of NF-κB-ζ (IκB-ζ) is constitutively expressed in HTLV-I-infected T cell lines and ATL cells, and Tax transactivates the IκB-ζ gene, mainly through NF-κB. Microarray analysis of IκB-ζ-expressing uninfected T cells demonstrated that IκB-ζ induced the expression of NF-κB. and interferon-regulatory genes such as B cell CLL/lymphoma 3 (Bcl3), guanylate-binding protein 1, and signal transducer and activator of transcription 1. The transcriptional activation domain, nuclear localization signal, and NF-κB-binding domain of IκB-ζ were required for Bcl3 induction, and IκB-ζ synergistically enhanced Tax-induced Bcl3 transactivation in an NF-κB-dependent manner. Interestingly, IκB-ζ inhibited Tax-induced NF-κB, AP-1 activation, and HTLV-I transcription. Furthermore, IκB-ζ interacted with Tax in vitro and this interaction was also observed in an HTLV-I-transformed T cell line. These results suggest that IκB-ζ modulates Tax-dependent and Tax-independent gene transcription in T cells. The function of IκB-ζ may be of significance in ATL genesis and pathogenesis of HTLV-I-associated diseases. PMID:24027435
Liu, Yan-Yan; Sun, Ling-Cong; Wei, Jing-Jing; Li, Dong; Yuan, Ye; Yan, Bin; Liang, Zhi-Hui; Zhu, Hui-Fen; Xu, Yong; Li, Bo; Song, Chuan-Wang; Liao, Sheng-Jun; Lei, Zhang; Zhang, Gui-Mei; Feng, Zuo-Hua
2010-09-01
Gr-1(+)CD11b(+)F4/80(+) cells play important roles in tumor development and have a negative effect on tumor immunotherapy. So far, the mechanisms underlying the regulation of their immunosuppressive phenotype by classical and alternative macrophage activation stimuli are not well elucidated. In this study, we found that molecules from necrotic tumor cells (NTC-Ms) stimulated Gr-1(+)CD11b(+)F4/80(+) cells to induce apoptosis of activated T cells but not nonstimulated T cells. The apoptosis-inducing capacity was determined by higher expression levels of arginase I and IL-10 relative to those of NO synthase 2 and IL-12 in Gr-1(+)CD11b(+)F4/80(+) cells, which were induced by NTC-Ms through TLR4 signaling. The apoptosis-inducing capacity of NTC-Ms-stimulated Gr-1(+)CD11b(+)F4/80(+) cells could be enhanced by IL-10. IFN-gamma may reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells only if their response to IFN-gamma was not attenuated. However, the potential of Gr-1(+)CD11b(+)F4/80(+) cells to express IL-12 in response to IFN-gamma could be attenuated by tumor, partially due to the existence of active STAT3 in Gr-1(+)CD11b(+)F4/80(+) cells and NTC-Ms from tumor. In this situation, IFN-gamma could not effectively reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells. Tumor immunotherapy with 4-1BBL/soluble programmed death-1 may significantly reduce, but not abolish the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells in local microenvironment. Blockade of TLR4 signaling could further reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells and enhance the suppressive effect of 4-1BBL/soluble form of programmed death-1 on tumor growth. These findings indicate the relationship of distinct signaling pathways with apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells and emphasize the importance of blocking TLR4 signaling to prevent the induction of T cell apoptosis by Gr-1(+)CD11b(+)F4/80(+) cells.
Zhou, Cheng; Huang, Shoucheng
2017-01-01
Selenium- (Se-) enriched polysaccharide SPMP-2a was extracted and purified from Pleurotus geesteranus. SPMP-2a is a white flocculent polysaccharide and soluble in water, with a molecular weight of 3.32 × 104 Da. Fourier transform infrared spectroscopy spectral analysis indicated that it belongs to an acid Se polysaccharide with α-D-glucopyranoside bond. The effects of Se polysaccharide SPMP-2a in P. geesteranus against hydrogen peroxide- (H2O2-) induced oxidative damage in human keratinocytes (HaCaT) cells were evaluated further. Reduced cell viability and elevated apoptotic rates in H2O2-treated HaCaT cells were proven by MTT and flow cytometry assays. Hoechst 33342 staining revealed chromatin condensations in the nuclei of HaCaT cells. However, with the addition of SPMP-2a, cell viability improved, nuclear condensation declined, and cell apoptotic rates dropped significantly. Ultrastructural observation consistently revealed that treatments with SPMP-2a reduced the number of swollen and vacuolar mitochondria in the H2O2-treated cells compared with the controls. Furthermore, SPMP-2a increased the superoxide dismutase (SOD) and catalase (CAT) activities and reduced reactive oxygen species (ROS) content. Western blot analysis showed that SPMP-2a treatment effectively increased B-cell lymphoma 2 (Bcl-2) protein expression. Therefore, SPMP-2a could improve cellular antioxidant enzyme activities, reduce ROS levels, and increase Bcl-2 protein expression levels, thereby reducing cell apoptosis and protecting HaCaT cells from H2O2-induced oxidative damage. PMID:28293636
Zhang, Xiang; Jiang, Zhengping; Gu, Yan; Liu, Yanfang; Cao, Xuetao; Han, Yanmei
2016-12-01
Kupffer cells, tissue-resident macrophage lineage cell, are enriched in vertebrate liver. The mouse F4/80 + Kupffer cells have been subclassified into two subpopulations according to their phenotype and function: CD68 + subpopulation with potent reactive oxygen species (ROS) production and phagocytic capacities, and CD11b + subpopulation with a potent capacity to produce T helper 1 cytokines. In addition, CD11b + Kupffer cells/macrophages may be migrated from the bone marrow or spleen, especially in inflammatory conditions of the liver. For analyzing diverse Kupffer cell subsets, we infected mice with Listeria monocytogenes and analyzed the phenotype variations of hepatic Kupffer cells. During L. monocytogenes infection, hepatic CD69 + Kupffer cells were significantly induced and expanded, and CD69 + Kupffer cells expressed higher level of CD11b, and particularly high level of membrane-bound TGF-β1 (mTGF-β1) but lower level of F4/80. We also found that clodronate liposome administration did not eliminate hepatic CD69 + Kupffer cell subset. We consider the hepatic CD69 + Kupffer cell population corresponds to CD11b + Kupffer cells, the bone marrow-derived population. Hepatic CD69 + Kupffer cells suppressed Ag-nonspecific and OVA-specific CD4 T cell proliferation through mTGF-β1 both in vitro and in vivo, meanwhile, they did not interfere with activation of CD4 T cells. Thus, we have identified a new subset of inflammation-induced CD69 + Kupffer cells which can feedback inhibit CD4 T cell response via cell surface TGF-β1 at the late stage of immune response against infection. CD69 + Kupffer cells may contribute to protect host from pathological injure by preventing overactivation of immune response.
Cruz-Adalia, Aránzazu; Ramirez-Santiago, Guillermo; Osuna-Pérez, Jesús; Torres-Torresano, Mónica; Zorita, Virgina; Martínez-Riaño, Ana; Boccasavia, Viola; Borroto, Aldo; Martínez Del Hoyo, Gloria; González-Granado, José María; Alarcón, Balbino; Sánchez-Madrid, Francisco; Veiga, Esteban
2017-11-17
Bacterial phagocytosis and antigen cross-presentation to activate CD8 + T cells are principal functions of professional antigen presenting cells. However, conventional CD4 + T cells also capture and kill bacteria from infected dendritic cells in a process termed transphagocytosis (also known as transinfection). Here, we show that transphagocytic T cells present bacterial antigens to naive CD8 + T cells, which proliferate and become cytotoxic in response. CD4 + T-cell-mediated antigen presentation also occurs in vivo in the course of infection, and induces the generation of central memory CD8 + T cells with low PD-1 expression. Moreover, transphagocytic CD4 + T cells induce protective anti-tumour immune responses by priming CD8 + T cells, highlighting the potential of CD4 + T cells as a tool for cancer immunotherapy.
ArtinM Mediates Murine T Cell Activation and Induces Cell Death in Jurkat Human Leukemic T Cells
Oliveira-Brito, Patrícia Kellen Martins; Gonçalves, Thiago Eleutério; Vendruscolo, Patrícia Edivânia; Roque-Barreira, Maria Cristina
2017-01-01
The recognition of cell surface glycans by lectins may be critical for the innate and adaptive immune responses. ArtinM, a d-mannose-binding lectin from Artocarpus heterophyllus, activates antigen-presenting cells by recognizing TLR2 N-glycans and induces Th1 immunity. We recently demonstrated that ArtinM stimulated CD4+ T cells to produce proinflammatory cytokines. Here, we further studied the effects of ArtinM on adaptive immune cells. We showed that ArtinM activates murine CD4+ and CD8+ T cells, augmenting their positivity for CD25, CD69, and CD95 and showed higher interleukin (IL)-2 and interferon (IFN)-γ production. The CD4+ T cells exhibited increased T-bet expression in response to ArtinM, and IL-2 production by CD4+ and CD8+ T cells depended on the recognition of CD3εγ-chain glycans by ArtinM. The ArtinM effect on aberrantly-glycosylated neoplastic lymphocytes was studied in Jurkat T cells, in which ArtinM induced IL-2, IFN-γ, and IL-1β production, but decreased cell viability and growth. A higher frequency of AnnexinV- and propidium iodide-stained cells demonstrated the induction of Jurkat T cells apoptosis by ArtinM, and this apoptotic response was reduced by caspases and protein tyrosine kinase inhibitors. The ArtinM effects on murine T cells corroborated with the immunomodulatory property of lectin, whereas the promotion of Jurkat T cells apoptosis may reflect a potential applicability of ArtinM in novel strategies for treating lymphocytic leukemia. PMID:28665310
Oteiza, Alexandra; Mechti, Nadir
2011-01-01
Activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway by the viral Tax oncoprotein plays a pivotal role in clonal expansion of human T-cell leukemia virus type 1 (HTLV-1)-infected cells. As the Forkhead box O (FoxO) tumor suppressors act as downstream effectors of PI3K/Akt, they represent good candidate targets whose dysregulation by Tax might be involved in HTLV-1-mediated activation and transformation of infected cells. In this report, we provide evidence showing that Tax induces a dose-dependent degradation of FoxO4 by the ubiquitin-proteasome pathway. Consistent with that, we demonstrate that Tax expression increases the interaction between FoxO4 and Mdm2 E3 ligase, leading to a strong FoxO4 polyubiquitination. These processes require the phosphorylation of FoxO4 by Akt, since a mutant of FoxO4 with mutations on its three Akt phosphorylation sites appears to be resistant to Tax-mediated degradation and ubiquitination. In addition, we show that Tax expression is associated with degradation and phosphorylation of endogenous FoxO4 in Jurkat T cells. Finally, we demonstrate that Tax represses FoxO4 transcriptional activity. Our study demonstrates that Tax can control FoxO4 protein stability and transcriptional activity and provides new insight into the subversion of cell signaling pathways during HTLV-1 infection. PMID:21525355
Hohl, Tobias M.; Collins, Nichole; Leiner, Ingrid; Gallegos, Alena; Saijo, Shinobu; Coward, Jesse W.; Iwakura, Yoichiro
2011-01-01
Pulmonary infection of mice with Aspergillus fumigatus induces concurrent T helper type 1 (Th1) and Th17 responses that depend on Toll-like receptor/MyD88 and Dectin-1, respectively. However, the mechanisms balancing Th1 and Th17 CD4 T cell populations during infection remain incompletely defined. In this study, we show that Dectin-1 deficiency disproportionally increases Th1 responses and decreases Th17 differentiation after A. fumigatus infection. Dectin-1 signaling in A. fumigatus–infected wild-type mice reduces IFN-γ and IL-12p40 expression in the lung, thereby decreasing T-bet expression in responding CD4 T cells and enhancing Th17 responses. Absence of IFN-γ or IL-12p35 in infected mice or T-bet in responding CD4 T cells enhances Th17 differentiation, independent of Dectin-1 expression, in A. fumigatus–infected mice. Transient deletion of monocyte-derived dendritic cells also reduces Th1 and boosts Th17 differentiation of A. fumigatus–specific CD4 T cells. Our findings indicate that Dectin-1–mediated signals alter CD4 T cell responses to fungal infection by decreasing the production of IL-12 and IFN-γ in innate cells, thereby decreasing T-bet expression in A. fumigatus–specific CD4 T cells and enabling Th17 differentiation. PMID:21242294
CD8+ T cells induce thyroid epithelial cell hyperplasia and fibrosis.
Yu, Shiguang; Fang, Yujiang; Sharav, Tumenjargal; Sharp, Gordon C; Braley-Mullen, Helen
2011-02-15
CD8(+) T cells can be important effector cells in autoimmune inflammation, generally because they can damage target cells by cytotoxicity. This study shows that activated CD8(+) T cells induce thyroid epithelial cell hyperplasia and proliferation and fibrosis in IFN-γ(-/-) NOD.H-2h4 SCID mice in the absence of CD4(+) T cells. Because CD8(+) T cells induce proliferation rather than cytotoxicity of target cells, these results describe a novel function for CD8(+) T cells in autoimmune disease. In contrast to the ability of purified CD8(+) T cells to induce thyrocyte proliferation, CD4(+) T cells or CD8 T cell-depleted splenocytes induced only mild thyroid lesions in SCID recipients. T cells in both spleens and thyroids highly produce TNF-α. TNF-α promotes proliferation of thyrocytes in vitro, and anti-TNF-α inhibits development of thyroid epithelial cell hyperplasia and proliferation in SCID recipients of IFN-γ(-/-) splenocytes. This suggests that targeting CD8(+) T cells and/or TNF-α may be effective for treating epithelial cell hyperplasia and fibrosis.
1996-01-01
Thymic selection of natural killer-1+ natural T cells that express alpha beta T cell receptors requires a conserved beta 2-microglobulin- associated molecule, presumably CD1d, displayed by CD4+8+ thymocytes. Here we demonstrate that positive selection of natural T cells occurs independent of transporters associated with antigen presentation-1 (TAP- 1) function. Moreover, natural T cells in TAP-1o/o mice are numerically expanded. Several H-2 class Ib molecules function in a TAP-independent manner, suggesting that if expressed in TAP-1o/o thymocytes, they could play a role in natural T cell development. Of these class Ib molecules, H-2TL is expressed by TAP-1o/o thymocytes. Moreover, we find that thymi of TL+ mice congenic or transgenic for H-2T18 also have a numerically expanded natural T cell repertoire compared with TL- mice. This expansion, as in TAP-1o/o thymi, is evident in each of the limited T cell receptor V beta chains expressed by natural T cells, suggesting that TL and CD1d impact similar repertoires. Thus TL, in addition to CD1d, plays a role in natural T cell development. PMID:8879233
Higuchi, Masaya; Takahashi, Masahiko; Tanaka, Yuetsu; Fujii, Masahiro
2014-12-01
Human T-cell leukemia virus type 1 (HTLV-1), an etiological agent of adult T-cell leukemia, immortalizes and transforms primary human T cells in vitro in both an interleukin (IL)-2-dependent and IL-2-independent manner. Expression of the HTLV-1 oncoprotein Tax transforms the growth of the mouse T-cell line CTLL-2 from being IL-2-dependent to IL-2-independent. Withdrawal of IL-2 from normal activated T cells induces apoptosis, which is mediated through the inducible expression of several proapoptotic proteins, including Bim. In this study, we found that Tax protects IL-2-depleted T cells against Bim-induced apoptosis. Withdrawal of IL-2 from CTLL-2 cells induced a prominent increase in the level of Bim protein in CTLL-2 cells, but not in Tax-transformed CTLL-2 cells. This inhibition of Bim in Tax-transformed CTLL-2 cells was mediated by two mechanisms: downregulation of Bim mRNA and posttranscriptional reduction of Bim protein. Transient expression of Tax in CTLL-2 cells also inhibited IL-2 depletion-induced expression of Bim, however, this decrease in Bim protein expression was not due to downregulation of Bim mRNA, thus indicating that Bim mRNA downregulation in Tax-transformed CTLL-2 occurs only after long-term expression of Tax. Transient expression of Tax in CTLL-2 cells also induced Erk activation, however, this was not involved in the reduction of Bim protein. Knockdown of Bim expression in CTLL-2 cells augmented Tax-induced IL-2-independent transformation. HTLV-1 infection of human T cells also reduced their levels of Bim protein, and restoring Bim expression in HTLV-1-infected cells reduced their proliferation by inducing apoptosis. Taken together, these results indicate that Tax-induced downregulation of Bim in HTLV-1-infected T cells promotes their IL-2-independent growth, thereby supporting the persistence of HTLV-1 infection in vivo. © 2014 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.
Higuchi, Masaya; Takahashi, Masahiko; Tanaka, Yuetsu; Fujii, Masahiro
2014-01-01
Human T-cell leukemia virus type 1 (HTLV-1), an etiological agent of adult T-cell leukemia, immortalizes and transforms primary human T cells in vitro in both an interleukin (IL)-2-dependent and IL-2-independent manner. Expression of the HTLV-1 oncoprotein Tax transforms the growth of the mouse T-cell line CTLL-2 from being IL-2-dependent to IL-2-independent. Withdrawal of IL-2 from normal activated T cells induces apoptosis, which is mediated through the inducible expression of several proapoptotic proteins, including Bim. In this study, we found that Tax protects IL-2-depleted T cells against Bim-induced apoptosis. Withdrawal of IL-2 from CTLL-2 cells induced a prominent increase in the level of Bim protein in CTLL-2 cells, but not in Tax-transformed CTLL-2 cells. This inhibition of Bim in Tax-transformed CTLL-2 cells was mediated by two mechanisms: downregulation of Bim mRNA and posttranscriptional reduction of Bim protein. Transient expression of Tax in CTLL-2 cells also inhibited IL-2 depletion–induced expression of Bim, however, this decrease in Bim protein expression was not due to downregulation of Bim mRNA, thus indicating that Bim mRNA downregulation in Tax-transformed CTLL-2 occurs only after long-term expression of Tax. Transient expression of Tax in CTLL-2 cells also induced Erk activation, however, this was not involved in the reduction of Bim protein. Knockdown of Bim expression in CTLL-2 cells augmented Tax-induced IL-2-independent transformation. HTLV-1 infection of human T cells also reduced their levels of Bim protein, and restoring Bim expression in HTLV-1-infected cells reduced their proliferation by inducing apoptosis. Taken together, these results indicate that Tax-induced downregulation of Bim in HTLV-1-infected T cells promotes their IL-2-independent growth, thereby supporting the persistence of HTLV-1 infection in vivo. PMID:25175936
Fantini, M C; Becker, C; Tubbe, I; Nikolaev, A; Lehr, H A; Galle, P; Neurath, M F
2006-05-01
The imbalance between effector and regulatory T cells plays a central role in the pathogenesis of inflammatory bowel diseases. In addition to the thymus, CD4+CD25+ regulatory T cells can be induced in the periphery from a population of CD25- T cells by treatment with transforming growth factor beta (TGF-beta). Here, we analysed the in vivo function of TGF-beta induced regulatory T (Ti-Treg) cells in experimental colitis. Ti-Treg cells were generated in cell culture in the presence or absence of TGF-beta and tested for their regulatory potential in experimental colitis using the CD4+CD62L+ T cell transfer model. Ti-Treg cells significantly suppressed Th1 mediated colitis on CD4+CD62L+ T cell transfer in vivo, as shown by high resolution endoscopy, histology, immunohistochemistry, and cytokine analysis. Further analysis of in vivo and in vitro expanded Ti-Treg cells showed that exogenous interleukin 2 (IL-2) was crucial for survival and expansion of these cells. Our data suggest that regulatory Ti-Treg cells expand by TGF-beta and exogenous IL-2 derived from effector T cells at the site of inflammation. In addition to Tr1 and thymic CD4+CD25+ T cells, peripheral Ti-Treg cells emerge as a class of regulatory T cells with therapeutic potential in T cell mediated chronic intestinal inflammation.
Ana o 1 and Ana o 2 cashew allergens share cross-reactive CD4+ T-cell epitopes with other tree nuts
Archila, Luis Diego; Chow, I-Ting; McGinty, John W.; Renand, Amedee; Jeong, David; Robinson, David; Farrington, Mary L.; Kwok, William.W.
2017-01-01
Background Allergies to cashew are increasing in prevalence, with clinical symptoms ranging from oral pruritus to fatal anaphylactic reaction. Yet, cashew-specific T-cell epitopes and T-cell cross-reactivity amongst cashew and other tree nut allergens in humans remain uncharacterized. Objectives In this study, we characterized cashew specific T-cell responses in cashew allergic subjects and examined cross-reactivity of these cashew specific cells toward other tree nut allergens. Methods CD154 up-regulation assay was used to determine immunodominance hierarchy among cashew major allergens at the T cell level. The phenotype, magnitude and functionality of cashew-specific T-cells was determined by utilizing ex vivo staining with MHC class II tetramers. Dual tetramer staining and proliferation experiments were used to determine cross-reactivity to other tree nuts. Results CD4+ T-cell responses were directed towards cashew allergens Ana o 1 and Ana o 2. Multiple Ana o 1 and Ana o 2 T-cell epitopes were then identified. These epitopes elicited either TH2 or TH2/TH17 responses in allergic subjects, which were either cashew unique epitope or cross-reactive epitopes. For clones that recognized the cross-reactive epitope, T-cell clones responded robustly to cashew, hazelnut and/or pistachio but not to walnut. Conclusions Phylogenetically diverse tree nut allergens can activate cashew reactive T-cells and elicit a TH2 type response at an epitope specific level. Clinical relevance Lack of cross-reactivity between walnut and cashew suggest that cashew peptide immunotherapy approach may not be most effective for walnut. PMID:27129138
Archila, L D; Chow, I-T; McGinty, J W; Renand, A; Jeong, D; Robinson, D; Farrington, M L; Kwok, W W
2016-06-01
Allergies to cashew are increasing in prevalence, with clinical symptoms ranging from oral pruritus to fatal anaphylactic reaction. Yet, cashew-specific T cell epitopes and T cell cross-reactivity amongst cashew and other tree nut allergens in humans remain uncharacterized. In this study, we characterized cashew-specific T cell responses in cashew-allergic subjects and examined cross-reactivity of these cashew-specific cells towards other tree nut allergens. CD154 up-regulation assay was used to determine immunodominance hierarchy among cashew major allergens at the T cell level. The phenotype, magnitude and functionality of cashew-specific T cells were determined by utilizing ex vivo staining with MHC class II tetramers. Dual tetramer staining and proliferation experiments were used to determine cross-reactivity to other tree nuts. CD4(+) T cell responses were directed towards cashew allergens Ana o 1 and Ana o 2. Multiple Ana o 1 and Ana o 2 T cell epitopes were then identified. These epitopes elicited either TH 2 or TH 2/TH 17 responses in allergic subjects, which were either cashew unique epitope or cross-reactive epitopes. For clones that recognized the cross-reactive epitope, T cell clones responded robustly to cashew, hazelnut and/or pistachio but not to walnut. Phylogenetically diverse tree nut allergens can activate cashew-reactive T cells and elicit a TH 2-type response at an epitope-specific level. Lack of cross-reactivity between walnut and cashew suggests that cashew peptide immunotherapy approach may not be most effective for walnut. © 2016 John Wiley & Sons Ltd.
Kido, Masahiro; Watanabe, Norihiko; Okazaki, Taku; Akamatsu, Takuji; Tanaka, Junya; Saga, Kazuyuki; Nishio, Akiyoshi; Honjo, Tasuku; Chiba, Tsutomu
2008-10-01
Because of the lack of animal models developing spontaneous autoimmune hepatitis (AIH), the molecular mechanisms involved in the development of AIH are still unclear. This study aims to examine the regulatory roles of naturally arising CD4(+)CD25(+) regulatory T (Treg) cells and programmed cell death 1 (PD-1)-mediated signaling in the development of AIH. To induce a concurrent loss of Treg cells and PD-1-mediated signaling, neonatal thymectomy (NTx), which severely reduces the number of Treg cells, was performed on PD-1(-/-) mice. After the NTx, we performed histologic examination, assessed autoantibody production and infiltrating cells in the liver, and conducted adoptive transfer experiments. In contrast to NTx mice and PD-1(-/-) mice, NTx-PD-1(-/-) mice produced antinuclear antibodies and developed fatal hepatitis characterized by a CD4(+) and CD8(+) T-cell infiltration invading the parenchyma with massive lobular necrosis. Induction of AIH in NTx-PD-1(-/-) mice was suppressed by transfer of Treg cells, even derived from PD-1(-/-) mice. Transfer of total but not CD4(+) T-cell-depleted splenocytes from NTx-PD-1(-/-) mice into RAG2(-/-) mice induced the development of severe hepatitis. In contrast, the transfer of CD8(+) T-cell-depleted splenocytes triggered only mononuclear infiltrates without massive necrosis of the parenchyma. NTx-PD-1(-/-) mice are the first mouse model of spontaneous fatal AIH. The concurrent loss of Treg cells and PD-1-mediated signaling can induce the development of fatal AIH. Autoreactive CD4(+) T cells are essential for induction of AIH, whereas CD8(+) T cells play an important role in progression to fatal hepatic damage.
Li, Jinbo; Wang, Qian; Yang, Renlei; Zhang, Jiaqi; Li, Xing; Zhou, Xichao; Miao, Dengshun
2017-05-01
Previous studies have shown that estrogen regulates bone homeostasis through regulatory effects on oxidative stress. However, it is unclear how estrogen deficiency triggers reactive oxygen species (ROS) accumulation. Recent studies provide evidence that the B lymphoma Mo-MLV insertion region 1 (BMI-1) plays a critical role in protection against oxidative stress and that this gene is directly regulated by estrogen via estrogen receptor (ER) at the transcriptional level. In this study, ovariectomized mice were given drinking water with/without antioxidant N-acetyl-cysteine (NAC, 1 mg/mL) supplementation, and compared with each other and with sham mice. Results showed that ovariectomy resulted in bone loss with increased osteoclast surface, increased ROS levels, T cell activation, and increased TNF and RANKL levels in serum and in CD4 T cells; NAC supplementation largely prevented these alterations. BMI-1 expression levels were dramatically downregulated in CD4 T cells from ovariectomized mice. We supplemented drinking water to BMI-1-deficient mice with/without NAC and compared them with each other and with wild-type (WT) mice. We found that BMI-1 deficiency mimicked alterations observed in ovariectomy whereas NAC supplementation reversed all alterations induced by BMI-1 deficiency. Because T cells are critical in mediating ovariectomy-induced bone loss, we further assessed whether BMI-1 overexpression in lymphocytes can protect against estrogen deficiency-induced osteoclastogenesis and bone loss by inhibiting oxidative stress, T cell activation, and RANKL production. When WT and Eμ-BMI-1 transgenic mice with BMI-1 specifically overexpressed in lymphocytes were ovariectomized and compared with each other and with WT sham mice, we found that BMI-1 overexpression in lymphocytes clearly reversed all alterations induced by ovariectomy. Results from this study indicate that estrogen deficiency downregulates BMI-1 and subsequently increases ROS, T cell activation, and
2013-01-01
Background Human T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia (ATL). Treatment options are limited and prophylactic agents are not available. We have previously demonstrated an essential role for CREB-regulating transcriptional coactivators (CRTCs) in HTLV-1 transcription. Results In this study we report on the negative regulatory role of LKB1 tumor suppressor and salt-inducible kinases (SIKs) in the activation of HTLV-1 long terminal repeats (LTR) by the oncoprotein Tax. Activation of LKB1 and SIKs effectively blunted Tax activity in a phosphorylation-dependent manner, whereas compromising these kinases, but not AMP-dependent protein kinases, augmented Tax function. Activated LKB1 and SIKs associated with Tax and suppressed Tax-induced LTR activation by counteracting CRTCs and CREB. Enforced expression of LKB1 or SIK1 in cells transfected with HTLV-1 molecular clone pX1MT repressed proviral transcription. On the contrary, depletion of LKB1 in pX1MT-transfected cells and in HTLV-1-transformed T cells boosted the expression of Tax. Treatment of HTLV-1 transformed cells with metformin led to LKB1/SIK1 activation, reduction in Tax expression, and inhibition of cell proliferation. Conclusions Our findings revealed a new function of LKB1 and SIKs as negative regulators of HTLV-1 transcription. Pharmaceutical activation of LKB1 and SIKs might be considered as a new strategy in anti-HTLV-1 and anti-ATL therapy. PMID:23577667
T cell priming versus T cell tolerance induced by synthetic peptides
1995-01-01
It is well known that synthetic peptides are able to both induce and tolerize T cells. We have examined the parameters leading either to priming or tolerance of CD8+ cytotoxic T lymphocytes (CTL) in vivo with a major histocompatibility complex class I (H-2 Db) binding peptide derived from the glycoprotein (GP aa33-41) of lymphocytic choriomeningitis virus (LCMV). By varying dose, route, and frequency of LCMV GP peptide application, we found that a single local subcutaneous injection of 50-500 micrograms peptide emulsified in incomplete Freund's adjuvant protected mice against LCMV infection, whereas repetitive and systemic intraperitoneal application of the same dose caused tolerance of LCMV-specific CTL. The peptide-induced tolerance was transient in euthymic mice but permanent in thymectomized mice. These findings are relevant for a selective use of peptides as a therapeutic approach: peptide-induced priming of T cells for vaccination and peptide-mediated T cell tolerance for intervention in immunopathologies and autoimmune diseases. PMID:7540654
Climent, Núria; Guerra, Susana; García, Felipe; Rovira, Cristina; Miralles, Laia; Gómez, Carmen Elena; Piqué, Núria; Gil, Cristina; Gatell, José María; Esteban, Mariano; Gallart, Teresa
2011-01-01
Currently, MVA virus vectors carrying HIV-1 genes are being developed as HIV-1/AIDS prophylactic/therapeutic vaccines. Nevertheless, little is known about the impact of these vectors on human dendritic cells (DC) and their capacity to present HIV-1 antigens to human HIV-specific T cells. This study aimed to characterize the interaction of MVA and MVA expressing the HIV-1 genes Env-Gag-Pol-Nef of clade B (referred to as MVA-B) in human monocyte-derived dendritic cells (MDDC) and the subsequent processes of HIV-1 antigen presentation and activation of memory HIV-1-specific T lymphocytes. For these purposes, we performed ex vivo assays with MDDC and autologous lymphocytes from asymptomatic HIV-infected patients. Infection of MDDC with MVA-B or MVA, at the optimal dose of 0.3 PFU/MDDC, induced by itself a moderate degree of maturation of MDDC, involving secretion of cytokines and chemokines (IL1-ra, IL-7, TNF-α, IL-6, IL-12, IL-15, IL-8, MCP-1, MIP-1α, MIP-1β, RANTES, IP-10, MIG, and IFN-α). MDDC infected with MVA or MVA-B and following a period of 48 h or 72 h of maturation were able to migrate toward CCL19 or CCL21 chemokine gradients. MVA-B infection induced apoptosis of the infected cells and the resulting apoptotic bodies were engulfed by the uninfected MDDC, which cross-presented HIV-1 antigens to autologous CD8+ T lymphocytes. MVA-B-infected MDDC co-cultured with autologous T lymphocytes induced a highly functional HIV-specific CD8+ T cell response including proliferation, secretion of IFN-γ, IL-2, TNF-α, MIP-1β, MIP-1α, RANTES and IL-6, and strong cytotoxic activity against autologous HIV-1-infected CD4+ T lymphocytes. These results evidence the adjuvant role of the vector itself (MVA) and support the clinical development of prophylactic and therapeutic anti-HIV vaccines based on MVA-B. PMID:21625608
Min, Arim; Lee, Young Ah; Kim, Kyeong Ah; Shin, Myeong Heon
2018-05-31
Trichomonas vaginalis is a sexually-transmitted protozoan parasite that causes vaginitis and cervicitis. Although mast cell activation is important for provoking tissue inflammation during infection with parasites, information regarding the signaling mechanisms in mast cell activation and T. vaginalis infection is limited. O-linked N-acetylglucosamine (O-GlcNAc) is a post-translational modification of serine and threonine residues that functions as a critical regulator of intracellular signaling, regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). We investigated if O-GlcNAcylation was associated with mast cell activation induced by T. vaginalis-derived secretory products (TvSP). Modified TvSP collected from live trichomonads treated with the 5-lipooxygenase inhibitor AA861 inhibited migration of mast cells. This result suggested that mast cell migration was caused by stimulation of T. vaginalis-secreted leukotrienes. Using the BLT1 antagonist U75302 or BLT1 siRNA, we found that migration of mast cells was evoked via LTB 4 receptor (BLT1). Furthermore, TvSP induced protein O-GlcNAcylation and OGT expression in HMC-1 cells, which was prevented by transfection with BLT1 siRNA. TvSP-induced migration, ROS generation, CD63 expression and IL-8 release were significantly suppressed by pretreatmemnt with OGT inhibitor ST045849 or OGT siRNA. These results suggested that BLT1-mediated OGlcNAcylation was important for mast cell activation during trichomoniasis. Copyright © 2018. Published by Elsevier Masson SAS.
Phosphoinositide 3–kinase γ participates in T cell receptor–induced T cell activation
Alcázar, Isabela; Marqués, Miriam; Kumar, Amit; Hirsch, Emilio; Wymann, Matthias; Carrera, Ana C.; Barber, Domingo F.
2007-01-01
Class I phosphoinositide 3–kinases (PI3Ks) constitute a family of enzymes that generates 3-phosphorylated polyphosphoinositides at the cell membrane after stimulation of protein tyrosine (Tyr) kinase–associated receptors or G protein–coupled receptors (GPCRs). The class I PI3Ks are divided into two types: class IA p85/p110 heterodimers, which are activated by Tyr kinases, and the class IB p110γ isoform, which is activated by GPCR. Although the T cell receptor (TCR) is a protein Tyr kinase–associated receptor, p110γ deletion affects TCR-induced T cell stimulation. We examined whether the TCR activates p110γ, as well as the consequences of interfering with p110γ expression or function for T cell activation. We found that after TCR ligation, p110γ interacts with Gαq/11, lymphocyte-specific Tyr kinase, and ζ-associated protein. TCR stimulation activates p110γ, which affects 3-phosphorylated polyphosphoinositide levels at the immunological synapse. We show that TCR-stimulated p110γ controls RAS-related C3 botulinum substrate 1 activity, F-actin polarization, and the interaction between T cells and antigen-presenting cells, illustrating a crucial role for p110γ in TCR-induced T cell activation. PMID:17998387
NF-κB signaling mechanisms in HTLV-1-induced adult T-cell leukemia/lymphoma.
Harhaj, Edward William; Giam, Chou-Zen
2018-05-03
The human T-cell leukemia virus type 1 (HTLV-1) is a complex deltaretrovirus linked to adult T-cell leukemia/lymphoma (ATLL), a fatal CD4+ malignancy in 3-5% of infected individuals. The HTLV-1 Tax regulatory protein plays indispensable roles in regulating viral gene expression and activating cellular signaling pathways that drive the proliferation and clonal expansion of T cells bearing HTLV-1 proviral integrations. Tax is a potent activator of NF-κB, a key signaling pathway that is essential for the survival and proliferation of HTLV-1 infected T cells. However, constitutive NF-κB activation by Tax also triggers a senescence response, suggesting the possibility that only T cells capable of overcoming NF-κB-induced senescence can selectively undergo clonal expansion after HTLV-1 infection. Tax expression is often silenced in the majority of ATLL due to genetic alterations in the tax gene or DNA hypermethylation of the 5'-LTR. Despite the loss of Tax, NF-κB activation remains persistently activated in ATLL due to somatic mutations in genes in the T/B-cell receptor (T/BCR) and NF-κB signaling pathways. In this review, we focus on the key events driving Tax-dependent and independent mechanisms of NF-κB activation during the multi-step process leading to ATLL. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
Marampon, F; Gravina, G L; Festuccia, C; Popov, V M; Colapietro, E A; Sanità, P; Musio, D; De Felice, F; Lenzi, A; Jannini, E A; Di Cesare, E; Tombolini, V
2016-04-01
Radiotherapy toxicity is related to oxidative stress-mediated endothelial dysfunction. Here, we investigated on radioprotective properties of Vitamin D (Vit.D) on human endothelial cells (HUVEC). HUVEC, pre-treated with Vit.D, were exposed to ionizing radiation (IR): ROS production, cellular viability, apoptosis, senescence and western blot for protein detection were performed. The role of MAPKs pathway was investigated by using U0126 (10 μM) MEKs/ERKs-, SB203580 (2.5 μM) p38-inhibitor or by over/expressing MKK6 p38-upstream activator. Vit.D reduced IR-induced ROS production protecting proliferating and quiescent HUVEC from cellular apoptosis or senescence, respectively, by regulating MAPKs pathways. In proliferating HUVEC, Vit.D prevented IR-induced apoptosis by activating ERKs while in quiescent HUVEC counteracted IR-induced senescence by inhibiting the p38-IR-induced activation. MEKs&ERKs inhibition in proliferating or MKK6/mediated p38 activation in quiescent HUVEC, respectively, reverted anti-apoptotic or anti-senescent Vit.D properties. SirT1 protein expression levels were up-regulated by Vit.D. ERKs inhibition blocked Vit.D-induced SirT1 protein up-regulation in proliferating cells. In quiescent HUVEC cells, p38 inhibition counteracted the IR-induced SirT1 protein down-regulation, while MKK6 transfection abrogated the Vit.D positive effects on SirT1 protein levels after irradiation. SirT1 inhibition by sirtinol blocked the Vit.D radioprotective effects. Vit.D protects HUVEC from IR induced/oxidative stress by positively regulating the MAPKs/SirT1 axis.
Tax, W J; Hermes, F F; Willems, R W; Capel, P J; Koene, R A
1984-09-01
In previous studies, it was shown that there is polymorphism in the mitogenic effect of mouse IgG1 monoclonal antibodies against the T3 antigen of human T cells. This polymorphism implies that IgG1 anti-T3 antibodies are not mitogenic for T cells from 30% of healthy individuals. The present results demonstrate that this polymorphism is caused by polymorphism of an Fc receptor for mouse IgG1, present on human monocytes. The Fc receptor for murine IgG1 could be detected by a newly developed rosetting assay on monocytes from all individuals responsive to the mitogenic effect of IgG1 anti-T3 antibodies. This Fc receptor was not detectable on monocytes from those individuals exhibiting no mitogenic responses to IgG1 anti-T3 monoclonal antibodies. Cross-linking of T3 antigens appears to be essential for antibody-induced mitosis of T cells, because mononuclear cells that did not proliferate in response to WT 31 (an IgG1 antibody against T3 antigen) showed a proliferative response to Sepharose beads coated with WT 31. The Fc receptor--if functionally present--may be involved in the cross-linking of T3 antigens through anti-T3 antibodies. Further evidence for the involvement of this Fc receptor in antibody-induced T cell proliferation was provided by inhibition studies. Immune complexes containing IgG1 antibodies were able to inhibit the proliferative response to IgG1 anti-T3 antibodies. This inhibition by immune complexes appears to be mediated through the monocyte Fc receptor for mouse IgG1. These findings are important for the interpretation of previously described inhibitory effects of anti-T cell monoclonal antibodies on T cell proliferation, and show that such inhibitory effects may be monocyte-mediated (via immune complexes) rather than caused by a direct involvement of the respective T cell antigens in T cell mitosis. The Fc receptor for mouse IgG1 plays a role in antibody-induced T cell proliferation. Its polymorphism may have important implications for the
T-cell receptor transfer for boosting HIV-1-specific T-cell immunity in HIV-1-infected patients.
Mummert, Christiane; Hofmann, Christian; Hückelhoven, Angela G; Bergmann, Silke; Mueller-Schmucker, Sandra M; Harrer, Ellen G; Dörrie, Jan; Schaft, Niels; Harrer, Thomas
2016-09-10
Strategies to cure HIV-1 infection require the eradication of viral reservoirs. An innovative approach for boosting the cytotoxic T-lymphocyte response is the transfer of T-cell receptors (TCRs). Previously, we have shown that electroporation of TCR-encoding mRNA is able to reprogram CD8 T cells derived from healthy donors. So far, it is unknown whether the transfer of HIV-1-specific TCRs is capable to reprogram CD8 T cells of HIV-1-infected patients. To assess the efficiency of TCR-transfer by mRNA electroporation and the functionality of reprogramed T cells in HIV-1-infected patients, we performed an in-vitro analysis of TCR-transfer into T cells from HIV-1-infected patients in various stages of disease and from healthy controls. Peripheral blood mononuclear cells from 16 HIV-1-infected patients (nine HLA-A02-positive, seven HLA-A02-negative) and from five healthy controls were electroporated with mRNA-constructs encoding TCRs specific for the HLA-A02/HIV-1-gag p17 epitope SLYNTVATL (SL9). Functionality of the TCRs was measured by γIFN-ELISpot assays. SL9/TCR transfection into peripheral blood mononuclear cells from both HLA-A02-positive and HLA-A02-negative HIV-1-infected patients and from healthy blood donors reprogramed T cells for recognition of SL9-presenting HLA-A02-positive cells in γIFN-ELISpot assays. SL9/TCR-transfer into T cells from an immunodeficient AIDS patient could induce recognition of SL9-expressing target cells only after reversion of T-cell dysfunction by antiretroviral therapy. The transfer of HIV-1-p17-specific TCRs into T cells is functional both in HIV-1-infected patients as well as in healthy blood donors. TCR-transfer is a promising method to boost the immune system against HIV-1.
Li, Wen; Green, William R
2011-12-01
LP-BM5 retrovirus induces a complex disease featuring an acquired immunodeficiency syndrome termed murine AIDS (MAIDS) in susceptible strains of mice, such as C57BL/6 (B6). CD4 T helper effector cells are required for MAIDS induction and progression of viral pathogenesis. CD8 T cells are not needed for viral pathogenesis, but rather, are essential for protection from disease in resistant strains, such as BALB/c. We have discovered an immunodominant cytolytic T lymphocyte (CTL) epitope encoded in a previously unrecognized LP-BM5 retroviral alternative (+1 nucleotide [nt]) gag translational open reading frame. CTLs specific for this cryptic gag epitope are the basis of protection from LP-BM5-induced immunodeficiency in BALB/c mice, and the inability of B6 mice to mount an anti-gag CTL response appears critical to the initiation and progression of LP-BM5-induced MAIDS. However, uninfected B6 mice primed by LP-BM5-induced tumors can generate CTL responses to an LP-BM5 retrovirus infection-associated epitope(s) that is especially prevalent on such MAIDS tumor cells, indicating the potential to mount a protective CD8 T-cell response. Here, we utilized this LP-BM5 retrovirus-induced disease system to test whether modulation of normal immune down-regulatory mechanisms can alter retroviral pathogenesis. Thus, following in vivo depletion of CD4 T regulatory (Treg) cells and/or selective interruption of PD-1 negative signaling in the CD8 T-cell compartment, retroviral pathogenesis was significantly decreased, with the combined treatment of CD4 Treg cell depletion and PD-1 blockade working in a synergistic fashion to substantially reduce the induction of MAIDS.
Newton, Darren J; Andrew, Elizabeth M; Dalton, Jane E; Mears, Rainy; Carding, Simon R
2006-02-01
Although gammadelta T cells are a common feature of many pathogen-induced immune responses, the factors that influence, promote, or regulate the response of individual gammadelta T-cell subsets to infection is unknown. Here we show that in the absence of Vgamma1+ T cells, novel subsets of gammadelta T cells, expressing T-cell receptor (TCR)-Vgamma chains that normally define TCRgammadelta+ dendritic epidermal T cells (DETCs) (Vgamma5+), intestinal intraepithelial lymphocytes (iIELs) (Vgamma7+), and lymphocytes associated with the vaginal epithelia (Vgamma6+), are recruited to the spleen in response to bacterial infection in TCR-Vgamma1-/- mice. By comparison of phenotype and structure of TCR-Vgamma chains and/or -Vdelta chains expressed by these novel subsets with those of their epithelium-associated counterparts, the Vgamma6+ T cells elicited in infected Vgamma1-/- mice were shown to be identical to those found in the reproductive tract, from where they are presumably recruited in the absence of Vgamma1+ T cells. By contrast, Vgamma5+ and Vgamma7+ T cells found in infected Vgamma1-/- mice were distinct from Vgamma5+ DETCs and Vgamma7+ iIELs. Functional analyses of the novel gammadelta T-cell subsets identified for infected Vgamma1-/- mice showed that whereas the Vgamma5+ and Vgamma7+ subsets may compensate for the absence of Vgamma1+ T cells by producing similar cytokines, they do not possess cytocidal activity and they cannot replace the macrophage homeostasis function of Vgamma1+ T cells. Collectively, these findings identify novel subsets of gammadelta T cells, the recruitment and activity of which is under the control of Vgamma1+ T cells.
Involvement of PINK1/parkin-mediated mitophagy in ZnO nanoparticle-induced toxicity in BV-2 cells.
Wei, Limin; Wang, Jianfeng; Chen, Aijie; Liu, Jia; Feng, Xiaoli; Shao, Longquan
2017-01-01
With the increasing application of zinc oxide nanoparticles (ZnO NPs) in biological materials, the neurotoxicity caused by these particles has raised serious concerns. However, the underlying molecular mechanisms of the toxic effect of ZnO NPs on brain cells remain unclear. Mitochondrial damage has been reported to be a factor in the toxicity of ZnO NPs. PINK1/parkin-mediated mitophagy is a newly emerging additional function of autophagy that selectively degrades impaired mitochondria. Here, a PINK1 gene knockdown BV-2 cell model was established to determine whether PINK1/parkin-mediated mitophagy was involved in ZnO NP-induced toxicity in BV-2 cells. The expression of total parkin, mito-parkin, cyto-parkin, and PINK1 both in wild type and PINK1 -/- BV-2 cells was evaluated using Western blot analysis after the cells were exposed to 10 μg/mL of 50 nm ZnO NPs for 2, 4, 8, 12, and 24 h. The findings suggested that the downregulation of PINK1 resulted in a significant reduction in the survival rate after ZnO NP exposure compared with that of control cells. ZnO NPs were found to induce the transportation of parkin from the cytoplasm to the mitochondria, implying the involvement of mitophagy in ZnO NP-induced toxicity. The deletion of the PINK1 gene inhibited the recruitment of parkin to the mitochondria, causing failure of the cell to trigger mitophagy. The present study demonstrated that apart from autophagy, PINK1/parkin-mediated mitophagy plays a protective role in ZnO NP-induced cytotoxicity.
Involvement of PINK1/parkin-mediated mitophagy in ZnO nanoparticle-induced toxicity in BV-2 cells
Wei, Limin; Wang, Jianfeng; Chen, Aijie; Liu, Jia; Feng, Xiaoli; Shao, Longquan
2017-01-01
With the increasing application of zinc oxide nanoparticles (ZnO NPs) in biological materials, the neurotoxicity caused by these particles has raised serious concerns. However, the underlying molecular mechanisms of the toxic effect of ZnO NPs on brain cells remain unclear. Mitochondrial damage has been reported to be a factor in the toxicity of ZnO NPs. PINK1/parkin-mediated mitophagy is a newly emerging additional function of autophagy that selectively degrades impaired mitochondria. Here, a PINK1 gene knockdown BV-2 cell model was established to determine whether PINK1/parkin-mediated mitophagy was involved in ZnO NP-induced toxicity in BV-2 cells. The expression of total parkin, mito-parkin, cyto-parkin, and PINK1 both in wild type and PINK1−/− BV-2 cells was evaluated using Western blot analysis after the cells were exposed to 10 μg/mL of 50 nm ZnO NPs for 2, 4, 8, 12, and 24 h. The findings suggested that the downregulation of PINK1 resulted in a significant reduction in the survival rate after ZnO NP exposure compared with that of control cells. ZnO NPs were found to induce the transportation of parkin from the cytoplasm to the mitochondria, implying the involvement of mitophagy in ZnO NP-induced toxicity. The deletion of the PINK1 gene inhibited the recruitment of parkin to the mitochondria, causing failure of the cell to trigger mitophagy. The present study demonstrated that apart from autophagy, PINK1/parkin-mediated mitophagy plays a protective role in ZnO NP-induced cytotoxicity. PMID:28331313
Virion encapsidated HIV-1 Vpr induces NFAT to prime non-activated T cells for productive infection
Höhne, Kristin; Businger, Ramona; van Nuffel, Anouk; Bolduan, Sebastian; Koppensteiner, Herwig; Baeyens, Ann; Vermeire, Jolien; Malatinkova, Eva; Verhasselt, Bruno; Schindler, Michael
2016-01-01
The majority of T cells encountered by HIV-1 are non-activated and do not readily allow productive infection. HIV-1 Vpr is highly abundant in progeny virions, and induces signalling and HIV-1 LTR transcription. We hence hypothesized that Vpr might be a determinant of non-activated T-cell infection. Virion-delivered Vpr activated nuclear factor of activated T cells (NFAT) through Ca2+ influx and interference with the NFAT export kinase GSK3β. This leads to NFAT translocation and accumulation within the nucleus and was required for productive infection of unstimulated primary CD4+ T cells. A mutagenesis approach revealed correlation of Vpr-mediated NFAT activation with its ability to enhance LTR transcription and mediate cell cycle arrest. Upon NFAT inhibition, Vpr did not augment resting T-cell infection, and showed reduced G2/M arrest and LTR transactivation. Altogether, Vpr renders unstimulated T cells more permissive for productive HIV-1 infection and stimulates activation of productively infected as well as virus-exposed T cells. Therefore, it could be involved in the establishment and reactivation of HIV-1 from viral reservoirs and might have an impact on the levels of immune activation, which are determinants of HIV-1 pathogenesis. PMID:27383627
Virion encapsidated HIV-1 Vpr induces NFAT to prime non-activated T cells for productive infection.
Höhne, Kristin; Businger, Ramona; van Nuffel, Anouk; Bolduan, Sebastian; Koppensteiner, Herwig; Baeyens, Ann; Vermeire, Jolien; Malatinkova, Eva; Verhasselt, Bruno; Schindler, Michael
2016-07-01
The majority of T cells encountered by HIV-1 are non-activated and do not readily allow productive infection. HIV-1 Vpr is highly abundant in progeny virions, and induces signalling and HIV-1 LTR transcription. We hence hypothesized that Vpr might be a determinant of non-activated T-cell infection. Virion-delivered Vpr activated nuclear factor of activated T cells (NFAT) through Ca(2+) influx and interference with the NFAT export kinase GSK3β. This leads to NFAT translocation and accumulation within the nucleus and was required for productive infection of unstimulated primary CD4(+) T cells. A mutagenesis approach revealed correlation of Vpr-mediated NFAT activation with its ability to enhance LTR transcription and mediate cell cycle arrest. Upon NFAT inhibition, Vpr did not augment resting T-cell infection, and showed reduced G2/M arrest and LTR transactivation. Altogether, Vpr renders unstimulated T cells more permissive for productive HIV-1 infection and stimulates activation of productively infected as well as virus-exposed T cells. Therefore, it could be involved in the establishment and reactivation of HIV-1 from viral reservoirs and might have an impact on the levels of immune activation, which are determinants of HIV-1 pathogenesis. © 2016 The Authors.
Tanaka, Yuetsu; Takahashi, Yoshiaki; Tanaka, Reiko; Kodama, Akira; Fujii, Hideki; Hasegawa, Atsuhiko; Kannagi, Mari; Ansari, Aftab A; Saito, Mineki
2014-06-01
Human T cell leukemia virus type-1 (HTLV-1) is prevalent worldwide with foci of high prevalence. However, to date no effective vaccine or drug against HTLV-1 infection has been developed. In efforts to define the role of antibodies in the control of HTLV-1 infection, we capitalized on the use of our previously defined anti-gp46 neutralizing monoclonal antibody (mAb) (clone LAT-27) and high titers of human anti-HTLV-1 IgG purified from HAM/TSP patients (HAM-IgG). LAT-27 and HAM-IgG completely blocked syncytium formation and T cell immortalization mediated by HTLV-1 in vitro. The addition of these antibodies to cultures of CD8(+) T cell-depleted peripheral blood mononuclear cells (PBMCs) from HAM/TSP patients at the initiation of culture not only decreased the numbers of Tax-expressing cells and the production of HTLV-1 p24 but also inhibited the spontaneous immortalization of T cells. Coculture of in vitro-HTLV-1-immortalized T cell lines with autologous PBMCs in the presence of LAT-27 or HAM-IgG, but not an F(ab')2 fragment of LAT-27 or nonneutralizing anti-gp46 mAbs, resulted in depletion of HTLV-1-infected cells. A 24-h (51)Cr release assay showed the presence of significant antibody-dependent cellular cytotoxicity (ADCC) activity in LAT-27 and HAM-IgG, but not F(ab')2 of LAT-27, resulting in the depletion of HTLV-1-infected T cells by autologous PBMCs. The depletion of natural killer (NK) cells from the effector PBMCs reduced this ADCC activity. Altogether, the present data demonstrate that the neutralizing and ADCC-inducing activities of anti-HTLV-1 antibodies are capable of reducing infection and eliminating HTLV-1-infected cells in the presence of autologous PBMCs.
Newton, Jared M; Flores-Arredondo, Jose H; Suki, Sarah; Ware, Matthew J; Krzykawska-Serda, Martyna; Agha, Mahdi; Law, Justin J; Sikora, Andrew G; Curley, Steven A; Corr, Stuart J
2018-02-22
Previous work using non-invasive radiofrequency field treatment (RFT) in cancer has demonstrated its therapeutic potential as it can increase intratumoral blood perfusion, localization of intravenously delivered drugs, and promote a hyperthermic intratumoral state. Despite the well-known immunologic benefits that febrile hyperthermia can induce, an investigation of how RFT could modulate the intra-tumoral immune microenvironment had not been studied. Thus, using an established 4T1 breast cancer model in immune competent mice, we demonstrate that RFT induces a transient, localized, and T-cell dependent intratumoral inflammatory response. More specifically we show that multi- and singlet-dose RFT promote an increase in tumor volume in immune competent Balb/c mice, which does not occur in athymic nude models. Further leukocyte subset analysis at 24, 48, and 120 hours after a single RFT show a rapid increase in tumoral trafficking of CD4+ and CD8+ T-cells 24 hours post-treatment. Additional serum cytokine analysis reveals an increase in numerous pro-inflammatory cytokines and chemokines associated with enhanced T-cell trafficking. Overall, these data demonstrate that non-invasive RFT could be an effective immunomodulatory strategy in solid tumors, especially for enhancing the tumoral trafficking of lymphocytes, which is currently a major hindrance of numerous cancer immunotherapeutic strategies.
Fisetin induces Sirt1 expression while inhibiting early adipogenesis in 3T3-L1 cells.
Kim, Sang Chon; Kim, Yoo Hoon; Son, Sung Wook; Moon, Eun-Yi; Pyo, Suhkneung; Um, Sung Hee
2015-11-27
Fisetin (3,7,3',4'-tetrahydroxyflavone) is a naturally found flavonol in many fruits and vegetables and is known to have anti-aging, anti-cancer and anti-viral effects. However, the effects of fisetin on early adipocyte differentiation and the epigenetic regulator controlling adipogenic transcription factors remain unclear. Here, we show that fisetin inhibits lipid accumulation and suppresses the expression of PPARγ in 3T3-L1 cells. Fisetin suppressed early stages of preadipocyte differentiation, and induced expression of Sirt1. Depletion of Sirt1 abolished the inhibitory effects of fisetin on intracellular lipid accumulation and on PPARγ expression. Mechanistically, fisetin facilitated Sirt1-mediated deacetylation of PPARγ and FoxO1, and enhanced the association of Sirt1 with the PPARγ promoter, leading to suppression of PPARγ transcriptional activity, thereby repressing adipogenesis. Lowering Sirt1 levels reversed the effects of fisetin on deacetylation of PPARγ and increased PPARγ transactivation. Collectively, our results suggest the effects of fisetin in increasing Sirt1 expression and in epigenetic control of early adipogenesis. Copyright © 2015 Elsevier Inc. All rights reserved.
NASA Technical Reports Server (NTRS)
Yamaguchi, T.; Chattopadhyay, N.; Kifor, O.; Butters, R. R. Jr; Sugimoto, T.; Brown, E. M.; O'Malley, B. W. (Principal Investigator)
1998-01-01
The calcium-sensing receptor (CaR) is a G protein-coupled receptor that plays key roles in extracellular calcium ion (Ca2+o) homeostasis in parathyroid gland and kidney. Osteoblasts appear at sites of osteoclastic bone resorption during bone remodeling in the "reversal" phase following osteoclastic resorption and preceding bone formation. Bone resorption produces substantial local increases in Ca2+o that could provide a signal for osteoblasts in the vicinity, leading us to determine whether such osteoblasts express the CaR. In this study, we used the mouse osteoblastic, clonal cell line MC3T3-E1. Both immunocytochemistry and Western blot analysis, using an antiserum specific for the CaR, detected CaR protein in MC3T3-E1 cells. We also identified CaR transcripts in MC3T3-E1 cells by Northern analysis using a CaR-specific riboprobe and by reverse transcription-polymerase chain reaction with CaR-specific primers, followed by nucleotide sequencing of the amplified products. Exposure of MC3T3-E1 cells to high Ca2+o (up to 4.8 mM) or the polycationic CaR agonists, neomycin and gadolinium (Gd3+), stimulated both chemotaxis and DNA synthesis in MC3T3-E1 cells. Therefore, taken together, our data strongly suggest that the osteoblastic cell line MC3T3-E1 possesses both CaR protein and mRNA very similar, if not identical, to those in parathyroid and kidney. Furthermore, the CaR in these osteoblasts could play a key role in regulating bone turnover by stimulating the proliferation and migration of such cells to sites of bone resorption as a result of local release of Ca2+o.
Song, Jia-Le; Zhao, Xin; Wang, Qiang; Zhang, Ting
2013-05-01
Reactive oxygen species (ROS)-induced pancreatic β cell death affects insulin secretion and is important in the pathogenesis of diabetes. Lagerstroemia speciosa, a traditional folk medicine, has been used for t he prevention and treatment of diabetes. However, whether Lagerstroemia speciosa has a cytoprotective effect on pancreatic β cells remains to be elucidated. The present study aimed to investigate the cytoprotective effects of hot water extracts from Lagerstroemia speciosa leaves (LWE) on 3-morpholinosydnonimine (SIN-1)-induced oxidative damage in Syrian hamster pancreatic insulinoma HIT-T15 cells. The HIT-T15 cells were first treated with SIN-1 (50 µM) for 24 h and then co-incubated with LWE for 48 h. SIN-1 significantly decreased HIT-T15 cell viability (P<0.05); however, LWE did not exert a significant cytotoxic effect and increased the viability of HIT-T15 cells in a dose‑dependent manner. To further investigate the protective effects of LWE on SIN-1‑induced oxidative stress in HIT-T15 cells, the cellular levels of ROS, lipid peroxidation and endogenous antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-px), were determined. LWE decreased the intracellular levels of ROS and lipid peroxidation, and increased the activities of antioxidant enzymes. These results suggest that LWE has a cytoprotective effect against SIN-1‑induced oxidative stress in HIT-T15 cells through the inhibition of lipid peroxidation, a decrease in ROS levels and an increase in antioxidant enzyme activity. In addition, LWE increased insulin secretion in SIN-1-treated HIT-T15 cells. Our results suggested that LWE were effective in the treatment of diabetes. Further studies are required to study the anti-diabetic molecular mechanism in a cell model.
Kozako, Tomohiro; Soeda, Shuhei; Yoshimitsu, Makoto; Arima, Naomichi; Kuroki, Ayako; Hirata, Shinya; Tanaka, Hiroaki; Imakyure, Osamu; Tone, Nanako; Honda, Shin-Ichiro; Soeda, Shinji
2016-05-01
Adult T-cell leukemia/lymphoma (ATL), an aggressive T-cell malignancy that develops after long-term infection with human T-cell leukemia virus (HTLV-1), requires new treatments. Drug repositioning, reuse of a drug previously approved for the treatment of another condition to treat ATL, offers the possibility of reduced time and risk. Among clinically available angiotensin II receptor blockers, telmisartan is well known for its unique ability to activate peroxisome proliferator-activated receptor-γ, which plays various roles in lipid metabolism, cellular differentiation, and apoptosis. Here, telmisartan reduced cell viability and enhanced apoptotic cells via caspase activation in ex vivo peripheral blood monocytes from asymptomatic HTLV-1 carriers (ACs) or via caspase-independent cell death in acute-type ATL, which has a poor prognosis. Telmisartan also induced significant growth inhibition and apoptosis in leukemia cell lines via caspase activation, whereas other angiotensin II receptor blockers did not induce cell death. Interestingly, telmisartan increased the LC3-II-enriched protein fraction, indicating autophagosome accumulation and autophagy. Thus, telmisartan simultaneously caused caspase activation and autophagy. A hypertension medication with antiproliferation effects on primary and leukemia cells is intriguing. Patients with an early diagnosis of ATL are generally monitored until the disease progresses; thus, suppression of progression from AC and indolent ATL to acute ATL is important. Our results suggest that telmisartan is highly effective against primary cells and leukemia cell lines in caspase-dependent and -independent manners, and its clinical use may suppress acute transformation and improve prognosis of patients with this mortal disease. This is the first report demonstrating a cell growth-inhibitory effect of telmisartan in fresh peripheral blood mononuclear cells from leukemia patients.
Choi, Eun-Sun; Chung, Taeho; Kim, Jun-Sung; Lee, Hakmo; Kwon, Ki Han; Cho, Nam-Pyo; Cho, Sung-Dae
2013-01-01
Mithramycin A (Mith) is an aureolic acid-type polyketide produced by various soil bacteria of the genus Streptomyces. Mith inhibits myeloid cell leukemia-1 (Mcl-1) to induce apoptosis in prostate cancer, but the molecular mechanism underlying this process has not been fully elucidated. The aim of this study was therefore to investigate the detailed molecular mechanism related to Mith-induced apoptosis in prostate cancer cells. Mith decreased the phosphorylation of mammalian target of rapamycin (mTOR) in both cell lines overexpressing phospho-mTOR compared to RWPE-1 human normal prostate epithelial cells. Mith significantly induced truncated Bid (tBid) and siRNA-mediated knock-down of Mcl-1 increased tBid protein levels. Moreover, Mith also inhibited the phosphorylation of mTOR on serine 2448 and Mcl-1, and increased tBid protein in prostate tumors in athymic nude mice bearing DU145 cells as xenografts. Thus, Mith acts as an effective tumor growth inhibitor in prostate cancer cells through the mTOR/Mcl-1/tBid signaling pathway. PMID:24062605
Cereseto, A; Diella, F; Mulloy, J C; Cara, A; Michieli, P; Grassmann, R; Franchini, G; Klotman, M E
1996-09-01
Human T-cell lymphotropic/leukemia virus type I (HTLV-I) is associated with T-cell transformation both in vivo and in vitro. Although some of the mechanisms responsible for transformation remain unknown, increasing evidence supports a direct role of viral as well as dysregulated cellular proteins in transformation. We investigated the potential role of the tumor suppressor gene p53 and of the p53-regulated gene, p21waf1/cip1 (wild-type p53 activated fragment 1/cycling dependent kinases [cdks] interacting protein 1), in HTLV-I-infected T cells. We have found that the majority of HTLV-I-infected T cells have the wild-type p53 gene. However, its function in HTLV-I-transformed cells appears to be impaired, as shown by the lack of appropriate p53-mediated responses to ionizing radiation (IR). Interestingly, the expression of the p53 inducible gene, p21waf1/cip1, is elevated at the messenger ribonucleic acid and protein levels in all HTLV-I-infected T-cell lines examined as well as in Taxl-1, a human T-cell line stably expressing Tax. Additionally, Tax induces upregulation of a p21waf1/cip1 promoter-driven luciferase gene in p53 null cells, and increases p21waf1/cip1 expression in Jurkat T cells. These findings suggest that the Tax protein is at least partially responsible for the p53-independent expression of p21waf1/cip1 in HTLV-I-infected cells. Dysregulation of p53 and p21waf1/cip1 proteins regulating cell-cycle progression, may represent an important step in HTLV-I-induced T-cell transformation.
Xue, Chunxue; Wen, Mingjie; Bao, Linlin; Li, Hui; Li, Fengdi; Liu, Meng; Lv, Qi; An, Yunqing; Zhang, Xulong; Cao, Bin
2017-01-01
The influenza A (H1N1) pdm09 virus remains a critical global health concern and causes high levels of morbidity and mortality. Severe acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are the major outcomes among severely infected patients. Our previous study found that interleukin (IL)-17A production by humans or mice infected with influenza A (H1N1) pdm09 substantially contributes to ALI and subsequent morbidity and mortality. However, the cell types responsible for IL-17A production during the early stage of severe influenza A (H1N1) pdm09 infection remained unknown. In this study, a mouse model of severe influenza A (H1N1) pdm09 infection was established. Our results show that, in the lungs of infected mice, the percentage of γδT cells, but not the percentages of CD4 + Th and CD8 + Tc cells, gradually increased and peaked at 3 days post-infection (dpi). Further analysis revealed that the Vγ4 + γδT subset, but not the Vγ1 + γδT subset, was significantly increased among the γδT cells. At 3 dpi, the virus induced significant increases in IL-17A in the bronchoalveolar lavage fluid (BALF) and serum. IL-17A was predominantly secreted by γδT cells (especially the Vγ4 + γδT subset), but not CD4 + Th and CD8 + Tc cells at the early stage of infection, and IL-1β and/or IL-23 were sufficient to induce IL-17A production by γδT cells. In addition to secreting IL-17A, γδT cells secreted interferon (IFN)-γ and expressed both an activation-associated molecule, natural killer group 2, member D (NKG2D), and an apoptosis-associated molecule, FasL. Depletion of γδT cells or the Vγ4 + γδT subset significantly rescued the virus-induced weight loss and improved the survival rate by decreasing IL-17A secretion and reducing immunopathological injury. This study demonstrated that, by secreting IL-17A, lung Vγ4 + γδT cells, at least, in part mediated influenza A (H1N1) pdm09-induced immunopathological injury. This mechanism might
Meyer Zu Horste, Gerd; Przybylski, Dariusz; Schramm, Markus A; Wang, Chao; Schnell, Alexandra; Lee, Youjin; Sobel, Raymond; Regev, Aviv; Kuchroo, Vijay K
2018-03-20
The death receptor Fas removes activated lymphocytes through apoptosis. Previous transcriptional profiling predicted that Fas positively regulates interleukin-17 (IL-17)-producing T helper 17 (Th17) cells. Here, we demonstrate that Fas promoted the generation and stability of Th17 cells and prevented their differentiation into Th1 cells. Mice with T-cell- and Th17-cell-specific deletion of Fas were protected from induced autoimmunity, and Th17 cell differentiation and stability were impaired. Fas-deficient Th17 cells instead developed a Th1-cell-like transcriptional profile, which a new algorithm predicted to depend on STAT1. Experimentally, Fas indeed bound and sequestered STAT1, and Fas deficiency enhanced IL-6-induced STAT1 activation and nuclear translocation, whereas deficiency of STAT1 reversed the transcriptional changes induced by Fas deficiency. Thus, our computational and experimental approach identified Fas as a regulator of the Th17-to-Th1 cell balance by controlling the availability of opposing STAT1 and STAT3 to have a direct impact on autoimmunity. Copyright © 2018. Published by Elsevier Inc.
Ding, Ling-Zhi; Teng, Xiao; Zhang, Zhao-Bo; Zheng, Chang-Jun; Chen, Shi-Hong
2018-05-01
Mangiferin is a xanthone glucoside, which possesses antioxidant, antiviral, antitumor and anti-inflammatory functions, and is associated with gene regulation. However, it remains unknown whether mangiferin protects osteoblasts, such as the MC3T3-E1 cell line, against glucocorticoid-induced damage. In the present study, MC3T3-E1 cells were treated with dexamethasone (Dex), which is a well-known synthetic glucocorticoid, in order to establish a glucocorticoid-induced cell injury model. After Dex and/or mangiferin treatment, cell viability, apoptosis and reactive oxygen species (ROS) production was measured by Cell Counting kit-8 (CCK-8) and flow cytometry, respectively, and the concentration of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and macrophage colony-stimulating factor (M-CSF) was measured by ELISA. The expression of bone morphogenetic protein 2 (BMP2), phosphorylated‑SMAD family member 1 (p-Smad-1), t-Smad-1, osterix (OSX), osteocalcin (OCN), osteoprotegerin (OPG), receptor activator of nuclear factor-κB (RANK), RANK ligand (RANKL), B‑cell lymphoma 2 (Bcl-2) and Bcl‑2‑associated X protein (Bax) was measured by real-time PCR and/or western blot analysis. The results indicated that pretreatment of MC3T3-E1 cells with mangiferin for 3 h prior to exposure to Dex for 48 h significantly attenuated Dex-induced injury and inflammation, as demonstrated by increased cell viability, and decreases in apoptosis, ROS generation, and the secretion of TNF-α, IL-6 and M-CSF. In addition, pretreatment with mangiferin markedly reduced Dex-induced BMP2 and p‑Smad-1 downregulation, and corrected the expression of differentiation‑ and apoptosis‑associated markers, including alkaline phosphatase, OSX, OCN, OPG, RANK, RANKL, Bcl-2 and Bax, which were altered by Dex treatment. Similar to the protective effects of mangiferin, overexpression of BMP2 suppressed not only Dex-induced cytotoxicity, but also ROS generation, and the secretion of TNF
Glucose-Sensing Receptor T1R3: A New Signaling Receptor Activated by Glucose in Pancreatic β-Cells.
Kojima, Itaru; Nakagawa, Yuko; Hamano, Kunihisa; Medina, Johan; Li, Longfei; Nagasawa, Masahiro
2015-01-01
Subunits of the sweet taste receptors T1R2 and T1R3 are expressed in pancreatic β-cells. Compared with T1R3, mRNA expression of T1R2 is considerably lower. At the protein level, expression of T1R2 is undetectable in β-cells. Accordingly, a major component of the sweet taste-sensing receptor in β-cells may be a homodimer of T1R3 rather than a heterodimer of T1R2/T1R3. Inhibition of this receptor by gurmarin or deletion of the T1R3 gene attenuates glucose-induced insulin secretion from β-cells. Hence the T1R3 homodimer functions as a glucose-sensing receptor (GSR) in pancreatic β-cells. When GSR is activated by the T1R3 agonist sucralose, elevation of intracellular ATP concentration ([ATP]i) is observed. Sucralose increases [ATP]i even in the absence of ambient glucose, indicating that sucralose increases [ATP]i not simply by activating glucokinase, a rate-limiting enzyme in the glycolytic pathway. In addition, sucralose augments elevation of [ATP]i induced by methylsuccinate, suggesting that sucralose activates mitochondrial metabolism. Nonmetabolizable 3-O-methylglucose also increases [ATP]i and knockdown of T1R3 attenuates elevation of [ATP]i induced by high concentration of glucose. Collectively, these results indicate that the T1R3 homodimer functions as a GSR; this receptor is involved in glucose-induced insulin secretion by activating glucose metabolism probably in mitochondria.
Park, Su-Hyung; Shin, Eui-Cheol; Capone, Stefania; Caggiari, Laura; De Re, Valli; Nicosia, Alfredo; Folgori, Antonella; Rehermann, Barbara
2012-01-01
Background & Aims T cells are an important component for development of a vaccine against hepatitis C virus (HCV), but little is known about the features of successful vaccine-induced T cells. Methods We compared the phenotype, function, and kinetics of vaccine-induced and infection-induced T cells in chimpanzees with HCV infection using multicolor flow cytometry and real-time PCR. Results In chimpanzees successfully vaccinated with recombinant adenovirus and DNA against HCV NS3-NS5, HCV-specific T cells appeared earlier, maintained better functionality, and persisted at higher frequencies, for a longer time after HCV-challenge, than those of mock-vaccinated chimpanzees. Vaccine-induced T cells displayed higher levels of CD127, a marker of memory precursors, and lower levels of programmed death (PD)-1 than infection-induced T cells. Vaccine-induced, but not infection-induced T cells, were multifunctional; their ability to secrete interferon-γ and tumor necrosis factor-α correlated with early expression of CD127 but not PD-1. Based on a comparison of vaccine-induced and infection-induced T cells from the same chimpanzee, the CD127+ memory precursor phenotype was induced by the vaccine itself, rather than by low viremia. In contrast, PD-1 induction correlated with viremia, and levels of intrahepatic PD-1, PD-L1, and 2,5-OAS-1 mRNAs correlated with peak titers of HCV. Conclusions Compared with infection, vaccination induced HCV-specific CD127+ T cells with high functionality that persisted at higher levels for a longer time. Control of viremia prevented upregulation of PD-1 on T cells, and induction of PD-1, PD-L1, and 2,5-OAS-1 in the liver. Early development of a memory T-cell phenotype and, via control of viremia, attenuation of the inhibitory PD1–PD-L1 pathway might be necessary components of successful vaccine-induced protection against HCV. PMID:22705008
Hydrophilic CeO2 nanocubes protect pancreatic β-cell line INS-1 from H2O2-induced oxidative stress
NASA Astrophysics Data System (ADS)
Lyu, Guang-Ming; Wang, Yan-Jie; Huang, Xue; Zhang, Huai-Yuan; Sun, Ling-Dong; Liu, Yan-Jun; Yan, Chun-Hua
2016-04-01
Oxidative stress plays a key role in the occurrence and development of diabetes. With their unique redox properties, CeO2 nanoparticles (nanoceria) exhibit promising potential for the treatment of diabetes resulting from oxidative stress. Here, we develop a novel preparation of hydrophilic CeO2 nanocubes (NCs) with two different sizes (5 nm and 25 nm) via an acetate assisted hydrothermal method. Dynamic light scattering, zeta potential measurements and thermogravimetric analyses were utilized to investigate the changes in the physico-chemical characteristics of CeO2 NCs when exposed to in vitro cell culture conditions. CCK-8 assays revealed that the CeO2 NCs did not impair cell proliferation in the pancreatic β-cell line INS-1 at the highest dose of 200 μg mL-1 over the time scale of 72 h, while being able to protect INS-1 cells from H2O2-induced cytotoxicity even after protein adsorption. It is also noteworthy that nanoceria with a smaller hydrodynamic radius exhibit stronger antioxidant and anti-apoptotic effects, which is consistent with their H2O2 quenching capability in biological systems. These findings suggest that nanoceria can be used as an excellent antioxidant for controlling oxidative stress-induced pancreatic β-cell damage.Oxidative stress plays a key role in the occurrence and development of diabetes. With their unique redox properties, CeO2 nanoparticles (nanoceria) exhibit promising potential for the treatment of diabetes resulting from oxidative stress. Here, we develop a novel preparation of hydrophilic CeO2 nanocubes (NCs) with two different sizes (5 nm and 25 nm) via an acetate assisted hydrothermal method. Dynamic light scattering, zeta potential measurements and thermogravimetric analyses were utilized to investigate the changes in the physico-chemical characteristics of CeO2 NCs when exposed to in vitro cell culture conditions. CCK-8 assays revealed that the CeO2 NCs did not impair cell proliferation in the pancreatic β-cell line INS-1 at
Protective effect of Edaravone against hypoxia-induced cytotoxicity in osteoblasts MC3T3-E1 cells.
Cao, Bo; Chai, Chunxiang; Zhao, Sishun
2015-12-01
Edaravone is a newly developed clinical medicine for the treatment of acute cerebral infarction. Reduced blood supply to bones (hypoxia) has been involved in the pathological development of osteoporosis. In this study, we investigated the effect of Edaravone and its latent mechanism on hypoxia-induced cell toxicity in MC3T3-E1 cells. Cell viability was determined by the 3-(4,5-dimethyl-thiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Intracellular reactive oxygen species (ROS) and nitric oxide (NO) were determined by the fluorescence dyes 2',7'-dichlorofluorescein diacetate (DCFH-DA) and 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM DA), respectively. mRNA and proteins were determined by real-time polymerase chain reaction and Western blot analysis, respectively. Edaravone significantly restored the hypoxia-induced reduction of MC3T3-E1 cell viability and inhibited lactate dehydrogenase release. In addition, we found that Edaravone inhibits the generation of ROS and NO. Hoechst staining results indicated that the nuclear condensation characteristic of apoptosis was increased in MC3T3-E1 cells after hypoxia exposure, which was significantly suppressed by Edaravone treatment. Mechanistically, we found that Edaravone markedly reduced the expression of cleaved caspase-3 and blunted the release of cytochrome c. These findings strongly suggested that Edaravone suppresses hypoxia-induced cytotoxicity in MC3T3-E1 cells. The pleiotropic effects of Edaravone on hypoxia exposure in osteoblasts suggest potential antiosteoporosis mechanisms of Edaravone. © 2015 International Union of Biochemistry and Molecular Biology.
Engineering antigens for in situ erythrocyte binding induces T-cell deletion.
Kontos, Stephan; Kourtis, Iraklis C; Dane, Karen Y; Hubbell, Jeffrey A
2013-01-02
Antigens derived from apoptotic cell debris can drive clonal T-cell deletion or anergy, and antigens chemically coupled ex vivo to apoptotic cell surfaces have been shown correspondingly to induce tolerance on infusion. Reasoning that a large number of erythrocytes become apoptotic (eryptotic) and are cleared each day, we engineered two different antigen constructs to target the antigen to erythrocyte cell surfaces after i.v. injection, one using a conjugate with an erythrocyte-binding peptide and another using a fusion with an antibody fragment, both targeting the erythrocyte-specific cell surface marker glycophorin A. Here, we show that erythrocyte-binding antigen is collected much more efficiently than free antigen by splenic and hepatic immune cell populations and hepatocytes, and that it induces antigen-specific deletional responses in CD4(+) and CD8(+) T cells. We further validated T-cell deletion driven by erythrocyte-binding antigens using a transgenic islet β cell-reactive CD4(+) T-cell adoptive transfer model of autoimmune type 1 diabetes: Treatment with the peptide antigen fused to an erythrocyte-binding antibody fragment completely prevented diabetes onset induced by the activated, autoreactive CD4(+) T cells. Thus, we report a translatable modular biomolecular approach with which to engineer antigens for targeted binding to erythrocyte cell surfaces to induce antigen-specific CD4(+) and CD8(+) T-cell deletion toward exogenous antigens and autoantigens.
The cathepsin B inhibitor, z-FA-CMK is toxic and readily induced cell death in human T lymphocytes
DOE Office of Scientific and Technical Information (OSTI.GOV)
Liow, K.Y.; Chow, S.C., E-mail: chow.sek.chuen@monash.edu
The cathepsin B inhibitor, benzyloxycarbonyl-phenylalanine-alanine-chloromethylketone (z-FA-CMK) was found to be toxic and readily induced cell death in the human T cell line, Jurkat, whereas two other analogs benzyloxycarbonyl-phenylalanine-alanine-fluoromethylketone (z-FA-FMK) and benzyloxycarbonyl-phenylalanine-alanine-diazomethylketone (z-FA-DMK) were not toxic. The toxicity of z-FA-CMK requires not only the CMK group, but also the presence of alanine in the P1 position and the benzyloxycarbonyl group at the N-terminal. Dose–response studies showed that lower concentrations of z-FA-CMK induced apoptosis in Jurkat T cells whereas higher concentrations induced necrosis. In z-FA-CMK-induced apoptosis, both initiator caspases (-8 and -9) and effector caspases (-3, -6 and -7) were processed tomore » their respective subunits in Jurkat T cells. However, only the pro-form of the initiator caspases were reduced in z-FA-CMK-induced necrosis and no respective subunits were apparent. The caspase inihibitor benzyloxycarbonyl-valine-alanine-aspartic acid-(O-methyl)-fluoromehylketone (z-VAD-FMK) inhibits apoptosis and caspase processing in Jurkat T cells treated with low concentration of z-FA-CMK but has no effect on z-FA-CMK-induced necrosis and the loss of initiator caspases. This suggests that the loss of initiator caspases in Jurkat T cells during z-FA-CMK-induced necrosis is not a caspase-dependent process. Taken together, we have demonstrated that z-FA-CMK is toxic to Jurkat T cells and induces apoptosis at low concentrations, while at higher concentrations the cells die of necrosis. - Highlights: • z-FA-CMK is toxic and induce cell death in the human T cells. • z-FA-CMK toxicity requires the CMK group, alanine and the benzyloxycarbonyl group. • z-FA-CMK induced apoptosis at low concentration and necrosis at high concentration.« less
Ando, Shotaro; Kawada, Jun-ichi; Watanabe, Takahiro; Suzuki, Michio; Sato, Yoshitaka; Torii, Yuka; Asai, Masato; Goshima, Fumi; Murata, Takayuki; Shimizu, Norio; Ito, Yoshinori; Kimura, Hiroshi
2016-01-01
Epstein-Barr virus (EBV) infects not only B cells, but also T cells and natural killer (NK) cells, and is associated with T or NK cell lymphoma. These lymphoid malignancies are refractory to conventional chemotherapy. We examined the activation of the JAK3/STAT5 pathway in EBV-positive and -negative B, T and NK cell lines and in cell samples from patients with EBV-associated T cell lymphoma. We then evaluated the antitumor effects of the selective JAK3 inhibitor, tofacitinib, against these cell lines in vitro and in a murine xenograft model. We found that all EBV-positive T and NK cell lines and patient samples tested displayed activation of the JAK3/STAT5 pathway. Treatment of these cell lines with tofacitinib reduced the levels of phospho-STAT5, suppressed proliferation, induced G1 cell-cycle arrest and decreased EBV LMP1 and EBNA1 expression. An EBV-negative NK cell line was also sensitive to tofacitinib, whereas an EBV-infected NK cell line was more sensitive to tofacitinib than its parental line. Tofacitinib significantly inhibited the growth of established tumors in NOG mice. These findings suggest that tofacitinib may represent a useful therapeutic agent for patients with EBV-associated T and NK cell lymphoma. PMID:27732937
Ando, Shotaro; Kawada, Jun-Ichi; Watanabe, Takahiro; Suzuki, Michio; Sato, Yoshitaka; Torii, Yuka; Asai, Masato; Goshima, Fumi; Murata, Takayuki; Shimizu, Norio; Ito, Yoshinori; Kimura, Hiroshi
2016-11-22
Epstein-Barr virus (EBV) infects not only B cells, but also T cells and natural killer (NK) cells, and is associated with T or NK cell lymphoma. These lymphoid malignancies are refractory to conventional chemotherapy. We examined the activation of the JAK3/STAT5 pathway in EBV-positive and -negative B, T and NK cell lines and in cell samples from patients with EBV-associated T cell lymphoma. We then evaluated the antitumor effects of the selective JAK3 inhibitor, tofacitinib, against these cell lines in vitro and in a murine xenograft model. We found that all EBV-positive T and NK cell lines and patient samples tested displayed activation of the JAK3/STAT5 pathway. Treatment of these cell lines with tofacitinib reduced the levels of phospho-STAT5, suppressed proliferation, induced G1 cell-cycle arrest and decreased EBV LMP1 and EBNA1 expression. An EBV-negative NK cell line was also sensitive to tofacitinib, whereas an EBV-infected NK cell line was more sensitive to tofacitinib than its parental line. Tofacitinib significantly inhibited the growth of established tumors in NOG mice. These findings suggest that tofacitinib may represent a useful therapeutic agent for patients with EBV-associated T and NK cell lymphoma.
Antioxidative potential of Duranta repens (Linn.) fruits against H2O2 induced cell death in vitro.
Khan, Md Asaduzzaman; Rahman, Mohammad Mijanur; Tania, Mousumi; Shoshee, Nusrat Fatima; Xu, Ai-hua; Chen, Han-chun
2013-01-01
The effects of Duranta repens fruits were investigated on H2O2 induced oxidative cell death to evaluate its antioxidative potential in vitro. HEK293T cells were treated with different concentrations [0-1000 µg/ ml] of ethanol extract (E-Ex) and methanol extract (M-Ex) of D. repens for 24h, and then treated with 100 µM H2O2 for 24h. Cell viability, antioxidant parameters of cells, and antioxidant constituents of the extracts were determined. Treatment with limited dose of E-Ex or M-Ex increased the survival rate of H2O2-treated HEK293T cells, however the extra-high dose showed growth inhibitory effect. Treatment with E-Ex or M-Ex protected cellular lipid per-oxidation. In vitro analyses showed the 2,2-diphenyl-1-picrylhydrazyl and H2O2 scavenging activities as well as reducing potential of the extracts. We report here that the limited dose of E-Ex and M-Ex possess antioxidative potential, which can protect H2O2-induced oxidative cell damage.
Liang, Hua; Deng, Liufu; Chmura, Steven; Burnette, Byron; Liadis, Nicole; Darga, Thomas; Beckett, Michael A.; Lingen, Mark W.; Witt, MaryEllyn; Weichselbaum, Ralph R.; Fu, Yang-Xin
2013-01-01
Local failures following radiation therapy are multifactorial and the contributions of the tumor and the host are complex. Current models of tumor equilibrium suggest that a balance exists between cell birth and cell death due to insufficient angiogenesis, immune effects, or intrinsic cellular factors. We investigated whether host immune responses contribute to radiation induced tumor equilibrium in animal models. We report an essential role for immune cells and their cytokines in suppressing tumor cell regrowth in two experimental animal model systems. Depletion of T cells or neutralization of interferon-gamma reversed radiation-induced equilibrium leading to tumor regrowth. We also demonstrate that PD-L1 blockade augments T cell responses leading to rejection of tumors in radiation induced equilibrium. We identify an active interplay between tumor cells and immune cells that occurs in radiation-induced tumor equilibrium and suggest a potential role for disruption of the PD-L1/PD-1 axis in increasing local tumor control. PMID:23630355
Prior irradiation results in elevated programmed cell death protein 1 (PD-1) in T cells.
Li, Deguan; Chen, Renxiang; Wang, Yi-Wen; Fornace, Albert J; Li, Heng-Hong
2018-05-01
In this study we addressed the question whether radiation-induced adverse effects on T cell activation are associated with alterations of T cell checkpoint receptors. Expression levels of checkpoint receptors on T cell subpopulations were analyzed at multiple post-radiation time points ranging from one to four weeks in mice receiving a single fraction of 1 or 4 Gy of γ-ray. T cell activation associated metabolic changes were assessed. Our results showed that prior irradiation resulted in significant elevated expression of programmed cell death protein 1 (PD-1) in both CD4+ and CD8+ populations, at all three post-radiation time points. T cells with elevated PD-1 mostly were either central memory or naïve cells. In addition, the feedback induction of PD-1 expression in activated T cells declined after radiation. Taken together, the elevated PD-1 level observed at weeks after radiation exposure is connected to T cell dysfunction. Recent preclinical and clinical studies have showed that a combination of radiotherapy and T cell checkpoint blockade immunotherapy including targeting the programmed death-ligand 1 (PD-L1)/PD-1 axis may potentiate the antitumor response. Understanding the dynamic changes in PD-1 levels in T cells after radiation should help in the development of a more effective therapeutic strategy.
Nagai, K; Takahashi, Y; Mikami, I; Fukusima, T; Oike, H; Kobori, M
2009-01-01
Background and purpose: Cell-to-cell interactions between mast cells and activated T cells are increasingly recognized as a possible mechanism in the aetiology of allergic or non-allergic inflammatory disorders. To determine the anti-allergic effect of fisetin, we examined the ability of fisetin to suppress activation of the human mast cell line, HMC-1, induced by activated Jurkat T cell membranes. Experimental approach: HMC-1 cells were incubated with or without fisetin for 15 min and then co-cultured with Jurkat T cell membranes activated by phorbol-12-myristate 13-acetate for 16 h. We determined gene expression in activated HMC-1 cells by DNA microarray and quantitative reverse transcription (RT)-PCR analysis. We also examined activation of the transcription factor NF-κB and MAP kinases (MAPKs) in activated HMC-1 cells. Key results: Fisetin suppresses cell spreading and gene expression in HMC-1 cells stimulated by activated T cell membranes. Additionally, we show that these stimulated HMC-1 cells expressed granzyme B. The stimulatory interaction also induced activation of NF-κB and MAPKs; these activations were suppressed by fisetin. Fisetin also reduced the amount of cell surface antigen CD40 and intercellular adhesion molecule-1 (ICAM-1) on activated HMC-1 cells. Conclusions and implications: Fisetin suppressed activation of HMC-1 cells by activated T cell membranes by interfering with cell-to-cell interaction and inhibiting the activity of NF-κB and MAPKs and thereby suppressing gene expression. Fisetin may protect against the progression of inflammatory diseases by limiting interactions between mast cells and activated T cells. PMID:19702784
Ueda, Norihiro; Uemura, Yasushi; Zhang, Rong; Kitayama, Shuichi; Iriguchi, Shoichi; Kawai, Yohei; Yasui, Yutaka; Tatsumi, Minako; Ueda, Tatsuki; Liu, Tian-Yi; Mizoro, Yasutaka; Okada, Chihiro; Watanabe, Akira; Nakanishi, Mahito; Senju, Satoru; Nishimura, Yasuharu; Kuzushima, Kiyotaka; Kiyoi, Hitoshi; Naoe, Tomoki; Kaneko, Shin
2018-06-05
CD4 + T helper (Th) cell activation is essential for inducing cytotoxic T lymphocyte (CTL) responses against malignancy. We reprogrammed a Th clone specific for chronic myelogenous leukemia (CML)-derived b3a2 peptide to pluripotency and re-differentiated the cells into original TCR-expressing T-lineage cells (iPS-T cells) with gene expression patterns resembling those of group 1 innate lymphoid cells. CD4 gene transduction into iPS-T cells enhanced b3a2 peptide-specific responses via b3a2 peptide-specific TCR. iPS-T cells upregulated CD40 ligand (CD40L) expression in response to interleukin-2 and interleukin-15. In the presence of Wilms tumor 1 (WT1) peptide, antigen-specific dendritic cells (DCs) conditioned by CD4-modified CD40L high iPS-T cells stimulated WT1-specific CTL priming, which eliminated WT1 peptide-expressing CML cells in vitro and in vivo. Thus, CD4 modification of CD40L high iPS-T cells generates innate lymphoid helper-like cells inducing bcr-abl-specific TCR signaling that mediates effectiveanti-leukemic CTL responses via DC maturation, showing potential for adjuvant immunotherapy against leukemia. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.
Sepsis-induced alteration in T-cell Ca(2+) signaling in neonatal rats.
Alattar, M H; Ravindranath, T M; Choudhry, M A; Muraskas, J K; Namak, S Y; Dallal, O; Sayeed, M M
2001-01-01
Sepsis-induced suppression in T-cell proliferation follows deranged Ca(2+) signaling in adult rats. In preliminary studies, we observed suppression in T-cell proliferation in septic neonatal rats as well. In this study, we assessed splenic T-cell cytosolic Ca(2+) concentration, [Ca(2+)](i), as its elevation plays an important role in T-cell proliferation. Also, we investigated the role of PGE(2) in sepsis-related changes in T-cell [Ca(2+)](i) in animals pretreated with cyclooxygenase-1 (COX-1) inhibitor (resveratrol) and cyclooxygenase-2 (COX-2) inhibitor (NS-398). Sepsis was induced in 15-day-old rat pups by intraperitoneal implantation of fecal pellets containing Escherichia coli and Bacteroides fragilis. The sham group consisted of pups implanted with sterile fecal pellets. Septic and sham pups were sacrificed 24 h after implantation and their spleens were removed. The spleens from sham and septic pups, along with spleens from unoperated control pups, were processed for single cell suspensions, and T cells were isolated using nylon wool columns. Fura-2 fluorophotometry was employed for the measurement of [Ca(2+)](i) (in nM units) in T cells stimulated with concanavalin A (ConA). Our results show that ConA-mediated T-cell [Ca(2+)](i) response is significantly suppressed in septic neonatal rats. Pretreatment of pups with COX-2, but not COX-1 inhibitor, prevented the decrease in the [Ca(2+)](i) response. These findings suggest that PGE(2) might induce the attenuation in T-cell Ca(2+) signaling during sepsis in neonatal rats. Copyright 2001 S. Karger AG, Basel
Weiss, Jonathan M.; Bilate, Angelina M.; Gobert, Michael; Ding, Yi; Curotto de Lafaille, Maria A.; Parkhurst, Christopher N.; Xiong, Huizhong; Dolpady, Jayashree; Frey, Alan B.; Ruocco, Maria Grazia; Yang, Yi; Floess, Stefan; Huehn, Jochen; Oh, Soyoung; Li, Ming O.; Niec, Rachel E.; Rudensky, Alexander Y.; Dustin, Michael L.; Littman, Dan R.
2012-01-01
Foxp3 activity is essential for the normal function of the immune system. Two types of regulatory T (T reg) cells express Foxp3, thymus-generated natural T reg (nT reg) cells, and peripherally generated adaptive T reg (iT reg) cells. These cell types have complementary functions. Until now, it has not been possible to distinguish iT reg from nT reg cells in vivo based solely on surface markers. We report here that Neuropilin 1 (Nrp1) is expressed at high levels by most nT reg cells; in contrast, mucosa-generated iT reg and other noninflammatory iT reg cells express low levels of Nrp1. We found that Nrp1 expression is under the control of TGF-β. By tracing nT reg and iT reg cells, we could establish that some tumors have a very large proportion of infiltrating iT reg cells. iT reg cells obtained from highly inflammatory environments, such as the spinal cords of mice with spontaneous autoimmune encephalomyelitis (EAE) and the lungs of mice with chronic asthma, express Nrp1. In the same animals, iT reg cells in secondary lymphoid organs remain Nrp1low. We also determined that, in spontaneous EAE, iT reg cells help to establish a chronic phase of the disease. PMID:22966001
Memory CD8+ T Cells Protect Dendritic Cells from CTL Killing1
Watchmaker, Payal B.; Urban, Julie A.; Berk, Erik; Nakamura, Yutaro; Mailliard, Robbie B.; Watkins, Simon C.; van Ham, S. Marieke; Kalinski, Pawel
2010-01-01
CD8+ T cells have been shown to be capable of either suppressing or promoting immune responses. To reconcile these contrasting regulatory functions, we compared the ability of human effector and memory CD8+ T cells to regulate survival and functions of dendritic cells (DC). We report that, in sharp contrast to the effector cells (CTLs) that kill DCs in a granzyme B- and perforin-dependent mechanism, memory CD8+ T cells enhance the ability of DCs to produce IL-12 and to induce functional Th1 and CTL responses in naive CD4+ and CD8+ T cell populations. Moreover, memory CD8+ T cells that release the DC-activating factor TNF-α before the release of cytotoxic granules induce DC expression of an endogenous granzyme B inhibitor PI-9 and protect DCs from CTL killing with similar efficacy as CD4+ Th cells. The currently identified DC-protective function of memory CD8+ T cells helps to explain the phenomenon of CD8+ T cell memory, reduced dependence of recall responses on CD4+ T cell help, and the importance of delayed administration of booster doses of vaccines for the optimal outcome of immunization. PMID:18322193
Shuda, Masahiro; Guastafierro, Anna; Geng, Xuehui; Shuda, Yoko; Ostrowski, Stephen M; Lukianov, Stefan; Jenkins, Frank J; Honda, Kord; Maricich, Stephen M; Moore, Patrick S; Chang, Yuan
2015-01-01
Merkel cell polyomavirus (MCV) causes the majority of human Merkel cell carcinomas (MCC) and encodes a small T (sT) antigen that transforms immortalized rodent fibroblasts in vitro. To develop a mouse model for MCV sT-induced carcinogenesis, we generated transgenic mice with a flox-stop-flox MCV sT sequence homologously recombined at the ROSA locus (ROSAsT), allowing Cre-mediated, conditional MCV sT expression. Standard tamoxifen (TMX) administration to adult UbcCreERT2; ROSAsT mice, in which Cre is ubiquitously expressed, resulted in MCV sT expression in multiple organs that was uniformly lethal within 5 days. Conversely, most adult UbcCreERT2; ROSAsT mice survived low-dose tamoxifen administration but developed ear lobe dermal hyperkeratosis and hypergranulosis. Simultaneous MCV sT expression and conditional homozygous p53 deletion generated multi-focal, poorly-differentiated, highly anaplastic tumors in the spleens and livers of mice after 60 days of TMX treatment. Mouse embryonic fibroblasts from these mice induced to express MCV sT exhibited anchorage-independent cell growth. To examine Merkel cell pathology, MCV sT expression was also induced during mid-embryogenesis in Merkel cells of Atoh1CreERT2/+; ROSAsT mice, which lead to significantly increased Merkel cell numbers in touch domes at late embryonic ages that normalized postnatally. Tamoxifen administration to adult Atoh1CreERT2/+; ROSAsT and Atoh1CreERT2/+; ROSAsT; p53flox/flox mice had no effects on Merkel cell numbers and did not induce tumor formation. Taken together, these results show that MCV sT stimulates progenitor Merkel cell proliferation in embryonic mice and is a bona fide viral oncoprotein that induces full cancer cell transformation in the p53-null setting.
Cusick, Matthew F.; Schiller, Jennifer J.; Gill, Joan C.; Eckels, David D.
2011-01-01
Regulatory T cell markers are increased in chronically infected individuals with the hepatitis C virus (HCV), but to date, the induction and maintenance of Tregs in HCV infection has not been clearly defined. In this paper, we demonstrate that naturally occurring viral variants suppress T cell responses to cognate NS3358-375 in an antigen-specific manner. Of four archetypal variants, S370P induced regulatory T cell markers in comparison to NS3358-375-stimulated CD4 T cells. Further, the addition of variant-specific CD4 T cells back into a polyclonal culture in a dose-dependent manner inhibited the T cell response. These results suggest that HCV is able to induce antigen-specific regulatory T cells to suppress the antiviral T cell response in an antigen-specific manner, thus contributing to a niche within the host that could be conducive to HCV persistence. PMID:21197453
PD-1 regulates extrathymic regulatory T-cell differentiation
Chen, Xiufen; Fosco, Dominick; Kline, Douglas E.; Meng, Liping; Nishi, Saki; Savage, Peter A.; Kline, Justin
2014-01-01
Regulatory T (Treg) cells and the programmed death-1/programmed death ligand-1 (PD-1/PD-L1) pathway are both critical for maintaining peripheral tolerance to self antigens. A significant subset of Treg cells constitutively expresses PD-1, which prompted an investigation into the role of PD-1/PD-L1 interactions in Treg-cell development, function and induction in vivo. The phenotype and abundance of Treg cells was not significantly altered in PD-1-deficient mice. The thymic development of polyclonal and monospecific Treg cells was not negatively impacted by PD-1 deficiency. The suppressive function of PD-1−/− Treg cells was similar to their PD-1+/+ counterparts both in vitro and in vivo. However, in three different in vivo experimental settings, PD-1−/− conventional CD4+ T cells demonstrated a strikingly diminished tendency toward differentiation into peripherally induced Treg (pTreg) cells. Our results demonstrate that PD-1 is dispensable for thymic (tTreg) Treg-cell development and suppressive function, but is critical for the extrathymic differentiation of pTreg cells in vivo. These data suggest that antibody blockade of the PD-1/PD-L1 pathway may augment T-cell responses by acting directly on conventional T cells, and also by suppressing the differentiation of pTreg cells. PMID:24975127
Altin, John A.; Daley, Stephen R.; Howitt, Jason; Rickards, Helen J.; Batkin, Alison K.; Horikawa, Keisuke; Prasad, Simon J.; Nelms, Keats A.; Kumar, Sharad; Wu, Lawren C.; Tan, Seong-Seng; Cook, Matthew C.; Goodnow, Christopher C.
2014-01-01
The NDFIP1 (neural precursor cell expressed, developmentally down-regulated protein 4 family-interacting protein 1) adapter for the ubiquitin ligase ITCH is genetically linked to human allergic and autoimmune disease, but the cellular mechanism by which these proteins enable foreign and self-antigens to be tolerated is unresolved. Here, we use two unique mouse strains—an Ndfip1-YFP reporter and an Ndfip1-deficient strain—to show that Ndfip1 is progressively induced during T-cell differentiation and activation in vivo and that its deficiency causes a cell-autonomous, Forkhead box P3-independent failure of peripheral CD4+ T-cell tolerance to self and exogenous antigen. In small cohorts of antigen-specific CD4+ cells responding in vivo, Ndfip1 was necessary for tolerogen-reactive T cells to exit cell cycle after one to five divisions and to abort Th2 effector differentiation, defining a step in peripheral tolerance that provides insights into the phenomenon of T-cell anergy in vivo and is distinct from the better understood process of Bcl2-interacting mediator of cell death-mediated apoptosis. Ndfip1 deficiency precipitated autoimmune pancreatic destruction and diabetes; however, this depended on a further accumulation of nontolerant anti-self T cells from strong stimulation by exogenous tolerogen. These findings illuminate a peripheral tolerance checkpoint that aborts T-cell clonal expansion against allergens and autoantigens and demonstrate how hypersensitive responses to environmental antigens may trigger autoimmunity. PMID:24520172
Polycomb (PcG) Proteins, BMI1 and SUZ12, Regulate Arsenic-induced Cell Transformation*
Kim, Hong-Gyum; Kim, Dong Joon; Li, Shengqing; Lee, Kun Yeong; Li, Xiang; Bode, Ann M.; Dong, Zigang
2012-01-01
Inorganic arsenic is a well-documented human carcinogen associated with cancers of the skin, lung, liver, and bladder. However, the underlying mechanisms explaining the tumorigenic role of arsenic are not well understood. The present study explored a potential mechanism of cell transformation induced by arsenic exposure. Exposure to a low dose (0.5 μm) of arsenic trioxide (As2O3) caused transformation of BALB/c 3T3 cells. In addition, in a xenograft mouse model, tumor growth of the arsenic-induced transformed cells was dramatically increased. In arsenic-induced transformed cells, polycomb group (PcG) proteins, including BMI1 and SUZ12, were activated resulting in enhanced histone H3K27 tri-methylation levels. On the other hand, tumor suppressor p16INK4a and p19ARF mRNA and protein expression were dramatically suppressed. Introduction of small hairpin (sh) RNA-BMI1 or -SUZ12 into BALB/c 3T3 cells resulted in suppression of arsenic-induced transformation. Histone H3K27 tri-methylation returned to normal in BMI1- or SUZ12-knockdown BALB/c 3T3 cells compared with BMI1- or SUZ12-wildtype cells after arsenic exposure. As a consequence, the expression of p16INK4a and p19ARF was recovered in arsenic-treated BMI1- or SUZ12-knockdown cells. Thus, arsenic-induced cell transformation was blocked by inhibition of PcG function. Taken together, these results strongly suggest that the polycomb proteins, BMI1 and SUZ12 are required for cell transformation induced by organic arsenic exposure. PMID:22843710
Andolfi, Grazia; Fousteri, Georgia; Rossetti, Maura; Magnani, Chiara F; Jofra, Tatiana; Locafaro, Grazia; Bondanza, Attilio; Gregori, Silvia; Roncarolo, Maria-Grazia
2012-01-01
Type 1 regulatory T (Tr1) cells are an inducible subset of CD4+ Tr cells characterized by high levels of interleukin (IL)-10 production and regulatory properties. Several protocols to generate human Tr1 cells have been developed in vitro. However, the resulting population includes a significant fraction of contaminating non-Tr1 cells, representing a major bottleneck for clinical application of Tr1 cell therapy. We generated an homogeneous IL-10–producing Tr1 cell population by transducing human CD4+ T cells with a bidirectional lentiviral vector (LV) encoding for human IL-10 and the marker gene, green fluorescent protein (GFP), which are independently coexpressed. The resulting GFP+ LV-IL-10–transduced human CD4+ T (CD4LV-IL-10) cells expressed, upon T-cell receptor (TCR) activation, high levels of IL-10 and concomitant low levels of IL-4, and markers associated with IL-10. Moreover, CD4LV-IL-10 T cells displayed typical Tr1 features: the anergic phenotype, the IL-10, and transforming growth factor (TGF)-β dependent suppression of allogeneic T-cell responses, and the ability to suppress in a cell-to-cell contact independent manner in vitro. CD4LV-IL-10 T cells were able to control xeno graft-versus-host disease (GvHD), demonstrating their suppressive function in vivo. These results show that constitutive over-expression of IL-10 in human CD4+ T cells leads to a stable cell population that recapitulates the phenotype and function of Tr1 cells. PMID:22692497
Distinct susceptibility of HIV vaccine vector-induced CD4 T cells to HIV infection
Niu, Qingli; Hou, Wei; Churchyard, Gavin; Nitayaphan, Sorachai; Pitisuthithum, Punnee; Rerks-Ngarm, Supachai; Franchini, Genoveffa
2018-01-01
The concerns raised from adenovirus 5 (Ad5)-based HIV vaccine clinical trials, where excess HIV infections were observed in some vaccine recipients, have highlighted the importance of understanding host responses to vaccine vectors and the HIV susceptibility of vector-specific CD4 T cells in HIV vaccination. Our recent study reported that human Ad5-specific CD4 T cells induced by Ad5 vaccination (RV156A trial) are susceptible to HIV. Here we further investigated the HIV susceptibility of vector-specific CD4 T cells induced by ALVAC, a canarypox viral vector tested in the Thai trial RV144, as compared to Ad5 vector-specific CD4 T cells in the HVTN204 trial. We showed that while Ad5 vector-specific CD4 T cells were readily susceptible to HIV, ALVAC-specific CD4 T cells in RV144 PBMC were substantially less susceptible to both R5 and X4 HIV in vitro. The lower HIV susceptibility of ALVAC-specific CD4 T cells was associated with the reduced surface expression of HIV entry co-receptors CCR5 and CXCR4 on these cells. Phenotypic analyses identified that ALVAC-specific CD4 T cells displayed a strong Th1 phenotype, producing higher levels of IFN-γ and CCL4 (MIP-1β) but little IL-17. Of interest, ALVAC and Ad5 vectors induced distinct profiles of vector-specific CD8 vs. CD4 T-cell proliferative responses in PBMC, with ALVAC preferentially inducing CD8 T-cell proliferation, while Ad5 vector induced CD4 T-cell proliferation. Depletion of ALVAC-, but not Ad5-, induced CD8 T cells in PBMC led to a modest increase in HIV infection of vector-specific CD4 T cells, suggesting a role of ALVAC-specific CD8 T cells in protecting ALVAC-specific CD4 T cells from HIV. Taken together, our data provide strong evidence for distinct HIV susceptibility of CD4 T cells induced by different vaccine vectors and highlight the importance of better evaluating anti-vector responses in HIV vaccination. PMID:29474461
New approaches to design HIV-1 T-cell vaccines.
Perrin, Hélène; Canderan, Glenda; Sékaly, Rafick-Pierre; Trautmann, Lydie
2010-09-01
Following the evidence that T-cell responses are crucial in the control of HIV-1 infection, vaccines targeting T-cell responses were tested in recent clinical trials. However, these vaccines showed a lack of efficacy. This review attempts to define the qualitative and quantitative features that are desirable for T-cell-induced responses by vaccines. We also describe strategies that could lead to achievement of this goal. Using the yellow fever vaccine as a benchmark of an efficient vaccine, recent studies identified factors of immune protection and more importantly innate immune pathways needed for the establishment of long-term protective adaptive immunity. To prevent or control HIV-1 infection, a vaccine must induce efficient and persistent antigen-specific T cells endowed with mucosal homing capacity. Such cells should have the capability to counteract HIV-1 diversity and its rapid spread from the initial site of infection. To achieve this goal, the activation of a diversified innate immune response is critical. New systems biology approaches will provide more precise correlates of immune protection that will pave the way for new approaches in T-cell-based vaccines.
Crouthamel, Matthew H.; Lau, Wei Ling; Leaf, Elizabeth M.; Chavkin, Nick; Wallingford, Mary C.; Peterson, Danielle F.; Li, Xianwu; Liu, Yonggang; Chin, Michael T.; Levi, Moshe; Giachelli, Cecilia M.
2014-01-01
Objective Elevated serum phosphate has emerged as a major risk factor for vascular calcification. The sodium-dependent phosphate cotransporter, PiT-1, was previously shown to be required for phosphate-induced osteogenic differentiation and calcification of cultured human VSMCs, but its importance in vascular calcification in vivo, as well as the potential role of its homologue, PiT-2, have not been determined. We investigated the in vivo requirement for PiT-1 in vascular calcification using a mouse model of chronic kidney disease, and the potential compensatory role of PiT-2 using in vitro knockdown and over-expression strategies. Approach and Results Mice with targeted deletion of PiT-1 in VSMCs were generated (PiT-1Δsm). PiT-1 mRNA levels were undetectable whereas PiT-2 mRNA levels were increased 2 fold in the vascular aortic media of PiT-1Δsm compared to PiT-1flox/flox control. When arterial medial calcification was induced in PiT-1Δsm and PiT-1flox/flox by chronic kidney disease followed by dietary phosphate loading, the degree of aortic calcification was not different between genotypes, suggesting compensation by PiT-2. Consistent with this possibility, VSMCs isolated from PiT-1Δsm mice had no PiT-1 mRNA expression, increased PiT-2 mRNA levels, and no difference in sodium-dependent phosphate uptake or phosphate-induced matrix calcification compared to PiT-1flox/flox VSMCs. Knockdown of PiT-2 decreased phosphate uptake and phosphate-induced calcification of PiT-1Δsm VSMCs. Furthermore, over-expression of PiT-2 restored these parameters in human PiT-1-deficient VSMCs. Conclusions PiT-2 can mediate phosphate uptake and calcification of VSMCs in the absence of PiT-1. Mechanistically, PiT-1 and PiT-2 appear to serve redundant roles in phosphate-induced calcification of vascular smooth muscle cells. PMID:23968976
Delano, Matthew J.; Scumpia, Philip O.; Weinstein, Jason S.; Coco, Dominique; Nagaraj, Srinivas; Kelly-Scumpia, Kindra M.; O'Malley, Kerri A.; Wynn, James L.; Antonenko, Svetlana; Al-Quran, Samer Z.; Swan, Ryan; Chung, Chun-Shiang; Atkinson, Mark A.; Ramphal, Reuben; Gabrilovich, Dmitry I.; Reeves, Wesley H.; Ayala, Alfred; Phillips, Joseph; LaFace, Drake; Heyworth, Paul G.; Clare-Salzler, Michael; Moldawer, Lyle L.
2007-01-01
Polymicrobial sepsis alters the adaptive immune response and induces T cell suppression and Th2 immune polarization. We identify a GR-1+CD11b+ population whose numbers dramatically increase and remain elevated in the spleen, lymph nodes, and bone marrow during polymicrobial sepsis. Phenotypically, these cells are heterogeneous, immature, predominantly myeloid progenitors that express interleukin 10 and several other cytokines and chemokines. Splenic GR-1+ cells effectively suppress antigen-specific CD8+ T cell interferon (IFN) γ production but only modestly suppress antigen-specific and nonspecific CD4+ T cell proliferation. GR-1+ cell depletion in vivo prevents both the sepsis-induced augmentation of Th2 cell–dependent and depression of Th1 cell–dependent antibody production. Signaling through MyD88, but not Toll-like receptor 4, TIR domain–containing adaptor-inducing IFN-β, or the IFN-α/β receptor, is required for complete GR-1+CD11b+ expansion. GR-1+CD11b+ cells contribute to sepsis-induced T cell suppression and preferential Th2 polarization. PMID:17548519
Rigas, Diamanda; Lewis, Gavin; Aron, Jennifer L; Wang, Bowen; Banie, Homayon; Sankaranarayanan, Ishwarya; Galle-Treger, Lauriane; Maazi, Hadi; Lo, Richard; Freeman, Gordon J; Sharpe, Arlene H; Soroosh, Pejman; Akbari, Omid
2017-05-01
Atopic diseases, including asthma, exacerbate type 2 immune responses and involve a number of immune cell types, including regulatory T (Treg) cells and the emerging type 2 innate lymphoid cells (ILC2s). Although ILC2s are potent producers of type 2 cytokines, the regulation of ILC2 activation and function is not well understood. In the present study, for the first time, we evaluate how Treg cells interact with pulmonary ILC2s and control their function. ILC2s and Treg cells were evaluated by using in vitro suppression assays, cell-contact assays, and gene expression panels. Also, human ILC2s and Treg cells were adoptively transferred into NOD SCID γC-deficient mice, which were given isotype or anti-inducible T-cell costimulator ligand (ICOSL) antibodies and then challenged with IL-33 and assessed for airway hyperreactivity. We show that induced Treg cells, but not natural Treg cells, effectively suppress the production of the ILC2-driven proinflammatory cytokines IL-5 and IL-13 both in vitro and in vivo. Mechanistically, our data reveal the necessity of inducible T-cell costimulator (ICOS)-ICOS ligand cell contact for Treg cell-mediated ILC2 suppression alongside the suppressive cytokines TGF-β and IL-10. Using a translational approach, we then demonstrate that human induced Treg cells suppress syngeneic human ILC2s through ICOSL to control airway inflammation in a humanized ILC2 mouse model. These findings suggest that peripheral expansion of induced Treg cells can serve as a promising therapeutic target against ILC2-dependent asthma. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.
Rook, Alain H; Gelfand, Joel M; Gelfand, Joel C; Wysocka, Maria; Troxel, Andrea B; Benoit, Bernice; Surber, Christian; Elenitsas, Rosalie; Buchanan, Marie A; Leahy, Deborah S; Watanabe, Rei; Kirsch, Ilan R; Kim, Ellen J; Clark, Rachael A
2015-09-17
Early-stage cutaneous T-cell lymphoma (CTCL) is a skin-limited lymphoma with no cure aside from stem cell transplantation. Twelve patients with stage IA-IIA CTCL were treated in a phase 1 trial of 0.03% and 0.06% topical resiquimod gel, a Toll-like receptor 7/8 agonist. Treated lesions significantly improved in 75% of patients and 30% had clearing of all treated lesions. Resiquimod also induced regression of untreated lesions. Ninety-two percent of patients had more than a 50% improvement in body surface area involvement by the modified Severity-Weighted Assessment Tool analysis and 2 patients experienced complete clearing of disease. Four of 5 patients with folliculotropic disease also improved significantly. Adverse effects were minor and largely skin limited. T-cell receptor sequencing and flow cytometry studies of T cells from treated lesions demonstrated decreased clonal malignant T cells in 90% of patients and complete eradication of malignant T cells in 30%. High responses were associated with recruitment and expansion of benign T-cell clones in treated skin, increased skin T-cell effector functions, and a trend toward increased natural killer cell functions. In patients with complete or near eradication of malignant T cells, residual clinical inflammation was associated with cytokine production by benign T cells. Fifty percent of patients had increased activation of circulating dendritic cells, consistent with a systemic response to therapy. In summary, topical resiquimod is safe and effective in early-stage CTCL and the first topical therapy to our knowledge that can induce clearance of untreated lesions and complete remissions in some patients. This trial was registered at www.clinicaltrials.gov as #NCT813320. © 2015 by The American Society of Hematology.
Overexpression of GATA-3 in T cells accelerates dextran sulfate sodium-induced colitis.
Okamura, Midori; Yoh, Keigyou; Ojima, Masami; Morito, Naoki; Takahashi, Satoru
2014-01-01
Ulcerative colitis (UC) is an inflammatory bowel disease, and its pathogenesis includes genetic, environmental, and immunological factors, such as T helper cells and their secreted cytokines. T helper cells are classified as Th1, Th2, and Th17 cells. However, it is unclear which T helper cells are important in UC. Dextran sulfate sodium (DSS)-induced colitis is a commonly used model of UC. In this study, we induced DSS colitis in Th1 dominant (T-bet transgenic (Tg)) mice, Th2 dominant (GATA-3 Tg) mice, and Th17 dominant (RORγt Tg) mice to elucidate the roles of T helper cell in DSS colitis. The results showed that GATA-3 Tg mice developed the most severe DSS colitis compared with the other groups. GATA-3 Tg mice showed a significant decreased in weight from day 1 to day 7, and an increased high score for the disease activity index compared with the other groups. Furthermore, GATA-3 Tg mice developed many ulcers in the colon, and many neutrophils and macrophages were detected on day 4 after DSS treatment. Measurement of GATA-3-induced cytokines demonstrated that IL-13 was highly expressed in the colon from DSS-induced GATA-3 Tg mice. In conclusion, GATA-3 overexpression in T-cells and IL-13 might play important roles in the development of DSS colitis.
Peckham, R K; Brill, R; Foster, D S; Bowen, A L; Leigh, J A; Coffey, T J; Flynn, R J
2014-06-25
IL-17 has emerged as a key player in the immune system, exhibiting roles in protection from infectious diseases and promoting inflammation in autoimmunity. Initially thought to be CD4 T-cell-derived, the sources of IL-17 are now known to be varied and belong to both the innate and adaptive arms of the immune system. Mechanisms for inducing IL-17 production in lymphoid cells are thought to rely on appropriate antigenic stimulation in the context of TGF-β1, IL-6 and/or IL-1β. Using culture protocols adapted from human studies, we have effectively induced both bovine CD4(+) and WC1(+) γδ T-cells to produce IL-17 termed Th17 and γδ17 cells, respectively. The negative regulatory effect of IFN-γ on mouse and human IL-17 production can be extended to the bovine model, as addition of IFN-γ decreases IL-17 production in both cell types. Furthermore we show that infection with the protozoan Neospora caninum will induce fibroblasts to secrete pro-IL-17 factors thereby inducing a γδ17 phenotype that preferentially kills infected target cells. Our study identifies two T-cell sources of IL-17, and is the first to demonstrate a protective effect of IL-17(+) T-cells in ruminants. Our findings offer further opportunities for future adjuvants or vaccines which could benefit from inducing these responses.
Identification of a G protein coupled receptor induced in activated T cells.
Kaplan, M H; Smith, D I; Sundick, R S
1993-07-15
Many genes are induced after T cell activation to make a cell competent for proliferation and ultimately, function. Many of these genes encode surface receptors for growth factors that signal a cell to proliferate. We have cloned a novel gene (clone 6H1) that codes for a member of the G protein-coupled receptor superfamily. This gene was isolated from a chicken activated T cell cDNA library by low level hybridization to mammalian IL-2 cDNA probes. The 308 amino acid open reading frame has seven hydrophobic, presumably transmembrane domains and a consensus site for interaction with G proteins. Tissue distribution studies suggest that gene expression is restricted to activated T cells. The message appears by 1 h after activation and is maintained for at least 45 h. Transcription of 6H1 is induced by a number of T cell stimuli and is inhibited by cyclosporin A, but not by cycloheximide. This is the first description of a member of this superfamily expressed specifically in activated T cells. The gene product may provide a link between T cell growth factors and G protein activation.
Latta, Markus; Mohan, Karkada; Issekutz, Thomas B
2007-01-01
Numerous chemokine receptors are increased in number on T cells in inflamed tissues. Our objective was to examine CXCR6 expression on lymphocytes during immune and inflammatory reactions and its potential for mediating T-cell recruitment. The cDNA for rat CXCR6 was cloned and monoclonal antibodies (mAbs) to CXCR6 were developed. CXCR6 was present on 4–6% of CD4 and CD8 T cells in blood, normal lymph nodes (LNs) and the spleen, primarily on memory T cells. In vitro antigen re-stimulation of LN T cells from animals with autoimmune arthritis and experimental autoimmune encephalomyelitis (EAE) increased the proportion of CXCR6+ T cells to 35–50% and anti-T-cell receptor (TCR) activation to 60–80%. In vivo, after antigen challenge of LNs there was only a small increase in CXCR6+ T cells on the lymphoblasts in the LNs, and a much higher percentage of T cells were CXCR6+ in virus-induced peritoneal exudates (∼47%) and in allergen-induced lung inflammation (33%). Chemotaxis of CXCR6-expressing inflammatory T cells to CXCL16 was poor, but that to CXCL10 was robust. We conclude that few T cells in normal and antigen-challenged LNs are CXCR6+, whereas a high proportion of in vitro activated T cells and T cells from inflammatory sites are CXCR6+, but these cells migrate poorly to CXCL16. This suggests that CXCR6 may contribute to T-cell positioning and activation, rather than recruitment. CXCR6 is also expressed on T cells not only in T helper type 1 (Th1) inflammation (arthritis and EAE) but also, as shown here, in Th2 inflammation, where it is increased after allergen challenge. PMID:17437534
Latta, Markus; Mohan, Karkada; Issekutz, Thomas B
2007-08-01
Numerous chemokine receptors are increased in number on T cells in inflamed tissues. Our objective was to examine CXCR6 expression on lymphocytes during immune and inflammatory reactions and its potential for mediating T-cell recruitment. The cDNA for rat CXCR6 was cloned and monoclonal antibodies (mAbs) to CXCR6 were developed. CXCR6 was present on 4-6% of CD4 and CD8 T cells in blood, normal lymph nodes (LNs) and the spleen, primarily on memory T cells. In vitro antigen re-stimulation of LN T cells from animals with autoimmune arthritis and experimental autoimmune encephalomyelitis (EAE) increased the proportion of CXCR6(+) T cells to 35-50% and anti-T-cell receptor (TCR) activation to 60-80%. In vivo, after antigen challenge of LNs there was only a small increase in CXCR6(+) T cells on the lymphoblasts in the LNs, and a much higher percentage of T cells were CXCR6(+) in virus-induced peritoneal exudates (approximately 47%) and in allergen-induced lung inflammation (33%). Chemotaxis of CXCR6-expressing inflammatory T cells to CXCL16 was poor, but that to CXCL10 was robust. We conclude that few T cells in normal and antigen-challenged LNs are CXCR6(+), whereas a high proportion of in vitro activated T cells and T cells from inflammatory sites are CXCR6(+), but these cells migrate poorly to CXCL16. This suggests that CXCR6 may contribute to T-cell positioning and activation, rather than recruitment. CXCR6 is also expressed on T cells not only in T helper type 1 (Th1) inflammation (arthritis and EAE) but also, as shown here, in Th2 inflammation, where it is increased after allergen challenge.
Ambler, Carrie A.; Watt, Fiona M.
2010-01-01
Notch signalling regulates epidermal differentiation and tumour formation via non-cell autonomous mechanisms that are incompletely understood. This study shows that epidermal Notch activation via a 4-hydroxy-tamoxifen-inducible transgene caused epidermal thickening, focal detachment from the underlying dermis and hair clumping. In addition, there was dermal accumulation of T lymphocytes and stromal cells, some of which localised to the blisters at the epidermal-dermal boundary. The T cell infiltrate was responsible for hair clumping but not for other Notch phenotypes. Notch-induced stromal cells were heterogeneous, expressing markers of neural crest, melanocytes, smooth muscle and peripheral nerve. Although Slug1 expression was expanded in the epidermis, the stromal cells did not arise through epithelial-mesenchymal transition. Epidermal Notch activation resulted in upregulation of jagged 1 in both epidermis and dermis. When Notch was activated in the absence of epidermal jagged 1, jagged 1 was not upregulated in the dermis, and epidermal thickening, blister formation, accumulation of T cells and stromal cells were inhibited. Gene expression profiling revealed that epidermal Notch activation resulted in upregulation of several growth factors and cytokines, including TNFα, the expression of which was dependent on epidermal jagged 1. We conclude that jagged 1 is a key mediator of non-cell autonomous Notch signalling in skin. PMID:20940224
Ambler, Carrie A; Watt, Fiona M
2010-11-01
Notch signalling regulates epidermal differentiation and tumour formation via non-cell autonomous mechanisms that are incompletely understood. This study shows that epidermal Notch activation via a 4-hydroxy-tamoxifen-inducible transgene caused epidermal thickening, focal detachment from the underlying dermis and hair clumping. In addition, there was dermal accumulation of T lymphocytes and stromal cells, some of which localised to the blisters at the epidermal-dermal boundary. The T cell infiltrate was responsible for hair clumping but not for other Notch phenotypes. Notch-induced stromal cells were heterogeneous, expressing markers of neural crest, melanocytes, smooth muscle and peripheral nerve. Although Slug1 expression was expanded in the epidermis, the stromal cells did not arise through epithelial-mesenchymal transition. Epidermal Notch activation resulted in upregulation of jagged 1 in both epidermis and dermis. When Notch was activated in the absence of epidermal jagged 1, jagged 1 was not upregulated in the dermis, and epidermal thickening, blister formation, accumulation of T cells and stromal cells were inhibited. Gene expression profiling revealed that epidermal Notch activation resulted in upregulation of several growth factors and cytokines, including TNFα, the expression of which was dependent on epidermal jagged 1. We conclude that jagged 1 is a key mediator of non-cell autonomous Notch signalling in skin.
Lavorgna, Alfonso; Matsuoka, Masao; Harhaj, Edward William
2014-01-01
Human T-cell leukemia virus type 1 (HTLV-1) infection is linked to the development of adult T-cell leukemia (ATL) and the neuroinflammatory disease HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP). The HTLV-1 Tax protein functions as a potent viral oncogene that constitutively activates the NF-κB transcription factor to transform T cells; however, the underlying mechanisms remain obscure. Here, using next-generation RNA sequencing we identified the IL-25 receptor subunit IL-17RB as an aberrantly overexpressed gene in HTLV-1 immortalized T cells. Tax induced the expression of IL-17RB in an IκB kinase (IKK) and NF-κB-dependent manner. Remarkably, Tax activation of the canonical NF-κB pathway in T cells was critically dependent on IL-17RB expression. IL-17RB and IL-25 were required for HTLV-1-induced immortalization of primary T cells, and the constitutive NF-κB activation and survival of HTLV-1 transformed T cells. IL-9 was identified as an important downstream target gene of the IL-17RB pathway that drives the proliferation of HTLV-1 transformed cells. Furthermore, IL-17RB was overexpressed in leukemic cells from a subset of ATL patients and also regulated NF-κB activation in some, but not all, Tax-negative ATL cell lines. Together, our results support a model whereby Tax instigates an IL-17RB-NF-κB feed-forward autocrine loop that is obligatory for HTLV-1 leukemogenesis. PMID:25340344
Choi, Eun Mi
2012-06-01
Antimycin A treatment of cells blocks the mitochondrial electron transport chain and leads to elevated ROS generation. In the present study, we investigated the protective effects of magnolol, a hydroxylated biphenyl compound isolated from Magnolia officinalis, on antimycin A-induced toxicity in osteoblastic MC3T3-E1 cells. Osteoblastic MC3T3-E1 cells were pre-incubated with magnolol before treatment with antimycin A. Cell viability and mineralization of osteoblasts were assessed by MTT assay and Alizarin Red staining, respectively. Mitochondrial dysfunction in cells was measured by mitochondrial membrane potential (MMP), complex IV activity, and ATP level. The cellular antioxidant effect of magnolol in osteoblastic MC3T3-E1 cells was assessed by measuring cardiolipin oxidation, mitochondrial superoxide levels, and nitrotyrosine content. Phosphorylated cAMP-response element-binding protein (CREB ) was evaluated using ELISA assay. Pretreatment with magnolol prior to antimycin A exposure significantly reduced antimycin A-induced osteoblast dysfunction by preventing MMP dissipation, ATP loss, and CREB inactivation. Magnolol also reduced cardiolipin peroxidation, mitochondrial superoxide, and nitrotyrosine production induced by antimycin A. These results suggest that magnolol has a protective effect against antimycin A-induced cell damage by its antioxidant effects and the attenuation of mitochondrial dysfunction. All these data indicate that magnolol may reduce or prevent osteoblast degeneration in osteoporosis or other degenerative disorders.
Ethylene Insensitivity Modulates Ozone-Induced Cell Death in Birch1
Vahala, Jorma; Ruonala, Raili; Keinänen, Markku; Tuominen, Hannele; Kangasjärvi, Jaakko
2003-01-01
We have used genotypic variation in birch (Betula pendula Roth) to investigate the roles of ozone (O3)-induced ethylene (ET), jasmonic acid, and salicylic acid in the regulation of tissue tolerance to O3. Of these hormones, ET evolution correlated best with O3-induced cell death. Disruption of ET perception by transformation of birch with the dominant negative mutant allele etr1-1 of the Arabidopsis ET receptor gene ETR1 or blocking of ET perception with 1-methylcyclopropene reduced but did not completely prevent the O3-induced cell death, when inhibition of ET biosynthesis with aminooxyacetic acid completely abolished O3 lesion formation. This suggests the presence of an ET-signaling-independent but ET biosynthesis-dependent component in the ET-mediated stimulation of cell death in O3-exposed birch. Functional ET signaling was required for the O3 induction of the gene encoding β-cyanoalanine synthase, which catalyzes detoxification of the cyanide formed during ET biosynthesis. The results suggest that functional ET signaling is required to protect birch from the O3-induced cell death and that a decrease in ET sensitivity together with a simultaneous, high ET biosynthesis can potentially cause cell death through a deficient detoxification of cyanide. PMID:12746524
Kefir induces cell-cycle arrest and apoptosis in HTLV-1-negative malignant T-lymphocytes
Maalouf, Katia; Baydoun, Elias; Rizk, Sandra
2011-01-01
Background: Adult lymphoblastic leukemia (ALL) is a malignancy that occurs in white blood cells. The overall cure rate in children is 85%, whereas it is only 40% in adults. Kefir is an important probiotic that contains many bioactive ingredients, which give it unique health benefits. It has been shown to control several cellular types of cancer. Purpose: The present study investigates the effect of a cell-free fraction of kefir on CEM and Jurkat cells, which are human T-lymphotropic virus type I (HTLV-1)-negative malignant T-lymphocytes. Methods: Cells were incubated with different kefir concentrations. The cytotoxicity of the compound was evaluated by determining the percentage viability of cells. The effect of all the noncytotoxic concentrations of kefir on the proliferation of CEM and Jurkat cells was then assessed. The levels of transforming growth factor-alpha (TGF-α), transforming growth factor- beta1 (TGF-β1), matrix metalloproteinase-2 (MMP-2), and MMP-9 mRNA upon kefir treatment were then analyzed using reverse transcriptase polymerase chain reaction (RT-PCR). Finally, the growth inhibitory effects of kefir on cell-cycle progression/apoptosis were assessed by Cell Death Detection (ELISA) and flow cytometry. Results: The maximum cytotoxicity recorded after 48-hours treatment with 80 μg/μL kefir was only 42% and 39% in CEM and Jurkat cells, respectively. The percent reduction in proliferation was very significant, and was dose-, and time-dependent. In both cell lines, kefir exhibited its antiproliferative effect by downregulating TGF-α and upregulating TGF-β1 mRNA expression. Upon kefir treatment, a marked increase in cell-cycle distribution was noted in the preG1 phase of CEM and Jurkat cells, indicating the proapoptotic effect of kefir, which was further confirmed by Cell Death Detection ELISA. However, kefir did not affect the mRNA expression of metalloproteinases needed for the invasion of leukemic cell lines. Conclusion: In conclusion, kefir is
Airway Hyperresponsiveness through Synergy of γδ T Cells and NKT Cells1
Jin, Niyun; Miyahara, Nobuaki; Roark, Christina L.; French, Jena D.; Aydintug, M. Kemal; Matsuda, Jennifer L.; Gapin, Laurent; O'Brien, Rebecca L.; Gelfand, Erwin W.; Born, Willi K.
2015-01-01
Mice sensitized and challenged with OVA were used to investigate the role of innate T cells in the development of allergic airway hyperresponsiveness (AHR). AHR, but not eosinophilic airway inflammation, was induced in T cell-deficient mice by small numbers of cotransferred γδ T cells and invariant NKT cells, whereas either cell type alone was not effective. Only Vγ1+Vδ5+ γδ T cells enhanced AHR. Surprisingly, OVA-specific αβ T cells were not required, revealing a pathway of AHR development mediated entirely by innate T cells. The data suggest that lymphocytic synergism, which is key to the Ag-specific adaptive immune response, is also intrinsic to T cell-dependent innate responses. PMID:17709511
Shi, Meiqing; Hao, Siguo; Su, Liping; Zhang, Xueshu; Yuan, Jinying; Guo, Xuling; Zheng, Changyu; Xiang, Jim
2005-08-01
The CXC chemokine SDF-1 has been characterized as a T-cell chemoattractant both in vitro and in vivo. To determine whether SDF-1 expression within tumors can influence tumor growth, we transfected an expression vector pCI-SDF-1 for SDF-1 into J558 myeloma cells and tested their ability to form tumors in BALB/c. Production of biologically active SDF-1 (1.2 ng/mL) was detected in the culture supernatants of cells transfected with the expression vector pCI-SDF-1. J558 cells gave rise to a 100% tumor incidence, whereas SDF-1-expressing J558/SDF-1 tumors invariably regressed in BALB/c mice and became infiltrated with CD4(+) and CD8(+) T cells. Regression of the J558/SDF-1 tumors was dependent on both CD4(+) and CD8(+) T-cells. Our data also indicate that TIT cells containing both CD4(+) and CD8(+) T-cells within J558/SDF-1 tumors express the SDF-1 receptor CXCR4, and that SDF-1 specifically chemoattracts these cells in vitro. Furthermore, immunization of mice with engineered J558/SDF-1 cells elicited the most potent protective immunity against 0.5 x 10(6) cells J558 tumor challenge in vivo, compared to immunization with the J558 alone, and this antitumor immunity mediated by J558/SDF-1 tumor cell vaccination in vivo appeared to be dependent on CD8(+) CTL. Thus, SDF-1 has natural adjuvant activities that may augment antitumor responses through their effects on T-cells and thereby could be important in gene transfer immunotherapies for some cancers.
Akhmetzyanova, Ilseyar; Zelinskyy, Gennadiy; Schimmer, Simone; Brandau, Sven; Altenhoff, Petra; Sparwasser, Tim; Dittmer, Ulf
2013-02-01
The important role of tumor-specific cytotoxic CD8(+) T cells is well defined in the immune control of the tumors, but the role of effector CD4(+) T cells is poorly understood. In the current research, we have used a murine retrovirus-induced tumor cell line of C57BL/6 mouse origin, namely FBL-3 cells, as a model to study basic mechanisms of immunological control and escape during tumor formation. This study shows that tumor-specific CD4(+) T cells are able to protect against virus-induced tumor cells. We show here that there is an expansion of tumor-specific CD4(+) T cells producing cytokines and cytotoxic molecule granzyme B (GzmB) in the early phase of tumor growth. Importantly, we demonstrate that in vivo depletion of regulatory T cells (Tregs) and CD8(+) T cells in FBL-3-bearing DEREG transgenic mice augments IL-2 and GzmB production by CD4(+) T cells and increases FV-specific CD4(+) T-cell effector and cytotoxic responses leading to the complete tumor regression. Therefore, the capacity to reject tumor acquired by tumor-reactive CD4(+) T cells largely depends on the direct suppressive activity of Tregs. We suggest that a cytotoxic CD4(+) T-cell immune response may be induced to enhance resistance against oncovirus-associated tumors.
IL-15 regulates memory CD8+ T cell O-glycan synthesis and affects trafficking
Nolz, Jeffrey C.; Harty, John T.
2014-01-01
Memory and naive CD8+ T cells exhibit distinct trafficking patterns. Specifically, memory but not naive CD8+ T cells are recruited to inflamed tissues in an antigen-independent manner. However, the molecular mechanisms that regulate memory CD8+ T cell trafficking are largely unknown. Here, using murine models of infection and T cell transfer, we found that memory but not naive CD8+ T cells dynamically regulate expression of core 2 O-glycans, which interact with P- and E-selectins to modulate trafficking to inflamed tissues. Following infection, antigen-specific effector CD8+ T cells strongly expressed core 2 O-glycans, but this glycosylation pattern was lost by most memory CD8+ T cells. After unrelated infection or inflammatory challenge, memory CD8+ T cells synthesized core 2 O-glycans independently of antigen restimulation. The presence of core 2 O-glycans subsequently directed these cells to inflamed tissue. Memory and naive CD8+ T cells exhibited the opposite pattern of epigenetic modifications at the Gcnt1 locus, which encodes the enzyme that initiates core 2 O-glycan synthesis. The open chromatin configuration in memory CD8+ T cells permitted de novo generation of core 2 O-glycans in a TCR-independent, but IL-15–dependent, manner. Thus, IL-15 stimulation promotes antigen-experienced memory CD8+ T cells to generate core 2 O-glycans, which subsequently localize them to inflamed tissues. These findings suggest that CD8+ memory T cell trafficking potentially can be manipulated to improve host defense and immunotherapy. PMID:24509081
Long, Adrienne H.; Haso, Waleed M.; Shern, Jack F.; Wanhainen, Kelsey M.; Murgai, Meera; Ingaramo, Maria; Smith, Jillian P.; Walker, Alec J.; Kohler, M. Eric; Venkateshwara, Vikas R.; Kaplan, Rosandra N.; Patterson, George H.; Fry, Terry J.; Orentas, Rimas J.; Mackall, Crystal L.
2015-01-01
Chimeric antigen receptors (CARs) targeting CD19 have mediated dramatic anti-tumor responses in hematologic malignancies, but tumor regression has rarely occurred using CARs targeting other antigens. It remains unknown whether the impressive effects of CD19 CARs relate to greater susceptibility of hematologic malignancies to CAR therapies, or superior functionality of the CD19 CAR itself. We discovered that tonic CAR CD3ζ phosphorylation, triggered by antigen-independent clustering of CAR scFvs, can induce early exhaustion of CAR T cells that limits anti-tumor efficacy. Such activation is present to varying degrees in all CARs studied, with the exception of the highly effective CD19 CAR. We further identify that CD28 costimulation augments, while 4-1BB costimulation ameliorates, exhaustion induced by persistent CAR signaling. Our results provide biological explanations for the dramatic anti-tumor effects of CD19 CARs and for the observations that CD19.BBz CAR T cells are more persistent than CD19.28z CAR T cells in clinical trials. PMID:25939063
Bertin, Samuel; Aoki-Nonaka, Yukari; Lee, Jihyung; de Jong, Petrus R; Kim, Peter; Han, Tiffany; Yu, Timothy; To, Keith; Takahashi, Naoki; Boland, Brigid S; Chang, John T; Ho, Samuel B; Herdman, Scott; Corr, Maripat; Franco, Alessandra; Sharma, Sonia; Dong, Hui; Akopian, Armen N; Raz, Eyal
2017-09-01
Transient receptor potential ankyrin-1 (TRPA1) and transient receptor potential vanilloid-1 (TRPV1) are calcium (Ca 2+ )-permeable ion channels mostly known as pain receptors in sensory neurons. However, growing evidence suggests their crucial involvement in the pathogenesis of IBD. We explored the possible contribution of TRPA1 and TRPV1 to T-cell-mediated colitis. We evaluated the role of Trpa1 gene deletion in two models of experimental colitis (ie, interleukin-10 knockout and T-cell-adoptive transfer models). We performed electrophysiological and Ca 2+ imaging studies to analyse TRPA1 and TRPV1 functions in CD4+ T cells. We used genetic and pharmacological approaches to evaluate TRPV1 contribution to the phenotype of Trpa1 -/- CD4+ T cells. We also analysed TRPA1 and TRPV1 gene expression and TRPA1 + TRPV1 + T cell infiltration in colonic biopsies from patients with IBD. We identified a protective role for TRPA1 in T-cell-mediated colitis. We demonstrated the functional expression of TRPA1 on the plasma membrane of CD4+ T cells and identified that Trpa1 -/- CD4+ T cells have increased T-cell receptor-induced Ca 2+ influx, activation profile and differentiation into Th1-effector cells. This phenotype was abrogated upon genetic deletion or pharmacological inhibition of the TRPV1 channel in mouse and human CD4+ T cells. Finally, we found differential regulation of TRPA1 and TRPV1 gene expression as well as increased infiltration of TRPA1 + TRPV1 + T cells in the colon of patients with IBD. Our study indicates that TRPA1 inhibits TRPV1 channel activity in CD4+ T cells, and consequently restrains CD4+ T-cell activation and colitogenic responses. These findings may therefore have therapeutic implications for human IBD. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.
Signalling through NK1.1 triggers NK cells to die but induces NK T cells to produce interleukin-4.
Asea, A; Stein-Streilein, J
1998-02-01
In vivo inoculation of specific antibody is an accepted protocol for elimination of specific cell populations. Except for anti-CD3 and anti-CD4, it is not known if the depleted cells are eliminated by signalling through the target molecule or through a more non-specific mechanism. C57BL/6 mice were inoculated with anti-natural killer (NK1.1) monoclonal antibody (mAb). Thereafter spleen cells were harvested, stained for both surface and intracellular markers, and analysed by flow cytometry. As early as 2 hr post inoculation, NK cells were signalled to become apoptotic while signalling through the NK1.1 molecule activated NK1.1+ T-cell receptor (TCR)+ (NK T) cells to increase in number, and produce interleukin-4 (IL-4). Anti NK1.1 mAb was less efficient at signalling apoptosis in NK cells when NK T-cell deficient [beta 2-microglobulin beta 2m-deficient] mice were used compared with wild type mice. Efficient apoptotic signalling was restored when beta 2m-deficient mice were reconstituted with NK T cells. NK-specific antibody best signals the apoptotic process in susceptible NK cells when resistant NK T cells are present, activated, and secrete IL-4.
Yin, Shanshan; Zhang, Jianmin; Mao, Yujia; Hu, Yu; Cui, Lianxian; Kang, Ning; He, Wei
2013-09-13
T cell antigen receptor γδ (TCRγδ) and natural killer group 2, member D (NKG2D) are two crucial receptors for γδT cell cytotoxicity. Compelling evidences suggest that γδT cell cytotoxicity is TCRγδ-dependent and can be co-stimulated by NKG2D. However, the molecular mechanism of underlying TCRγδ-dependent activation of γδT cells remains unclear. In this study we demonstrated that TCRγδ but not NKG2D engagement induced lytic granule polarization and promoted γδT cell cytotoxicity. TCRγδ activation alone was sufficient to trigger Vav1-dependent phospholipase C-γ1 signaling, resulting in lytic granule polarization and effective killing, whereas NKG2D engagement alone failed to trigger cytotoxicity-related signaling to overcome the inhibitory effect of Cbl-b; therefore, NKG2D engagement alone could not induce effective killing. However, NKG2D ligation augmented the activation of γδT cell cytotoxicity through the Vav1-phospholipase C-γ1 pathway. Vav1 overexpression or Cbl-b knockdown not only enhanced TCRγδ activation-initiated killing but also enabled NKG2D activation alone to induce γδT cell cytotoxicity. Taken together, our results suggest that the activation of γδT cell cytotoxicity requires a strong activation signal to overcome the inhibitory effect of Cbl-b. Our finding provides new insights into the molecular mechanisms underlying the initiation of γδT cell cytotoxicity and likely implications for optimizing γδT cell-based cancer immunotherapy.
The interplay of T1- and T2-relaxation on T1-weighted MRI of hMSCs induced by Gd-DOTA-peptides.
Cao, Limin; Li, Binbin; Yi, Peiwei; Zhang, Hailu; Dai, Jianwu; Tan, Bo; Deng, Zongwu
2014-04-01
Three Gd-DOTA-peptide complexes with different peptide sequence are synthesized and used as T1 contrast agent to label human mesenchymal stem cells (hMSCs) for magnetic resonance imaging study. The peptides include a universal cell penetrating peptide TAT, a linear MSC-specific peptide EM7, and a cyclic MSC-specific peptide CC9. A significant difference in labeling efficacy is observed between the Gd-DOTA-peptides as well as a control Dotarem. All Gd-DOTA-peptides as well as Dotarem induce significant increase in T1 relaxation rate which is in favor of T1-weighted MR imaging. Gd-DOTA-CC9 yields the maximum labeling efficacy but poor T1 contrast enhancement. Gd-DOTA-EM7 yields the minimum labeling efficacy but better T1 contrast enhancement. Gd-DOTA-TAT yields a similar labeling efficacy as Gd-DOTA-CC9 and similar T1 contrast enhancement as Gd-DOTA-EM7. The underlying mechanism that governs T1 contrast enhancement effect is discussed. Our results suggest that T1 contrast enhancement induced by Gd-DOTA-peptides depends not only on the introduced cellular Gd content, but more importantly on the effect that Gd-DOTA-peptides exert on the T1-relaxation and T2-relaxation processes/rates. Both T1 and particularly T2 relaxation rate have to be taken into account to interpret T1 contrast enhancement. In addition, the interpretation has to be based on cellular instead of aqueous longitudinal and transverse relaxivities of Gd-DOTA-peptides. Copyright © 2014 Elsevier Ltd. All rights reserved.
Probiotic Bifidobacterium breve induces IL-10-producing Tr1 cells in the colon.
Jeon, Seong Gyu; Kayama, Hisako; Ueda, Yoshiyasu; Takahashi, Takuya; Asahara, Takashi; Tsuji, Hirokazu; Tsuji, Noriko M; Kiyono, Hiroshi; Ma, Ji Su; Kusu, Takashi; Okumura, Ryu; Hara, Hiromitsu; Yoshida, Hiroki; Yamamoto, Masahiro; Nomoto, Koji; Takeda, Kiyoshi
2012-01-01
Specific intestinal microbiota has been shown to induce Foxp3(+) regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103(+) dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103(+) DCs from Il10(-/-), Tlr2(-/-), and Myd88(-/-) mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103(+) DCs failed to induce IL-10 production from co-cultured Il27ra(-/-) T cells. B. breve treatment of Tlr2(-/-) mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103(+) DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4(+) T cells from wild-type mice, but not Il10(-/-) mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.
DOE Office of Scientific and Technical Information (OSTI.GOV)
Sheu, C.W.; Moreland, F.M.; Dunkel, V.C.
1987-01-01
The response of BALB/3T3 clone A31-1-1 cells to chemically induced morphological transformation was evaluated using 3-methylcholanthrene (MCA). Stock cultures were initiated from cryopreserved cells, grown in T25 flasks containing 5 ml of medium, and replated at subconfluency. Serially transferred cells were then subjected to transformation assay. After 24-hr seeding, cells were incubated 48 hr with MCA in a 5% CO/sub 2/ incubator. They were then rinsed and incubated for an additional 4 weeks with twice weekly medium change. Type III foci were scored after fixation and staining with Giemsa. With serial passage from the frozen state, cells of passages 3-14more » had a low level of spontaneous transformation; zero to 6 type III foci per 20 dishes were counted. In the MCA-treated cultures the number of transformed foci, however, increased with passage. Such passage-related sensitivity to MCA was demonstrated for cells cultured in two batches of sera: one from MA Bioproducts (Lot no. 2E052) and the other from Armour Pharmaceuticals (Lot no. Y65801). The passage-related increase in number of transformed foci was not related to doubling time, cloning efficiency, or MCA-induced growth inhibition.« less
Zeng, Zhiwen; Wang, Haitao; Shang, Fu; Zhou, Lihua; Little, Peter J; Quirion, Remi; Zheng, Wenhua
2016-03-01
Lithium is currently used in the treatment of mental illness. We have previously reported that lithium stimulated the protein kinase B/Forkhead box O1 (Akt/FoxO1) pathway in rats. However, little information is available regarding its neuroprotective role of this pathway and underlying mechanisms. PC12 cells treated with serum deprivation were used as a toxicity model to study the protective effect of lithium and its underlying mechanisms. Cell viability was determined by methyl thiazolyl tetrazolium assay and Hoechst staining. FoxO1 subcellular location and its overexpression were used to study the underlying mechanisms. Various pathway inhibitors were used to investigate the possible pathways, while the phosphorylation of Akt and FoxO1 was analyzed by Western blot. Lithium pretreatment dose-dependently reduced PC12 cell apoptosis induced by serum starvation. The protective effect of lithium was abolished by LY294002, a PI3K-specific inhibitor, and Akt inhibitor Akt inhibitor VIII, whereas mitogen-activated protein kinase kinase (MEK kinase) inhibitor U0126 had no effect. Lithium induced the phosphorylation of Akt and FoxO1 in a time- and concentration-dependent manner. Lithium-induced phosphorylation of Akt and FoxO1 is mediated by the PI3K/Akt pathway. Serum deprivation caused nuclear translocation of FoxO1 while application of lithium reversed the effect of serum deprivation. Moreover, overexpression of FoxO1 enhanced cell apoptosis induced by serum withdrawal. Finally, lithium was found to reduce the exogenous and endogenous FoxO1 protein levels in PC12 cells in a concentration-dependent fashion. The protective effect of lithium against serum starvation cell death is mediated by the PI3K/Akt/FoxO1 pathway.
T-bet expression by Th cells promotes type 1 inflammation but is dispensable for colitis.
Zimmermann, J; Kühl, A A; Weber, M; Grün, J R; Löffler, J; Haftmann, C; Riedel, R; Maschmeyer, P; Lehmann, K; Westendorf, K; Mashreghi, M-F; Löhning, M; Mack, M; Radbruch, A; Chang, H D
2016-11-01
The transcription factor T-bet is highly expressed by Th cells isolated from the inflamed intestine of Crohn's disease patients, and has been regarded a critical driver of murine T cell-induced colitis. However, we show here that T-bet expression by Th cells is not required for the manifestation of T-cell-induced colitis in the presence of segmented filamentous bacteria and Helicobacter hepaticus. T-bet expression by Th cells controls their survival and localization, their repertoire of chemokine and chemokine receptor expression, the accumulation of monocytes and macrophages in the inflamed colon, and their differentiation to the M1 type, i.e., type 1 inflammation. Nevertheless, T-bet-deficient Th cells efficiently induce colitis, as reflected by weight loss, diarrhea, and colon histopathology. T-bet-deficient Th cells differentiate into Th1/17 cells, able to express IFN-γ and IL-17A upon restimulation. While neutralization of IL-17A exacerbated colitis induced by wild-type or T-bet-deficient Th cells, neutralization of IFN-γ completely abolished colitis.
Regulatory T cells ameliorate tissue plasminogen activator-induced brain haemorrhage after stroke.
Mao, Leilei; Li, Peiying; Zhu, Wen; Cai, Wei; Liu, Zongjian; Wang, Yanling; Luo, Wenli; Stetler, Ruth A; Leak, Rehana K; Yu, Weifeng; Gao, Yanqin; Chen, Jun; Chen, Gang; Hu, Xiaoming
2017-07-01
Delayed thrombolytic treatment with recombinant tissue plasminogen activator (tPA) may exacerbate blood-brain barrier breakdown after ischaemic stroke and lead to lethal haemorrhagic transformation. The immune system is a dynamic modulator of stroke response, and excessive immune cell accumulation in the cerebral vasculature is associated with compromised integrity of the blood-brain barrier. We previously reported that regulatory T cells, which function to suppress excessive immune responses, ameliorated blood-brain barrier damage after cerebral ischaemia. This study assessed the impact of regulatory T cells in the context of tPA-induced brain haemorrhage and investigated the underlying mechanisms of action. The number of circulating regulatory T cells in stroke patients was dramatically reduced soon after stroke onset (84 acute ischaemic stroke patients with or without intravenous tPA treatment, compared to 115 age and gender-matched healthy controls). Although stroke patients without tPA treatment gradually repopulated the numbers of circulating regulatory T cells within the first 7 days after stroke, post-ischaemic tPA treatment led to sustained suppression of regulatory T cells in the blood. We then used the murine suture and embolic middle cerebral artery occlusion models of stroke to investigate the therapeutic potential of adoptive regulatory T cell transfer against tPA-induced haemorrhagic transformation. Delayed administration of tPA (10 mg/kg) resulted in haemorrhagic transformation in the ischaemic territory 1 day after ischaemia. When regulatory T cells (2 × 106/mouse) were intravenously administered immediately after delayed tPA treatment in ischaemic mice, haemorrhagic transformation was significantly decreased, and this was associated with improved sensorimotor functions. Blood-brain barrier disruption and tight junction damages were observed in the presence of delayed tPA after stroke, but were mitigated by regulatory T cell transfer. Mechanistic
De Ford, C; Heidersdorf, B; Haun, F; Murillo, R; Friedrich, T; Borner, C; Merfort, I
2016-01-01
T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy that preferentially affects children and adolescents. Over 50% of human T-ALLs possess activating mutations of Notch1. The clerodane diterpene casearin J (CJ) is a natural product that inhibits the sarcoendoplasmatic reticulum calcium ATPase (SERCA) pump and induces cell death in leukemia cells, but the molecular mechanism of cytotoxicity remains poorly understood. Here we show that owing to SERCA pump inhibition, CJ induces depletion of the endoplasmic reticulum calcium pools, oxidative stress, and apoptosis via the intrinsic signaling pathway. Moreover, Notch1 signaling is reduced in T-ALL cells with auto-activating mutations in the HD-domain of Notch1, but not in cells that do not depend on Notch1 signaling. CJ also provoked a slight activation of NF-κB, and consistent with this notion a combined treatment of CJ and the NF-κB inhibitor parthenolide (Pt) led to a remarkable synergistic cell death in T-ALL cells. Altogether, our data support the concept that inhibition of the SERCA pump may be a novel strategy for the treatment of T-ALL with HD-domain-mutant Notch1 receptors and that additional treatment with the NF-κB inhibitor parthenolide may have further therapeutic benefits. PMID:26821066
Bae, Jooeun; Prabhala, Rao; Voskertchian, Annie; Brown, Andrew; Maguire, Craig; Richardson, Paul; Dranoff, Glen; Anderson, Kenneth C.; Munshi, Nikhil C.
2014-01-01
We evaluated a cocktail of HLA-A2-specific peptides including heteroclitic XBP1 US184-192 (YISPWILAV), heteroclitic XBP1 SP367-375 (YLFPQLISV), native CD138260-268 (GLVGLIFAV) and native CS1239-247 (SLFVLGLFL), for their ability to elicit multipeptide specific cytotoxic T lymphocytes (MP-CTL) using T cells from smoldering multiple myeloma (SMM) patients. Our results demonstrate that MP-CTL generated from SMM patients’ T cells show effective anti-MM responses including CD137 (4-1BB) upregulation, CTL proliferation, IFN-γ production, and degranulation (CD107a) in an HLA-A2-restricted and peptide-specific manner. Phenotypically, we observed increased total CD3+CD8+ T cells (>80%) and cellular activation (CD69+) within the memory SMM MP-CTL (CD45RO+/CD3+CD8+) subset after repeated multipeptide stimulation. Importantly, SMM patients could be categorized into distinct groups by their level of MP-CTL expansion and anti-tumor activity. In high responders, the effector memory (CCR7-CD45RO+/CD3+CD8+) T cell subset was enriched, while the remaining responders’ CTL contained a higher frequency of the terminal effector (CCR7-CD45RO-/CD3+CD8+) subset. These results suggest that this multipeptide cocktail has the potential to induce effective and durable memory MP-CTL in SMM patients. Therefore, our findings provide the rationale for clinical evaluation of a therapeutic vaccine to prevent or delay progression of SMM to active disease. PMID:24935722
Ding, Zhi-Chun; Lu, Xiaoyun; Yu, Miao; Lemos, Henrique; Huang, Lei; Chandler, Phillip; Liu, Kebin; Walters, Matthew; Krasinski, Antoni; Mack, Matthias; Blazar, Bruce R; Mellor, Andrew L; Munn, David H; Zhou, Gang
2014-07-01
In recent years, immune-based therapies have become an increasingly attractive treatment option for patients with cancer. Cancer immunotherapy is often used in combination with conventional chemotherapy for synergistic effects. The alkylating agent cyclophosphamide (CTX) has been included in various chemoimmunotherapy regimens because of its well-known immunostimulatory effects. Paradoxically, cyclophosphamide can also induce suppressor cells that inhibit immune responses. However, the identity and biologic relevance of these suppressor cells are poorly defined. Here we report that cyclophosphamide treatment drives the expansion of inflammatory monocytic myeloid cells (CD11b(+)Ly6C(hi)CCR2(hi)) that possess immunosuppressive activities. In mice with advanced lymphoma, adoptive transfer (AT) of tumor-specific CD4(+) T cells following cyclophosphamide treatment (CTX+CD4 AT) provoked a robust initial antitumor immune response, but also resulted in enhanced expansion of monocytic myeloid cells. These therapy-induced monocytes inhibited long-term tumor control and allowed subsequent relapse by mediating functional tolerization of antitumor CD4(+) effector cells through the PD-1-PD-L1 axis. PD-1/PD-L1 blockade after CTX+CD4 AT therapy led to persistence of CD4(+) effector cells and durable antitumor effects. Depleting proliferative monocytes by administering low-dose gemcitabine effectively prevented tumor recurrence after CTX+CD4 AT therapy. Similarly, targeting inflammatory monocytes by disrupting the CCR2 signaling pathway markedly potentiated the efficacy of cyclophosphamide-based therapy. Besides cyclophosphamide, we found that melphalan and doxorubicin can also induce monocytic myeloid suppressor cells. These findings reveal a counter-regulation mechanism elicited by certain chemotherapeutic agents and highlight the importance of overcoming this barrier to prevent late tumor relapse after chemoimmunotherapy. ©2014 American Association for Cancer Research.
Notch ligands Delta1 and Jagged1 transmit distinct signals to T-cell precursors
Lehar, Sophie M.; Dooley, James; Farr, Andrew G.; Bevan, Michael J.
2009-01-01
Signaling through the Notch pathway plays an essential role in inducing T-lineage commitment and promoting the maturation of immature thymocytes. Using an in vitro culture system, we show that 2 different classes of Notch ligands, Jagged1 or Delta1, transmit distinct signals to T-cell progenitors. OP9 stromal cells expressing either Jagged1 or Delta1 inhibit the differentiation of DN1 thymocytes into the B-cell lineage, but only the Delta1-expressing stromal cells promote the proliferation and maturation of T-cell progenitors through the early double-negative (DN) stages of thymocyte development. Whereas the majority of bone marrow-derived stem cells do not respond to Jagged1 signals, T-cell progenitors respond to Jagged1 signals during a brief window of their development between the DN1 and DN3 stages of thymic development. During these stages, Jagged1 signals can influence the differentiation of immature thymocytes along the natural killer (NK) and γδ T-cell lineages. PMID:15486060
Santana, Vinicius Canato; Almeida, Rafael Ribeiro; Ribeiro, Susan Pereira; Ferreira, Luís Carlos de Souza; Kalil, Jorge; Rosa, Daniela Santoro; Cunha-Neto, Edecio
2015-01-01
T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity. PMID:26602876
Park, Jeongho; Goergen, Craig J; HogenEsch, Harm; Kim, Chang H
2016-03-01
Short-chain fatty acids (SCFAs) are major products of gut microbial fermentation and profoundly affect host health and disease. SCFAs generate IL-10(+) regulatory T cells, which may promote immune tolerance. However, SCFAs can also induce Th1 and Th17 cells upon immunological challenges and, therefore, also have the potential to induce inflammatory responses. Because of the seemingly paradoxical SCFA activities in regulating T cells, we investigated, in depth, the impact of elevated SCFA levels on T cells and tissue inflammation in mice. Orally administered SCFAs induced effector (Th1 and Th17) and regulatory T cells in ureter and kidney tissues, and they induced T cell-mediated ureteritis, leading to kidney hydronephrosis (hereafter called acetate-induced renal disease, or C2RD). Kidney hydronephrosis in C2RD was caused by ureteral obstruction, which was, in turn, induced by SCFA-induced inflammation in the ureteropelvic junction and proximal ureter. Oral administration of all major SCFAs, such as acetate, propionate, and butyrate, induced the disease. We found that C2RD development is dependent on mammalian target of rapamycin activation, T cell-derived inflammatory cytokines such as IFN-γ and IL-17, and gut microbiota. Young or male animals were more susceptible than old or female animals, respectively. However, SCFA receptor (GPR41 or GPR43) deficiency did not affect C2RD development. Thus, SCFAs, when systemically administered at levels higher than physiological levels, cause dysregulated T cell responses and tissue inflammation in the renal system. The results provide insights into the immunological and pathological effects of chronically elevated SCFAs. Copyright © 2016 by The American Association of Immunologists, Inc.
Racila, E; Hsueh, R; Marches, R; Tucker, T F; Krammer, P H; Scheuermann, R H; Uhr, J W
1996-01-01
Signal transduction initiated by crosslinking of antigen-specific receptors on T- and B-lymphoma cells induces apoptosis. In T-lymphoma cells, such crosslinking results in upregulation of the APO-1 ligand, which then interacts with induced or constitutively expressed APO-1, thereby triggering apoptosis. Here we show that crosslinking the membrane immunoglobulin on human lymphoma cells (Daudi) (that constitutively express APO-1) does not induce synthesis of APO-1 ligand. Further, a noncytotoxic fragment of anti-APO-1 antibody that blocks T-cell-receptor-mediated apoptosis in T-lymphoma cells does not block anti-mu-induced apoptosis. Hence, in B-lymphoma cells, apoptosis induced by signaling via membrane IgM is not mediated by the APO-1 ligand. Images Fig. 2 Fig. 3 PMID:8700902
Probiotic Bifidobacterium breve Induces IL-10-Producing Tr1 Cells in the Colon
Ueda, Yoshiyasu; Takahashi, Takuya; Asahara, Takashi; Tsuji, Hirokazu; Tsuji, Noriko M.; Kiyono, Hiroshi; Ma, Ji Su; Kusu, Takashi; Okumura, Ryu; Hara, Hiromitsu; Yoshida, Hiroki; Yamamoto, Masahiro; Nomoto, Koji; Takeda, Kiyoshi
2012-01-01
Specific intestinal microbiota has been shown to induce Foxp3+ regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103+ dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103+ DCs from Il10 −/−, Tlr2 −/−, and Myd88 −/− mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103+ DCs failed to induce IL-10 production from co-cultured Il27ra −/− T cells. B. breve treatment of Tlr2 −/− mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103+ DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4+ T cells from wild-type mice, but not Il10 −/− mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells. PMID:22693446
Tano, Tomoyuki; Okamoto, Masato; Kan, Shin; Bando, Takashi; Goda, Hiroyuki; Nakashiro, Koh-ichi; Shimodaira, Shigetaka; Koido, Shigeo; Homma, Sadamu; Fujita, Tomonobu; Sato, Mitsunobu; Yamashita, Naomi; Hamakawa, Hiroyuki; Kawakami, Yutaka
2013-01-01
Eighty-one patients with oral squamous cell carcinoma (OSCC) received oral fluoropyrimidine UFT and radiotherapy (RT) with or without an immunotherapeutic agent OK-432. Both overall survival and progression-free survival of patients who received RT + UFT + OK-432 were significantly longer than those of patients who received RT + UFT (P = .0075 and P = .0175, respectively). Clinical response was also more favorable in RT + UFT + OK-432 group than in RT + UFT group (P = .0066). Next, in vitro experiments were conducted to examine the effect of 5-fluorouracil (5-FU) and X-ray irradiation in OK-432-induced immunity. Human peripheral blood mononuclear cells stimulated with OK-432 produced helper T cell 1 (Th1)-type cytokines as well as interleukin-10 (IL-10) and transforming growth factor-β (TGF-β), which are produced by Th2 and regulatory T cells (Tregs), respectively, and are inhibitory in antitumor immunity. OK-432-induced IL-10 and TGF-β but not Th1 cytokines were significantly inhibited by 5-FU and/or X-ray. 5-FU and X-ray also inhibited the expression of mRNAs for GATA-3 and Foxp3, which are transcription factors for Th2 and Tregs, respectively, but not for T-bet, a transcription factor for Th1. In addition, 5-FU and X-ray decreased the expression of mRNAs for suppressor of cytokine signaling 1 (SOCS1) and SOCS3. Antisense oligonucleotides for SOCS1 and SOCS3 markedly reduced OK-432-induced IL-10 and TGF-β. This is the first report clearly demonstrating that OK-432-based immunotherapy significantly enhanced the therapeutic effects of chemoradiotherapy in patients with OSCC as well as elucidating the mechanism of the synergistic effect of immunochemoradiotherapy in which 5-FU and radiation enhanced OK-432-induced Th1 response mediated by the inhibition of SOCS1 and SOCS3 gene expression. PMID:23814492
Eomesodermin Promotes the Development of Type-1 Regulatory T (TR1) Cells
Zhang, Ping; Lee, Jason S.; Gartlan, Kate H.; Schuster, Iona S; Comerford, Iain; Varelias, Antiopi; Ullah, Md Ashik; Vuckovic, Slavica; Koyama, Motoko; Kuns, Rachel D.; Locke, Kelly R.; Beckett, Kirrilee J.; Olver, Stuart D.; Samson, Luke D.; de Oca, Marcela Montes; de Labastida Rivera, Fabian; Clouston, Andrew D.; Belz, Gabrielle T.; Blazar, Bruce R.; MacDonald, Kelli P.; McColl, Shaun R.; Thomas, Ranjeny; Engwerda, Christian R.; Degli-Esposti, Mariapia A.; Kallies, Axel; Tey, Siok-Keen; Hill, Geoffrey R.
2017-01-01
Type-1 regulatory T (TR1) cells are Foxp3-negative IL-10-producing CD4+ T cells with potent immune suppressive properties but their requirements for lineage development have remained elusive. Here we show that TR1 cells constitute the most abundant regulatory population after allogeneic bone marrow transplantation (BMT), express the transcription factor Eomesodermin (Eomes) and are critical for the prevention of graft-versus-host disease (GVHD). We demonstrate that Eomes is required for TR1 cell differentiation during which it acts in concert with the transcription factor B-lymphocyte-induced maturation protein-1 (Blimp-1) by transcriptionally activating IL-10 expression and repressing differentiation into other Th lineages. We further show that Eomes induction in TR1 cells requires T-bet and donor macrophage-derived IL-27. We thus define the cellular and transcriptional control of TR1 cell differentiation during bone marrow transplantation, opening new avenues to therapeutic manipulation. PMID:28738016
TNF-α blockade induces IL-10 expression in human CD4+ T cells
NASA Astrophysics Data System (ADS)
Evans, Hayley G.; Roostalu, Urmas; Walter, Gina J.; Gullick, Nicola J.; Frederiksen, Klaus S.; Roberts, Ceri A.; Sumner, Jonathan; Baeten, Dominique L.; Gerwien, Jens G.; Cope, Andrew P.; Geissmann, Frederic; Kirkham, Bruce W.; Taams, Leonie S.
2014-02-01
IL-17+ CD4+ T (Th17) cells contribute to the pathogenesis of several human inflammatory diseases. Here we demonstrate that TNF inhibitor (TNFi) drugs induce the anti-inflammatory cytokine IL-10 in CD4+ T cells including IL-17+ CD4+ T cells. TNFi-mediated induction of IL-10 in IL-17+ CD4+ T cells is Treg-/Foxp3-independent, requires IL-10 and is overcome by IL-1β. TNFi-exposed IL-17+ CD4+ T cells are molecularly and functionally distinct, with a unique gene signature characterized by expression of IL10 and IKZF3 (encoding Aiolos). We show that Aiolos binds conserved regions in the IL10 locus in IL-17+ CD4+ T cells. Furthermore, IKZF3 and IL10 expression levels correlate in primary CD4+ T cells and Aiolos overexpression is sufficient to drive IL10 in these cells. Our data demonstrate that TNF-α blockade induces IL-10 in CD4+ T cells including Th17 cells and suggest a role for the transcription factor Aiolos in the regulation of IL-10 in CD4+ T cells.
Dobashi, H; Seki, S; Habu, Y; Ohkawa, T; Takeshita, S; Hiraide, H; Sekine, I
1999-08-01
Although bacterial superantigens have been well characterized as potent stimulators of T cells, their role in natural killer (NK)-type cells remains largely unknown. In the present study, we examined the effect of bacterial superantigens on mouse liver NK cells and NK1.1 Ag(+) (NK1(+)) T cells. C57BL/6 mice were intravenously injected with staphylococcal enterotoxin B (SEB) or streptococcal pyrogenic exotoxin A (SPE-A), and mononuclear cells (MNC) of various organs were obtained from mice 4 hours after being injected with superantigen. MNC were cultured for 48 hours, and interferon gamma (IFN-gamma) levels of supernatants were measured. The antitumor cytotoxicities of the liver and spleen MNC were also evaluated 24 hours after the mice were injected with superantigen. Liver MNC produced more IFN-gamma than did splenocytes, and peripheral blood and lung MNC did not produce any detectable IFN-gamma. In addition, liver MNC acquired a potent antitumor cytotoxicity by the SEB injection, and both NK cells and NK1(+)T cells but not cluster of differentiation (CD)8(+) T cells were responsible for the cytotoxicity as demonstrated by either in vivo or in vitro cell depletion experiments, and the NK-type cells were partly responsible for the increased serum IFN-gamma. Activation of liver NK-type cells was also supported by the fact that liver NK cells proportionally increased and NK1(+) T cells augmented their CD11a expressions after SEB injection. The pretreatment of mice with anti-IFN-gamma Ab and/or with anti-interleukin-12 (IL-12) Ab diminished the SEB-induced cytotoxicity of liver MNC. Furthermore, the in vivo depletion of Kupffer cells decreased the SEB-induced cytotoxicity of liver MNC. Consistent with these results, liver MNC stimulated with superantigens in the presence of Kupffer cells in vitro produced a greater amount of IFN-gamma than did the liver MNC without Kupffer cells or splenocytes. Our results suggest that bacterial superantigen-primed Kupffer cells
Klemke, Claus-Detlev; Brenner, Dirk; Weiss, Eva-Maria; Schmidt, Marc; Leverkus, Martin; Gülow, Karsten; Krammer, Peter H
2009-05-15
Restimulation of previously activated T cells via the T-cell receptor (TCR) leads to activation-induced cell death (AICD), which is, at least in part, dependent on the death receptor CD95 (APO-1, FAS) and its natural ligand (CD95L). Here, we characterize cutaneous T-cell lymphoma (CTCL) cells (CTCL tumor cell lines and primary CTCL tumor cells from CTCL patients) as AICD resistant. We show that CTCL cells have elevated levels of the CD95-inhibitory protein cFLIP. However, cFLIP is not responsible for CTCL AICD resistance. Instead, our data suggest that reduced TCR-proximal signaling in CTCL cells is responsible for the observed AICD resistance. CTCL cells exhibit no PLC-gamma1 activity, resulting in an impaired Ca(2+)release and reduced generation of reactive oxygen species upon TCR stimulation. Ca(2+) and ROS production are crucial for up-regulation of CD95L and reconstitution of both signals resulted in AICD sensitivity of CTCL cells. In accordance with these data, CTCL tumor cells from patients with Sézary syndrome do not up-regulate CD95L upon TCR-stimulation and are therefore resistant to AICD. These results show a novel mechanism of AICD resistance in CTCL that could have future therapeutic implications to overcome apoptosis resistance in CTCL patients.
Song, K-J; Jang, Y S; Lee, Y A; Kim, K A; Lee, S K; Shin, M H
2011-07-01
Naegleria fowleri, a free-living amoeba, is the causative pathogen of primary amoebic meningoencephalitis in humans and experimental mice. N. fowleri is capable of destroying tissues and host cells through lytic necrosis. However, the mechanism by which N. fowleri induces host cell death is unknown. Electron microscopy indicated that incubation of Jurkat T cells with N. fowleri trophozoites induced necrotic morphology of the Jurkat T cells. N. fowleri also induced cytoskeletal protein cleavage, extensive poly (ADP-ribose) polymerase hydrolysis and lactate dehydrogenase (LDH) release. Although no activation of caspase-3 was observed in Jurkat T cells co-incubated with amoebae, intracellular reactive oxygen species (ROS) were strongly generated by NADPH oxidase (NOX). Pretreating cells with necroptosis inhibitor necrostatin-1 or NOX inhibitor diphenyleneiodonium chloride (DPI) strongly inhibited amoeba-induced ROS generation and Jurkat cell death, whereas pan-caspase inhibitor z-VAD-fmk did not. N. fowleri-derived secretory products (NfSP) strongly induced intracellular ROS generation and cell death. Necroptotic effects of NfSP were effectively inhibited by pretreating NfSP with proteinase K. Moreover, NfSP-induced LDH release and intracellular ROS accumulation were inhibited by pretreating Jurkat T cells with DPI or necrostatin-1. These results suggest that N. fowleri induces ROS-dependent necroptosis in Jurkat T cells. © 2011 Blackwell Publishing Ltd.
Deletion of BCG Hip1 protease enhances dendritic cell and CD4 T cell responses.
Bizzell, Erica; Sia, Jonathan Kevin; Quezada, Melanie; Enriquez, Ana; Georgieva, Maria; Rengarajan, Jyothi
2018-04-01
Dendritic cells (DCs) play a key role in the generation of CD4 T cell responses to pathogens. Mycobacterium tuberculosis (Mtb) harbors immune evasion mechanisms that impair DC responses and prevent optimal CD4 T cell immunity. The vaccine strain Mycobacterium bovis Bacille Calmette-Guérin (BCG) shares many of the immune evasion proteins utilized by Mtb, but the role of these proteins in DC and T cell responses elicited by BCG is poorly understood. We previously reported that the Mtb serine protease, Hip1, promotes sub-optimal DC responses during infection. Here, we tested the hypothesis that BCG Hip1 modulates DC functions and prevents optimal antigen-specific CD4 T cell responses that limit the immunogenicity of BCG. We generated a strain of BCG lacking hip1 (BCGΔhip1) and show that it has superior capacity to induce DC maturation and cytokine production compared with the parental BCG. Furthermore, BCGΔhip1-infected DCs were more effective at driving the production of IFN-γ and IL-17 from antigen-specific CD4 T cells in vitro. Mucosal transfer of BCGΔhip1-infected DCs into mouse lungs induced robust CD4 T cell activation in vivo and generated antigen-specific polyfunctional CD4 T cell responses in the lungs. Importantly, BCGΔhip1-infected DCs enhanced control of pulmonary bacterial burden following Mtb aerosol challenge compared with the transfer of BCG-infected DCs. These results reveal that BCG employs Hip1 to impair DC activation, leading to attenuated lung CD4 T cell responses with limited capacity to control Mtb burden after challenge. ©2017 Society for Leukocyte Biology.
Lin, Ze-Min; Yang, Xiao-Qian; Zhu, Feng-Hua; He, Shi-Jun; Tang, Wei; Zuo, Jian-Ping
2016-01-01
SM934 is an artemisinin analogue with immunosuppressive properties and potent therapeutic activity against lupus-like diseases in autoimmune mice. In this report, the therapeutic efficacy and underlying mechanisms of SM934 on rheumatoid arthritis (RA) was investigated using collagen-induced arthritis (CIA) in DBA/1J mice. We demonstrated that SM934 treatment alleviate the severity of arthritis in CIA mice with established manifestations. The therapeutic benefits were associated with ameliorated joint swelling and reduced extent of bone erosion and destruction. Further, administration of SM934 diminished the development of T follicular helper (Tfh) cells and Th17 cells and suppressed the production of pathogenic antibodies, without altering the proportion of germinal center B cells. Ex vivo, SM934 treatment inhibited the bovine type II collagen (CII) induced proliferation and inflammatory cytokines secretion of CII -reactive T cells. In vitro, SM934 impeded the polarization of naïve CD4+ T cells into Tfh cells and the expression of its transcript factor Bcl-6. Moreover, SM934 decreased the IL-21-producing CD4+ T cells and dampened the IL-21 downstream signaling through STAT3. These finding offered the convincing evidence that artemisinin derivative might attenuate RA by simultaneously interfering with the generation of Tfh cells and Th17 cells as well as the subsequent antibody-mediated immune responses. PMID:27897259
Milasta, Sandra; Dillon, Christopher P; Sturm, Oliver E; Verbist, Katherine C; Brewer, Taylor L; Quarato, Giovanni; Brown, Scott A; Frase, Sharon; Janke, Laura J; Perry, S Scott; Thomas, Paul G; Green, Douglas R
2016-01-19
The role of apoptosis inducing factor (AIF) in promoting cell death versus survival remains controversial. We report that the loss of AIF in fibroblasts led to mitochondrial electron transport chain defects and loss of proliferation that could be restored by ectopic expression of the yeast NADH dehydrogenase Ndi1. Aif-deficiency in T cells led to decreased peripheral T cell numbers and defective homeostatic proliferation, but thymic T cell development was unaffected. In contrast, Aif-deficient B cells developed and functioned normally. The difference in the dependency of T cells versus B cells on AIF for function and survival correlated with their metabolic requirements. Ectopic Ndi1 expression rescued homeostatic proliferation of Aif-deficient T cells. Despite its reported roles in cell death, fibroblasts, thymocytes and B cells lacking AIF underwent normal death. These studies suggest that the primary role of AIF relates to complex I function, with differential effects on T and B cells. Copyright © 2016 Elsevier Inc. All rights reserved.
SLP-76-ADAP adaptor module regulates LFA-1 mediated costimulation and T cell motility.
Wang, Hongyan; Wei, Bin; Bismuth, Georges; Rudd, Christopher E
2009-07-28
Although adaptor ADAP (FYB) and its binding to SLP-76 has been implicated in TcR-induced "inside-out" signaling for LFA-1 activation in T cells, little is known regarding its role in LFA-1-mediated "outside-in" signaling. In this study, we demonstrate that ADAP and SLP-76-ADAP binding are coupled to LFA-1 costimulation of IL-2 production, F-actin clustering, cell polarization, and T cell motility. LFA-1 enhancement of anti-CD3-induced IL-2 production was completely dependent on SLP-76-ADAP binding. Further, anti-CD3 was found to require CD11a ligation by antibody or ICAM1 to cause T cell polarization. ADAP augmented this polarization induced by anti-CD3/CD11a, but not by anti-CD3 alone. ADAP expression with LFA-1 ligation alone was sufficient to polarize T cells directly and to increase T cell motility whereas the loss of ADAP in ADAP-/- primary T cells reduced motility. A mutant lacking SLP-76-binding sites (M12) blocked LFA-1 costimulation of IL-2 production, polarization, and motility. LFA-1-ADAP polarization was also dependent on src kinases, Rho GTPases, phospholipase C, and phosphoinositol 3-kinase. Our findings provide evidence of an obligatory role for the SLP-76-ADAP module in LFA-1-mediated costimulation in T cells.
[TNF-α, diabetes type 1 and regulatory T cells].
Ryba, Monika; Myśliwska, Jolanta
2010-01-01
Recent studies on animal models of diabetes as well as human regulatory T cells have shown that α impairs the ability of these cells to prevent the disease. NOD mice treated with α had decreased frequency of regulatory T cells, whereas anti-TNF administration induced the increase in the number of these cells and disease prevention. The action of α also influenced the suppressive potential of Tregs. Increased susceptibility of Tregs to the modulatory effects of α involves signaling through TNFR2 that is expressed on the surface of this cell population. It seems that α neutralization may rescue regulatory T cells and restore their function in several autoimmune and inflammatory diseases. This review describes recent data concerning regulatory T cells in the context of inflammation that is present during diabetes type 1. It describes how TNF contributes to the pathogenesis of type 1 diabetes, what is the impact of this cytokine on regulatory T cell population and therapeutic effects that result from its neutralization in several inflammatory and autoimmune diseases.
Role of T-bet, the master regulator of Th1 cells, in the cytotoxicity of murine CD4+ T cells.
Eshima, Koji; Misawa, Kana; Ohashi, Chihiro; Iwabuchi, Kazuya
2018-05-01
Although CD4 + T cells are generally regarded as helper T cells, some activated CD4 + T cells have cytotoxic properties. Given that CD4 + cytotoxic T lymphocytes (CTLs) often secrete IFN-γ, CTL activity among CD4 + T cells may be attributable to Th1 cells, where a T-box family molecule, T-bet serves as the "master regulator". However, although the essential contribution of T-bet to expression of IFN-γ has been well-documented, it remains unclear whether T-bet is involved in CD4 + T cell-mediated cytotoxicity. In this study, to investigate the ability of T-bet to confer cytolytic activity on CD4 + T cells, the T-bet gene (Tbx21) was introduced into non-cytocidal CD4 + T cell lines and their cytolytic function analyzed. Up-regulation of FasL (CD178), which provided the transfectant with cytotoxicity, was observed in Tbx21transfected CD4 + T cells but not in untransfected parental cells. In one cell line, T-bet transduction also induced perforin gene (Prf1) expression and Tbx21 transfectants efficiently killed Fas - target cells. Although T-bet was found to repress up-regulation of CD40L (CD154), which controls FasL-mediated cytolysis, the extent of CD40L up-regulation on in vitro-differentiated Th1 cells was similar to that on Th2 cells, suggesting the existence of a compensatory mechanism. These results collectively indicate that T-bet may be involved in the expression of genes, such as FasL and Prf1, which confer cytotoxicity on Th1 cells. © 2018 The Societies and John Wiley & Sons Australia, Ltd.
Reprogramming T cell Lymphocytes to Induced Pluripotent Stem Cells
NASA Astrophysics Data System (ADS)
Bared, Kalia
The discovery of induced pluripotent stem cells (iPSC) provided a novel technology for the study of development and pharmacology and complement embryonic stem cells (ES) for cell therapy applications. Though iPSC are derived from adult tissue they are comparable to ES cells in their behavior; multi-lineage differentiation and self-renewal. This makes iPSC research appealing because they can be studied in great detail and expanded in culture broadly. Fibroblasts were the first cell type reprogrammed to an iPSC using a retrovirus vector, since then alternative cell types including lymphocytes have been used to generate iPSC. Different types of vectors have also been developed to enhance iPSC formation and quality. However, specific T lymphocyte subsets have not been shown to reprogram to a pluripotent state to date. Here, we proposed to derive iPSC from peripheral blood effector and central memory T cells, reasoning that the resultant iPSC will maintain the epigenetic memory of a T lymphocyte, including the T cell receptor (TCR) gene rearrangement. This epigenetic memory will enable the differentiation and expansion of T cell iPSC into professional T cells containing a specific TCR. These could then be used for cell therapy to target specific antigens, as well as to improve culture techniques to expand T cells
Cuda, Carla M.; Zeumer, Leilani; Sobel, Eric S.; Croker, Byron P.; Morel, Laurence
2010-01-01
The NZM2410-derived Sle1a lupus susceptibility locus induces activated autoreactive CD4+ T cells and reduces the number and function of Foxp3+ regulatory T cells. In this study, we first showed that Sle1a contributes to autoimmunity by increasing anti-nuclear antibody production when expressed on either NZB or NZW heterozygous genomes, and by enhancing the chronic graft vs. host disease response indicating an expansion of the autoreactive B cell pool. Screening two non-overlapping recombinants, the Sle1a.1 and Sle1a.2 intervals that cover the entire Sle1a locus, revealed that both Sle1a.1 and Sle1a.2 were necessary for the full Sle1a phenotype. Sle1a.1, and to a lesser extent Sle1a.2, significantly affected CD4+ T cell activation as well as Treg differentiation and function. Sle1a.2 also increased the production of autoreactive B cells. Since the Sle1a.1 and Sle1a.2 intervals contain only one and 15 known genes, respectively, this study considerably reduces the number of candidate genes responsible for the production of autoreactive T cells. These results also demonstrate that the Sle1 locus is an excellent model for the genetic architecture of lupus, in which a major obligate phenotype results from the co-expression of multiple genetic variants with individual weak effects. PMID:20445563
An inducible caspase 9 safety switch for T-cell therapy
Straathof, Karin C.; Pulè, Martin A.; Yotnda, Patricia; Dotti, Gianpietro; Vanin, Elio F.; Brenner, Malcolm K.; Heslop, Helen E.; Spencer, David M.; Rooney, Cliona M.
2005-01-01
The efficacy of adoptive T-cell therapy as treatment for malignancies may be enhanced by genetic modification of infused cells. However, oncogenic events due to vector/transgene integration, and toxicities due to the infused cells themselves, have tempered enthusiasm. A safe and efficient means of removing aberrant cells in vivo would ameliorate these concerns. We describe a “safety switch” that can be stably and efficiently expressed in human T cells without impairing phenotype, function, or antigen specificity. This reagent is based on a modified human caspase 9 fused to a human FK506 binding protein (FKBP) to allow conditional dimerization using a small molecule pharmaceutical. A single 10-nM dose of synthetic dimerizer drug induces apoptosis in 99% of transduced cells selected for high transgene expression in vitro and in vivo. This system has several advantages over currently available suicide genes. First, it consists of human gene products with low potential immunogenicity. Second, administration of dimerizer drug has no effects other than the selective elimination of transduced T cells. Third, inducible caspase 9 maintains function in T cells overexpressing antiapoptotic molecules. These characteristics favor incorporation of inducible caspase 9 as a safety feature in human T-cell therapies. PMID:15728125
Tian, Jiangang; Gu, Liufang; Adams, Andrew; Wang, Xueliang; Huang, Ruizhe
2018-06-02
To determine the protective effects of Pellino-1 against H 2 O 2 -induced apoptosis in periodontal ligament stem cells (PDLSC). We demonstrated that H 2 O 2 decreases PDLSC viability by 40 and 50% with the concentrations of 400 and 500 μM, respectively, with an observed downregulation of Pellino-1 mRNA and protein; we further concluded that overexpression of Pellino-1 significantly lowers 8-hydroxy-2'-deoxyguanosine levels by 10% and upregulates superoxide dismutase 1, glutathione peroxidase levels, and catalase mRNA levels by 200, 40, and 250%, respectively. More importantly, we found that overexpression of Pellino-1 inhibited H 2 O 2 -induced cellular apoptosis through the activation of the NF-κB signaling pathway. Pellino-1 may be critically important for cell survival in the presence of oxidative elements; activation of the NF-κB signaling cascade was required for the overexpression of Pellino-1 to protect the cells from H 2 O 2 -induced apoptosis.
Chen, Ying; Liu, Fangwei; Weng, Dong; Song, Laiyu; Li, Cuiying; Tang, Wen; Yu, Ye; Dai, Wujing; Chen, Jie
2013-01-01
1,3-β-glucan is considered a fungal biomarker and exposure to this agent can induce lung inflammation. Complement activation plays an important role in early immune responses to β-glucan. Previous studies showed that T-regulatory cells (Tregs) regulated 1,3-β-glucan-induced lung inflammation by modulating the maintenance of immune homeostasis in the lung. Both interleukin (IL)-17 and TH17 cells play pivotal roles in inflammation associated with lung disease and share reciprocal developmental pathways with Tregs. However, the effect of Tregs on IL-17 and TH17 responses in 1,3-β-glucan-induced lung inflammation remains unclear. In this study, mice were exposed to 1,3-β-glucan by intratracheal instillation. To investigate the effects of Tregs on IL-17 and TH17 cells in the induced lung inflammation, a Treg-depleted mice model was generated by administration of anti-CD25 mAb. The results indicated that Treg-depleted mice showed more severe pathological inflammatory changes in lung tissues. Tregs depletion reduced IL-17 expression in these tissues, and increased those of TH1 cytokines. The expression of IL-17 increased at the early phase of the inflammation response. There were no significant effects of the Tregs on expression of RORγt and IL-6 or the amount of CD4(+)IL-17(+) cells in the lungs. When taken together, the late phase of the 1,3-β-glucan-induced inflammatory response in the mice was primarily mediated by TH1 cytokines rather than IL-17. In contrast, the early phase of the inflammatory response might be mediated in part by IL-17 along with activated complement. Tregs might be required for IL-17 expression during the late phase inflammatory response in mice. The increased IL-17 mRNA observed during the 1,3-β-glucan induced inflammatory response were attributed to cells other than TH17 cells.
Yamada, Takayuki; Ezura, Yoichi; Hayata, Tadayoshi; Moriya, Shuichi; Shirakawa, Jumpei; Notomi, Takuya; Arayal, Smriti; Kawasaki, Makiri; Izu, Yayoi; Harada, Kiyoshi; Noda, Masaki
2015-06-01
β adrenergic stimulation suppresses bone formation in vivo while its actions in osteoblastic differentiation are still incompletely understood. We therefore examined the effects of β2 adrenergic stimulation on osteoblast-like MC3T3-E1 cells focusing on BMP-induced alkaline phosphatase expression. Morphologically, isoproterenol treatment suppresses BMP-induced increase in the numbers of alkaline phosphatase-positive small foci in the cultures of MC3T3-E1 cells. Biochemically, isoproterenol treatment suppresses BMP-induced enzymatic activity of alkaline phosphatase in a dose-dependent manner. Isoproterenol suppression of alkaline phosphatase activity is observed even when the cells are treated with high concentrations of BMP. With respect to cell density, isoproterenol treatment tends to suppress BMP-induced increase in alkaline phosphatase expression more in osteoblasts cultured at higher cell density. In terms of treatment protocol, continuous isoproterenol treatment is compared to cyclic treatment. Continuous isoproterenol treatment is more suppressive against BMP-induced increase in alkaline phosphatase expression than cyclic regimen. At molecular level, isoproterenol treatment suppresses BMP-induced enhancement of alkaline phosphatase mRNA expression. Regarding the mode of isoproterenol action, isoproterenol suppresses BMP-induced BRE-luciferase activity. These data indicate that isoproterenol regulates BMP-induced alkaline phosphatase expression in osteoblast-like MC3T3E1 cells. © 2014 Wiley Periodicals, Inc.
Fernández, Dolores; García-Gómez, Concepción; Babín, Mar
2013-05-01
Zinc oxide nanoparticles (ZnO-NPs) are inevitably released into the environment and are potentially dangerous for aquatic life. However, the potential mechanisms of cytotoxicity of zinc nanoparticles remain unclear. Studying the toxicity of ZnO-NPs with In vitro systems will help to determine their interactions with cellular biomolecules. The aim of this study was to evaluate the cytotoxic potentials of ZnO-NPs in established fish cell lines (RTG-2, RTH-149 and RTL-W1) and compare them with those of bulk ZnO and Zn(2+) ions. Membrane function (CFDA-AM assay), mitochondrial function (MTT assay), cell growth (KBP assay), cellular stress (β-galactosidase assay), reductase enzyme activity (AB assay), reactive oxygen species (ROS), total glutathione cellular content (tGSH assay) and glutathione S-transferase (GST) activities were assessed for all cell lines. ZnO-NPs cytotoxicity was greater than those of bulk ZnO and Zn(2+). ZnO-NPs induced oxidative stress is dependent on their dose. Low cost tests, such as CFDA-AM, ROS, GST activity and tGSH cell content test that use fish cell lines, may be used to detect oxidative stress and redox status changes. Particle dissolution of the ZnO-NPs did not appear to play an important role in the observed toxicity in this study. Published by Elsevier B.V.
Programmed death-1 controls T cell survival by regulating oxidative metabolism1
Tkachev, Victor; Goodell, Stefanie; Opipari, Anthony W.; Hao, Ling-Yang; Franchi, Luigi; Glick, Gary D.; Ferrara, James L.M.; Byersdorfer, Craig A.
2015-01-01
The co-inhibitory receptor programmed death-1 (PD-1) maintains immune homeostasis by negatively regulating T cell function and survival. Blockade of PD-1 increases the severity of graft-versus-host disease (GVHD), but the interplay between PD-1 inhibition and T cell metabolism is not well studied. We found that both murine and human alloreactive T cells concomitantly up-regulated PD-1 expression and increased levels of reactive oxygen species (ROS) following allogeneic bone marrow transplantation. This PD-1HiROSHi phenotype was specific to alloreactive T cells and was not observed in syngeneic T cells during homeostatic proliferation. Blockade of PD-1 signaling decreased both mitochondrial H2O2 and total cellular ROS levels and PD-1 driven increases in ROS were dependent upon the oxidation of fatty acids, as treatment with etomoxir nullified changes in ROS levels following PD-1 blockade. Downstream of PD-1, elevated ROS levels impaired T cell survival in a process reversed by anti-oxidants. Furthermore, PD-1 driven changes in ROS were fundamental to establishing a cell’s susceptibility to subsequent metabolic inhibition, as blockade of PD-1 decreased the efficacy of later F1F0-ATP synthase modulation. These data indicate that PD-1 facilitates apoptosis in alloreactive T cells by increasing reactive oxygen species in a process dependent upon the oxidation of fat. In addition, blockade of PD-1 undermines the potential for subsequent metabolic inhibition, an important consideration given the increasing use of anti-PD-1 therapies in the clinic. PMID:25972478
Chandrasekaran, Murugesan; Pandurangan, Muthuraman
2016-07-01
The zinc oxide (ZnO) nanoparticle has been widely used in biomedical applications and cancer therapy and has been reported to induce a selective cytotoxic effect on cancer cell proliferation. The present study investigated the cytotoxicity of ZnO nanoparticles against co-cultured C2C12 myoblastoma cancer cells and 3T3-L1 adipocytes. Our results showed that the ZnO nanoparticles could be cytotoxic to C2C12 myoblastoma cancer cells than 3T3-L1 cells. The messenger RNA (mRNA) expressions of p53 and bax were significantly increased 114.3 and 118.2 % in the C2C12 cells, whereas 42.5 and 40 % were increased in 3T3-L1 cells, respectively. The mRNA expression of bcl-2 was reduced 38.2 and 28.5 % in the C2C12 and 3T3-L1 cells, respectively, whereas the mRNA expression of caspase-3 was increased 80.7 and 51.6 % in the C2C12 and 3T3-L1 cells, respectively. The protein expressions of p53, bax, and caspase-3 were significantly increased 40, 81.8, and 80 % in C2C12 cells, whereas 20.3, 28.2, and 37.9 % were increased in 3T3-L1 cells, respectively. The mRNA expression of bcl-2 was significantly reduced 32.2 and 22.7 % in C2C12 and 3T3-L1 cells, respectively. Caspase-3 enzyme activity and reactive oxygen species (ROS) were increased in co-cultured C2C12 cells compared to 3T3-L1 cells. Taking all these data together, it may suggest that ZnO nanoparticles severely induce apoptosis in C2C12 myoblastoma cancer cells than 3T3-L1 cells.
Ndhlovu, L C; Snyder-Cappione, J E; Carvalho, K I; Leal, F E; Loo, C P; bruno, F R; Jha, A R; Devita, D; Hasenkrug, A M; Barbosa, H M R; Segurado, A C; Nixon, D F; Murphy, E L; Kallas, E G
2009-01-01
Human T lymphotropic virus type 1 (HTLV-1) infects 10–20 million people worldwide. The majority of infected individuals are asymptomatic; however, approximately 3% develop the debilitating neurological disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). There is also currently no cure, vaccine or effective therapy for HTLV-1 infection, and the mechanisms for progression to HAM/TSP remain unclear. NK T cells are an immunoregulatory T cell subset whose frequencies and effector functions are associated critically with immunity against infectious diseases. We hypothesized that NK T cells are associated with HAM/TSP progression. We measured NK T cell frequencies and absolute numbers in individuals with HAM/TSP infection from two cohorts on two continents: São Paulo, Brazil and San Francisco, CA, USA, and found significantly lower levels when compared with healthy subjects and/or asymptomatic carriers. Also, the circulating NK T cell compartment in HAM/TSP subjects is comprised of significantly more CD4+ and fewer CD8+ cells than healthy controls. These findings suggest that lower numbers of circulating NK T cells and enrichment of the CD4+ NK T subset are associated with HTLV-1 disease progression. PMID:19778295
Van Snick, J; Houssiau, F; Proost, P; Van Damme, J; Renauld, J C
1996-09-15
We have previously reported that cytokines such as IL-9, IL-4, and IL-6 protect murine thymic lymphoma cell lines against dexamethasone-induced apoptosis. A similar activity, which could not be ascribed to any of these factors, was found in a number of human T cell supernatants that enabled mouse BW5147 thymic lymphoma not only to escape apoptosis but also to maintain proliferation. The protein responsible for this activity was purified to homogeneity from the culture medium of activated leukemic T cells and was found to be identical with the I-309 chemokine. Half-maximal anti-apoptotic activity was obtained with approximately 1 ng/ml, a concentration considerably lower than that required for the monocyte chemotactic activity of this molecule, as measured on THP-1 cells. The purified I-309 also improved the survival of two other mouse thymic lymphoma cell lines. This activity was as potent as that of IL-9, which was the strongest anti-apoptotic factor found to date for these cells. Similar results were obtained for BW5147 cells with recombinant I-309 and with T cell activation gene-3, the murine homologue of I-309, but not with other members of the chemokine family, including IL-8, neutrophil-activating peptide-2, granulocyte chemotactic protein-2, macrophage inflammatory protein-1a, RANTES (regulated upon activation, normal T cell expressed and secreted), monocyte chemotactic protein-1 (MCP-1), and MCP-2. MCP-3, however, showed a minor, but significant effect in this model. Unlike that of IL-9, the activity of I-309 was completely inhibited in the presence of pertussis toxin, indicating the involvement of a G protein in this process.
CD1b-autoreactive T cells contribute to hyperlipidemia-induced skin inflammation in mice
Bagchi, Sreya; He, Ying; Zhang, Hong; Cao, Liang; Van Rhijn, Ildiko; Moody, D. Branch; Gudjonsson, Johann E.
2017-01-01
A large proportion of human T cells are autoreactive to group 1 CD1 proteins, which include CD1a, CD1b, and CD1c. However, the physiological role of the CD1 proteins remains poorly defined. Here, we have generated a double-transgenic mouse model that expresses human CD1b and CD1c molecules (hCD1Tg) as well as a CD1b-autoreactive TCR (HJ1Tg) in the ApoE-deficient background (hCD1Tg HJ1Tg Apoe–/– mice) to determine the role of CD1-autoreactive T cells in hyperlipidemia-associated inflammatory diseases. We found that hCD1Tg HJ1Tg Apoe–/– mice spontaneously developed psoriasiform skin inflammation characterized by T cell and neutrophil infiltration and a Th17-biased cytokine response. Anti–IL-17A treatment ameliorated skin inflammation in vivo. Additionally, phospholipids and cholesterol preferentially accumulated in diseased skin and these autoantigens directly activated CD1b-autoreactive HJ1 T cells. Furthermore, hyperlipidemic serum enhanced IL-6 secretion by CD1b+ DCs and increased IL-17A production by HJ1 T cells. In psoriatic patients, the frequency of CD1b-autoreactive T cells was increased compared with that in healthy controls. Thus, this study has demonstrated the pathogenic role of CD1b-autoreactive T cells under hyperlipidemic conditions in a mouse model of spontaneous skin inflammation. As a large proportion of psoriatic patients are dyslipidemic, this finding is of clinical significance and indicates that self-lipid–reactive T cells might serve as a possible link between hyperlipidemia and psoriasis. PMID:28463230
Gargett, Tessa; Yu, Wenbo; Dotti, Gianpietro; Yvon, Eric S; Christo, Susan N; Hayball, John D; Lewis, Ian D; Brenner, Malcolm K; Brown, Michael P
2016-01-01
Chimeric antigen receptor (CAR) T cells have shown great promise in the treatment of hematologic malignancies but more variable results in the treatment of solid tumors and the persistence and expansion of CAR T cells within patients has been identified as a key correlate of antitumor efficacy. Lack of immunological “space”, functional exhaustion, and deletion have all been proposed as mechanisms that hamper CAR T-cell persistence. Here we describe the events following activation of third-generation CAR T cells specific for GD2. CAR T cells had highly potent immediate effector functions without evidence of functional exhaustion in vitro, although reduced cytokine production reversible by PD-1 blockade was observed after longer-term culture. Significant activation-induced cell death (AICD) of CAR T cells was observed after repeated antigen stimulation, and PD-1 blockade enhanced both CAR T-cell survival and promoted killing of PD-L1+ tumor cell lines. Finally, we assessed CAR T-cell persistence in patients enrolled in the CARPETS phase 1 clinical trial of GD2-specific CAR T cells in the treatment of metastatic melanoma. Together, these data suggest that deletion also occurs in vivo and that PD-1-targeted combination therapy approaches may be useful to augment CAR T-cell efficacy and persistence in patients. PMID:27019998
Sun, Y; Qian, C; Peng, D; Prieto, J
2000-01-01
To investigate the cooperative effect of B7-1 and IL-12 in the induction of antitumor activity, we have developed retroviral vectors encoding human B7-1, murine IL-12, or both B7-1 and IL-12 coordinately. Murine transformed liver cells (BNL) were engineered to stably express B7-1, IL-12, or both by infection with corresponding retroviruses. No tumor was observed in 20, 75, and 95% of mice receiving, respectively, B7-1-, IL-12-, and B7-1/IL-12-modified tumor cells after 250 days of inoculation. In contrast, injection of parental BNL or BNL/Neo cells resulted in lethal tumor progression in all mice. Protection against rechallenge with parental tumor cells was observed only in mice who had rejected BNL/IL-12, but not in animals that rejected BNL/B7-1 or BNL/B7-1-IL-12. Growth of parental tumor cells was significantly delayed by simultaneous injection in a distant site of irradiated tumor cells engineered to express IL-12 or both B7-1 and IL-12 but not B7-1 alone. BNL/B7-1 and BNL/B7-1-IL-12 showed similar efficacy in these experiments. Antitumor immunity induced by B7, with or without IL-12, was found to depend mainly on CD4+ T cells with a minor contribution of a non-T cell mechanism; whereas the effect of IL-12 was dependent on CD8+ T cells and on non-T cell effectors. Immunization of mice with IL-12-modified BNL cells induced secretion of a Thl pattern of cytokines while immunization with cells expressing both IL-12 and B7-1 resulted in inhibition of IFN-gamma production. Immunization with BNL/B7-1-IL-12 cells in the presence of anti-human B7-1 MAb resulted in restoration of IFN-gamma production to the levels found in animals injected with BNL/IL-12 cells. To summarize, in our model coexpression of B7-1 and IL-12 in tumor cells does not result in improved antitumoral activity as compared with expression of IL-12 alone. This may be related to the fact that B7-1 changes the mechanisms of antitumor immunity and inhibits IFN-gamma production induced by IL-12 in vivo.
Gechev, Tsanko; Mehterov, Nikolay; Denev, Iliya; Hille, Jacques
2013-01-01
A genetic approach is described to isolate mutants more tolerant to oxidative stress. A collection of T-DNA activation tag Arabidopsis thaliana mutant lines was screened for survivors under conditions that trigger H2O2-induced cell death. Oxidative stress was induced by applying the catalase (CAT) inhibitor aminotriazole (AT) in the growth media, which results in decrease in CAT enzyme activity, H2O2 accumulation, and subsequent plant death. One mutant was recovered from the screening and named oxr1 (oxidative stress resistant 1). The location of the T-DNA insertion was identified by TAIL-PCR. Oxr1 exhibited lack of cell death symptoms and more fresh weight and chlorophyll content compared to wild type. The lack of cell death correlated with more prominent induction of anthocyanins synthesis in oxr1. These results demonstrate the feasibility of AT as a screening agent for the isolation of oxidative stress-tolerant mutants and indicate a possible protective role for anthocyanins against AT-induced cell death. The chapter includes protocols for ethyl methanesulfonate mutagenesis, mutant screening using AT, T-DNA identification by TAIL-PCR, CAT activity measurements, and determination of malondialdehyde, chlorophyll, and anthocyanins. Copyright © 2013 Elsevier Inc. All rights reserved.
Yang, Nianlan; Baban, Babak; Isales, Carlos M; Shi, Xing-Ming
2015-09-01
Bone marrow is a reservoir for regulatory T (T(reg)) cells, but how T(reg) cells are regulated in that environment remains poorly understood. We show that expression of glucocorticoid (GC)-induced leucine zipper (GILZ) in bone marrow mesenchymal lineage cells or bone marrow-derived mesenchymal stem cells (BMSCs) increases the production of T(reg) cells via a mechanism involving the up-regulation of developmental endothelial locus-1 (Del-1), an endogenous leukocyte-endothelial adhesion inhibitor. We found that the expression of Del-1 is increased ∼4-fold in the bone tissues of GILZ transgenic (Tg) mice, and this increase is coupled with a significant increase in the production of IL-10 (2.80 vs. 0.83) and decrease in the production of IL-6 (0.80 vs. 2.33) and IL-12 (0.25 vs. 1.67). We also show that GILZ-expressing BMSCs present antigen in a way that favors T(reg) cells. These results indicate that GILZ plays a critical role mediating the crosstalk between BMSCs and T(reg) in the bone marrow microenvironment. These data, together with our previous findings that overexpression of GILZ in BMSCs antagonizes TNF-α-elicited inflammatory responses, suggest that GILZ plays important roles in bone-immune cell communication and BMSC immune suppressive functions. © FASEB.
Kondo, Taisuke; Imura, Yuuki; Chikuma, Shunsuke; Hibino, Sana; Omata-Mise, Setsuko; Ando, Makoto; Akanuma, Takashi; Iizuka, Mana; Sakai, Ryota; Morita, Rimpei; Yoshimura, Akihiko
2018-05-23
Adoptive T cell therapy is an effective strategy for cancer immunotherapy. However, infused T cells frequently become functionally exhausted, and consequently offer a poor prognosis after transplantation into patients. Adoptive transfer of tumor antigen-specific stem cell memory T (T SCM ) cells is expected to overcome this shortcoming since T SCM cells are close to naïve T cells, but are also highly proliferative, long-lived, and produce a large number of effector T cells in response to antigen stimulation. We previously reported that activated effector T cells can be converted into T SCM -like cells (iT SCM ) by co-culturing with OP9 cells expressing Notch ligand, Delta-like 1 (OP9-hDLL1). Here we show the methodological parameters of human CD8 + iT SCM cell generation and their application to adoptive cancer immunotherapy. Regardless of the stimulation by anti-CD3/CD28 antibodies or by antigen-presenting cells, human iT SCM cells were more efficiently induced from central memory type T cells than from effector memory T cells. During the induction phase by co-culture with OP9-hDLL1 cells, IL-7 and IL-15 (but not IL-2 or IL-21) could efficiently generate iT SCM cells. Epstein Barr (EB) virus-specific iT SCM cells showed much stronger antitumor potentials than conventionally activated T cells did in humanized EB virus transformed-tumor model mice. Thus, adoptive T cell therapy with iT SCM offers a promising therapeutic strategy for cancer immunotherapy. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
Nishida, Atsushi; Nagahama, Kiyotaka; Imaeda, Hirotsugu; Ogawa, Atsuhiro; Lau, Cindy W; Kobayashi, Taku; Hisamatsu, Tadakazu; Preffer, Frederic I; Mizoguchi, Emiko; Ikeuchi, Hiroki; Hibi, Toshifumi; Fukuda, Minoru; Andoh, Akira; Blumberg, Richard S; Mizoguchi, Atsushi
2012-12-17
Immune responses are modified by a diverse and abundant repertoire of carbohydrate structures on the cell surface, which is known as the glycome. In this study, we propose that a unique glycome that can be identified through the binding of galectin-4 is created on local, but not systemic, memory CD4+ T cells under diverse intestinal inflammatory conditions, but not in the healthy state. The colitis-associated glycome (CAG) represents an immature core 1-expressing O-glycan. Development of CAG may be mediated by down-regulation of the expression of core-2 β1,6-N-acetylglucosaminyltransferase (C2GnT) 1, a key enzyme responsible for the production of core-2 O-glycan branch through addition of N-acetylglucosamine (GlcNAc) to a core-1 O-glycan structure. Mechanistically, the CAG seems to contribute to super raft formation associated with the immunological synapse on colonic memory CD4+ T cells and to the consequent stabilization of protein kinase C θ activation, resulting in the stimulation of memory CD4+ T cell expansion in the inflamed intestine. Functionally, CAG-mediated CD4+ T cell expansion contributes to the exacerbation of T cell-mediated experimental intestinal inflammations. Therefore, the CAG may be an attractive therapeutic target to specifically suppress the expansion of effector memory CD4+ T cells in intestinal inflammation such as that seen in inflammatory bowel disease.
Inducible nitric oxide synthase in T cells regulates T cell death and immune memory
Vig, Monika; Srivastava, Smita; Kandpal, Usha; Sade, Hadassah; Lewis, Virginia; Sarin, Apurva; George, Anna; Bal, Vineeta; Durdik, Jeannine M.; Rath, Satyajit
2004-01-01
The progeny of T lymphocytes responding to immunization mostly die rapidly, leaving a few long-lived survivors functioning as immune memory. Thus, control of this choice of death versus survival is critical for immune memory. There are indications that reactive radicals may be involved in this death pathway. We now show that, in mice lacking inducible nitric oxide synthase (iNOS), higher frequencies of both CD4 and CD8 memory T cells persist in response to immunization, even when iNOS+/+ APCs are used for immunization. Postactivation T cell death by neglect is reduced in iNOS–/– T cells, and levels of the antiapoptotic proteins Bcl-2 and Bcl-xL are increased. Inhibitors of the iNOS-peroxynitrite pathway also enhance memory responses and block postactivation death by neglect in both mouse and human T cells. However, early primary immune responses are not enhanced, which suggests that altered survival, rather than enhanced activation, is responsible for the persistent immunity observed. Thus, in primary immune responses, iNOS in activated T cells autocrinely controls their susceptibility to death by neglect to determine the level of persisting CD4 and CD8 T cell memory, and modulation of this pathway can enhance the persistence of immune memory in response to vaccination. PMID:15199408
Jiang, Xianzhi; Alami Chentoufi, Aziz; Hsiang, Chinhui; Carpenter, Dale; Osorio, Nelson; BenMohamed, Lbachir; Fraser, Nigel W.; Jones, Clinton; Wechsler, Steven L.
2011-01-01
The herpes simplex virus type 1 (HSV-1) latency-associated transcript (LAT) is the only HSV-1 gene transcript abundantly expressed throughout latency. LAT null mutants have a significantly reduced reactivation phenotype. LAT's antiapoptosis activity is the major LAT factor involved in supporting the wild-type reactivation phenotype. During HSV-1 latency, some ganglionic neurons are surrounded by CD8 T cells, and it has been proposed that these CD8 T cells help maintain HSV-1 latency by suppressing viral reactivations. Surprisingly, despite injection of cytotoxic lytic granules by these CD8 T cells into latently infected neurons, neither apoptosis nor neuronal cell death appears to occur. We hypothesized that protection of latently infected neurons against cytotoxic CD8 T-cell killing is due to LAT's antiapoptosis activity. Since CD8 T-cell cytotoxic lytic granule-mediated apoptosis is critically dependent on granzyme B (GrB), we examined LAT's ability to block GrB-induced apoptosis. We report here that (i) LAT can interfere with GrB-induced apoptosis in cell cultures, (ii) LAT can block GrB-induced cleavage (activation) of caspase-3 both in cell culture and in a cell-free in vitro cell extract assay, and (iii) LAT can protect C1300 and Neuro2A cells from cytotoxic CD8 T-cell killing in vitro. These findings support the hypothesis that LAT's antiapoptosis activity can protect latently infected neurons from being killed by CD8 T-cell lytic granules in vivo. PMID:21177822
SPAK kinase is a substrate and target of PKCθ in T-cell receptor-induced AP-1 activation pathway
Li, Yingqiu; Hu, Junru; Vita, Randi; Sun, Binggang; Tabata, Hiroki; Altman, Amnon
2004-01-01
Protein kinase C-θ (PKCθ) plays an important role in T-cell activation via stimulation of AP-1 and NF-κB. Here we report the isolation of SPAK, a Ste20-related upstream mitogen-activated protein kinase (MAPK), as a PKCθ-interacting kinase. SPAK interacted with PKCθ (but not with PKCα) via its 99 COOH-terminal residues. TCR/CD28 costimulation enhanced this association and stimulated the catalytic activity of SPAK. Recombinant SPAK was phosphorylated on Ser-311 in its kinase domain by PKCθ, but not by PKCα. The magnitude and duration of TCR/CD28-induced endogenous SPAK activation were markedly impaired in PKCθ-deficient T cells. Transfected SPAK synergized with constitutively active PKCθ to activate AP-1, but not NF-κB. This synergistic activity, as well as the receptor-induced SPAK activation, required the PKCθ-interacting region of SPAK, and Ser-311 mutation greatly reduced these activities of SPAK. Conversely, a SPAK-specific RNAi or a dominant-negative SPAK mutant inhibited PKCθ- and TCR/CD28-induced AP-1, but not NF-κB, activation. These results define SPAK as a substrate and target of PKCθ in a TCR/CD28-induced signaling pathway leading selectively to AP-1 (but not NF-κB) activation. PMID:14988727
Sasaki, Takanori; Kanaseki, Takayuki; Shionoya, Yosuke; Tokita, Serina; Miyamoto, Sho; Saka, Eri; Kochin, Vitaly; Takasawa, Akira; Hirohashi, Yoshihiko; Tamura, Yasuaki; Miyazaki, Akihiro; Torigoe, Toshihiko; Hiratsuka, Hiroyoshi; Sato, Noriyuki
2016-04-01
Hypoxia and glucose deprivation are often observed in the microenvironment surrounding solid tumors in vivo. However, how they interfere with MHC class I antigen processing and CD8(+) T-cell responses remains unclear. In this study, we analyzed the production of antigenic peptides presented by classical MHC class I in mice, and showed that it is quantitatively decreased in the cells exposed to either hypoxia or glucose deprivation. In addition, we unexpectedly found increased surface expression of HLA-E in human and Qa-1 in mouse tumor cells exposed to combined oxygen and glucose deprivation. The induced Qa-1 on the stressed tumor model interacted with an inhibitory NKG2/CD94 receptor on activated CD8(+) T cells and attenuated their specific response to the antigen. Our results thus suggest that microenvironmental stresses modulate not only classical but also nonclassical MHC class I presentation, and confer the stressed cells the capability to escape from the CD8(+) T-cell recognition. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
γδ T cells protect against LPS-induced lung injury
Wehrmann, Fabian; Lavelle, James C.; Collins, Colm B.; Tinega, Alex N.; Thurman, Joshua M.; Burnham, Ellen L.; Simonian, Philip L.
2016-01-01
γδ T lymphocytes are a unique T cell population with important anti-inflammatory capabilities. Their role in acute lung injury, however, is poorly understood but may provide significant insight into lung-protective mechanisms occurring after injury. In a murine model of lung injury, wild-type C57BL/6 and TCRδ−/− mice were exposed to Escherichia coli LPS, followed by analysis of γδ T cell and macrophage subsets. In the absence of γδ T cells, TCRδ−/− mice developed increased inflammation and alveolar-capillary leak compared with wild-type C57BL/6 mice after LPS exposure that correlated with expansion of distinct macrophage populations. Classically activated M1 macrophages were increased in the lung of TCRδ−/− mice at d 1, 4, and 7 after LPS exposure that peaked at d 4 and persisted at d 7 compared with wild-type animals. In response to LPS, Vγ1 and Vγ7 γδ T cells were expanded in the lung and expressed IL-4. Coculture experiments showed decreased expression of TNF-α by resident alveolar macrophages in the presence of γδ T cells that was reversed in the presence of an anti-IL-4-blocking antibody. Treatment of mice with rIL4 resulted in reduced numbers of M1 macrophages, inflammation, and alveolar-capillary leak. Therefore, one mechanism by which Vγ1 and Vγ7 γδ T cells protect against LPS-induced lung injury is through IL-4 expression, which decreases TNF-α production by resident alveolar macrophages, thus reducing accumulation of M1 macrophages, inflammation, and alveolar-capillary leak. PMID:26428678
Khanna, S; Sen, C K; Roy, S; Christen, M O; Packer, L
1998-07-01
The protective effects of anethole dithiolethione (ADT) against H2O2- or 4-hydroxynonenal (HNE)-induced cytotoxicity in human Jurkat T cells were investigated. Jurkat T cells were pretreated with ADT (10-50 microM) for 18 hr and then challenged with H202 or HNE for up to 4 hr. Cytotoxicity was assessed by measuring: 1) leakage of lactate dehydrogenase from cells to medium; and 2) exclusion of the DNA intercalating fluorescent probe propidium iodide by viable cells. Pretreatment of cells with ADT (10 or 25 microM) for 18 hr significantly protected cells against H202- or HNE-induced cytotoxicity. Treatment of cells with ADT (10-50 microM) for 72 hr significantly increased the activities of catalase and glutathione reductase. The maximum effect of ADT treatment on the activity of these enzymes was observed when cells were treated with 25 microM of ADT for 72 hr. A significant increase in cellular GSH was observed in cells that were treated with ADT for 72 hr. Using monobromobimane as a thiol probe, we consistently observed that cells pretreated for 18 hr with ADT (25 or 50 microM) had also increased total thiol content. Exposure of Jurkat T cells to H202 or HNE resulted in a time-dependent decrease in cellular GSH. ADT (10-50 microM, 18 hr) pretreatment circumvented H202-dependent lowering of cellular GSH. In conclusion, ADT proved to be a potent cytoprotective thiol antioxidant with multifaceted mechanisms of action, suggesting that the drug has a remarkable therapeutic potential.
Pappritz, Kathleen; Savvatis, Konstantinos; Miteva, Kapka; Kerim, Bahtiyar; Dong, Fengquan; Fechner, Henry; Müller, Irene; Brandt, Christine; Lopez, Begoña; González, Arantxa; Ravassa, Susana; Klingel, Karin; Diez, Javier; Reinke, Petra; Volk, Hans-Dieter; Van Linthout, Sophie; Tschöpe, Carsten
2018-06-04
Regulatory T (T reg ) cells offer new therapeutic options for controlling undesired systemic and local immune responses. The aim of the current study was to determine the impact of therapeutic T reg administration on systemic and cardiac inflammation and remodeling in coxsackievirus B3 (CVB3) -induced myocarditis. Therefore, syngeneic T reg cells were applied intravenously in CVB3-infected mice 3 d after infection. Compared with CVB3 + PBS mice, CVB3 + T reg mice exhibited lower left ventricular (LV) chemokine expression, accompanied by reduced cardiac presence of proinflammatory Ly6C high CCR2 high Cx3Cr1 low monocytes and higher retention of proinflammatory Ly6C mid CCR2 high Cx3Cr1 low monocytes in the spleen. In addition, splenic myelopoiesis was reduced in CVB3 + T reg compared with CVB3 + PBS mice. Coculture of T reg cells with splenocytes isolated from mice 3 d post-CVB3 infection further demonstrated the ability of T reg cells to modulate monocyte differentiation in favor of the anti-inflammatory Ly6C low CCR2 low Cx3Cr1 high subset. T reg -mediated immunomodulation was paralleled by lower collagen 1 protein expression and decreased levels of soluble and insoluble collagen in LV of CVB3 + T reg compared with CVB3 + PBS mice. In agreement with these findings, LV systolic and diastolic function was improved in CVB3 + T reg mice compared with CVB3 + PBS mice. In summary, adoptive T reg transfer in the inflammatory phase of viral-induced myocarditis protects the heart against inflammatory damage and fibrosis via modulation of monocyte subsets.-Pappritz, K., Savvatis, K., Miteva, K., Kerim, B., Dong, F., Fechner, H., Müller, I., Brandt, C., Lopez, B., González, A., Ravassa, S., Klingel, K., Diez, J., Reinke, P., Volk, H.-D., Van Linthout, S., Tschöpe, C. Immunomodulation by adoptive regulatory T-cell transfer improves Coxsackievirus B3-induced myocarditis.
Yoo, Jae-Myung; Lee, Youn-Sun; Choi, Heon-Kyo; Lee, Yong-Moon; Hong, Jin-Tae; Yun, Yeo-Pyo; Oh, Seikwan; Yoo, Hwan-Soo
2005-03-01
Oxidative stress has been reported to elevate ceramide level during cell death. The purpose of the present study was to modulate cell death in relation to cellular glutathione (GSH) level and GST (glutathione S-transferase) expression by regulating the sphingolipid metabolism. LLC-PK1 cells were treated with H2O2 in the absence of serum to induce cell death. Subsequent to exposure to H2O2, LLC-PK1 cells were treated with desipramine, sphingomyelinase inhibitor, and N-acetylcysteine (NAC), GSH substrate. Based on comparative visual observation with H2O2-treated control cells, it was observed that 0.5 microM of desipramine and 25 mM of NAC exhibited about 90 and 95% of cytoprotection, respectively, against H2O2-induced cell death. Desipramine and NAC lowered the release of LDH activity by 36 and 3%, respectively, when compared to 71% in H2O2-exposed cells. Cellular glutathione level in 500 microM H2O2-treated cells was reduced to 890 pmol as compared to control level of 1198 pmol per mg protein. GST P1-1 expression was decreased in H2O2-treated cells compared to healthy normal cells. In conclusion, it has been inferred that H2O2-induced cell death is closely related to cellular GSH level and GST P1-1 expression in LLC-PK1 cells and occurs via ceramide elevation by sphingomyelinase activation.
TCF1 and LEF1 act as T-cell intrinsic HTLV-1 antagonists by targeting Tax.
Ma, Guangyong; Yasunaga, Jun-ichirou; Akari, Hirofumi; Matsuoka, Masao
2015-02-17
Human T-cell leukemia virus type 1 (HTLV-1) is a delta-type retrovirus that induces malignant and inflammatory diseases during its long persistence in vivo. HTLV-1 can infect various kinds of cells; however, HTLV-1 provirus is predominantly found in peripheral CD4 T cells in vivo. Here we find that TCF1 and LEF1, two Wnt transcription factors that are specifically expressed in T cells, inhibit viral replication through antagonizing Tax functions. TCF1 and LEF1 can each interact with Tax and inhibit Tax-dependent viral expression and activation of NF-κB and AP-1. As a result, HTLV-1 replication is suppressed in the presence of either TCF1 or LEF1. On the other hand, T-cell activation suppresses the expression of both TCF1 and LEF1, and this suppression enables Tax to function as an activator. We analyzed the thymus of a simian T-cell leukemia virus type 1 (STLV-1) infected Japanese macaque, and found a negative correlation between proviral load and TCF1/LEF1 expression in various T-cell subsets, supporting the idea that TCF1 and LEF1 negatively regulate HTLV-1 replication and the proliferation of infected cells. Thus, this study identified TCF1 and LEF1 as Tax antagonistic factors in vivo, a fact which may critically influence the peripheral T-cell tropism of this virus.
Xu, Aizhang; Freywald, Andrew; Xie, Yufeng; Li, Zejun; Xiang, Jim
2017-01-01
Whether inflation of CD8 + memory T (mT) cells, which is often derived from repeated prime-boost vaccinations or chronic viral infections in the elderly, would affect late CD8 + T-cell immunity is a long-standing paradox. We have previously established an animal model with mT-cell inflation by transferring ConA-stimulated monoclonal CD8 + T cells derived from Ova-specific T-cell-receptor transgenic OTI mice into irradiation-induced lymphopenic B6 mice. In this study, we also established another two animal models with mT-cell inflation by transferring, 1) ConA-stimulated monoclonal CD8 + T cells derived from lymphocytic choriomeningitis virus glycoprotein-specific T-cell-receptor transgenic P14 mice, and 2) ConA-stimulated polyclonal CD8 + T cells derived from B6.1 mice into B6 mice with irradiation-induced lymphopenia. We vaccinated these mice with recombinant Ova-expressing Listeria monocytogenes and Ova-pulsed dendritic cells, which stimulated CD4 + T cell-independent and CD4 + T-cell-dependent CD8 + T-cell responses, respectively, and assessed Ova-specific CD8 + T-cell responses by flow cytometry. We found that Ova-specific CD8 + T-cell responses derived from the latter but not the former vaccination were significantly reduced in mice with CD8 + mT-cell inflation compared to wild-type B6 mice. We determined that naïve CD8 + T cells purified from splenocytes of mice with mT-cell inflation had defects in cell proliferation upon stimulation in vitro and in vivo and upregulated T-cell anergy-associated Itch and GRAIL molecules. Taken together, our data reveal that CD8 + mT-cell inflation renders compromised CD4 + T-cell-dependent CD8 + T-cell immunity via naïve T-cell anergy, and thus show promise for the design of efficient vaccines for elderly patients with CD8 + mT-cell inflation.
Komohara, Yoshihiro; Ma, Chaoya; Yano, Hiromu; Pan, Cheng; Horlad, Hasita; Saito, Yoichi; Ohnishi, Koji; Fujiwara, Yukio; Okuno, Yutaka; Nosaka, Kisato; Shimosaki, Shunsuke; Morishita, Kazuhiro; Matsuoka, Masao; Wakayama, Tomohiko; Takeya, Motohiro
2017-07-05
Cell adhesion molecule 1 (CADM1) is a cell adhesion molecule that is expressed in brain, liver, lung, testis, and some kinds of cancer cells including adult T-cell leukemia/lymphoma (ATLL). Recent studies have indicated the involvement of CADM1 in cell-cell contact between cytotoxic T-lymphocytes and virus infected cells. We previously reported that cell-cell interaction between lymphoma cells and macrophages induces lymphoma cell proliferation. In the present study, we investigated whether CADM1 is associated with cell-cell interaction between several human lymphoma cell lines and macrophages.CADM1 expression was observed in the ATLL cell lines, ATN-1, ATL-T, and ATL-35T, and in the B cell lymphoma cell lines, TL-1, DAUDI, and SLVL, using western blotting. Significant cell-cell interaction between macrophages and ATN-1, ATL-T, ATL-35T and MT-2, DAUDI, and SLVL cells, as assessed by induction of cell proliferation, was observed. Immunohistochemical analysis of human biopsy samples indicated CADM1 expression in 10 of 14 ATLL cases; however, no case of follicular lymphoma or diffuse large B-cell lymphoma was positive for CADM1. Finally, the interaction of macrophages with cells of the CADM1-negative ED ATLL cell line and CADM1-transfected ED cells was tested. However, significant cell-cell interaction between macrophage and CADM1-transfected ED cells was not observed. We conclude that CADM1 was not associated with cell-cell interaction between lymphoma cells and macrophages, although CADM1 may be a useful marker of ATLL for diagnostic procedures.
Intrinsic role of FoxO3a in the development of CD8+ T cell memory
Tzelepis, Fanny; Joseph, Julie; Haddad, Elias K.; MacLean, Susanne; Dudani, Renu; Agenes, Fabien; Peng, Stanford L.; Sekaly, Rafick-Pierre; Sad, Subash
2013-01-01
CD8+ T cells undergo rapid expansion during infection with intracellular pathogens, which is followed by swift and massive culling of primed CD8+ T cells. The mechanisms that govern the massive contraction and maintenance of primed CD8+ T cells are not clear. We show here that the transcription factor, FoxO3a does not influence antigen-presentation and the consequent expansion of CD8+ T cell response during Listeria monocytogenes (LM) infection, but plays a key role in the maintenance of memory CD8+ T cells. The effector function of primed CD8+ T cells as revealed by cytokine secretion and CD107a degranulation was not influenced by inactivation of FoxO3a. Interestingly, FoxO3a-deficient CD8+ T cells displayed reduced expression of pro-apoptotic molecules BIM and PUMA during the various phases of response, and underwent reduced apoptosis in comparison to WT cells. A higher number of memory precursor effector cells (MPECs) and memory subsets were detectable in FoxO3a-deficient mice compared to WT mice. Furthermore, FoxO3a-deficient memory CD8+ T cells upon transfer into normal or RAG1-deficient mice displayed enhanced survival. These results suggest that FoxO3a acts in a cell intrinsic manner to regulate the survival of primed CD8+ T cells. PMID:23277488
Werthmöller, N; Frey, B; Wunderlich, R; Fietkau, R; Gaipl, U S
2015-01-01
One prerequisite that radiotherapy (RT) and chemotherapy (CT) result in anti-tumor immune responses is triggering of immunogenic cell death forms such as necroptosis. The latter is inducible by inhibition of apoptosis with the pan-caspase inhibitor zVAD-fmk. The design of multimodal therapies that overcome melanoma's resistance to apoptosis is a big challenge of oncoimmunology. As hints exist that immune stimulation by hyperthermia (HT) augments the efficacy of melanoma therapies and that tumors can be sensitized for RT with zVAD-fmk, we asked whether combinations of RT with dacarbazine (DTIC) and/or HT induce immunogenic melanoma cell death and how this is especially influenced by zVAD-fmk. Necroptosis was inducible in poorly immunogenic B16-F10 melanoma cells and zVAD-fmk generally increased melanoma cell necrosis concomitantly with the release of HMGB1. Supernatants (SNs) of melanoma cells whose cell death was modulated with zVAD-fmk induced an upregulation of the activation markers CD86 and MHCII on macrophages. The same was seen on dendritic cells (DCs), but only when zVAD-fmk was added to multimodal tumor treatments including DTIC. DCs of MyD88 KO mice and DCs incubated with SNs containing apyrase did not increase the expression of these activation markers on their surface. The in vivo experiments revealed that zVAD-fmk decreases the tumor growth significantly and results in a significantly reduced tumor infiltration of Tregs when added to multimodal treatment of the tumor with RT, DTIC and HT. Further, a significantly increased DC and CD8+ T-cell infiltration into the tumor and in the draining lymph nodes was induced, as well as an increased expression of IFNγ by CD8+ T cells. However, zVAD-fmk did not further reduce tumor growth in MyD88 KO mice, mice treated with apyrase or RAG KO mice. We conclude that HMGB1, nucleotides and CD8+ T cells mediate zVAD-fmk induced anti-melanoma immune reactions in multimodal therapy settings. PMID:25973681
Lee, Mina; Sung, Sang Hyun
2016-01-01
Background: Obesity causes or aggravates many health problems, both independently and in association with several pathological disorders, including Type II diabetes, hypertension, atherosclerosis, and cancer. Therefore, we screened small compounds isolated from natural products for the development of anti-obesity drugs. Objective: The purpose of this study was to investigate the anti-adipogenic activities of platyphylloside, diarylheptanoid isolated from Betula platyphylla, which was selected based on the screening using 3T3-L1 cells. Materials and Methods: To evaluate the inhibition of adipocyte differentiation and lipolysis, lipid contents of BPP on were measured using Oil Red O staining in 3T3-L1 cells. The mRNA and protein expression levels of various adipokines were measured by Quantitative real-time PCR and Western blotting analysis, respectively. Results: Platyphylloside showed significant inhibitory activity on adipocyte differentiation in 3T3-L1 cells and suppressed adipocyte differentiation even in the presence of troglitazone, a PPARγ agonist. Platyphylloside might suppress adipocyte differentiation through PPARγ, C/EBPα, and SREBP1-induced adipogenesis, which is synergistically associated with downstream adipocyte-specific gene promoters such as aP2, FAS, SCD-1, LPL, and Adiponectin. In addition, platyphylloside affected lipolysis by down-regulating perilipin and HSL and up-regulating TNFα. Conclusion: Taken together, the results reveal that platyphylloside has anti-adipogenic activity and highlight its potential in the prevention and treatment of obesity. SUMMARY The extract of B. platyphylla bark and its isolate, BPP, had anti-adipogenic activity in 3T3-L1 cells via suppression of adipocyte differentiation from preadipocytes.Treatment with BPP significantly down-regulated the expression of PPARγ, C/EBP, C/EBPβ, C/EBPδ, SREBP1c, SCD-1, FAS, aP2 and LPL.BPP induced a lipolytic response in mature adipocytes via up-regulation krof TNFá and down
Poulin, B; Rich, N; Mas, J L; Kordon, C; Enjalbert, A; Drouva, S V
1998-07-25
Exposure of the gonadotrope cells to gonadotropin-releasing hormone (GnRH) reduces their responsiveness to a new GnRH stimulation (homologous desensitization). The time frame as well as the mechanisms underlying this phenomenon are yet unclear. We studied in a gonadotrope cell line (alphaT3-1) the effects of short as well as long term GnRH pretreatments on the GnRH-induced phospholipases-C (PLC), -A2 (PLA2) and -D (PLD) activities, by measuring the production of IP3, total inositol phosphates (IPs), arachidonic acid (AA) and phosphatidylethanol (PEt) respectively. We demonstrated that although rapid desensitization of GnRH-induced IP3 formation did not occur in these cells, persistent stimulation of cells with GnRH or its analogue resulted in a time-dependent attenuation of GnRH-elicited IPs formation. GnRH-induced IPs desensitization was potentiated after direct activation of PKC by the phorbol ester TPA, suggesting the involvement of distinct mechanisms in the uncoupling exerted by either GnRH or TPA on GnRH-stimulated PI hydrolysis. The levels of individual phosphoinositides remained unchanged under any desensitization condition applied. Interestingly, while the GnRH-induced PLA2 activity was rapidly desensitized (2.5 min) after GnRH pretreatments, the neuropeptide-evoked PLD activation was affected at later times, indicating an important time-dependent contribution of these enzymatic activities in the sequential events underlying the GnRH-induced homologous desensitization processes in the gonadotropes. Under GnRH desensitization conditions, TPA was still able to induce PLD activation and to further potentiate the GnRH-evoked PLD activity. AlphaT3-1 cells possess several PKC isoforms which, except PKCzeta, were differentially down-regulated by TPA (PKCalpha, betaII, delta, epsilon, eta) or GnRH (PKCbetaII, delta, epsilon, eta). In spite of the presence of PKC inhibitors or down-regulation of PKC isoforms by TPA, the desensitizing effect of the neuropeptide on
Antigen-specific, CD4+CD25+ regulatory T cell clones induced in Peyer's patches.
Tsuji, Noriko M; Mizumachi, Koko; Kurisaki, Jun-Ichi
2003-04-01
Since intestine is exposed to numerous exogenous antigens such as food and commensal bacteria, the organ bears efficient mechanisms for establishment of tolerance and induction of regulatory T cells (T(reg)). Intestinal and inducible T(reg) include T(r)1-like and T(h)3 cells whose major effector molecules are IL-10 and transforming growth factor (TGF)-beta. These antigen-specific T(reg) are expected to become clinical targets to modify the inflammatory immune response associated with allergy, autoimmune diseases and transplantation. In the present study, we characterized the antigen-specific T(reg) induced in the intestine by orally administering high-dose beta-lactoglobulin (BLG) to BALB/c mice. Seven days after feeding, only Peyer's patch (PP) cells among different organs exerted significant suppressive effect on antibody production upon in vitro BLG stimulation. This suppressive effect was also prominent in six BLG-specific CD4(+) T cell clones (OPP1-6) established from PP from mice orally administered with high doses of BLG and was partially reversed by antibodies to TGF-beta. Intravenous transfer of OPP2 efficiently suppressed BLG-specific IgG1 production in serum following immunization, indicating the role of such T(reg) in the systemic tolerance after oral administration of antigen (oral tolerance). OPP clones secrete TGF-beta, IFN-gamma and low levels of IL-10, a cytokine pattern similar to that secreted by anergic T cells. OPP clones bear a CD4(+)CD25(+) phenotype and show significantly lower proliferative response compared to T(h)0 clones. This lower response is recovered by the addition of IL-2. Thus, antigen-specific CD4(+)CD25(+) T(reg), which have characteristics of anergic cells and actively suppress antibody production are induced in PP upon oral administration of protein antigen.
α-lipoic acid inhibits high glucose-induced apoptosis in HIT-T15 cells.
Yang, Yi; Wang, Weiping; Liu, Yinan; Guo, Ting; Chen, Ping; Ma, Kangtao; Zhou, Chunyan
2012-06-01
High blood glucose plays an important role in the pathogenesis of diabetes. α-lipoic acid (LA) has been used to prevent and treat diabetes, and is thought to act by increasing insulin sensitivity in many tissues. However, whether LA also has a cytoprotective effect on pancreatic islet beta cells remains unclear. In this study, we assessed whether LA could inhibit apoptosis in beta cells exposed to high glucose concentrations. HIT-T15 pancreatic beta cells were treated with 30 mmol/L glucose in the presence or absence of 0.5 mmol/L LA for 8 days. LA significantly reduced the numbers of apoptotic HIT-T15 cells and inhibited the cell overgrowth normally induced by high glucose treatment. Additionally, LA inhibited insulin expression and secretion in HIT-T15 cells induced by high glucose. Further study demonstrated that LA upregulated Pdx1 and Bcl2 gene expression, reduced Bax gene expression, and promoted phosphorylation of Akt in HIT-T15 cells treated with high glucose. Intriguingly, knockdown of Pdx1 expression partially offset the anti-apoptotic effect of LA. However, inhibition of Akt by PI3K/AKT antagonist LY294002 only slightly reversed the anti-apoptosis effect of LA and mildly decreased the gene expression level of Pdx1 (P > 0.05). Moreover, LA only slightly attenuated reactive oxygen species (ROS) production and augmented mitochondrial membrane potential. Therefore, our data suggest that α-lipoic acid can effectively attenuate high glucose-induced HIT-T15 cell apoptosis probably by increasing Pdx1 expression. These findings provide a new interpretation on the role of LA in the treatment of diabetes. © 2012 The Authors Development, Growth & Differentiation © 2012 Japanese Society of Developmental Biologists.
DOE Office of Scientific and Technical Information (OSTI.GOV)
Benveniste, E.; Stevens, R.H.
1983-04-01
Tetanus toxoid immunization of humans generates circulating B cells which secrete IgG anti-tetanus toxoid antibodies (IgG-Tet) when stimulated in vitro with T cells and pokeweed mitogen (PWM). A unique property of these cells is the inhibition of maturation into antibody-secreting plasma cells following a 1-hr in vitro pulse with tetanus toxoid. Studies were undertaken to determine if different T-cell subsets could modulate the in vitro generated B-cell unresponsive state. The addition of OKT4+/OKT8- cells to antigen-treated B cells resulted in a partial reversal of the antigen-induced inhibition of IgG-Tet synthesis. The addition of OKT4-/OKT8+ cells to the treated B cellsmore » caused a suppression of IgG-Tet synthesis comparable to that seen in cultures containing unfractionated T cells. These results indicate that (1) the B-cell unresponsive state generated by antigen treatment is not absolute, (2) the degree of B-cell unresponsiveness results from a balance of suppressor and helper signals, and (3) T-suppressor cells need to be present to induce and maintain the B-cell unresponsive state.« less
Schmetterer, Klaus G; Haiderer, Daniela; Leb-Reichl, Victoria M; Neunkirchner, Alina; Jahn-Schmid, Beatrice; Küng, Hans J; Schuch, Karina; Steinberger, Peter; Bohle, Barbara; Pickl, Winfried F
2011-01-01
Regulatory T (Treg) cells establish and maintain tolerance to self-antigens and many foreign antigens, such as allergens, by suppressing effector T-cell proliferation and function. We have previously shown that human T-cell receptor (TCR) αβ-chains specific for allergen-derived epitopes confer allergen specificity on peripheral blood T cells of individuals with and without allergy. To study the feasibility of generating allergen-specific human Treg cells by retroviral transduction of a transcription unit encoding forkhead box protein 3 (FOXP3) and allergen-specific TCR αβ-chains. cDNAs encoding the α and β-chains of a Bet v 1(142-153)-specific TCR (TCR alpha variable region 6/TCR beta variable region 20) and human FOXP3 were linked via picornaviral 2A sequences and expressed as single translational unit from an internal ribosomal entry site-green fluorescence protein-containing retroviral vector. Retrovirally transduced peripheral blood T cells were tested for expression of transgenes, Treg phenotype, and regulatory capacity toward allergen-specific effector T cells. Transduced T cells displayed a Treg phenotype with clear-cut upregulation of CD25, CD39, and cytotoxic T-lymphocyte antigen 4. The transduced cells were hyporesponsive in cytokine production and secretion and, like naturally occurring Treg cells, did not proliferate after antigen-specific or antigen-mimetic stimulation. However, proliferation was inducible upon exposure to exogenous IL-2. In coculture experiments, TRAV6(+)TRBV20(+)FOXP3(+) transgenic T cells, unlike FOXP3(+) single transgenic T cells or naturally occurring Treg cells, highly significantly suppressed T cell cytokine production and proliferation of corresponding allergen-specific effector T cells in an allergen-specific, dose-dependent manner. We demonstrate a transgenic approach to engineer human allergen-specific Treg cells that exert their regulatory function in an activation-dependent manner. Customized Treg cells might become
Calcium signaling properties of a thyrotroph cell line, mouse TαT1 cells.
Tomić, Melanija; Bargi-Souza, Paula; Leiva-Salcedo, Elias; Nunes, Maria Tereza; Stojilkovic, Stanko S
2015-12-01
TαT1 cells are mouse thyrotroph cell line frequently used for studies on thyroid-stimulating hormone beta subunit gene expression and other cellular functions. Here we have characterized calcium-signaling pathways in TαT1 cells, an issue not previously addressed in these cells and incompletely described in native thyrotrophs. TαT1 cells are excitable and fire action potentials spontaneously and in response to application of thyrotropin-releasing hormone (TRH), the native hypothalamic agonist for thyrotrophs. Spontaneous electrical activity is coupled to small amplitude fluctuations in intracellular calcium, whereas TRH stimulates both calcium mobilization from intracellular pools and calcium influx. Non-receptor-mediated depletion of intracellular pool also leads to a prominent facilitation of calcium influx. Both receptor and non-receptor stimulated calcium influx is substantially attenuated but not completely abolished by inhibition of voltage-gated calcium channels, suggesting that depletion of intracellular calcium pool in these cells provides a signal for both voltage-independent and -dependent calcium influx, the latter by facilitating the pacemaking activity. These cells also express purinergic P2Y1 receptors and their activation by extracellular ATP mimics TRH action on calcium mobilization and influx. The thyroid hormone triiodothyronine prolongs duration of TRH-induced calcium spikes during 30-min exposure. These data indicate that TαT1 cells are capable of responding to natively feed-forward TRH signaling and intrapituitary ATP signaling with acute calcium mobilization and sustained calcium influx. Amplification of TRH-induced calcium signaling by triiodothyronine further suggests the existence of a pathway for positive feedback effects of thyroid hormones probably in a non-genomic manner. Published by Elsevier Ltd.
Li, Xiuying; Xu, Zhuo; Bai, Jinping; Yang, Shuyuan; Zhao, Shuli; Zhang, Yingjie; Chen, Xiaodong
2016-01-01
It has been reported that human mesenchymal stem cells are able to inhibit T lymphocyte activation; however, the discrepancy among different sources of MSCs is not well documented. In this study, we have compared the MSCs from bone marrow (BM), adipose tissue (AT), placenta (PL), and umbilical cord (UC) to determine which one displayed the most efficient immunosuppressive effects on phytohemagglutinin-induced T cell proliferation. Among them we found that hUC-MSC has the strongest effects on inhibiting T cell proliferation and is chosen to do the further study. We observed that T lymphocyte spontaneously released abundant IFN-γ. And IFN-γ secreted by T lymphocyte could induce the expression of indoleamine 2, 3-dioxygenase (IDO) in hUC-MSCs. IDO was previously reported to induce T lymphocyte apoptosis and cell cycle arrest in S phase. When cocultured with hUC-MSCs, T lymphocyte expression of caspase 3 was significantly increased, while Bcl2 and CDK4 mRNA expression decreased dramatically. Addition of 1-methyl tryptophan (1-MT), an IDO inhibitor, restored T lymphocyte proliferation, reduced apoptosis, and induced resumption of the cell cycle. In addition, the changes in caspase 3, CDK4, and Bcl2 expression were reversed by 1-MT. These findings demonstrate that hUC-MSCs induce T lymphocyte apoptosis and cell cycle arrest by expressing abundant IDO and provide an explanation for some of the immunomodulatory effects of MSCs. PMID:27418932
SATB1 Expression Governs Epigenetic Repression of PD-1 in Tumor-Reactive T Cells.
Stephen, Tom L; Payne, Kyle K; Chaurio, Ricardo A; Allegrezza, Michael J; Zhu, Hengrui; Perez-Sanz, Jairo; Perales-Puchalt, Alfredo; Nguyen, Jenny M; Vara-Ailor, Ana E; Eruslanov, Evgeniy B; Borowsky, Mark E; Zhang, Rugang; Laufer, Terri M; Conejo-Garcia, Jose R
2017-01-17
Despite the importance of programmed cell death-1 (PD-1) in inhibiting T cell effector activity, the mechanisms regulating its expression remain poorly defined. We found that the chromatin organizer special AT-rich sequence-binding protein-1 (Satb1) restrains PD-1 expression induced upon T cell activation by recruiting a nucleosome remodeling deacetylase (NuRD) complex to Pdcd1 regulatory regions. Satb1 deficienct T cells exhibited a 40-fold increase in PD-1 expression. Tumor-derived transforming growth factor β (Tgf-β) decreased Satb1 expression through binding of Smad proteins to the Satb1 promoter. Smad proteins also competed with the Satb1-NuRD complex for binding to Pdcd1 enhancers, releasing Pdcd1 expression from Satb1-mediated repression, Satb1-deficient tumor-reactive T cells lost effector activity more rapidly than wild-type lymphocytes at tumor beds expressing PD-1 ligand (CD274), and these differences were abrogated by sustained CD274 blockade. Our findings suggest that Satb1 functions to prevent premature T cell exhaustion by regulating Pdcd1 expression upon T cell activation. Dysregulation of this pathway in tumor-infiltrating T cells results in diminished anti-tumor immunity. Copyright © 2017 Elsevier Inc. All rights reserved.
Wang, Sufan; Wan, Ting; Ye, Mingtong; Qiu, Yun; Pei, Lei; Jiang, Rui; Pang, Nengzhi; Huang, Yuanling; Liang, Baoxia; Ling, Wenhua; Lin, Xiaojun; Zhang, Zhenfeng; Yang, Lili
2018-07-01
Nicotinamide riboside (NR) is a nicotinamide adenine dinucleotide (NAD + ) precursor which is present in foods such as milk and beer. It was reported that NR can prevent obesity, increase longevity, and promote liver regeneration. However, whether NR can prevent ethanol-induced liver injuries is not known. This study aimed to explore the effect of NR on ethanol induced liver injuries and the underlying mechanisms. We fed C57BL/6 J mice with Lieber-DeCarli ethanol liquid diet with or without 400 mg/kg·bw NR for 16 days. Liver injuries and SirT1-PGC-1α-mitochondrial function were analyzed. In in vitro experiments, HepG2 cells (CYP2E1 over-expressing cells) were incubated with ethanol ± 0.5 mmol/L NR. Lipid accumulation and mitochondrial function were compared. SirT1 knockdown in HepG2 cells were further applied to confirm the role of SirT1 in the protection of NR on lipid accumulation. We found that ethanol significantly decreased the expression and activity of hepatic SirT1 and induced abnormal expression of enzymes of lipid metabolism in mice. Both in vivo and in vitro experiments showed that NR activated SirT1 through increasing NAD + levels, decreased oxidative stress, increased deacetylation of PGC-1α and mitochondrial function. In SirT1 knockdown HepG2 cells, NR lost its ability in enhancing mitochondrial function, and its protection against lipid accumulation induced by ethanol. NR can protect against ethanol induced liver injuries via replenishing NAD + , reducing oxidative stress, and activating SirT1-PGC-1α-mitochondrial biosynthesis. Our data indicate that SirT1 plays an important role in the protection of NR against lipid accumulation and mitochondrial dysfunctions induced by ethanol. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.
Xing, Zhiguo; Ni, Yaping; Zhao, Junjie; Ma, Xudong
2017-05-01
Glioblastoma multiforme is a type of central nervous system tumor with extremely poor prognosis. Previously, hydrogen peroxide (H 2 O 2 ), which promotes the oxidative stress response, has been reported to induce the apoptosis of glioma cells. Recently, secreted frizzled-related protein 1 (SFRP1) has been shown to be associated with various types of malignant tumors and with H 2 O 2 -induced oxidative stress in cardiomyocytes by negatively regulating the Wnt signaling pathway. This study aimed to explore SFRP1 expression and its roles in H 2 O 2 -induced apoptosis in human glioma cells. We found that the SFRP1 level was decreased in several human glioma cell lines, including U87, U251, and SW1783 cells. In U251 cells, SFRP1 could function as a cancer suppressor gene, and the growth of U251 cells could be inhibited not only by H 2 O 2 but also by the overexpression of SFRP1. Furthermore, we demonstrated that H 2 O 2 -induced SFRP1 gene demethylation partially contributed to H 2 O 2 -induced U251 cell apoptosis, which was verified by studies using an SFRP inhibitor (WAY-316606). Our research identified that H 2 O 2 -induced SFRP1 gene demethylation contributes to H 2 O 2 -induced apoptosis in human U251 glioma cells.
Shankar, Esaki Muthu; Che, Karlhans Fru; Messmer, Davorka; Lifson, Jeffrey D; Larsson, Marie
2011-01-01
Accumulating evidence indicates that immune impairment in persistent viral infections could lead to T-cell exhaustion. To evaluate the potential contribution of induction of negative costimulatory molecules to impaired T-cell responses, we primed naïve T cells with mature monocyte-derived dendritic cells (MDDCs) pulsed with HIV-1 in vitro. We used quantitative real-time polymerase chain reaction and flow cytometry, respectively, to compare the gene and surface-protein expression profiles of naïve T cells primed with HIV-pulsed or mock-pulsed DCs. We detected elevated expressions of negative costimulatory molecules, including lymphocyte activation gene-3 (LAG-3), CD160, cytolytic T-lymphocyte antigen-4 (CTLA-4), T-cell immunoglobulin mucin-containing domain-3 (TIM-3), programmed death-1 (PD-1) and TRAIL (tumor necrosis-factor–related apoptosis-inducing ligand) in T cells primed by HIV-pulsed DCs. The PD-1+ T-cell population also coexpressed TIM-3, LAG-3, and CTLA-4. Interestingly, we also found an increase in gene expression of the transcriptional repressors Blimp-1 (B-lymphocyte–induced maturation protein-1) and Foxp3 (forkhead transcription factor) in T-cells primed by HIV-pulsed DCs; Blimp-1 expression was directly proportional to the expression of the negative costimulatory molecules. Furthermore, levels of the effector cytokines interleukin-2, tumor necrosis factor-α and interferon-γ, and perforin and granzyme B were decreased in T-cell populations primed by HIV-pulsed DCs. In conclusion, in vitro priming of naïve T-cells with HIV-pulsed DC leads to expansion of T cells with coexpression of a broad array of negative costimulatory molecules and Blimp-1, with potential deleterious consequences for T-cell responses. PMID:21103670
DOE Office of Scientific and Technical Information (OSTI.GOV)
Trinks, Cecilia, E-mail: Cecilia.trinks@liu.se; Severinsson, Emelie A., E-mail: Emelie.severinsson@liu.se; Holmlund, Birgitta, E-mail: Birgitta.holmlund@lio.se
2011-07-08
Highlights: {yields} Canertinib induces caspase-mediated apoptosis in T-cell leukemia cells in vitro. {yields} Canertinib mediates activation of the intrinsic apoptotic pathway. {yields} Canertinib induces apoptosis in an ErbB receptor independent manner. {yields} Lymphocyte specific proteins as well as survival kinases are inhibited. {yields} Canertinib may act as a multi-kinase inhibiting drug in human T-cell malignancies. -- Abstract: Canertinib is a novel ErbB-receptor inhibitor currently in clinical development for the treatment of solid tumors overexpressing ErbB-receptors. We have recently demonstrated that canertinib displays anti-proliferative and pro-apoptotic effects in human myeloid leukemia cells devoid of ErbB-receptors. The mechanism mediating these effects aremore » however unknown. In this study, we show that canertinib is able to act as a multi-kinase inhibitor by inhibition of several intracellular kinases involved in T-cell signaling such as Akt, Erk1/2 and Zap-70, and reduced Lck protein expression in the human T-cell leukemia cell line Jurkat. Treatment with canertinib at a concentration of 2 {mu}M caused accumulation of Jurkat cells in the G{sub 1} cell cycle phase and increased doses induced apoptosis in a time-dependent manner. Apoptotic signs of treated cells were detected by Annexin V staining and cleavage of PARP, caspase-3, -8, -9, -10 and Bid. A subset of the pro-apoptotic signals mediated by canertinib could be significantly reduced by specific caspase inhibitors. Taken together, these results demonstrate the dual ability of canertinib to downregulate important signaling pathways and to activate caspase-mediated intrinsic apoptosis pathway in human T-cell leukemia cells.« less
Hoshino, Katsuaki; Kashiwamura, Shin-ichiro; Kuribayashi, Kozo; Kodama, Taku; Tsujimura, Tohru; Nakanishi, Kenji; Matsuyama, Tomohiro; Takeda, Kiyoshi; Akira, Shizuo
1999-01-01
T1/ST2, an orphan receptor with homology with the interleukin (IL)-1 receptor family, is expressed constitutively and stably on the surface of T helper type 2 (Th2) cells, but not on Th1 cells. T1/ST2 is also expressed on mast cells, which are critical for Th2-mediated effector responses. To evaluate whether T1/ST2 is required for Th2 responses and mast cell function, we have generated T1/ST2-deficient (T1/ST2−/−) mice and examined the roles of T1/ST2. Naive CD4+ T cells isolated from T1/ST2−/− mice developed to Th2 cells in response to IL-4 in vitro. T1/ST2−/− mice showed normal Th2 responses after infection with the helminthic parasite Nippostrongylus brasiliensis as well as in the mouse model of allergen-induced airway inflammation. In addition, differentiation and function of bone marrow–derived cultured mast cells were unaffected. These findings demonstrate that T1/ST2 does not play an essential role in development and function of Th2 cells and mast cells. PMID:10562328
DCAF1 controls T-cell function via p53-dependent and -independent mechanisms.
Guo, Zengli; Kong, Qing; Liu, Cui; Zhang, Song; Zou, Liyun; Yan, Feng; Whitmire, Jason K; Xiong, Yue; Chen, Xian; Wan, Yisong Y
2016-01-05
On activation, naive T cells grow in size and enter cell cycle to mount immune response. How the fundamental processes of T-cell growth and cell cycle entry are regulated is poorly understood. Here we report that DCAF1 (Ddb1-cullin4-associated-factor 1) is essential for these processes. The deletion of DCAF1 in T cells impairs their peripheral homeostasis. DCAF1 is upregulated on T-cell receptor activation and critical for activation-induced T-cell growth, cell cycle entry and proliferation. In addition, DCAF1 is required for T-cell expansion and function during anti-viral and autoimmune responses in vivo. DCAF1 deletion leads to a drastic stabilization of p53 protein, which can be attributed to a requirement of DCAF1 for MDM2-mediated p53 poly-ubiquitination. Importantly, p53 deletion rescues the cell cycle entry defect but not the growth defect of DCAF1-deficient cells. Therefore, DCAF1 is vital for T-cell function through p53-dependent and -independent mechanisms.
DCAF1 controls T-cell function via p53-dependent and -independent mechanisms
Guo, Zengli; Kong, Qing; Liu, Cui; Zhang, Song; Zou, Liyun; Yan, Feng; Whitmire, Jason K.; Xiong, Yue; Chen, Xian; Wan, Yisong Y.
2016-01-01
On activation, naive T cells grow in size and enter cell cycle to mount immune response. How the fundamental processes of T-cell growth and cell cycle entry are regulated is poorly understood. Here we report that DCAF1 (Ddb1–cullin4-associated-factor 1) is essential for these processes. The deletion of DCAF1 in T cells impairs their peripheral homeostasis. DCAF1 is upregulated on T-cell receptor activation and critical for activation-induced T-cell growth, cell cycle entry and proliferation. In addition, DCAF1 is required for T-cell expansion and function during anti-viral and autoimmune responses in vivo. DCAF1 deletion leads to a drastic stabilization of p53 protein, which can be attributed to a requirement of DCAF1 for MDM2-mediated p53 poly-ubiquitination. Importantly, p53 deletion rescues the cell cycle entry defect but not the growth defect of DCAF1-deficient cells. Therefore, DCAF1 is vital for T-cell function through p53-dependent and -independent mechanisms. PMID:26728942
Dimethyl fumarate–induced lymphopenia in MS due to differential T-cell subset apoptosis
Ghadiri, Mahtab; Rezk, Ayman; Li, Rui; Evans, Ashley; Luessi, Felix; Zipp, Frauke; Giacomini, Paul S.; Antel, Jack
2017-01-01
Objective: To examine the mechanism underlying the preferential CD8+ vs CD4+ T-cell lymphopenia induced by dimethyl fumarate (DMF) treatment of MS. Methods: Total lymphocyte counts and comprehensive T-cell subset analyses were performed in high-quality samples obtained from patients with MS prior to and serially following DMF treatment initiation. Random coefficient mixed-effects analysis was used to model the trajectory of T-cell subset losses in vivo. Survival and apoptosis of distinct T-cell subsets were assessed following in vitro exposure to DMF. Results: Best-fit modeling indicated that the DMF-induced preferential reductions in CD8+ vs CD4+ T-cell counts nonetheless followed similar depletion kinetics, suggesting a similar rather than distinct mechanism involved in losses of both the CD8+ and CD4+ T cells. In vitro, DMF exposure resulted in dose-dependent reductions in T-cell survival, which were found to reflect apoptotic cell death. This DMF-induced apoptosis was greater for CD8+ vs CD4+, as well as for memory vs naive, and conventional vs regulatory T-cell subsets, a pattern which mirrored preferential T-cell subset losses that we observed during in vivo treatment of patients. Conclusions: Differential apoptosis mediated by DMF may underlie the preferential lymphopenia of distinct T-cell subsets, including CD8+ and memory T-cell subsets, seen in treated patients with MS. This differential susceptibility of distinct T-cell subsets to DMF-induced apoptosis may contribute to both the safety and efficacy profiles of DMF in patients with MS. PMID:28377940
Kobayashi, Hiroya; Ngato, Toshihiro; Sato, Keisuke; Aoki, Naoko; Kimura, Shoji; Tanaka, Yuetsu; Aizawa, Hitoshi; Tateno, Masatoshi; Celis, Esteban
2006-01-01
Purpose Adult T-cell leukemia/lymphoma induced by human T-cell leukemia virus type 1 (HTLV-1) is usually a fatal lymphoproliferative malignant disease. HTLV-1 Tax protein plays a critical role in HTLV-1-associated leukemogenesis and is an attractive target for vaccine development. Although HTLV-1Tax is the most dominant antigen for HTLV-1-specific CD8+ CTLs in HTLV-1-infected individuals, few epitopes recognized by CD4+ helper T lymphocytes in HTLV-1Tax protein have been described.The aim of the present study was to study T-helper-cell responses to HTLV-1 Tax and to identify naturally processed MHC class II – restricted epitopes that could be used for vaccine development. Experimental Design An MHC class II binding peptide algorithm was used to predict potential T-helper cell epitope peptides from HTLV-1 Tax. We assessed the ability of the corresponding peptides to elicit helper T-cell responses by in vitro vaccination of purified CD4+ T lymphocytes. Results Peptides Tax191–205 and Tax305–319 were effective in inducingT-helper-cell responses. Although Tax191–205 was restricted by the HLA-DR1 and DR9 alleles, responses to Tax305–319 were restricted by either DR15 or DQ9. Both these epitopes were found to be naturally processed by HTLV-1+ T-cell lymphoma cells and by autologous antigen-presenting cells that were pulsed with HTLV-1Tax+ tumor lysates. Notably, the two newly identified helper T-cell epitopes are found to lie proximal to known CTL epitopes, which will facilitate the development of prophylactic peptide – based vaccine capable of inducing simultaneous CTL andT-helper responses. Conclusion Our data suggest that HTLV-1 Tax protein could serve as tumor-associated antigen for CD4+ helper T cells and that the present epitopes might be used for T-cell-based immunotherapy against tumors expressing HTLV-1. PMID:16778109
Pathogenic bacteria induce colonic PepT1 expression: an implication in host defense response
Nguyen, Hang Thi Thu; Dalmasso, Guillaume; Powell, Kimberly R.; Yan, Yutao; Bhatt, Shantanu; Kalman, Daniel; Sitaraman, Shanthi; Merlin, Didier
2009-01-01
Background & Aims Expression of the di/tripeptide transporter PepT1 has been observed in the colon under inflammatory conditions, however, the inducing factors and underlying mechanisms remain unknown. Here, we address the effects of pathogenic bacteria on colonic PepT1 expression together with its functional consequences. Methods Human colonic HT29-Cl.19A cells were infected with the attaching and effacing (A/E) enteropathogenic E. coli (EPEC). Wild-type and PepT1 transgenic mice or cultured colonic tissues derived from these mice were infected with Citrobacter rodentium, a murine A/E pathogen related to EPEC. Results EPEC induced PepT1 expression and activity in HT29-Cl.19A cells by intimately attaching to host cells through lipid rafts. Induction of PepT1 expression by EPEC required the transcription factor Cdx2. PepT1 expression reduced binding of EPEC to lipid rafts, as well as activation of NF-κB and MAP kinase and production of IL-8. Accordingly, ex vivo and in vivo experiments revealed that C. rodentium induced colonic PepT1 expression and that, compared to their wild-type counterparts, PepT1 transgenic mice infected with C. rodentium exhibited decreased bacterial colonization, production of pro-inflammatory cytokines, and neutrophil infiltration into the colon. Conclusions Our findings demonstrate a molecular mechanism underlying the regulation of colonic PepT1 expression under pathological conditions and reveal a novel role for PepT1 in host defense via its capacity to modulate bacterial-epithelial interactions and intestinal inflammation. PMID:19549526
CCR6 and NK1.1 distinguish between IL-17A and IFN-gamma-producing gammadelta effector T cells.
Haas, Jan D; González, Frano H Malinarich; Schmitz, Susanne; Chennupati, Vijaykumar; Föhse, Lisa; Kremmer, Elisabeth; Förster, Reinhold; Prinz, Immo
2009-12-01
Gammadelta T cells are a potent source of innate IL-17A and IFN-gamma, and they acquire the capacity to produce these cytokines within the thymus. However, the precise stages and required signals that guide this differentiation are unclear. Here we show that the CD24(low) CD44(high) effector gammadelta T cells of the adult thymus are segregated into two lineages by the mutually exclusive expression of CCR6 and NK1.1. Only CCR6+ gammadelta T cells produced IL-17A, while NK1.1+ gammadelta T cells were efficient producers of IFN-gamma but not of IL-17A. Their effector phenotype correlated with loss of CCR9 expression, particularly among the NK1.1+ gammadelta T cells. Accordingly, both gammadelta T-cell subsets were rare in gut-associated lymphoid tissues, but abundant in peripheral lymphoid tissues. There, they provided IL-17A and IFN-gamma in response to TCR-specific and TCR-independent stimuli. IL-12 and IL-18 induced IFN-gamma and IL-23 induced IL-17A production by NK1.1+ or CCR6+ gammadelta T cells, respectively. Importantly, we show that CCR6+ gammadelta T cells are more responsive to TCR stimulation than their NK1.1+ counterparts. In conclusion, our findings support the hypothesis that CCR6+ IL-17A-producing gammadelta T cells derive from less TCR-dependent selection events than IFN-gamma-producing NK1.1+ gammadelta T cells.
Linhart, B; Narayanan, M; Focke-Tejkl, M; Wrba, F; Vrtala, S; Valenta, R
2014-02-01
Vaccines consisting of allergen-derived peptides lacking IgE reactivity and allergen-specific T cell epitopes bound to allergen-unrelated carrier molecules have been suggested as candidates for allergen-specific immunotherapy. To study whether prophylactic and therapeutic vaccination with carrier-bound peptides from the major birch pollen allergen Bet v 1 lacking allergen-specific T cell epitopes has influence on Bet v 1-specific T cell responses. Three Bet v 1-derived peptides, devoid of Bet v 1-specific T cell epitopes, were coupled to KLH and adsorbed to aluminium hydroxide to obtain a Bet v 1-specific allergy vaccine. Groups of BALB/c mice were immunized with the peptide vaccine before or after sensitization to Bet v 1. Bet v 1- and peptide-specific antibody responses were analysed by ELISA. T cell and cytokine responses to Bet v 1, KLH, and the peptides were studied in proliferation assays. The effects of peptide-specific and allergen-specific antibodies on T cell responses and allergic lung inflammation were studied using specific antibodies. Prophylactic and therapeutic vaccination with carrier-bound Bet v 1 peptides induced a Bet v 1-specific IgG antibody response without priming/boosting of Bet v 1-specific T cells. Prophylactic and therapeutic vaccination of mice with the peptide vaccine induced Bet v 1-specific antibodies which suppressed Bet v 1-specific T cell responses and allergic lung inflammation. Vaccination with carrier-bound allergen-derived peptides lacking allergen-specific T cell epitopes induces allergen-specific IgG antibodies which suppress allergen-specific T cell responses and allergic lung inflammation. © 2013 John Wiley & Sons Ltd.
Xu, Aizhang; Bhanumathy, Kalpana Kalyanasundaram; Wu, Jie; Ye, Zhenmin; Freywald, Andrew; Leary, Scot C; Li, Rongxiu; Xiang, Jim
2016-01-01
Lymphopenia promotes naïve T-cell homeostatic proliferation and adoptive effector T-cell survival and memory formation. IL-7 plays a critical role in homeostatic proliferation, survival and memory formation of naïve T-cells in lymphopenia, and its underlying molecular mechanism has also been well studied. However, the mechanism for adoptively transferred effector T-cell survival and memory formation is not fully understood. Here, we transferred in vitro-activated transgenic OT-I CD8(+) effector T-cells into irradiation (600 rads)-induced lymphopenic C57BL/6, IL-7 knockout (KO) and IL-15 KO mice, and investigated the survival and memory formation of transferred T-cells in lymphopenia. We demonstrate that transferred T-cells prolong their survival and enhance their memory in lymphopenic mice, in a manner that depends on IL-15 signaling, but not IL-7. We determine that in vitro stimulation of naïve or effector T-cells with IL-7 and IL-15 reduces IL-7Rα, and increases and/or maintains IL-15Rβ expression, respectively. Consistent with these findings, the expression of IL-7Rα and IL-15Rβ is down- and up-regulated, respectively, in vivo on transferred T-cells in an early phase post T-cell transfer in lymphopenia. We further show that in vitro IL-15 restimulation-induced memory T-cells (compared to IL-2 restimulation-induced effector T-cells) and in vivo transferred T-cells in irradiated IL-15-sufficient C57BL/6 mice (compared to IL-15-deficient IL-15 KO mice) have increased mitochondrial content, but less NADH and lower mitochondrial potential (ΔΨm), and demonstrate greater phosphorylation of signal transducers and activators of transcription-5 (STAT5) and Unc-51-like kinase-1 (ULK1), and higher expression of B-cell leukemia/lymphoma-2 (Bcl2) and memory-, autophagy- and mitochondrial biogenesis-related molecules. Irradiation-induced lymphopenia promotes effector T-cell survival via IL-15 signaling the STAT5/Bcl2 pathway, enhances T-cell memory formation via IL
Zheng, Min; Zhang, Qinggao; Joe, Yeonsoo; Lee, Bong Hee; Ryu, Do Gon; Kwon, Kang Beom; Ryter, Stefan W; Chung, Hun Taeg
2013-03-01
Curcumin, a natural polyphenolic antioxidant compound, exerts well-known anti-inflammatory and immunomodulatory effects, the latter which can influence the activation of immune cells including T cells. Furthermore, curcumin can inhibit the expression of pro-inflammatory cytokines and chemokines, through suppression of the NF-κB signaling pathway. The beneficial effects of curcumin in diseases such as arthritis, allergy, asthma, atherosclerosis, diabetes and cancer may be due to its immunomodulatory properties. We studied the potential of curcumin to modulate CD4+ T cells-mediated autoimmune disease, by examining the effects of this compound on human CD4+ lymphocyte activation. Stimulation of human T cells with PHA or CD3/CD28 induced IL-2 mRNA expression and activated the endoplasmic reticulum (ER) stress response. The treatment of T cells with curcumin induced the unfolded protein response (UPR) signaling pathway, initiated by the phosphorylation of PERK and IRE1. Furthermore, curcumin increased the expression of the ER stress associated transcriptional factors XBP-1, cleaved p50ATF6α and C/EBP homologous protein (CHOP) in human CD4+ and Jurkat T cells. In PHA-activated T cells, curcumin further enhanced PHA-induced CHOP expression and reduced the expression of the anti-apoptotic protein Bcl-2. Finally, curcumin treatment induced apoptotic cell death in activated T cells via eliciting an excessive ER stress response, which was reversed by the ER-stress inhibitor 4-phenylbutyric acid or transfection with CHOP-specific siRNA. These results suggest that curcumin can impact both ER stress and mitochondria functional pathways, and thereby could be used as a promising therapy in the context of Th1-mediated autoimmune diseases. Copyright © 2013 Elsevier B.V. All rights reserved.
Watanabe, Toshiki
2017-03-02
Adult T-cell leukemia (ATL) is an aggressive T-cell malignancy caused by human T-cell leukemia virus type 1 (HTLV-1) that develops through a multistep carcinogenesis process involving 5 or more genetic events. We provide a comprehensive overview of recently uncovered information on the molecular basis of leukemogenesis in ATL. Broadly, the landscape of genetic abnormalities in ATL that include alterations highly enriched in genes for T-cell receptor-NF-κB signaling such as PLCG1 , PRKCB , and CARD11 and gain-of function mutations in CCR4 and CCR7 Conversely, the epigenetic landscape of ATL can be summarized as polycomb repressive complex 2 hyperactivation with genome-wide H3K27 me3 accumulation as the basis of the unique transcriptome of ATL cells. Expression of H3K27 methyltransferase enhancer of zeste 2 was shown to be induced by HTLV-1 Tax and NF-κB. Furthermore, provirus integration site analysis with high-throughput sequencing enabled the analysis of clonal composition and cell number of each clone in vivo, whereas multicolor flow cytometric analysis with CD7 and cell adhesion molecule 1 enabled the identification of HTLV-1-infected CD4 + T cells in vivo. Sorted immortalized but untransformed cells displayed epigenetic changes closely overlapping those observed in terminally transformed ATL cells, suggesting that epigenetic abnormalities are likely earlier events in leukemogenesis. These new findings broaden the scope of conceptualization of the molecular mechanisms of leukemogenesis, dissecting them into immortalization and clonal progression. These recent findings also open a new direction of drug development for ATL prevention and treatment because epigenetic marks can be reprogrammed. Mechanisms underlying initial immortalization and progressive accumulation of these abnormalities remain to be elucidated. © 2017 by The American Society of Hematology.
Human Atopic Dermatitis Skin-derived T Cells can Induce a Reaction in Mouse Keratinocytes in vivo.
Martel, B C; Blom, L; Dyring-Andersen, B; Skov, L; Thestrup-Pedersen, K; Skov, S; Skak, K; Poulsen, L K
2015-08-01
In atopic dermatitis (AD), the inflammatory response between skin-infiltrating T cells and keratinocytes is fundamental to the development of chronic lesional eczema. The aim of this study was to investigate whether skin-derived T cells from AD patients could induce an inflammatory response in mice through keratinocyte activation and consequently cause the development of eczematous lesions. Punch biopsies of the lesional skin from AD patients were used to establish skin-derived T cell cultures, which were transferred to NOD.Cg-Prkd(scid) Il2rg(tm1Sug) /JicTac (NOG) mice. We found that the subcutaneous injection of the human AD skin-derived T cells resulted in the migration of the human T cells from subcutis to the papillary dermis followed by the development of erythema and oedema in the mouse skin. Furthermore, the human T cells induced a transient proliferative response in the mouse keratinocytes shown as increased numbers of Ki-67(+) keratinocytes and increased epidermal thickness. Out of six established AD skin-derived T cell cultures, two were superior at inducing a skin reaction in the mice, and these cultures were found to contain >10% CCR10(+) T cells compared to <2% for the other cultures. In comparison, blood-derived in vitro-differentiated Th2 cells only induced a weak response in a few of the mice. Thus, we conclude that human AD skin-derived T cells can induce a reaction in the mouse skin through the induction of a proliferative response in the mouse keratinocytes. © 2015 The Foundation for the Scandinavian Journal of Immunology.
Leukocyte-specific protein 1 regulates T-cell migration in rheumatoid arthritis
Hwang, Seong-Hye; Jung, Seung-Hyun; Lee, Saseong; Choi, Susanna; Yoo, Seung-Ah; Park, Ji-Hwan; Hwang, Daehee; Shim, Seung Cheol; Sabbagh, Laurent; Kim, Ki-Jo; Park, Sung Hwan; Cho, Chul-Soo; Kim, Bong-Sung; Leng, Lin; Montgomery, Ruth R.; Bucala, Richard; Chung, Yeun-Jun; Kim, Wan-Uk
2015-01-01
Copy number variations (CNVs) have been implicated in human diseases. However, it remains unclear how they affect immune dysfunction and autoimmune diseases, including rheumatoid arthritis (RA). Here, we identified a novel leukocyte-specific protein 1 (LSP1) deletion variant for RA susceptibility located in 11p15.5. We replicated that the copy number of LSP1 gene is significantly lower in patients with RA, which correlates positively with LSP1 protein expression levels. Differentially expressed genes in Lsp1-deficient primary T cells represent cell motility and immune and cytokine responses. Functional assays demonstrated that LSP1, induced by T-cell receptor activation, negatively regulates T-cell migration by reducing ERK activation in vitro. In mice with T-cell–dependent chronic inflammation, loss of Lsp1 promotes migration of T cells into the target tissues as well as draining lymph nodes, exacerbating disease severity. Moreover, patients with RA show diminished expression of LSP1 in peripheral T cells with increased migratory capacity, suggesting that the defect in LSP1 signaling lowers the threshold for T-cell activation. To our knowledge, our work is the first to demonstrate how CNVs result in immune dysfunction and a disease phenotype. Particularly, our data highlight the importance of LSP1 CNVs and LSP1 insufficiency in the pathogenesis of RA and provide previously unidentified insights into the mechanisms underlying T-cell migration toward the inflamed synovium in RA. PMID:26554018
Liu, Kan; Zhao, Chaofei; Chen, Jianwen; Wu, Shengpan; Yao, Yuanxin; Wu, Chong; Luo, Guoxiong; Zhang, Xu
2016-06-01
Objective To establish selenoprotein P, plasma 1 (SEPP1) gene recombinant lentiviral vector and investigate the effect of SEPP1 on the proliferation of human clear cell renal cell carcinoma (ccRCC) cells. Methods cDNA sequence of SEPP1 was cloned from the total cDNA of HEK293T cells by PCR. Then, the cDNA fragment was combined with the pLV-EGFP(2A)Puro vector and the constructed plasmid pLV-EGFP(2A)Puro-SEPP1 was transfected into HEK293T cells for packaging the virus. Forty-eight hours after transfected with the virus supernatant, the level of SEPP1 protein in 769-P and 786-O cells were tested by Western blotting. Cells were divided into recombinant lentivirus-infected cells, empty vector lentivirus-infected cells and the blank control cells. Cell proliferation rate was detected by MTS assay, colony forming ability was evaluated by plate clony formation assay and cell cycle change was assayed by flow cytometry after transfected with pLV-EGFP(2A)Puro-SEPP1 or empty pLV-EGFP(2A)Puro vector. Results Enzyme digestion analysis and DNA sequencing showed that the recombinant plasmid pLV-EGFP(2A)Puro-SEPP1 was constructed successfully. After being infected by the virus supernatant, the 786-O and 769-P cells expressed EGFP. Compared with the empty vector group and the blank control group, expression level of SEPP1 in the experimental group was much higher. The cell proliferative ability was inhibited in the cells overexpressing SEPP1, and the colony forming ability of SEPP1-overexpressed cells evidently decreased. Cell cycle was arrested in G2/M phase in 786-O cells overexpressing SEPP1. Conclusion The recombinant plasmid pLV-EGFP(2A)Puro-SEPP1 has been constructed successfully. Overexpression of SEPP1 could significantly reduce the proliferation rate of 786-O and 769P cells, and cause G2/M phase arrest of 786-O cells.
Olds, Peter; Park, Andrew; Schlesinger, Sarah J.
2011-01-01
Foxp3+ regulatory T cells (T reg cells) effectively suppress immunity, but it is not determined if antigen-induced T reg cells (iT reg cells) are able to persist under conditions of inflammation and to stably express the transcription factor Foxp3. We used spleen cells to stimulate the mixed leukocyte reaction (MLR) in the presence of transforming growth factor β (TGF-β) and retinoic acid. We found that the CD11chigh dendritic cell fraction was the most potent at inducing high numbers of alloreactive Foxp3+ cells. The induced CD4+CD25+Foxp3+ cells appeared after extensive proliferation. When purified from the MLR, iT reg cells suppressed both primary and secondary MLR in vitro in an antigen-specific manner. After transfer into allogeneic mice, iT reg cells persisted for 6 mo and prevented graft versus host disease (GVHD) caused by co-transferred CD45RBhi T cells. Similar findings were made when iT reg cells were transferred after onset of GVHD. The CNS2 intronic sequence of the Foxp3 gene in the persisting iT reg cells was as demethylated as the corresponding sequence of naturally occurring T reg cells. These results indicate that induced Foxp3+ T reg cells, after proliferating and differentiating into antigen-specific suppressive T cells, can persist for long periods while suppressing a powerful inflammatory disease. PMID:22084406
Egan, Charlotte E; Dalton, Jane E; Andrew, Elizabeth M; Smith, Judith E; Gubbels, Marc-Jan; Striepen, Boris; Carding, Simon R
2005-12-15
gammadelta T cells are a diverse population of T cells that are widely distributed and are a common feature of pathogen-induced immune responses. It is not clear, however, whether different populations of gammadelta T cells have specific functions, and what factors determine the functional properties of individual populations. A murine model of peroral Toxoplasma gondii infection was used to determine the contribution Vgamma1+ intestinal intraepithelial lymphocytes (IELs) vs systemic Vgamma1+ T cells make to the acute and chronic stages of the host immune response, and whether the macrophage cytocidal activity of Vgamma1+ T cells described in bacterial infections is seen in other, unrelated infectious disease models. In response to oral infection with virulent type 1 or avirulent type II strains of T. gondii, TCR-delta-/- mice rapidly developed severe ileitis. In contrast, in mice deficient in Vgamma1+ T cells and IELs and wild-type mice, inflammation was delayed in onset and less severe. The protective effect of (Vgamma1-) IELs to Toxoplasma infection was unrelated to their cytolytic and cytokine (Th1)-producing capabilities. Systemic Vgamma1+ T cells were shown to play an essential role in limiting parasite growth and inflammation in peripheral tissues and, in particular, in the CNS, that was associated with their ability to efficiently kill parasite-elicited and infected macrophages. These findings suggest that macrophage cytocidal activity of Vgamma1+ T cells may be a universal feature of pathogen-induced immune responses and that microenvironmental factors influence the involvement and function of gammadelta T cells in the host response to infection.
Yoon, Yeo Cho; Kim, Sung-Hee; Kim, Min Jung; Yang, Hye Jeong; Rhyu, Mee-Ra; Park, Jae-Ho
2015-01-01
This study investigated the effects of an ethanol extract of black pepper and its constituent, piperine, on odorant-induced signal transduction in non-chemosensory cells. An ethanol extract of black pepper decreased eugenol-induced cAMP and calcium levels in preadipocyte 3T3-L1 cells with no toxicity. Phosphorylation of CREB (cAMP response element-binding protein) was down-regulated by the black pepper extract. The concentration (133.8 mg/g) and retention time (5.5 min) of piperine in the ethanol extract were quantified using UPLC-MS/MS. Pretreatment with piperine decreased eugenol-induced cAMP and calcium levels in 3T3-L1 cells. Piperine also decreased the phosphorylation of CREB, which is up-regulated by eugenol. These results suggest that piperine inhibits the eugenol-induced signal transduction pathway through modulation of cAMP and calcium levels and phosphorylation of CREB in non-chemosensory cells.
NASA Astrophysics Data System (ADS)
Yan, Li; Liu, Xiao; Liu, Wei-Xia; Tan, Xiao-Qiu; Xiong, Fei; Gu, Ning; Hao, Wei; Gao, Xue; Cao, Ji-Min
2015-12-01
Superparamagnetic iron oxide nanoparticles (SPIONs) are promising nanomaterials in medical practice due to their special magnetic characteristics and nanoscale size. However, their potential impacts on immune cells are not well documented. This study aims to investigate the effects of Fe2O3 nanoparticles (Fe2O3-NPs) on the electrophysiology of Kv1.3 channels in Jurkat T cells. Using the whole-cell patch-clamp technique, we demonstrate that incubation of Jurkat cells with Fe2O3-NPs dose- and time-dependently decreased the current density and shifted the steady-state inactivation curve and the recovery curve of Kv1.3 channels to a rightward direction. Fe2O3-NPs increased the NADP level but decreased the NADPH level of Jurkat cells. Direct induction of NADPH into the cytosole of Jurkat cells via the pipette abolished the rightward shift of the inactivation curve. In addition, transmission electron microscopy showed that Fe2O3-NPs could be endocytosed by Jurkat cells with relatively low speed and capacity. Fe2O3-NPs did not significantly affect the viability of Jurkat cells, but suppressed the expressions of certain cytokines (TNFα, IFNγ and IL-2) and interferon responsive genes (IRF-1 and PIM-1), and the time courses of Fe2O3-NPs endocytosis and effects on the expressions of cytokines and interferon responsive genes were compatible. We conclude that Fe2O3-NPs can be endocytosed by Jurkat cells and act intracellularly. Fe2O3-NPs decrease the current density and delay the inactivation and recovery kinetics of Kv1.3 channels in Jurkat cells by oxidizing NADPH and therefore disrupting the redox activity of the Kvβ2 auxiliary subunit, and as a result, lead to changes of the Kv1.3 channel function. These results suggest that iron oxide nanoparticles may affect T cell function by disturbing the activity of Kv1.3 channels. Further, the suppressing effects of Fe2O3-NPs on the expressions of certain inflammatory cytokines and interferon responsive genes suggest that iron
DOE Office of Scientific and Technical Information (OSTI.GOV)
Liu, Qingqing; Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qi-xiu Road, Nantong 226001, Jiangsu Province; Tao, Tao
As an essential post-translational modification, O-GlcNAcylation has been thought to be able to modulate various nuclear and cytoplasmic proteins and is emerging as a key regulator of multiple biological processes, such as transcription, cell growth, signal transduction, and cell motility. Recently, authoritative glycomics analyses have reported extensive crosstalk between O-GlcNAcylation and phosphorylation, which always dynamically interplay with each other and regulate signaling, transcription, and other cellular processes. Also, plentiful studies have shown close correlation between YB-1 phosphorylation and tumorigenesis. Therefore, our study aimed to determine whether YB-1 was O-GlcNAc modified and whether such modification could interact with its phosphorylation duringmore » the process of HCC development. Western blot and immunohistochemistry were firstly conducted to reveal obvious up-regulation of YB-1, OGT and O-GlcNAc modification in HCC tissues. What is more, not only YB-1 was identified to be O-GlcNAcylated but hyper-O-GlcNAcylation was demonstrated to facilitate HCC cell proliferation in a YB-1 dependent manner. Moreover, we detected four specific O-GlcNAc sites and confirmed T126A to be the most effective mutant in HCC cell proliferation via close O-GlcNAcylation-phosphorylation interaction. Even more interestingly, we discovered that T126A-induced HCC cell retardation and subdued transcriptional activity of YB-1 could be partially reversed by T126A/S102E mutant. From all above, it is not difficult to find that glycosylated-YB-1 mainly enhanced cell proliferation through congenerous actions with YB-1 phosphorylation and thus played indispensable roles in fine-tuning cell proliferation and procession of HCC. - Highlights: • YB-1 and OGT are associated with HCC prognosis. • YB-1 is O-GlcNAc modified in HCC. • Hyper-O-GlcNAcylation promotes HCC cell proliferation in dependent of YB-1. • The proliferating role of O-GlcNAcylation is based on Ser
Watkins, Stephan; Pichler, Werner J.
2013-01-01
T cell receptors (TCR) containing Vβ20-1 have been implicated in a wide range of T cell mediated disease and allergic reactions, making it a target for understanding these. Mechanics of T cell receptors are largely unexplained by static structures available from x-ray crystallographic studies. A small number of molecular dynamic simulations have been conducted on TCR, however are currently lacking either portions of the receptor or explanations for differences between binding and non-binding TCR recognition of respective peptide-HLA. We performed molecular dynamic simulations of a TCR containing variable domain Vβ20-1, sequenced from drug responsive T cells. These were initially from a patient showing maculopapular eruptions in response to the sulfanilamide-antibiotic sulfamethoxazole (SMX). The CDR2β domain of this TCR was found to dock SMX with high affinity. Using this compound as a perturbation, overall mechanisms involved in responses mediated by this receptor were explored, showing a chemical action on the TCR free from HLA or peptide interaction. Our simulations show two completely separate modes of binding cognate peptide-HLA complexes, with an increased affinity induced by SMX bound to the Vβ20-1. Overall binding of the TCR is mediated through a primary recognition by either the variable β or α domain, and a switch in recognition within these across TCR loops contacting the peptide and HLA occurs when SMX is present in the CDR2β loop. Large binding affinity differences are induced by summed small amino acid changes primarily by SMX modifying only three critical CDR2β loop amino acid positions. These residues, TYRβ57, ASPβ64, and LYSβ65 initially hold hydrogen bonds from the CDR2β to adjacent CDR loops. Effects from SMX binding are amplified and traverse longer distances through internal TCR hydrogen bonding networks, controlling the overall TCR conformation. Thus, the CDR2β of Vβ20-1 acts as a ligand controlled switch affecting overall TCR
IL-1β and IL-23 Promote Extrathymic Commitment of CD27+CD122− γδ T Cells to γδT17 Cells
2017-01-01
γδT17 cells are a subset of γδ T cells committed to IL-17 production and are characterized by the expression of IL-23R and CCR6 and lack of CD27 expression. γδT17 cells are believed to arise within a narrow time window during prenatal thymic development. In agreement with this concept, we show in this study that adult Rag1−/− recipient mice of Il23rgfp/+ (IL-23R reporter) bone marrow selectively lack IL-23R+ γδT17 cells. Despite their absence in secondary lymphoid tissues during homeostasis, γδT17 cells emerge in bone marrow chimeric mice upon induction of skin inflammation by topical treatment with imiquimod cream (Aldara). We demonstrate that IL-1β and IL-23 together are able to promote the development of bona fide γδT17 cells from peripheral CD122−IL-23R− γδ T cells, whereas CD122+ γδ T cells fail to convert into γδT17 cells and remain stable IFN-γ producers (γδT1 cells). IL-23 is instrumental in expanding extrathymically generated γδT17 cells. In particular, TCR-Vγ4+ chain–expressing CD122−IL-23R− γδ T cells are induced to express IL-23R and IL-17 outside the thymus during skin inflammation. In contrast, TCR-Vγ1+ γδ T cells largely resist this process because prior TCR engagement in the thymus has initiated their commitment to the γδT1 lineage. In summary, our data reveal that the peripheral pool of γδ T cells retains a considerable degree of plasticity because it harbors “naive” precursors, which can be induced to produce IL-17 and replenish peripheral niches that are usually occupied by thymus-derived γδT17 cells. PMID:28855314
Zhang, Xiaoli; Rocha-Ferreira, Eridan; Li, Tao; Vontell, Regina; Jabin, Darakhshan; Hua, Sha; Zhou, Kai; Nazmi, Arshed; Albertsson, Anna-Maj; Sobotka, Kristina; Ek, Joakim; Thornton, Claire; Hagberg, Henrik; Mallard, Carina; Leavenworth, Jianmei W; Zhu, Changlian; Wang, Xiaoyang
2017-12-20
Infection and sepsis are associated with brain white matter injury in preterm infants and the subsequent development of cerebral palsy. In the present study, we used a neonatal mouse sepsis-induced white matter injury model to determine the contribution of different T cell subsets (αβT cells and γδT cells) to white matter injury and consequent behavioral changes. C57BL/6J wild-type (WT), T cell receptor (TCR) δ-deficient (Tcrd -/- , lacking γδT cells), and TCRα-deficient (Tcra -/- , lacking αβT cells) mice were administered with lipopolysaccharide (LPS) at postnatal day (PND) 2. Brain myelination was examined at PNDs 12, 26, and 60. Motor function and anxiety-like behavior were evaluated at PND 26 or 30 using DigiGait analysis and an elevated plus maze. White matter development was normal in Tcrd -/- and Tcrα -/- compared to WT mice. LPS exposure induced reductions in white matter tissue volume in WT and Tcrα -/- mice, but not in the Tcrd -/- mice, compared with the saline-treated groups. Neither LPS administration nor the T cell deficiency affected anxiety behavior in these mice as determined with the elevated plus maze. DigiGait analysis revealed motor function deficiency after LPS-induced sepsis in both WT and Tcrα -/- mice, but no such effect was observed in Tcrd -/- mice. Our results suggest that γδT cells but not αβT cells contribute to sepsis-induced white matter injury and subsequent motor function abnormalities in early life. Modulating the activity of γδT cells in the early stages of preterm white matter injury might represent a novel therapeutic strategy for the treatment of perinatal brain injury.
Jubin, Virginie; Ventre, Erwan; Leverrier, Yann; Djebali, Sophia; Mayol, Katia; Tomkowiak, Martine; Mafille, Julien; Teixeira, Marie; Teoh, Denise Y-L; Lina, Bruno; Walzer, Thierry; Arpin, Christophe; Marvel, Jacqueline
2012-06-01
Besides the classically described subsets of memory CD8 T cells generated under infectious conditions, are T inflammatory memory cells generated under sterile priming conditions, such as sensitization to allergens. Although not fully differentiated as pathogen-induced memory cells, they display memory properties that distinguish them from naive CD8 T cells. Given these memory cells are generated in an antigen-specific context that is devoid of pathogen-derived danger signals and CD4 T cell help, we herein questioned whether they maintained their activation and differentiation potential, could be recruited in an immune response directed against a pathogen expressing their cognate antigen and further differentiate in fully competent secondary memory cells. We show that T inflammatory memory cells can indeed take part to the immune response triggered by a viral infection, differentiate into secondary effectors and further generate typical central memory CD8 T cells and effector memory CD8 T cells. Furthermore, the secondary memory cells they generate display a functional advantage over primary memory cells in their capacity to produce TNF-α and the XCL1 chemokine. These results suggest that cross-reactive stimulations and differentiation of cells directed against allergens or self into fully competent pathogen-induced memory cells might have incidences in inflammatory immuno-pathologies.
Mouse glucocorticoid-induced tumor necrosis factor receptor ligand is costimulatory for T cells
Tone, Masahide; Tone, Yukiko; Adams, Elizabeth; Yates, Stephen F.; Frewin, Mark R.; Cobbold, Stephen P.; Waldmann, Herman
2003-01-01
Recently, agonist antibodies to glucocorticoid-induced tumor necrosis factor receptor (GITR) (tumor necrosis factor receptor superfamily 18) have been shown to neutralize the suppressive activity of CD4+CD25+ regulatory T cells. It was anticipated that this would be the role of the physiological ligand. We have identified and expressed the gene for mouse GITR ligand and have confirmed that its interaction with GITR reverses suppression by CD4+CD25+ T cells. It also, however, provides a costimulatory signal for the antigen-driven proliferation of naïve T cells and polarized T helper 1 and T helper 2 clones. RT-PCR and mAb staining revealed mouse GITR ligand expression in dendritic cells, macrophages, and B cells. Expression was controlled by the transcription factor NF-1 and potentially by alternative splicing of mRNA destabilization sequences. PMID:14608036
Chen, Dandan; Tang, Ping; Liu, Linxiang; Wang, Fang; Xing, Haizhou; Sun, Ling; Jiang, Zhongxing
2018-05-21
This study aims to explore the effect of bone marrow mesenchymal stem cells (BMSCs) on multiple myeloma (MM) development and the underlying mechanism. BMSCs from C57BL/6 J mice were isolated and the third passage was used for subsequent experiments. Additionally, a series of in vitro transwell coculture assays were performed to explore the effects of BMSCs on the proliferation of MM cells 5TGM1 and CD4 + T cells. Furthermore, a 5TGM1-induced MM mice model was established. Moreover, PD-L1 shRNA was transfected into BMSCs to investigate whether PD-1/PD-L1 pathway involved in BMSCs-mediated regulation of T cells and MM growth. Data revealed that BMSCs significantly promoted 5TGM1 proliferation in a dose-dependent manner. Furthermore, BMSCs administration exerted stimulatory effects on MM development in terms of shortening the mouse survival rate, promoting tumor growth, and enhancing inflammatory infiltration in the MM model mice. Moreover, BMSCs decreased the percentage of Th1 and Th17 cells, whereas increased that of Th2 and Treg cells. Their corresponding cytokines of these T cell subsets showed similar alteration in the presence of BMSCs. Additionally, BMSCs significantly suppressed CD4 + T cell proliferation. We also found that PD-L1 shRNA inhibited 5TGM1 proliferation likely through activation of CD4 + T cells. Further in vivo experiments confirmed that PD-L1 inhibition attenuated BMSCs-induced MM growth, inflammation infiltration and imbalance of Th1/Th2 and Th17/Treg. In summary, our findings demonstrated that BMSCs promoted cell proliferation of MM through inhibiting T cell immune responses via PD-1/PD-L1 pathway.
SirT1 mediates hyperbaric oxygen preconditioning-induced ischemic tolerance in rat brain
Yan, Wenjun; Fang, Zongping; Yang, Qianzi; Dong, Hailong; Lu, Yan; Lei, Chong; Xiong, Lize
2013-01-01
Our previous studies have shown that hyperbaric oxygen preconditioning (HBO-PC) induces tolerance to cerebral ischemia/reperfusion (I/R). This study aimed to investigate whether SirT1, a class III histone deacetylase, is involved in neuroprotection elicited by HBO-PC in animal and cell culture models of ischemia. Rats were subjected to middle cerebral artery occlusion for 120 minutes after HBO-PC (once a day for 5 days). Primary cultured cortical neurons were exposed to 2 hours of HBO-PC after 2 hours of oxygen–glucose deprivation (OGD). We showed that HBO-PC increased SirT1 protein and mRNA expression, promoted neurobehavioral score, reduced infarct volume, and improved morphology at 24 hours and 7 days after cerebral I/R. Neuroprotection of HBO-PC was attenuated by SirT1 inhibitor EX527 and SirT1 knockdown by short interfering RNA (siRNA), whereas it was mimicked by SirT1 activator resveratrol. Furthermore, HBO-PC enhanced SirT1 expression and cell viability and reduced lactate dehydrogenase release 24 hours after OGD/re-oxygenation. The neuroprotective effect of HBO-PC was emulated through upregulating SirT1 and, reversely, attenuated through downregulating SirT1. The modulation of SirT1 was made by adenovirus infection carrying SirT1 or SirT1 siRNA. Besides, SirT1 increased B-cell lymphoma 2 (Bcl-2) expression and decrease cleaved caspase 3. These results indicate that SirT1 mediates HBO-PC-induced tolerance to cerebral I/R through inhibition of apoptosis. PMID:23299244
Roux, Clémence; Saviane, Gaëlle; Pini, Jonathan; Belaïd, Nourhène; Dhib, Gihen; Voha, Christine; Ibáñez, Lidia; Boutin, Antoine; Mazure, Nathalie M.; Wakkach, Abdelilah; Blin-Wakkach, Claudine; Rouleau, Matthieu
2018-01-01
Despite mesenchymal stromal cells (MSCs) are considered as a promising source of cells to modulate immune functions on cells from innate and adaptive immune systems, their clinical use remains restricted (few number, limited in vitro expansion, absence of a full phenotypic characterization, few insights on their in vivo fate). Standardized MSCs derived in vitro from human-induced pluripotent stem (huIPS) cells, remediating part of these issues, are considered as well as a valuable tool for therapeutic approaches, but their functions remained to be fully characterized. We generated multipotent MSCs derived from huiPS cells (huiPS-MSCs), and focusing on their immunosuppressive activity, we showed that human T-cell activation in coculture with huiPS-MSCs was significantly reduced. We also observed the generation of functional CD4+ FoxP3+ regulatory T (Treg) cells. Further tested in vivo in a model of human T-cell expansion in immune-deficient NSG mice, huiPS-MSCs immunosuppressive activity prevented the circulation and the accumulation of activated human T cells. Intracytoplasmic labeling of cytokines produced by the recovered T cells showed reduced percentages of human-differentiated T cells producing Th1 inflammatory cytokines. By contrast, T cells producing IL-10 and FoxP3+-Treg cells, absent in non-treated animals, were detected in huiPS-MSCs treated mice. For the first time, these results highlight the immunosuppressive activity of the huiPS-MSCs on human T-cell stimulation with a concomitant generation of human Treg cells in vivo. They may favor the development of new tools and strategies based on the use of huiPS cells and their derivatives for the induction of immune tolerance. PMID:29422893
Roux, Clémence; Saviane, Gaëlle; Pini, Jonathan; Belaïd, Nourhène; Dhib, Gihen; Voha, Christine; Ibáñez, Lidia; Boutin, Antoine; Mazure, Nathalie M; Wakkach, Abdelilah; Blin-Wakkach, Claudine; Rouleau, Matthieu
2017-01-01
Despite mesenchymal stromal cells (MSCs) are considered as a promising source of cells to modulate immune functions on cells from innate and adaptive immune systems, their clinical use remains restricted (few number, limited in vitro expansion, absence of a full phenotypic characterization, few insights on their in vivo fate). Standardized MSCs derived in vitro from human-induced pluripotent stem (huIPS) cells, remediating part of these issues, are considered as well as a valuable tool for therapeutic approaches, but their functions remained to be fully characterized. We generated multipotent MSCs derived from huiPS cells (huiPS-MSCs), and focusing on their immunosuppressive activity, we showed that human T-cell activation in coculture with huiPS-MSCs was significantly reduced. We also observed the generation of functional CD4 + FoxP3 + regulatory T (Treg) cells. Further tested in vivo in a model of human T-cell expansion in immune-deficient NSG mice, huiPS-MSCs immunosuppressive activity prevented the circulation and the accumulation of activated human T cells. Intracytoplasmic labeling of cytokines produced by the recovered T cells showed reduced percentages of human-differentiated T cells producing Th1 inflammatory cytokines. By contrast, T cells producing IL-10 and FoxP3 + -Treg cells, absent in non-treated animals, were detected in huiPS-MSCs treated mice. For the first time, these results highlight the immunosuppressive activity of the huiPS-MSCs on human T-cell stimulation with a concomitant generation of human Treg cells in vivo . They may favor the development of new tools and strategies based on the use of huiPS cells and their derivatives for the induction of immune tolerance.
Said, Sawsan Sudqi; Barut, Guliz Tuba; Mansur, Nesteren; Korkmaz, Asli; Sayi-Yazgan, Ayca
2018-04-01
Regulatory B cells (Bregs) play a crucial role in immunological tolerance primarily through the production of IL-10 in many diseases including autoimmune disorders, allergy, infectious diseases, and cancer. To date, various Breg subsets with overlapping phenotypes have been identified. However, the roles of Bregs in Helicobacter infection are largely unknown. In the present study, we investigate the phenotype and function of Helicobacter -stimulated B cells. Our results demonstrate that Helicobacter felis -stimulated IL-10- producing B cells (Hf stim - IL-10 + B) are composed of B10 and Transitional 2 Marginal Zone Precursor (T2-MZP) cells with expression of CD9, Tim-1, and programmed death 1 (PD-1). On the other hand, Helicobacter felis -stimulated IL-10- nonproducing B (Hf stim - IL-10 - B) cells are mainly marginal zone (MZ) B cells that express PD-L1 and secrete TGF-β, IL-6, and TNF-α, and IgM and IgG2b. Furthermore, we show that both Hf stim - IL-10 + B cells and Hf stim - IL-10 - B cells induce CD49b + LAG-3 + Tr1 cells. Here, we describe a novel mechanism for PD-1/PD-L1- driven B cell-dependent Tr1 cell differentiation. Finally, we explore the capability of Hf stim - IL-10 - B cells to induce Th17 cell differentiation, which we find to be dependent on TGF-β. Taken together, the current study demonstrates that Hf stim - B cells induce Tr1 cells through the PD-1/PD-L1 axis and Th17 cells by secreting TGF-β. Copyright © 2018 Elsevier Ltd. All rights reserved.
Neuroligin-3 protects retinal cells from H2O2-induced cell death via activation of Nrf2 signaling.
Li, Xiu-Miao; Huang, Dan; Yu, Qing; Yang, Jian; Yao, Jin
2018-05-25
Intensified oxidative stress can cause severe damage to human retinal pigment epithelium (RPE) cells and retinal ganglion cells (RGCs). The potential effect of neuroligin-3 (NLGN3) against the process is studied here. Our results show that NLGN3 efficiently inhibited hydrogen peroxide (H 2 O 2 )-induced death and apoptosis in human RPE cells and RGCs. H 2 O 2 -induced reactive oxygen species (ROS) production, lipid peroxidation and DNA damage in retinal cells were alleviated by NLGN3. NLGN3 activated nuclear-factor-E2-related factor 2 (Nrf2) signaling, enabling Nrf2 protein stabilization, nuclear translocation and expression of key anti-oxidant enzymes (HO1, NOQ1 and GCLC) in RPE cells and RGCs. Further results demonstrate that NLGN3 activated Akt-mTORC1 signaling in retinal cells. Conversely, Akt-mTORC1 inhibitors (RAD001 and LY294002) reduced NLGN3-induced HO1, NOQ1 and GCLC mRNA expression. Significantly, Nrf2 silencing by targeted shRNAs reversed NLGN3-induced retinal cytoprotection against H 2 O 2 . We conclude that NLGN3 activates Nrf2 signaling to protect human retinal cells from H 2 O 2 . NLGN3 could be further tested as a valuable retinal protection agent. Copyright © 2018 Elsevier Inc. All rights reserved.
Ren, Boxue; Li, Defang; Si, Lingling; Ding, Yangfang; Han, Jichun; Chen, Xiaoyu; Zheng, Qiusheng
2018-04-01
Emerging evidence showed that alteronol has a potential antitumour effect in several tumour cells. However, the antitumour effect of alteronol on breast cancer has not been reported. This study investigated the mechanisms of alteronol-induced cell proliferation inhibition in human breast cancer T47D cells. After treatment with alteronol, T47D cell proliferation was examined by MTT assay. The cell cycle distribution, cell apoptosis, reactive oxygen species level and mitochondrial membrane potential were evaluated via flow cytometry. Next, the protein levels of cyclin B1, cdc2, p21, p-cyclin B1, p-cdc2, p53, Bax, Bcl-2 and cytochrome c were analysed using Western blot analysis. Meanwhile, the mRNA levels of cyclin B1, cdc2, p21 and p53 were examined by qRT-PCR. Our data showed that alteronol inhibited the proliferation of T47D cells via inducing G2-phase arrest and cell apoptosis. Compared with control group, alteronol significantly increased ROS level and triggered mitochondrial dysfunction in alteronol-treated T47D cells. Further studies showed that the mRNA and protein levels of cdc2 and cyclin B1 were downregulated, while the mRNA and protein levels of p21, p53, p-cyclin B1, p-cdc2 and cytochrome c were upregulated. In addition, the expression level of Bax was increased, and the expression level of Bcl-2 was decreased. Alteronol induced T47D cell cycle arrest and cell apoptosis through increasing ROS production and triggering mitochondrial dysfunction, and subsequently inhibiting T47D cell proliferation. © 2018 Royal Pharmaceutical Society.
CHARACTERIZATION OF NORMAL HUMAN LUNG LYMPHOCYTES AND INTERLEUKIN-2-INDUCED LUNG T CELL LINES
Lymphocytes from the lower respiratory tract were obtained by bronchoalveolar lavage of healthy, non-smoking individuals. arious monoclonal antibodies characterizing activated T cells, helper-inducer and suppressor-inducer T cell subsets, and naive versus memory cells were used t...
DOE Office of Scientific and Technical Information (OSTI.GOV)
Takaoka, Yuki; Kawamoto, Seiji, E-mail: skawa@hiroshima-u.ac.jp; Katayama, Akiko
2013-02-08
Highlights: ► Anti-histone H1 autoantibody (anti-H1) acts on T cells to inhibit their activation. ► Anti-H1 suppresses T cell activation in Treg cell-dependent and -independent manners. ► Suboptimal dose of anti-H1 enhances suppressor function of Treg cells. ► High dose of anti-H1 directly inhibits T cell receptor signaling. -- Abstract: Induction of anti-nuclear antibodies against DNA or histones is a hallmark of autoimmune disorders, but their actual contribution to disease predisposition remains to be clarified. We have previously reported that autoantibodies against histone H1 work as a critical graft survival factor in a rat model of tolerogeneic liver transplantation. Heremore » we show that an immunosuppressive anti-histone H1 monoclonal antibody (anti-H1 mAb) acts directly on T cells to inhibit their activation in response to T cell receptor (TCR) ligation. Intriguingly, the T cell activation inhibitory activity of anti-H1 mAb under suboptimal dosages required regulatory T (Treg) cells, while high dose stimulation with anti-H1 mAb triggered a Treg cell-independent, direct negative regulation of T cell activation upon TCR cross-linking. In the Treg cell-dependent mode of immunosuppressive action, anti-H1 mAb did not induce the expansion of CD4{sup +}Foxp3{sup +} Treg cells, but rather potentiated their regulatory capacity. These results reveal a previously unappreciated T cell regulatory role of anti-H1 autoantibody, whose overproduction is generally thought to be pathogenic in the autoimmune settings.« less
Role of interleukin 1 in antigen-specific T cell proliferation.
Chu, E; Rosenwasser, L J; Dinarello, C A; Lareau, M; Geha, R S
1984-03-01
The role of interleukin 1 (IL 1) in human antigen-specific T cell proliferation was examined. Nylon wool-purified T cells proliferated in the presence of autologous monocytes (Mo.) pulsed for 18 h with tetanus toxoid (TT) antigen (Mo.TT). Irradiation of Mo.TT with ultraviolet (UV) light (72 J/m2) abolished their capacity to support T cell proliferation and drastically reduced their capacity to secrete IL 1 after stimulation with Staphylococcus albus. The defect in antigen presentation induced by UV irradiation of Mo.TT was reversed in a dose-dependent manner by the addition of two different preparations containing human interleukin 1 (IL 1). The first preparation consisted of supernatants of Mo. stimulated with Con A for 18 hr and in which Con A activity was blocked by alpha-D-methyl-mannoside (Mo.-Con A-Sup). The second preparation consisted of human IL 1 partially purified from supernatants of human peripheral blood mononuclear cells stimulated with S. albus. This IL 1 copurified with human leukocyte pyrogen (LP) and was termed IL 1/LP. Both IL 1-containing preparations enhanced the response of C57BL/6 mouse thymocytes to phytohemagglutinin. A rabbit antibody to human IL 1/LP inhibited the capacity of T cells to proliferate in response to Mo.TT and inhibited the capacity of Mo.-Con A-Sup to reconstitute the T cell response to UV-irradiated Mo.TT. IL 1/LP was not necessary for T cells to recognize the immunogenic moiety presented by Mo., because monolayers of UV-irradiated Mo.TT were equivalent to monolayers of unirradiated MO.TT in their capacity to adsorb TT-reactive T cells specifically. Furthermore, the addition of rabbit antibody to IL 1/LP did not interfere with the capacity of UV-irradiated Mo.TT to adsorb TT-reactive T cells. The results obtained in this study indicate that IL 1 is involved in optimal antigen-driven proliferation of human T lymphocytes.
Kang, Min-Jung; Song, Eun-Jung; Kim, Bo-Yeon; Kim, Dong-Jae; Park, Jong-Hwan
2014-12-01
Although Helicobacter pylori have been known to induce vascular endothelial growth factor (VEGF) production in gastric epithelial cells, the precise mechanism for cellular signaling is incompletely understood. In this study, we investigated the role of bacterial virulence factor and host cellular signaling in VEGF production of H. pylori-infected gastric epithelial cells. We evaluated production of VEGF, activation of nuclear factor nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinases (MAPKs) and hypoxia-inducible factor-1α (HIF-1α) stabilization in gastric epithelial cells infected with H. pylori WT or isogenic mutants deficient in type IV secretion system (T4SS). H. pylori induced VEGF production in gastric epithelial cells via both T4SS-dependent and T4SS-independent pathways, although T4SS-independent pathway seems to be the dominant signaling. The inhibitor assay implicated that activation of NF-κB and MAPKs is dispensable for H. pylori-induced VEGF production in gastric epithelial cells. H. pylori led to HIF-1α stabilization in gastric epithelial cells independently of T4SS, NF-κB, and MAPKs, which was essential for VEGF production in these cells. N-acetyl-cysteine (NAC), a reactive oxygen species (ROS) inhibitor, treatment impaired H. pylori-induced HIF-1α stabilization and VEGF production in gastric epithelial cells. We defined the important role of ROS-HIF-1α axis in VEGF production of H. pylori-infected gastric epithelial cells, and bacterial T4SS has a minor role in H. pylori-induced VEGF production of gastric epithelial cells. © 2014 John Wiley & Sons Ltd.
Gualde, N; Goodwin, J S
1984-04-01
Unfractionated human T cells exposed to 10-50 rad of X irradiation incorporated less [3H]thymidine than nonirradiated T cells when subsequently cultured with PHA or Con A. The cytotoxic/suppressor T-cell subset, isolated as either OKT8(+) or OKT4(-) cells, demonstrated significantly enhanced [3H]thymidine incorporation in PHA- or Con A-stimulated cultures after exposure to 10-50 rad, compared to unirradiated cells, while the proliferation of the OKT4(+) helper/inducer subset was inhibited by low dose irradiation. It has been previously reported that approximately 30% of the cytotoxic/suppressor subset also stains with OKM1. When the cytotoxic/suppressor subset was further subdivided into OKT4(-), OKM1(+), and OKT4(-), OKM1(-) cells, proliferation of the OKT4(-), OKM1(+) population was inhibited by exposure to 25 rad while proliferation of the OKT4(-), OKM1(-) population was stimulated. The increase in proliferation of the cytotoxic/suppressor T-cell subset after low dose irradiation is paralleled by an increase in suppressor activity of these cells. T cells exposed to 25 rad and then cultured with Con A for 48 hr caused greater inhibition of IgG production when added to fresh autologous lymphocytes stimulated by pokeweed mitogen than did unirradiated cells. Thus, low dose irradiation enhances both the proliferation and function of the human suppressor T-cell subset.
NoxO1 Controls Proliferation of Colon Epithelial Cells.
Moll, Franziska; Walter, Maria; Rezende, Flávia; Helfinger, Valeska; Vasconez, Estefania; De Oliveira, Tiago; Greten, Florian R; Olesch, Catherine; Weigert, Andreas; Radeke, Heinfried H; Schröder, Katrin
2018-01-01
Reactive oxygen species (ROS) produced by enzymes of the NADPH oxidase family serve as second messengers for cellular signaling. Processes such as differentiation and proliferation are regulated by NADPH oxidases. In the intestine, due to the exceedingly fast and constant renewal of the epithelium both processes have to be highly controlled and balanced. Nox1 is the major NADPH oxidase expressed in the gut, and its function is regulated by cytosolic subunits such as NoxO1. We hypothesize that the NoxO1-controlled activity of Nox1 contributes to a proper epithelial homeostasis and renewal in the gut. NoxO1 is highly expressed in the colon. Knockout of NoxO1 reduces the production of superoxide in colon crypts and is not subsidized by an elevated expression of its homolog p47phox. Knockout of NoxO1 increases the proliferative capacity and prevents apoptosis of colon epithelial cells. In mouse models of dextran sulfate sodium (DSS)-induced colitis and azoxymethane/DSS induced colon cancer, NoxO1 has a protective role and may influence the population of natural killer cells. NoxO1 affects colon epithelium homeostasis and prevents inflammation.
Abacavir, an anti-HIV-1 drug, targets TDP1-deficient adult T cell leukemia.
Tada, Kohei; Kobayashi, Masayuki; Takiuchi, Yoko; Iwai, Fumie; Sakamoto, Takashi; Nagata, Kayoko; Shinohara, Masanobu; Io, Katsuhiro; Shirakawa, Kotaro; Hishizawa, Masakatsu; Shindo, Keisuke; Kadowaki, Norimitsu; Hirota, Kouji; Yamamoto, Junpei; Iwai, Shigenori; Sasanuma, Hiroyuki; Takeda, Shunichi; Takaori-Kondo, Akifumi
2015-04-01
Adult T cell leukemia (ATL) is an aggressive T cell malignancy caused by human T cell leukemia virus type 1 (HTLV-1) and has a poor prognosis. We analyzed the cytotoxic effects of various nucleoside analog reverse transcriptase inhibitors (NRTIs) for HIV-1 on ATL cells and found that abacavir potently and selectively kills ATL cells. Although NRTIs have minimal genotoxicities on host cells, the therapeutic concentration of abacavir induced numerous DNA double-strand breaks (DSBs) in the chromosomal DNA of ATL cells. DSBs persisted over time in ATL cells but not in other cell lines, suggesting impaired DNA repair. We found that the reduced expression of tyrosyl-DNA phosphodiesterase 1 (TDP1), a repair enzyme, is attributable to the cytotoxic effect of abacavir on ATL cells. We also showed that TDP1 removes abacavir from DNA ends in vitro. These results suggest a model in which ATL cells with reduced TDP1 expression are unable to excise abacavir incorporated into genomic DNA, leading to irreparable DSBs. On the basis of the above mechanism, we propose abacavir as a promising chemotherapeutic agent for ATL.
Böhnlein, E; Siekevitz, M; Ballard, D W; Lowenthal, J W; Rimsky, L; Bogérd, H; Hoffman, J; Wano, Y; Franza, B R; Greene, W C
1989-04-01
The human immunodeficiency virus type 1 (HIV-1) preferentially infects CD4+ T lymphocytes and may exist as a latent provirus within these cells for extended periods. The transition to a productive retroviral infection results in T-cell death and clinically may lead to the acquired immune deficiency syndrome. Accelerated production of infectious HIV-1 virions appears to be closely linked to a heightened state of T-cell activation. The transactivator (Tax) protein of the type I human T-cell leukemia virus (HTLV-I) can produce such an activated T-cell phenotype and augments activity of the HIV-1 long terminal repeat. One Tax-responsive region within the HIV-1 long terminal repeat has been mapped to a locus composed of two 10-base-pair direct repeats sharing homology with the binding site for the eucaryotic transcription factor NF-kappaB (GGGACTTTCC). Tax-expressing Jurkat T cells contain one or more inducible cellular proteins that specifically associate with the HIV-1 enhancer at these binding sites. Microscale DNA affinity precipitation assays identified a Tax-inducible 86-kilodalton protein, HIVEN86A, as one of these HIV-1 enhancer-binding factors. The interaction of HIVEN86A, and presumably other cellular proteins, with the HIV-1 enhancer appears functionally important as oligonucleotides corresponding to this enhancer were sufficient to impart Tax inducibility to an unresponsive heterologous promoter. These findings suggest that the Tax-inducible cellular protein HIVEN86A plays an important role in the transcriptional activation of the HIV-1 enhancer. These specific protein-DNA interactions may also be important for the transition of HIV-1 from a latent to a productive mode of infection. Furthermore, these findings highlight an intriguing biological interplay between HTLV-1 and HIV-1 through a cellular transcriptional pathway that is normally involved in T-cell activation and growth.
PD-1 expression on dendritic cells suppresses CD8+ T cell function and antitumor immunity.
Lim, Tong Seng; Chew, Valerie; Sieow, Je Lin; Goh, Siting; Yeong, Joe Poh-Sheng; Soon, Ai Ling; Ricciardi-Castagnoli, Paola
2016-03-01
Programmed death one (PD-1) is a well-established co-inhibitory regulator that suppresses proliferation and cytokine production of T cells. Despite remarkable progress in delineating the functional roles of PD-1 on T lymphocytes, little is known about the regulatory role of PD-1 expressed on myeloid cells such as dendritic cells (DCs). Here, we show that CD8 + T cells can be more potently activated to secrete IL-2 and IFNγ by PD-1-deficient DCs compared to wild-type DCs. Adoptive transfer of PD-1-deficient DCs demonstrated their superior capabilities in inducing antigen-specific CD8 + T cell proliferation in vivo . In addition, we provide first evidence demonstrating the existence of peripheral blood DCs and CD11c + tumor-infiltrating myeloid cells that co-express PD-1 in patients with hepatocellular carcinoma (HCC). The existence of PD-1-expressing HCC-infiltrating DCs (HIDCs) was further supported in a mouse model of HCC. Intratumoral transfer of PD-1-deficient DCs rendered recipient mice resistant to the growth of HCC by promoting tumor-infiltrating CD8 + effector T cells to secrete perforin and granzyme B. This novel finding provides a deeper understanding of the role of PD-1 in immune regulation and has significant implications for cancer immunotherapies targeting PD-1.
T Cell CX3CR1 Mediates Excess Atherosclerotic Inflammation in Renal Impairment
Dong, Lei; Nordlohne, Johannes; Ge, Shuwang; Hertel, Barbara; Melk, Anette; Rong, Song; Haller, Hermann
2016-01-01
Reduced kidney function increases the risk for atherosclerosis and cardiovascular death. Leukocytes in the arterial wall contribute to atherosclerotic plaque formation. We investigated the role of fractalkine receptor CX3CR1 in atherosclerotic inflammation in renal impairment. Apoe−/− (apolipoprotein E) CX3CR1−/− mice with renal impairment were protected from increased aortic atherosclerotic lesion size and macrophage accumulation. Deficiency of CX3CR1 in bone marrow, only, attenuated atherosclerosis in renal impairment in an independent atherosclerosis model of LDL receptor–deficient (LDLr−/−) mice as well. Analysis of inflammatory leukocytes in atherosclerotic mixed bone-marrow chimeric mice (50% wild-type/50% CX3CR1−/− bone marrow into LDLr−/− mice) showed that CX3CR1 cell intrinsically promoted aortic T cell accumulation much more than CD11b+CD11c+ myeloid cell accumulation and increased IL-17-producing T cell counts. In vitro, fewer TH17 cells were obtained from CX3CR1−/− splenocytes than from wild-type splenocytes after polarization with IL-6, IL-23, and TGFβ. Polarization of TH17 or TREG cells, or stimulation of splenocytes with TGFβ alone, increased T cell CX3CR1 reporter gene expression. Furthermore, TGFβ induced CX3CR1 mRNA expression in wild-type cells in a dose- and time-dependent manner. In atherosclerotic LDLr−/− mice, CX3CR1+/− T cells upregulated CX3CR1 and IL-17A production in renal impairment, whereas CX3CR1−/− T cells did not. Transfer of CX3CR1+/− but not Il17a−/− T cells into LDLr−/−CX3CR1−/− mice increased aortic lesion size and aortic CD11b+CD11c+ myeloid cell accumulation in renal impairment. In summary, T cell CX3CR1 expression can be induced by TGFβ and is instrumental in enhanced atherosclerosis in renal impairment. PMID:26449606
Tano, Tomoyuki; Okamoto, Masato; Kan, Shin; Bando, Takashi; Goda, Hiroyuki; Nakashiro, Koh-ichi; Shimodaira, Shigetaka; Koido, Shigeo; Homma, Sadamu; Fujita, Tomonobu; Sato, Mitsunobu; Yamashita, Naomi; Hamakawa, Hiroyuki; Kawakami, Yutaka
2013-07-01
Eighty-one patients with oral squamous cell carcinoma (OSCC) received oral fluoropyrimidine UFT and radiotherapy (RT) with or without an immunotherapeutic agent OK-432. Both overall survival and progression-free survival of patients who received RT + UFT + OK-432 were significantly longer than those of patients who received RT + UFT (P = .0075 and P = .0175, respectively). Clinical response was also more favorable in RT + UFT + OK-432 group than in RT + UFT group (P = .0066). Next, in vitro experiments were conducted to examine the effect of 5-fluorouracil (5-FU) and X-ray irradiation in OK-432-induced immunity. Human peripheral blood mononuclear cells stimulated with OK-432 produced helper T cell 1 (Th1)-type cytokines as well as interleukin-10 (IL-10) and transforming growth factor-β (TGF-β), which are produced by Th2 and regulatory T cells (Tregs), respectively, and are inhibitory in antitumor immunity. OK-432-induced IL-10 and TGF-β but not Th1 cytokines were significantly inhibited by 5-FU and/or X-ray. 5-FU and X-ray also inhibited the expression of mRNAs for GATA-3 and Foxp3, which are transcription factors for Th2 and Tregs, respectively, but not for T-bet, a transcription factor for Th1. In addition, 5-FU and X-ray decreased the expression of mRNAs for suppressor of cytokine signaling 1 (SOCS1) and SOCS3. Antisense oligonucleotides for SOCS1 and SOCS3 markedly reduced OK-432-induced IL-10 and TGF-β. This is the first report clearly demonstrating that OK-432-based immunotherapy significantly enhanced the therapeutic effects of chemoradiotherapy in patients with OSCC as well as elucidating the mechanism of the synergistic effect of immunochemoradiotherapy in which 5-FU and radiation enhanced OK-432-induced Th1 response mediated by the inhibition of SOCS1 and SOCS3 gene expression.
Shrestha, Anup; Nepal, Saroj; Kim, Mi Jin; Chang, Jae Hoon; Kim, Sang-Hyun; Jeong, Gil-Saeng; Jeong, Chul-Ho; Park, Gyu Hwan; Jung, Sunghee; Lim, Jaecheong; Cho, Eunha; Lee, Soyoung; Park, Pil-Hoon
2016-02-01
Adiponectin predominantly secreted from adipose tissue has exhibited potent anti-proliferative properties in cancer cells via modulating cell cycle and apoptosis. FoxO3A, a Forkhead box O member of the transcription factor, plays a critical role in modulating expression of genes involved in cell death and/or survival. In this study, we investigated the role of FoxO3A signaling in anti-cancer activities of adiponectin. Herein, we have shown that treatment with globular adiponectin (gAcrp) increases p27 but decreases cyclinD1 expression in human hepatoma (HepG2) and breast (MCF-7) cancer cells. Gene ablation of FoxO3A prevented gAcrp-induced increase in p27 and decreased in cyclin D1 expression, and further ameliorated cell cycle arrest by gAcrp, indicating a critical role of FoxO3A in gAcrp-induced cell cycle arrest of cancer cells. Moreover, treatment with gAcrp also induced caspase-3/7 activation and increased Fas ligand (FasL) expression in both HepG2 and MCF-7 cells. Transfection with FoxO3A siRNA inhibited gAcrp-induced caspase-3/7 activation and FasL expression, suggesting that FoxO3A signaling also plays an important role in gAcrp-induced apoptosis of cancer cells. We also found that gene silencing of AMPK prevented gAcrp-induced nuclear translocation of FoxO3A in HepG2 and MCF-7 cells. In addition, suppression of AMPK also blocked gAcrp-induced cell cycle arrest and further attenuated gAcrp-induced caspase-3/7 activation, indicating that AMPK signaling plays a pivotal role in both gAcrp-induced cell cycle arrest and apoptosis via acting as an upstream signaling of FoxO3A. Taken together, our findings demonstrated that AMPK/FoxO3A axis plays a cardinal role in anti-proliferative effect of adiponectin in cancer cells. © 2015 Wiley Periodicals, Inc.
Wang, Yan; Nag, Mukta; Tuohy, Joanne L; De Paris, Kristina; Fogle, Jonathan E
2018-03-01
Polyfunctional CD8 + T cells play a critical role in controlling viremia during AIDS lentiviral infections. However, for most HIV-infected individuals, virus-specific CD8 + T cells exhibit loss of polyfunctionality, including loss of IL2, TNFα, and IFNγ. Using the feline immunodeficiency virus (FIV) model for AIDS lentiviral persistence, our laboratory has demonstrated that FIV-activated Treg cells target CD8 + T cells, leading to a reduction in IL2 and IFNγ production. Furthermore, we have demonstrated that Treg cells induce expression of the repressive transcription factor, Foxp3, in CD8 + T cells. Based upon these findings, we asked if Treg-induced Foxp3 could bind to the IL2, TNFα, and IFNγ promoter regions in virus-specific CD8 + T cells. Following coculture with autologous Treg cells, we demonstrated decreased mRNA levels of IL2 and IFNγ at weeks 4 and 8 postinfection and decreased TNFα at week 4 postinfection in virus-specific CD8 + T cells. We also clearly demonstrated Treg cell-induced Foxp3 expression in virus-specific CD8 + T cells at weeks 1, 4, and 8 postinfection. Finally, we documented Foxp3 binding to the IL2, TNFα, and IFNγ promoters at 8 weeks and 6 months postinfection in virus-specific CD8 + T cells following Treg cell coculture. In summary, the results here clearly demonstrate that Foxp3 inhibits IL2, TNFα, and IFNγ transcription by binding to their promoter regions in lentivirus-specific CD8 + T cells. We believe this is the first description of this process during the course of AIDS lentiviral infection.
T-cell differentiation of multipotent hematopoietic cell line EML in the OP9-DL1 coculture system.
Kutlesa, Snjezana; Zayas, Jennifer; Valle, Alexandra; Levy, Robert B; Jurecic, Roland
2009-08-01
Multipotent hematopoietic cell line EML can differentiate into myeloid, erythroid, megakaryocytic, and B-lymphoid lineages, but it remained unknown whether EML cells have T-cell developmental potential as well. The goal of this study was to determine whether the coculture with OP9 stromal cells expressing Notch ligand Delta-like 1 (OP9-DL1) could induce differentiation of EML cells into T-cell lineage. EML cells were cocultured with control OP9 or OP9-DL1 stromal cells in the presence of cytokines (stem cell factor, interleukin-7, and Fms-like tyrosine kinase 3 ligand). Their T-cell lineage differentiation was assessed through flow cytometry and reverse transcription polymerase chain reaction expression analysis of cell surface markers and genes characterizing and associated with specific stages of T-cell development. The phenotypic, molecular, and functional analysis has revealed that in EML/OP9-DL1 cocultures with cytokines, but not in control EML/OP9 cocultures, EML cell line undergoes T-cell lineage commitment and differentiation. In OP9-DL1 cocultures, EML cell line has differentiated into cells that 1) resembled double-negative, double-positive, and single-positive stages of T-cell development; 2) initiated expression of GATA-3, Pre-Talpha, RAG-1, and T-cell receptor-Vbeta genes; and 3) produced interferon-gamma in response to T-cell receptor stimulation. These results support the notion that EML cell line has the capacity for T-cell differentiation. Remarkably, induction of T-lineage gene expression and differentiation of EML cells into distinct stages of T-cell development were very similar to previously described T-cell differentiation of adult hematopoietic stem cells and progenitors in OP9-DL1 cocultures. Thus, EML/OP9-DL1 coculture could be a useful experimental system to study the role of particular genes in T-cell lineage specification, commitment, and differentiation.
Zhang, Zheng; Xu, Xiangsheng; Lu, Jiyun; Zhang, Shuye; Gu, Lanlan; Fu, Junliang; Jin, Lei; Li, Haiying; Zhao, Min; Zhang, Jiyuan; Wu, Hao; Su, Lishan; Fu, Yang-Xin
2011-01-01
Background. Nonspecific T-cell hyperactivation is the main driving force for human immunodeficiency virus (HIV)–1 disease progression, but the reasons why the excess immune response is not properly shut off are poorly defined. Methods. Eighty-five HIV-1–infected individuals were enrolled to characterize B and T lymphocyte attenuator (BTLA) expression and function. Infection and blockade assays were used to dissect the factors that influenced BTLA signaling in vitro. Results. BTLA expression on overall CD4+ and CD8+ T cells was progressively decreased in HIV-1 infection, which was directly correlated with disease progression and CD4+ T-cell differentiation and activation. BTLA+CD4+ T cells from HIV-1–infected patients also displayed an altered immune status, which was indicated by reduced expression of naive markers but increased activation and exhaustion markers. Cross-linking of BTLA can substantially decrease CD4+ T-cell activation in vitro. This responsiveness of CD4+ T cells to BTLA-mediated inhibitory signaling was further found to be impaired in HIV-1–infected patients. Furthermore, HIV-1 NL4-3 down-regulated BTLA expression on CD4+ T cells dependent on plasmacytoid dendritic cell (pDC)-derived interferon (IFN)-α. Blockade of IFN-α or depletion of pDCs prevents HIV-1-induced BTLA down-regulation. Conclusions. HIV-1 infection potentially impairs BTLA-mediated signaling dependent on pDC-derived IFN-α, which may contribute to broad T-cell hyperactivation induced by chronic HIV-1 infection. PMID:21592997
Dimitrova, Petya; Alipieva, Kalina; Grozdanova, Tsvetinka; Simova, Svetlana; Bankova, Vassya; Georgiev, Milen I; Popova, Milena P
2018-01-01
The Verbascum species are widely used traditional herb remedies against respiratory, inflammatory conditions and disorders. In the present study methanol extract of the aerial parts of the endemic Verbascum nobile Velen, was investigated and two novel iridoid glycosides 1 and 2, together with nine known constituents: iridoids, phenylethanoids, and saponins characteristic of Verbascum genus were identified. Further, the biological activity of the extract and selected isolated compounds on concanavalin (Con A)-induced T cell proliferation and activation of human Jurkat T cell line and splenic murine CD3 T cells was evaluated. T cell growth was studied by colorimetric-based WST proliferation assay while DNA content, cell cycling, dynamic of cell proliferation, expression of activation markers, intracellular expression of cytokine IFN-γ, and phosphorylation of ERK were analyzed by flow cytometry. Caspase-mediated apoptosis resulting in a poly (ADP-ribose) polymerase (PARP) cleavage was assessed by colorimetric in-cell kit. It was found that the extract, and all tested compounds (1, 2, 3 and 9) inhibited lectin-induced cell growth of Jurkat T cell line. The novel compounds decreased the frequencies of cells in S phase without causing a significant cell cycle arrest at G1 phase, caspases-mediated apoptosis and/or a profound change in the dynamic of splenic murine CD3 + T cell proliferation. Both compounds showed stronger inhibitory effect on Con A-induced ERK phosphorylation than the known bioactive compounds 3 and 9, and suppressed the expression of early activation marker CD69, the intracellular level of IFN-γ, and the generation of CD3 + IFN-γ + effectors. Our data suggest that the novel iridoid glycosides might have a potential to modulate T cell-related pathologies. Copyright © 2017 Elsevier Ltd. All rights reserved.
Edwards, Justin P.; Fujii, Hodaka; Zhou, Angela X.; Creemers, John; Unutmaz, Derya; Shevach, Ethan M.
2013-01-01
GARP/LRRC32 has previously been defined as a marker of activated human regulatory T-cells (Tregs) that is responsible for surface localization of latent TGF-β1. We find that GARP and latent TGF-β1 are also found on mouse Tregs activated via TCR stimulation, but in contrast to human Tregs, GARP is also expressed at a low level on resting Tregs. The expression of GARP can be upregulated on mouse Tregs by IL-2 or IL-4 exposure in the absence of TCR signaling. GARP is expressed at a low level on Tregs within the thymus and Treg precursors from the thymus concomitantly express GARP and Foxp3 upon exposure to IL-2. The expression of GARP is independent of TGF-β1 and TGF-β1 loading into GARP and is independent of furin-mediated processing of pro-TGF-β1 to latent TGF-β1. Specific deletion of GARP in CD4+ T cells results in lack of expression of latent-TGF-β1 on activated Tregs. GARP-deficient Tregs develop normally, are present in normal numbers in peripheral tissues, and are fully competent suppressors of the activation of T conventional cells in vitro. Activated Tregs expressing GARP/latent-TGF-β1 complexes are potent inducers of Th17 differentiation in the presence of exogenous IL-6 and inducers of Treg in the presence of IL-2. Induction of both Th17 producing cells and Treg is preferentially induced by Tregs expressing the latent-TGF-β1/GARP complex on their cell surface rather than by secreted latent-TGF-β1. PMID:23645881
Suzuki, Tetsuya; Kuramoto, Yoshie; Kamiya, Hiroyuki
2018-05-21
O 6 -Methylguanine ( O 6 -MeG) is a damaged base produced by methylating reagents. The Werner syndrome protein (WRN) is a cancer-related human DNA helicase. The effects of WRN reduction on O 6 -MeG-caused mutagenesis were assessed by an siRNA-mediated knockdown in human U2OS cells, using a shuttle plasmid with a single O 6 -MeG base in the supF gene. The plasmid DNA was replicated in the cells, isolated, and electroporated into an Escherichia coli indicator strain. The lowered amount of WRN increased the frequency of mutations induced by O 6 -MeG, mainly G:C → A:T substitution. The increased mutation rate suggested that the cancer-related WRN suppresses the G:C → A:T substitution by O 6 -MeG in human cells.
IL-7–dependent STAT1 activation limits homeostatic CD4+ T cell expansion
Le Saout, Cecile; Luckey, Megan A.; Villarino, Alejandro V.; Smith, Mindy; Hasley, Rebecca B.; Myers, Timothy G.; Imamichi, Hiromi; Park, Jung-Hyun; O’Shea, John J.; Lane, H. Clifford
2017-01-01
IL-7 regulates homeostatic mechanisms that maintain the overall size of the T cell pool throughout life. We show that, under steady-state conditions, IL-7 signaling is principally mediated by activation of signal transducers and activators of transcription 5 (STAT5). In contrast, under lymphopenic conditions, there is a modulation of STAT1 expression resulting in an IL-7–dependent STAT1 and STAT5 activation. Consequently, the IL-7–induced transcriptome is altered with enrichment of IFN-stimulated genes (ISGs). Moreover, STAT1 overexpression was associated with reduced survival in CD4+ T cells undergoing lymphopenia-induced proliferation (LIP). We propose a model in which T cells undergoing LIP upregulate STAT1 protein, “switching on” an alternate IL-7–dependent program. This mechanism could be a physiological process to regulate the expansion and size of the CD4+ T cell pool. During HIV infection, the virus could exploit this pathway, leading to the homeostatic dysregulation of the T cell pools observed in these patients. PMID:29202461
DOE Office of Scientific and Technical Information (OSTI.GOV)
Hara, T.; Jung, L.K.L.; FU, S.M.
1986-03-01
With human T cells activated for 12 hours by 12-o-tetradecanoyl phorbol-13-acetate (TPA) as immunogen, an IgG/sub 2a/ monoclonal antibody, mAb Ea 1, has been generated to a 60KD phosphorylated protein with 32KD and 28KD subunits. The antigen, Ea 1, is readily detected on 60% of isolated thymocytes by indirect immunofluorescence. A low level of Ea 1 expression is detectable on 2-6% of blood lymphocytes. Isolated T cells have been induced to express Ea 1 by TPA, mitogens and anitgens. TPA activated T cells express Ea 1 as early as 1 hour after activation. By 4 hours, greater than 95% ofmore » the T cells stain with mAb Ea 1. About 50% of the PHA or Con A activated T cells express Ea 1 with a similar kinetics. Ea 1 expression proceeds that of IL-2 receptor in these activation processes. T cells activated by soluble antigens (tetanus toxoid and PPD) and alloantigens in MLR also express Ea 1 after a long incubation. About 20% of the T cells stain for Ea 1 at day 6. Ea 1 expression is not limited to activated T cells. B cells activated by TPA or anti-IgM Ab plus B cell growth factor express Ea 1. The kinetics of Ea 1 expression is slower and the staining is less intense. Repeated attempts to detect Ea 1 on resting and activated monocytes and granulocytes have not been successful. Ea 1 expression is due to de novo synthesis for its induction is blocked by cycloheximide and actinomycin D. Ea 1 is the earliest activation antigen detectable to-date.« less
Intravenous apoptotic spleen cell infusion induces a TGF-beta-dependent regulatory T-cell expansion
Kleinclauss, François M.; Perruche, Sylvain; Masson, Emeline; De Carvalho Bittencourt, Marcelo; Biichle, Sabeha; Remy-Martin, Jean-Paul; Ferrand, Christophe; Martin, Mael; Bittard, Hugues; Chalopin, Jean-Marc; Seilles, Estelle; Tiberghien, Pierre; Saas, Philippe
2006-01-01
Apoptotic leukocytes are endowed with immunomodulatory properties that can be used to enhance hematopoietic engraftment and prevent graft-versus-host disease. This apoptotic cell-induced tolerogenic effect is mediated by host macrophages and not recipient dendritic cells or donor phagocytes present in the bone marrow graft as evidenced by selective cell depletion and trafficking experiments. Furthermore, apoptotic cell infusion is associated with TGF-β-dependent donor CD4+CD25+ T cell expansion. Such cells have a regulatory phenotype (CD62Lhigh and intracellular CTLA-4+), express high levels of Foxp3 mRNA and exert ex vivo suppressive activity through a cell-to-cell contact mechanism. In vivo CD25 depletion after apoptotic cell infusion prevents the apoptotic spleen cell-induced beneficial effects on engraftment and graft-versus-host disease occurrence. This highlights the role of regulatory T cells in the tolerogenic effect of apoptotic spleen cell infusion. This novel association between apoptosis and regulatory T cell expansion may also contribute to preventing deleterious auto-immune responses during normal turnover. PMID:15962005
PD-1 and PD-L1 Up-regulation Promotes T-cell Apoptosis in Gastric Adenocarcinoma.
Chiu, Ying-Ming; Tsai, Chung-Lin; Kao, Jung-Ta; Hsieh, Chin-Tung; Shieh, Dong-Chen; Lee, Yi-Ju; Tsay, Gregory J; Cheng, Ken-Sheng; Wu, Yi-Ying
2018-04-01
The programmed death 1 (PD-1) receptor and its ligand (PD-L1) play pivotal roles in regulating host immune responses. However, the inhibitory effects of this pathway on the function of tumor infiltrating T lymphocytes in gastric adenocarcinoma patients are not well-defined. We characterized the expression of PD-1 and PD-L1 in peripheral blood and tumor infiltrating cells and analyzed the association between PD-1/PD-L1 expression and disease progression in a cohort of 60 patients with Helicobacter pylori infection, including 18 with gastric adenocarcinoma, 23 with gastritis, and 19 asymptomatic controls. Relative to controls, the expression of PD-1 on peripheral blood and tumor infiltrating T cells increased with disease progression. In vitro, T cells induced PD-L1 expression on primary gastric adenocarcinoma epithelial cells in an IFN-γ-dependent manner, which in turn promoted T cells apoptosis. Blocking of PD-L1 reversed this effect. This study provides evidence for a new therapeutic target in gastric adenocarcinoma patients. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.
Coombs, Melanie R Power; Harrison, Megan E; Hoskin, David W
2016-10-01
Programmed death ligand 1 (PD-L1) is expressed by many cancer cell types, as well as by activated T cells and antigen-presenting cells. Constitutive and inducible PD-L1 expression contributes to immune evasion by breast cancer (BC) cells. We show here that the dietary phytochemical apigenin inhibited interferon (IFN)-γ-induced PD-L1 upregulation by triple-negative MDA-MB-468 BC cells, HER2(+) SK-BR-3 BC cells, and 4T1 mouse mammary carcinoma cells, as well as human mammary epithelial cells, but did not affect constitutive PD-L1 expression by triple-negative MDA-MB-231 BC cells. IFN-β-induced expression of PD-L1 by MDA-MB-468 cells was also inhibited by apigenin. In addition, luteolin, the major metabolite of apigenin, inhibited IFN-γ-induced PD-L1 expression by MDA-MB-468 cells. Apigenin-mediated inhibition of IFN-γ-induced PD-L1 expression by MDA-MB-468 and 4T1 cells was associated with reduced phosphorylation of STAT1, which was early and transient at Tyr701 and sustained at Ser727. Apigenin-mediated inhibition of IFN-γ-induced PD-L1 expression by MDA-MB-468 cells also increased proliferation and interleukin-2 synthesis by PD-1-expressing Jurkat T cells that were co-cultured with MDA-MB-468 cells. Apigenin therefore has the potential to increase the vulnerability of BC cells to T cell-mediated anti-tumor immune responses. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.
Sphingosine kinase-1 mediates androgen-induced osteoblast cell growth
DOE Office of Scientific and Technical Information (OSTI.GOV)
Martin, Claire; Universite de Toulouse, UPS, IPBS, Toulouse F-31000; Lafosse, Jean-Michel
Herein we report that the lipid kinase sphingosine kinase-1 (SphK1) is instrumental in mediating androgen-induced cell proliferation in osteoblasts. Dihydrotestosterone (DHT) triggered cell growth in steroid-deprived MC3T3 cells, which was associated with a rapid stimulation of SphK1 and activation of both Akt and ERK signaling pathways. This mechanism relied on functional androgen receptor/PI3K/Akt nongenotropic signaling as pharmacological antagonists could block SphK1 stimulation by DHT and its consequences. Finally, SphK1 inhibition not only abrogated DHT-induced ERK activation but also blocked cell proliferation, while ERK inhibition had no impact, suggesting that SphK1 was critical for DHT signaling yet independently of the ERK.
Lynch, Adam; Hawk, William; Nylen, Emily; Ober, Sean; Autin, Pierre; Barber, Amorette
2017-11-01
Adoptive transfer of T cells is a promising cancer therapy and expression of chimeric antigen receptors can enhance tumour recognition and T-cell effector functions. The programmed death protein 1 (PD1) receptor is a prospective target for a chimeric antigen receptor because PD1 ligands are expressed on many cancer types, including lymphoma. Therefore, we developed a murine chimeric PD1 receptor (chPD1) consisting of the PD1 extracellular domain fused to the cytoplasmic domain of CD3ζ. Additionally, chimeric antigen receptor therapies use various co-stimulatory domains to enhance efficacy. Hence, the inclusion of a Dap10 or CD28 co-stimulatory domain in the chPD1 receptor was compared to determine which domain induced optimal anti-tumour immunity in a mouse model of lymphoma. The chPD1 T cells secreted pro-inflammatory cytokines and lysed RMA lymphoma cells. Adoptive transfer of chPD1 T cells significantly reduced established tumours and led to tumour-free survival in lymphoma-bearing mice. When comparing chPD1 receptors containing a Dap10 or CD28 domain, both receptors induced secretion of pro-inflammatory cytokines; however, chPD1-CD28 T cells also secreted anti-inflammatory cytokines whereas chPD1-Dap10 T cells did not. Additionally, chPD1-Dap10 induced a central memory T-cell phenotype compared with chPD1-CD28, which induced an effector memory phenotype. The chPD1-Dap10 T cells also had enhanced in vivo persistence and anti-tumour efficacy compared with chPD1-CD28 T cells. Therefore, adoptive transfer of chPD1 T cells could be a novel therapy for lymphoma and inclusion of the Dap10 co-stimulatory domain in chimeric antigen receptors may induce a preferential cytokine profile and T-cell differentiation phenotype for anti-tumour therapies. © 2017 John Wiley & Sons Ltd.
NiO nanoparticles induce apoptosis through repressing SIRT1 in human bronchial epithelial cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Duan, Wei-Xia; He, Min-Di; Mao, Lin
2015-07-15
With application of nano-sized nickel-containing particles (Nano-Ni) expanding, the health concerns about their adverse effects on the pulmonary system are increasing. However, the mechanisms for the pulmonary toxicity of these materials remain unclear. In the present study, we focused on the impacts of NiO nanoparticles (NiONPs) on sirtuin1 (SIRT1), a NAD-dependent deacetylase, and investigated whether SIRT1 was involved in NiONPs-induced apoptosis. Although the NiONPs tended to agglomerate in fluid medium, they still entered into the human bronchial epithelial cells (BEAS-2B) and released Ni{sup 2+} inside the cells. NiONPs at doses of 5, 10, and 20 μg/cm{sup 2} inhibited the cellmore » viability. NiONPs' produced cytotoxicity was demonstrated through an apoptotic process, indicated by increased numbers of Annexin V positive cells and caspase-3 activation. The expression of SIRT1 was markedly down-regulated by the NiONPs, accompanied by the hyperacetylation of p53 (tumor protein 53) and overexpression of Bax (Bcl-2-associated X protein). However, overexpression of SIRT1 through resveratrol treatment or transfection clearly attenuated the NiONPs-induced apoptosis and activation of p53 and Bax. Our results suggest that the repression of SIRT1 may underlie the NiONPs-induced apoptosis via p53 hyperacetylation and subsequent Bax activation. Because SIRT1 participates in multiple biologic processes by deacetylation of dozens of substrates, this knowledge of the impact of NiONPs on SIRT1 may lead to an improved understanding of the toxic mechanisms of Nano-Ni and provide a molecular target to antagonize Nano-Ni toxicity. - Highlights: • NiONPs were taken up by BEAS-2B cells and released Ni{sup 2+}. • NiONPs produced cytotoxicity was demonstrated through an apoptotic process. • NiONPs repressed SIRT1 expression and activated p53 and Bax. • Overexpression of SIRT1 attenuated NiONPs-induced apoptosis via deacetylation p53.« less
Romain, P L; Morimoto, C; Daley, J F; Palley, L S; Reinherz, E L; Schlossman, S F
1984-01-01
To characterize the responding T cells in the autologous mixed lymphocyte reaction (AMLR), T cells were fractionated into purified subpopulations employing monoclonal antibodies and a variety of separation techniques including fluorescence-activated cell sorting. It was found that isolated T4 cells, but not T8 cells, proliferated in response to autologous non-T cells. More importantly, within the T4 subset, the autoreactive population was greatly enriched in a fraction reactive with an autoantibody from patients with juvenile chronic arthritis (JRA) or the monoclonal antibody anti-TQ1. Although T8 cells themselves were unable to proliferate in the AMLR, they could be induced to respond in the presence of either T4 cells or exogenous IL-2 containing medium. This was demonstrated by direct measurement of tritiated thymidine uptake by T8 cells during the course of the AMLR as well as by analysis of their relative DNA content. Taken together, these data indicate that the AMLR represents a complex pattern of immune responsiveness distinct from that observed in response to soluble antigen or alloantigen. The precise function of this T-cell circuit remains to be determined.
Kwon, Hye-Sook; Brent, Michael M.; Getachew, Ruth; Jayakumar, Prerana; Chen, Lin-Feng; Schnolzer, Martina; McBurney, Michael W.; Marmorstein, Ronen; Greene, Warner C.; Ott, Melanie
2009-01-01
Summary Symptoms of T-cell hyperactivation shape the course and outcome of HIV-1 infection, but the mechanism(s) underlying this chronic immune activation are not well understood. We find that the viral transactivator Tat promotes hyperactivation of T cells by blocking the nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase SIRT1. Tat directly interacts with the deacetylase domain of SIRT1 and blocks the ability of SIRT1 to deacetylate lysine 310 in the p65 subunit of NF-κB. Because acetylated p65 is more active as a transcription factor, Tat hyperactivates the expression of NF-κB-responsive genes, a function lost in SIRT1−/− cells. These results support a model where the normal function of SIRT1 as a negative regulator of T-cell activation is suppressed by Tat during HIV infection. These events likely contribute to the state of immune cell hyperactivation found in HIV-infected individuals. PMID:18329615
Zhang, Han; Sonoda, Koh-Hei; Hijioka, Kuniaki; Qiao, Hong; Oshima, Yuji; Ishibashi, Tatsuro
2009-04-17
Ocular neovascularization (NV) is the primary cause of blindness in a wide range of ocular diseases. The exact mechanism underlying the pathogenesis of ocular NV is not yet well understood, and so there is no satisfactory therapy for ocular NV. Here, we describe a strategy targeting Flk-1, a self-antigen overexpressed on proliferating endothelial cells in ocular NV, by antiangiogenic immunotherapy-DNA vaccine and adoptive T cell therapy. An oral DNA vaccine encoding Flk-1 carried by attenuated Salmonella typhimurium markedly suppressed development of laser-induced choroidal NV. We further demonstrated that adoptive transfer of vaccine-induced CD8+ T cells reduced pathological preretinal NV, with a concomitant facilitation of physiological revascularization after oxygen-induced retinal vessel obliteration. However, physiological retinal vascular development was unaffected in neonatal mice transferred with vaccine-induced CD8+ T cells. These findings suggested that antiangiogenic immunotherapy targeting Flk-1 such as vaccination and adoptive immunotherapy may contribute to future therapies for ocular NV.
DOE Office of Scientific and Technical Information (OSTI.GOV)
Gualde, N.; Goodwin, J.S.
1984-04-01
Unfractionated human T cells exposed to 10-50 rad of X irradiation incorporated less (/sup 3/H)thymidine than nonirradiated T cells when subsequently cultured with PHA or Con A. The cytotoxic/suppressor T-cell subset, isolated as either OKT8(+) or OKT4(-) cells, demonstrated significantly enhanced (/sup 3/H)thymidine incorporation in PHA- or Con A-stimulated cultures after exposure to 10-50 rad, compared to unirradiated cells, while the proliferation of the OKT4(+) helper/inducer subset was inhibited by low dose irradiation. It has been previously reported that approximately 30% of the cytotoxic/suppressor subset also stains with OKM1. When the cytotoxic/suppressor subset was further subdivided into OKT4(-), OKM1(+), andmore » OKT4(-), OKM1(-) cells, proliferation of the OKT4(-), OKM1(+) population was inhibited by exposure to 25 rad while proliferation of the OKT4(-), OKM1(-) population was stimulated. The increase in proliferation of the cytotoxic/suppressor T-cell subset after low dose irradiation is paralleled by an increase in suppressor activity of these cells. T cells exposed to 25 rad and then cultured with Con A for 48 hr caused greater inhibition of IgG production when added to fresh autologous lymphocytes stimulated by pokeweed mitogen than did unirradiated cells. Thus, low dose irradiation enhances both the proliferation and function of the human suppressor T-cell subset.« less
Chen, Wanjun; Jin, Wenwen; Wahl, Sharon M.
1998-01-01
Evidence indicates that cytotoxic T lymphocyte–associated antigen 4 (CTLA-4) may negatively regulate T cell activation, but the basis for the inhibitory effect remains unknown. We report here that cross-linking of CTLA-4 induces transforming growth factor β (TGF-β) production by murine CD4+ T cells. CD4+ T helper type 1 (Th1), Th2, and Th0 clones all secrete TGF-β after antibody cross-linking of CTLA-4, indicating that induction of TGF-β by CTLA-4 signaling represents a ubiquitous feature of murine CD4+ T cells. Stimulation of the CD3–T cell antigen receptor complex does not independently induce TGF-β, but is required for optimal CTLA-4–mediated TGF-β production. The consequences of cross-linking of CTLA-4, together with CD3 and CD28, include inhibition of T cell proliferation and interleukin (IL)-2 secretion, as well as suppression of both interferon γ (Th1) and IL-4 (Th2). Moreover, addition of anti–TGF-β partially reverses this T cell suppression. When CTLA-4 was cross-linked in T cell populations from TGF-β1 gene–deleted (TGF-β1−/−) mice, the T cell responses were only suppressed 38% compared with 95% in wild-type mice. Our data demonstrate that engagement of CTLA-4 leads to CD4+ T cell production of TGF-β, which, in part, contributes to the downregulation of T cell activation. CTLA-4, through TGF-β, may serve as a counterbalance for CD28 costimulation of IL-2 and CD4+ T cell activation. PMID:9815262
Parasitic Nematode-Induced CD4+Foxp3+T Cells Can Ameliorate Allergic Airway Inflammation
Kang, Shin Ae; Park, Mi-Kyung; Cho, Min Kyoung; Park, Sang Kyun; Jang, Min Seong; Yang, Bo-Gie; Jang, Myoung Ho; Kim, Dong-Hee; Yu, Hak Sun
2014-01-01
Background The recruitment of CD4+CD25+Foxp3+T (Treg) cells is one of the most important mechanisms by which parasites down-regulate the immune system. Methodology/Principal Findings We compared the effects of Treg cells from Trichinella spiralis-infected mice and uninfected mice on experimental allergic airway inflammation in order to understand the functions of parasite-induced Treg cells. After four weeks of T. spiralis infection, we isolated Foxp3-GFP-expressing cells from transgenic mice using a cell sorter. We injected CD4+Foxp3+ cells from T. spiralis-infected [Inf(+)Foxp3+] or uninfected [Inf(-)Foxp3+] mice into the tail veins of C57BL/6 mice before the induction of inflammation or during inflammation. Inflammation was induced by ovalbumin (OVA)-alum sensitization and OVA challenge. The concentrations of the Th2-related cytokines IL-4, IL-5, and IL-13 in the bronchial alveolar lavage fluid and the levels of OVA-specific IgE and IgG1 in the serum were lower in mice that received intravenous application of Inf(+)Foxp3+ cells [IV(inf):+(+) group] than in control mice. Some features of allergic airway inflammation were ameliorated by the intravenous application of Inf(-)Foxp3+ cells [IV(inf):+(-) group], but the effects were less distinct than those observed in the IV(inf):+(+) group. We found that Inf(+)Foxp3+ cells migrated to inflammation sites in the lung and expressed higher levels of Treg-cell homing receptors (CCR5 and CCR9) and activation markers (Klrg1, Capg, GARP, Gzmb, OX40) than did Inf(-)Foxp3+ cells. Conclusion/Significance T. spiralis infection promotes the proliferation and functional activation of Treg cells. Parasite-induced Treg cells migrate to the inflammation site and suppress immune responses more effectively than non-parasite-induced Treg cells. The adoptive transfer of Inf(+)Foxp3+ cells is an effective method for the treatment and prevention of allergic airway diseases in mice and is a promising therapeutic approach for the treatment
Baicalein induces cell death in murine T cell lymphoma via inhibition of thioredoxin system.
Patwardhan, Raghavendra S; Pal, Debojyoti; Checker, Rahul; Sharma, Deepak; Sandur, Santosh K
2017-10-01
We have earlier demonstrated the radioprotective potential of baicalein using murine splenic lymphocytes. Here, we have studied the effect of baicalein on murine T cell lymphoma EL4 cells and investigated the underlying mechanism of action. We observed that baicalein induced a dose dependent cell death in EL4 cells in vitro and significantly reduced the frequency of cancer stem cells. Previously, we have reported that murine and human T cell lymphoma cells have increased oxidative stress tolerance capacity due to active thioredoxin system. Hence, we monitored the effect of baicalein on thioredoxin system in EL4 cells. Docking studies revealed that baicalein could bind to the active site of thioredoxin reductase. Baicalein treatment led to significant reduction in the activity of thioredoxin reductase and nuclear levels of thioredoxin-1 thereby increasing ASK1 levels and caspase-3 activity. Interestingly, CRISPR-Cas9 based knock-out of ASK1 or over-expression of thioredoxin-1 abolished anti-tumor effects of baicalein in EL4 cells. Further, baicalein administration significantly reduced intra-peritoneal tumor burden of EL4 cells in C57BL/6 mice. Thus, our study describes anti-tumor effects of baicalein in EL4 cells via inhibition of thioredoxin system. Copyright © 2017 Elsevier Ltd. All rights reserved.
Peptide-Induced Antiviral Protection by Cytotoxic T Cells
NASA Astrophysics Data System (ADS)
Schulz, Manfred; Zinkernagel, Rolf M.; Hengartner, Hans
1991-02-01
A specific antiviral cytotoxic immune response in vivo could be induced by the subcutaneous injection of the T-cell epitope of the lymphocytic choriomeningitis virus (LCMV) nucleoprotein as an unmodified free synthetic peptide (Arg-Pro-Gln-Ala-Ser-Gly-Val-Tyr-Met-Gly-Asn-Leu-Thr-Ala-Gln) emulsified in incomplete Freund's adjuvant. This immunization rendered mice into a LCMV-specific protective state as shown by the inhibition of LCMV replication in spleens of such mice. The protection level of these mice correlated with the ability to respond to the peptide challenge by CD8^+ virus-specific cytotoxic T cells. This is a direct demonstration that peptide vaccines can be antivirally protective in vivo, thus encouraging further search for appropriate mixtures of stable peptides that may be used as T-cell vaccines.
IGF-1 protects intestinal epithelial cells from oxidative stress-induced apoptosis.
Baregamian, Naira; Song, Jun; Jeschke, Marc G; Evers, B Mark; Chung, Dai H
2006-11-01
Reactive oxygen species (ROS) are involved in the pathogenesis of necrotizing enterocolitis (NEC) in premature infants. We have recently found that activation of multiple cellular signaling transduction pathways occurs during ROS-induced intestinal cell apoptosis; the phosphatidylinositol 3-kinase (PI3-K) pathway plays an anti-apoptotic role during this process. Insulin-like growth factor (IGF)-1 activates PI3-K pathway to promote cell survival; however, the effects of IGF-1 treatment during gut injury are not clearly defined. The purpose of this study was to determine whether IGF-1 protects intestinal cells from ROS-induced apoptosis. Rat intestinal epithelial (RIE)-1 cells were treated with either IGF-1 (100 nm), hydrogen peroxide (H2O2; 500 microm), or combination. Western blotting was performed to assess phosphorylation of Akt, a downstream effector of PI3-K. Cell Death Detection ELISA, DCHF, and JC-1 assays were performed to demonstrate protective effects of IGF-1. Wortmannin, an inhibitor of PI3-K, was used to show PI3-K-dependent mechanism of action for IGF-1. H2O2 treatment resulted in increased intestinal epithelial cell apoptosis with intracellular ROS generation and mitochondrial membrane depolarization; IGF-1 pre-treatment attenuated this response without affecting ROS production. H2O2-induced phosphorylation of Akt was further increased with IGF-1 treatment; wortmannin abolished these effects in RIE-1 cells. PI3-K pathway is activated during ROS-induced intestinal epithelial cell injury; IGF-1 exerted an anti-apoptotic effect during this response by PI3-K activation. A better understanding of the exact role of IGF-1-mediated activation of PI3-K may allow us to facilitate the development of novel therapy against NEC.
Sierro, Sophie R; Donda, Alena; Perret, Rachel; Guillaume, Philippe; Yagita, Hideo; Levy, Frédéric; Romero, Pedro
2011-08-01
In the last two decades, anti-cancer vaccines have yielded disappointing clinical results despite the fact that high numbers of self/tumor-specific T cells can be elicited in immunized patients. Understanding the reasons behind this lack of efficacy is critical in order to design better treatment regimes. Recombinant lentivectors (rLVs) have been successfully used to induce antigen-specific T cells to foreign or mutated tumor antigens. Here, we show that rLV expressing a murine nonmutated self/tumor antigen efficiently primes large numbers of self/tumor-specific CD8(+) T cells. In spite of the large number of tumor-specific T cells, however, no anti-tumor activity could be measured in a therapeutic setting, in mice vaccinated with rLV. Accumulating evidence shows that, in the presence of malignancies, inhibition of T-cell activity may predominate overstimulation. Analysis of tumor-infiltrating lymphocytes revealed that specific anti-tumor CD8(+) T cells fail to produce cytokines and express high levels of inhibitory receptors such as programmed death (PD)-1. Association of active immunization with chemotherapy or antibodies that block inhibitory pathways often leads to better anti-tumor effects. We show here that combining rLV vaccination with either cyclophosphamide or PD-1 and PD-L1 blocking antibodies enhances rLV vaccination efficacy and improves anti-tumor immunity. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
T-kininogen induces endothelial cell proliferation.
Pérez, Viviana; Leiva-Salcedo, Elías; Acuña-Castillo, Claudio; Aravena, Mauricio; Gómez, Christian; Sabaj, Valeria; Colombo, Alicia; Nishimura, Sumiyo; Pérez, Claudio; Walter, Robin; Sierra, Felipe
2006-03-01
Basal proliferation of endothelial cells increases with age, and this might play a role in the etiology of age-related vascular diseases, as well as angiogenesis. Serum kininogen levels increase during aging in rats and humans, and T-kininogen (T-KG) can affect proliferative homeostasis in several cell models. Both kinins and kininogens have been shown previously to be angiogenic through activation of endothelial cell proliferation, and here we show that exposure of endothelial cells to T-KG results in vigorous cell proliferation, accompanied by ERK/AKT activation. In our experiments, the proliferative response requires B1 and B2 kinin receptors, even though kinins are not released from the precursor. We hypothesize that the age-related increase in T-KG could play a significant role in the age-related dysregulation of vascular physiology and function.
Cherpes, Thomas L.; Busch, James L.; Sheridan, Brian S.; Harvey, Stephen A. K.; Hendricks, Robert L.
2008-01-01
Clinical research suggests hormonal contraceptive use is associated with increased frequencies of herpes simplex virus (HSV) reactivation and shedding. We examined the effects of medroxyprogesterone acetate (MPA), the compound most commonly used for injectable hormonal contraception, on HSV-1 reactivation and CD8+ T cell function in murine trigeminal ganglia (TG). In ex vivo TG cultures, MPA dramatically inhibited canonical CD8+ T cell effector functions, including IFN-γ production and lytic granule release, and increased HSV-1 reactivation from latency. In vivo, MPA treatment of latently infected ovariectomized mice inhibited IFN-γ production and lytic granule release by TG resident CD8+ T cells stimulated directly ex vivo. RNA specific for the essential immediate early viral gene ICP4 as well as viral genome DNA copy number were increased in mice that received MPA during latency, suggesting that treatment increased in vivo reactivation. The increase in HSV-1 copy number appeared to be the result of a two-tine effect, as MPA induced higher reactivation frequencies from latently infected explanted TG neurons in the presence or absence of CD45+ cells. Our data suggest hormonal contraceptives that contain MPA may promote increased frequency of HSV reactivation from latency through the combinatory effects of inhibiting protective CD8+ T cell responses and by a leukocyte-independent effect on infected neurons. PMID:18606648
Blutt, Sarah E.; Warfield, Kelly L.; Estes, Mary K.; Conner, Margaret E.
2008-01-01
Correlates of protection from rotavirus infection are controversial. We compared the roles of B and T lymphocytes in protective immunity induced either by intranasally administered nonreplicating viruslike particles or inactivated virus or by orally administered murine rotavirus. We found that protection induced by nonreplicating vaccines requires CD4+ T cells and CD40/CD40L. In contrast, T cells were not required for short-term protective immunity induced by infection, but both T-cell-dependent and -independent mechanisms contributed to long-term maintenance of protection. Our findings indicate that more than one marker of protective immunity exists and that these markers depend on the vaccine that is administered. PMID:18184712
Annexin A1, Annexin A2, and Dyrk 1B are upregulated during GAS1-induced cell cycle arrest.
Pérez-Sánchez, Gilberto; Jiménez, Adriana; Quezada-Ramírez, Marco A; Estudillo, Enrique; Ayala-Sarmiento, Alberto E; Mendoza-Hernández, Guillermo; Hernández-Soto, Justino; Hernández-Hernández, Fidel C; Cázares-Raga, Febe E; Segovia, Jose
2018-05-01
GAS1 is a pleiotropic protein that has been investigated because of its ability to induce cell proliferation, cell arrest, and apoptosis, depending on the cellular or the physiological context in which it is expressed. At this point, we have information about the molecular mechanisms by which GAS1 induces proliferation and apoptosis; but very few studies have been focused on elucidating the mechanisms by which GAS1 induces cell arrest. With the aim of expanding our knowledge on this subject, we first focused our research on finding proteins that were preferentially expressed in cells arrested by serum deprivation. By using a proteomics approach and mass spectrometry analysis, we identified 17 proteins in the 2-DE protein profile of serum deprived NIH3T3 cells. Among them, Annexin A1 (Anxa1), Annexin A2 (Anxa2), dual specificity tyrosine-phosphorylation-regulated kinase 1B (Dyrk1B), and Eukaryotic translation initiation factor 3, F (eIf3f) were upregulated at transcriptional the level in proliferative NIH3T3 cells. Moreover, we demonstrated that Anxa1, Anxa2, and Dyrk1b are upregulated at both the transcriptional and translational levels by the overexpression of GAS1. Thus, our results suggest that the upregulation of Anxa1, Anxa2, and Dyrk1b could be related to the ability of GAS1 to induce cell arrest and maintain cell viability. Finally, we provided further evidence showing that GAS1 through Dyrk 1B leads not only to the arrest of NIH3T3 cells but also maintains cell viability. © 2017 Wiley Periodicals, Inc.
Lu, C; Li, J-Y; Ge, Z; Zhang, L; Zhou, G-P
2013-01-01
Although the intensification of therapy for children with T-cell acute lymphoblastic leukemia (T-ALL) has substantially improved clinical outcomes, T-ALL remains an important challenge in pediatric oncology. Here, we report that the cooperative synergy between prostate apoptosis response factor-4 (Par-4) and THAP1 induces cell cycle and apoptosis regulator 1 (CCAR1) gene expression and cellular apoptosis in human T-ALL cell line Jurkat cells, CEM cells and primary cultured neoplastic T lymphocytes from children with T-ALL. Par-4 and THAP1 collaborated to activate the promoter of CCAR1 gene. Mechanistic investigations revealed that Par-4 and THAP1 formed a protein complex by the interaction of their carboxyl termini, and THAP1 bound to CCAR1 promoter though its zinc-dependent DNA-binding domain at amino terminus. Par-4/THAP1 complex and Notch3 competitively bound to CCAR1 promoter and competitively modulated alternative pre-mRNA splicing of CCAR1, which resulted in two different transcripts and played an opposite role in T-ALL cell survival. Despite Notch3 induced a shift splicing from the full-length isoform toward a shorter form of CCAR1 mRNA by splicing factor SRp40 and SRp55, Par-4/THAP1 complex strongly antagonized this inductive effect. Our finding revealed a mechanistic rationale for Par-4/THAP1-induced apoptosis in T-ALL cells that would be of benefit to develop a new therapy strategy for T-ALL. PMID:23975424
Le Buanec, Hélène; Gougeon, Marie-Lise; Mathian, Alexis; Lebon, Pierre; Dupont, Jean-Michel; Peltre, Gabriel; Hemon, Patrice; Schmid, Michel; Bizzini, Bernard; Künding, Thomas; Burny, Arsène; Bensussan, Armand; Amoura, Zahir; Gallo, Robert C.; Zagury, Daniel
2011-01-01
Immune suppressive activities exerted by regulatory T-cell subsets have several specific functions, including self-tolerance and regulation of adaptive immune reactions, and their dysfunction can lead to autoimmune diseases and contribute to AIDS and cancer. Two functionally distinct regulatory T-cell subsets are currently identified in peripheral tissues: thymus-developed natural T regulatory cells (nTregs) controlling self-tolerance and antiinflammatory IL-10–secreting type 1 regulatory T cells (Tr1) derived from Ag-stimulated T cells, which regulate inflammation-dependent adaptive immunity and minimize immunopathology. We establish herein that cell contact-mediated nTreg regulatory function is inhibited by inflammation, especially in the presence of the complement C3b receptor (CD46). Instead, as with other T-cell subsets, the latter inflammatory conditions of stimulation skew nTreg differentiation to Tr1 cells secreting IL-10, an effect potentiated by IFN-α. The clinical relevance of these findings was verified in a study of 152 lupus patients, in which we showed that lupus nTreg dysfunction is not due to intrinsic defects but is rather induced by C3b stimulation of CD46 and IFN-α and that these immune components of inflammation are directly associated with active lupus. These results provide a rationale for using anti–IFN-α Ab immunotherapy in lupus patients. PMID:22065791
Seo, Young-Jin; Kim, Kui-Jin; Choi, Jia; Koh, Eun-Jeong; Lee, Boo-Yong
2018-06-01
Obesity predisposes animals towards the metabolic syndrome and diseases such as type 2 diabetes, atherosclerosis, and cardiovascular disease. Spirulina maxima is a microalga with anti-oxidant, anti-cancer, and neuroprotective activities, but the anti-obesity effect of Spirulina maxima 70% ethanol extract (SM70EE) has not yet been fully established. We investigated the effect of SM70EE on adipogenesis, lipogenesis, and browning using in vitro and in vivo obesity models. SM70EE treatment reduced lipid droplet accumulation by the oil red O staining method and downregulated the adipogenic proteins C/EBPα, PPARγ, and aP2, and the lipogenic proteins SREBP1, ACC, FAS, LPAATβ, Lipin1, and DGAT1 by western blot analysis. In addition, the index components of SM70EE, chlorophyll a, and C-phycocyanin, reduced adipogenesis and lipogenesis protein levels in 3T3-L1 and C3H10T1/2 cells. High-fat diet (HFD)-fed mice administered with SM70EE demonstrated smaller adipose depots and lower blood lipid concentrations than control HFD-fed mice. The lower body mass gain in treated SM70EE-administrated mice was associated with lower protein expression of adipogenesis factors and higher expression of AMPKα-induced adipose browning proteins PRDM16, PGC1α, and UCP1. SM70EE administration ameliorates obesity, likely by reducing adipogenesis and activating the thermogenic program, in 3T3-L1 cells and HFD-induced obese mice.
Targeting LAG-3 and PD-1 to Enhance T Cell Activation by Antigen-Presenting Cells
Lichtenegger, Felix S.; Rothe, Maurine; Schnorfeil, Frauke M.; Deiser, Katrin; Krupka, Christina; Augsberger, Christian; Schlüter, Miriam; Neitz, Julia; Subklewe, Marion
2018-01-01
Immune checkpoint inhibition has been shown to successfully reactivate endogenous T cell responses directed against tumor-associated antigens, resulting in significantly prolonged overall survival in patients with various tumor entities. For malignancies with low endogenous immune responses, this approach has not shown a clear clinical benefit so far. Therapeutic vaccination, particularly dendritic cell (DC) vaccination, is a strategy to induce T cell responses. Interaction of DCs and T cells is dependent on receptor–ligand interactions of various immune checkpoints. In this study, we analyzed the influence of blocking antibodies targeting programmed cell death protein 1 (PD-1), HVEM, CD244, TIM-3, and lymphocyte activation gene 3 (LAG-3) on the proliferation and cytokine secretion of T cells after stimulation with autologous TLR-matured DCs. In this context, we found that LAG-3 blockade resulted in superior T cell activation compared to inhibition of other pathways, including PD-1/PD-L1. This result was consistent across different methods to measure T cell stimulation (proliferation, IFN-γ secretion), various stimulatory antigens (viral and bacterial peptide pool, specific viral antigen, specific tumor antigen), and seen for both CD4+ and CD8+ T cells. Only under conditions with a weak antigenic stimulus, particularly when combining antigen presentation by peripheral blood mononuclear cells with low concentrations of peptides, we observed the highest T cell stimulation with dual blockade of LAG-3 and PD-1 blockade. We conclude that priming of novel immune responses can be strongly enhanced by blockade of LAG-3 or dual blockade of LAG-3 and PD-1, depending on the strength of the antigenic stimulus. PMID:29535740
Targeting LAG-3 and PD-1 to Enhance T Cell Activation by Antigen-Presenting Cells.
Lichtenegger, Felix S; Rothe, Maurine; Schnorfeil, Frauke M; Deiser, Katrin; Krupka, Christina; Augsberger, Christian; Schlüter, Miriam; Neitz, Julia; Subklewe, Marion
2018-01-01
Immune checkpoint inhibition has been shown to successfully reactivate endogenous T cell responses directed against tumor-associated antigens, resulting in significantly prolonged overall survival in patients with various tumor entities. For malignancies with low endogenous immune responses, this approach has not shown a clear clinical benefit so far. Therapeutic vaccination, particularly dendritic cell (DC) vaccination, is a strategy to induce T cell responses. Interaction of DCs and T cells is dependent on receptor-ligand interactions of various immune checkpoints. In this study, we analyzed the influence of blocking antibodies targeting programmed cell death protein 1 (PD-1), HVEM, CD244, TIM-3, and lymphocyte activation gene 3 (LAG-3) on the proliferation and cytokine secretion of T cells after stimulation with autologous TLR-matured DCs. In this context, we found that LAG-3 blockade resulted in superior T cell activation compared to inhibition of other pathways, including PD-1/PD-L1. This result was consistent across different methods to measure T cell stimulation (proliferation, IFN-γ secretion), various stimulatory antigens (viral and bacterial peptide pool, specific viral antigen, specific tumor antigen), and seen for both CD4 + and CD8 + T cells. Only under conditions with a weak antigenic stimulus, particularly when combining antigen presentation by peripheral blood mononuclear cells with low concentrations of peptides, we observed the highest T cell stimulation with dual blockade of LAG-3 and PD-1 blockade. We conclude that priming of novel immune responses can be strongly enhanced by blockade of LAG-3 or dual blockade of LAG-3 and PD-1, depending on the strength of the antigenic stimulus.
Conserved region C functions to regulate PD-1 expression and subsequent CD8 T cell memory1
Bally, Alexander P. R.; Tang, Yan; Lee, Joshua T.; Barwick, Benjamin G.; Martinez, Ryan; Evavold, Brian D.; Boss, Jeremy M.
2016-01-01
Expression of programmed death 1 (PD-1) on CD8 T cells promotes T cell exhaustion during chronic antigen exposure. During acute infections, PD-1 is transiently expressed and has the potential to modulate CD8 T cell memory formation. Conserved Region C (CR-C), a promoter proximal cis-regulatory element that is critical to PD-1 expression in vitro, responds to NFATc1, FoxO1, and/or NF-κB signaling pathways. Here, a CR-C knockout mouse (CRC−) was established to determine its role on PD-1 expression and corresponding effects on T cell function in vivo. Deletion of CR-C decreased PD-1 expression on CD4 T cells and antigen-specific CD8 T cells during acute and chronic lymphocytic choriomeningitis virus (LCMV) challenges, but did not affect the ability to clear an infection. Following acute LCMV infection, memory CD8 T cells in the CRC− mouse were formed in greater numbers, were more functional, and were more effective at responding to a melanoma tumor than wild-type memory cells. These data implicate a critical role for CR-C in governing PD-1 expression, and a subsequent role in guiding CD8 T cell differentiation. The data suggest the possibility that titrating PD-1 expression during CD8 T cell activation could have important ramifications in vaccine development and clinical care. PMID:27895178
PD-1 expression on dendritic cells suppresses CD8+ T cell function and antitumor immunity
Lim, Tong Seng; Chew, Valerie; Sieow, Je Lin; Goh, Siting; Yeong, Joe Poh-Sheng; Soon, Ai Ling; Ricciardi-Castagnoli, Paola
2016-01-01
ABSTRACT Programmed death one (PD-1) is a well-established co-inhibitory regulator that suppresses proliferation and cytokine production of T cells. Despite remarkable progress in delineating the functional roles of PD-1 on T lymphocytes, little is known about the regulatory role of PD-1 expressed on myeloid cells such as dendritic cells (DCs). Here, we show that CD8+ T cells can be more potently activated to secrete IL-2 and IFNγ by PD-1-deficient DCs compared to wild-type DCs. Adoptive transfer of PD-1-deficient DCs demonstrated their superior capabilities in inducing antigen-specific CD8+ T cell proliferation in vivo. In addition, we provide first evidence demonstrating the existence of peripheral blood DCs and CD11c+ tumor-infiltrating myeloid cells that co-express PD-1 in patients with hepatocellular carcinoma (HCC). The existence of PD-1-expressing HCC-infiltrating DCs (HIDCs) was further supported in a mouse model of HCC. Intratumoral transfer of PD-1-deficient DCs rendered recipient mice resistant to the growth of HCC by promoting tumor-infiltrating CD8+ effector T cells to secrete perforin and granzyme B. This novel finding provides a deeper understanding of the role of PD-1 in immune regulation and has significant implications for cancer immunotherapies targeting PD-1. PMID:27141339
Elevated Cyclic AMP Levels in T Lymphocytes Transformed by Human T-Cell Lymphotropic Virus Type 1▿
Kress, Andrea K.; Schneider, Grit; Pichler, Klemens; Kalmer, Martina; Fleckenstein, Bernhard; Grassmann, Ralph
2010-01-01
Human T-cell lymphotropic virus type 1 (HTLV-1), the cause of adult T-cell leukemia/lymphoma (ATLL), transforms CD4+ T cells to permanent growth through its transactivator Tax. HTLV-1-transformed cells share phenotypic properties with memory and regulatory T cells (T-reg). Murine T-reg-mediated suppression employs elevated cyclic AMP (cAMP) levels as a key regulator. This led us to determine cAMP levels in HTLV-1-transformed cells. We found elevated cAMP concentrations as a consistent feature of all HTLV-1-transformed cell lines, including in vitro-HTLV-1-transformed, Tax-transformed, and patient-derived cells. In transformed cells with conditional Tax expression, high cAMP levels coincided with the presence of Tax but were lost without it. However, transient ectopic expression of Tax alone was not sufficient to induce cAMP. We found specific downregulation of the cAMP-degrading phosphodiesterase 3B (PDE3B) in HTLV-1-transformed cells, which was independent of Tax in transient expression experiments. This is in line with the notion that PDE3B transcripts and cAMP levels are inversely correlated. Overexpression of PDE3B led to a decrease of cAMP in HTLV-1-transformed cells. Decreased expression of PDE3B was associated with inhibitory histone modifications at the PDE3B promoter and the PDE3B locus. In summary, Tax transformation and its continuous expression contribute to elevated cAMP levels, which may be regulated through PDE3B suppression. This shows that HTLV-1-transformed cells assume biological features of long-lived T-cell populations that potentially contribute to viral persistence. PMID:20573814
Co-Expansion of Cytokine-Induced Killer Cells and Vγ9Vδ2 T Cells for CAR T-Cell Therapy
Chen, Can; Tan, Wee-Kiat; Chi, Zhixia; Xu, Xue-Hu; Wang, Shu
2016-01-01
Gamma delta (γδ) T cells and cytokine-induced killer (CIK) cells, which are a heterogeneous population of T lymphocytes and natural killer T (NKT) cells, have been separately expanded ex vivo and shown to be capable of targeting and mediating cytotoxicity against various tumor cells in a major histocompatibility complex-unrestricted manner. However, the co-expansion and co-administration of these immune cells have not been explored. In this study we describe an efficient method to expand simultaneously both CIK and Vγ9Vδ2 T cells, termed as CIKZ cells, from human peripheral blood mononuclear cells (PBMCs) using Zometa, interferon-gamma (IFN-γ), interleukin 2 (IL-2), anti-CD3 antibody and engineered K562 feeder cells expressing CD64, CD137L and CD86. A 21-day culture of PBMCs with this method yielded nearly 20,000-fold expansion of CIKZ cells with γδ T cells making up over 20% of the expanded population. The expanded CIKZ cells exhibited antitumor cytotoxicity and could be modified to express anti-CD19 chimeric antigen receptor (CAR), anti-CEA CAR, and anti-HER2 CAR to enhance their specificity and cytotoxicity against CD19-, CEA-, or HER2-positive tumor cells. The tumor inhibitory activity of anti-CD19 CAR-modified CIKZ cells was further demonstrated in vivo in a Raji tumor mouse model. The findings herein substantiate the feasibility of co-expanding CIK and γδ cells for adoptive cellular immunotherapy applications such as CAR T-cell therapy against cancer. PMID:27598655
Lu, Peiyuan; Youngblood, Benjamin A.; Austin, James W.; Rasheed Mohammed, Ata Ur; Butler, Royce; Ahmed, Rafi
2014-01-01
Programmed cell death 1 (PD-1) is an inhibitory immune receptor that regulates T cell function, yet the molecular events that control its expression are largely unknown. We show here that B lymphocyte–induced maturation protein 1 (Blimp-1)–deficient CD8 T cells fail to repress PD-1 during the early stages of CD8 T cell differentiation after acute infection with lymphocytic choriomeningitis virus (LCMV) strain Armstrong. Blimp-1 represses PD-1 through a feed-forward repressive circuit by regulating PD-1 directly and by repressing NFATc1 expression, an activator of PD-1 expression. Blimp-1 binding induces a repressive chromatin structure at the PD-1 locus, leading to the eviction of NFATc1 from its site. These data place Blimp-1 at an important phase of the CD8 T cell effector response and provide a molecular mechanism for its repression of PD-1. PMID:24590765
The ion channel TRPV1 regulates the activation and proinflammatory properties of CD4+ T cells
Bertin, Samuel; Aoki-Nonaka, Yukari; de Jong, Petrus Rudolf; Stanwood, Shawna R.; Srikanth, Sonal; Lee, Jihyung; To, Keith; Abramson, Lior; Yu, Timothy; Han, Tiffany; Touma, Ranim; Li, Xiangli; González-Navajas, José M.; Herdman, Scott; Corr, Maripat; Fu, Guo; Dong, Hui; Gwack, Yousang; Franco, Alessandra; Jefferies, Wilfred A.; Raz, Eyal
2016-01-01
TRPV1 is a Ca2+-permeable channel mostly studied as a pain receptor in sensory neurons. However, its role in other cell types is poorly understood. Here, we demonstrate that TRPV1 is functionally expressed in CD4+ T cells where it acts as a non-store-operated Ca2+ channel and contributes to T cell receptor (TCR)-induced Ca2+ influx, TCR signaling and T cell activation. In models of T cell-mediated colitis, TRPV1 promotes colitogenic T cell responses and intestinal inflammation. Furthermore, genetic and pharmacological inhibition of TRPV1 in human CD4+ T cells recapitulates the phenotype of murine Trpv1−/− CD4+ T cells. These findings suggest that TRPV1 inhibition could represent a new therapeutic strategy to restrain proinflammatory T cell responses. PMID:25282159
Establishment of stable cell line for inducing KAP1 protein expression.
Liu, Xiaoyan; Khan, Md Asaduzzaman; Cheng, Jingliang; Wei, Chunli; Zhang, Lianmei; Fu, Junjiang
2015-06-01
Generation of the stable cell lines is a highly efficient tool in functional studies of certain genes or proteins, where the particular genes or proteins are inducibly expressed. The KRAB-associated protein-1 (KAP1) is an important transcription regulatory protein, which is investigated in several molecular biological studies. In this study, we have aimed to generate a stable cell line for inducing KAP1 expression. The recombinant plasmid pcDNA5/FRT/TO-KAP1 was constructed at first, which was then transfected into Flp-In™T-REx™-HEK293 cells to establish an inducible pcDNA5/FRT/TO-KAP1-HEK293 cell line. The Western blot analysis showed that the protein level of KAP1 is over-expressed in the established stable cell line by doxycycline induction, both dose and time dependently. Thus we have successfully established stable pcDNA5/FRT/TO-KAP1-HEK293 cell line, which can express KAP1 inducibly. This inducible cell line might be very useful for KAP1 functional studies.
Blockade of PD-1/PD-L1 Promotes Adoptive T-Cell Immunotherapy in a Tolerogenic Environment
Kenna, Tony J.; Galea, Ryan; Large, Justin; Yagita, Hideo; Steptoe, Raymond J.
2015-01-01
Adoptive cellular immunotherapy using in vitro expanded CD8+ T cells shows promise for tumour immunotherapy but is limited by eventual loss of function of the transferred T cells through factors that likely include inactivation by tolerogenic dendritic cells (DC). The co-inhibitory receptor programmed death-1 (PD-1), in addition to controlling T-cell responsiveness at effector sites in malignancies and chronic viral diseases is an important modulator of dendritic cell-induced tolerance in naive T cell populations. The most potent therapeutic capacity amongst CD8+ T cells appears to lie within Tcm or Tcm-like cells but memory T cells express elevated levels of PD-1. Based on established trafficking patterns for Tcm it is likely Tcm-like cells interact with lymphoid-tissue DC that present tumour-derived antigens and may be inherently tolerogenic to develop therapeutic effector function. As little is understood of the effect of PD-1/PD-L1 blockade on Tcm-like CD8+ T cells, particularly in relation to inactivation by DC, we explored the effects of PD-1/PD-L1 blockade in a mouse model where resting DC tolerise effector and memory CD8+ T cells. Blockade of PD-1/PD-L1 promoted effector differentiation of adoptively-transferred Tcm-phenotype cells interacting with tolerising DC. In tumour-bearing mice with tolerising DC, effector activity was increased in both lymphoid tissues and the tumour-site and anti-tumour activity was promoted. Our findings suggest PD-1/PD-L1 blockade may be a useful adjunct for adoptive immunotherapy by promoting effector differentiation in the host of transferred Tcm-like cells. PMID:25741704
Blockade of PD-1/PD-L1 promotes adoptive T-cell immunotherapy in a tolerogenic environment.
Blake, Stephen J P; Ching, Alan L H; Kenna, Tony J; Galea, Ryan; Large, Justin; Yagita, Hideo; Steptoe, Raymond J
2015-01-01
Adoptive cellular immunotherapy using in vitro expanded CD8+ T cells shows promise for tumour immunotherapy but is limited by eventual loss of function of the transferred T cells through factors that likely include inactivation by tolerogenic dendritic cells (DC). The co-inhibitory receptor programmed death-1 (PD-1), in addition to controlling T-cell responsiveness at effector sites in malignancies and chronic viral diseases is an important modulator of dendritic cell-induced tolerance in naive T cell populations. The most potent therapeutic capacity amongst CD8+ T cells appears to lie within Tcm or Tcm-like cells but memory T cells express elevated levels of PD-1. Based on established trafficking patterns for Tcm it is likely Tcm-like cells interact with lymphoid-tissue DC that present tumour-derived antigens and may be inherently tolerogenic to develop therapeutic effector function. As little is understood of the effect of PD-1/PD-L1 blockade on Tcm-like CD8+ T cells, particularly in relation to inactivation by DC, we explored the effects of PD-1/PD-L1 blockade in a mouse model where resting DC tolerise effector and memory CD8+ T cells. Blockade of PD-1/PD-L1 promoted effector differentiation of adoptively-transferred Tcm-phenotype cells interacting with tolerising DC. In tumour-bearing mice with tolerising DC, effector activity was increased in both lymphoid tissues and the tumour-site and anti-tumour activity was promoted. Our findings suggest PD-1/PD-L1 blockade may be a useful adjunct for adoptive immunotherapy by promoting effector differentiation in the host of transferred Tcm-like cells.
Vav1 GEF activity is required for T cell mediated allograft rejection.
Haubert, Dirk; Li, Jianping; Saveliev, Alexander; Calzascia, Thomas; Sutter, Esther; Metzler, Barbara; Kaiser, Daniel; Tybulewicz, Victor L J; Weckbecker, Gisbert
2012-06-01
The GDP exchange factor (GEF) Vav1 is a central signal transducer downstream of the T cell receptor and has been identified as a key factor for T cell activation in the context of allograft rejection. Vav1 has been shown to transduce signals both dependent and independent of its GEF function. The most promising approach to disrupt Vav1 activity by pharmacological inhibition would be to target its GEF function. However, the contribution of Vav1 GEF activity for allogeneic T cell activation has not been clarified yet. To address this question, we used knock-in mice bearing a mutated Vav1 with disrupted GEF activity but intact GEF-independent functions. T cells from these mice showed strongly reduced proliferation and activation in response to allogeneic stimulation. Furthermore, lack of Vav1 GEF activity strongly abrogated the in vivo expansion of T cells in a systemic graft-versus-host model. In a cardiac transplantation model, mice with disrupted Vav1 GEF activity show prolonged allograft survival. These findings demonstrate a strong requirement for Vav1 GEF activity for allogeneic T cell activation and graft rejection suggesting that disruption of Vav1 GEF activity alone is sufficient to induce significant immunosuppression. Copyright © 2012 Elsevier B.V. All rights reserved.
Vav1 GEF activity is required for T cell mediated allograft rejection
Haubert, Dirk; Li, Jianping; Saveliev, Alexander; Calzascia, Thomas; Sutter, Esther; Metzler, Barbara; Kaiser, Daniel; Tybulewicz, Victor L.J.; Weckbecker, Gisbert
2012-01-01
The GDP exchange factor (GEF) Vav1 is a central signal transducer downstream of the T cell receptor and has been identified as a key factor for T cell activation in the context of allograft rejection. Vav1 has been shown to transduce signals both dependent and independent of its GEF function. The most promising approach to disrupt Vav1 activity by pharmacological inhibition would be to target its GEF function. However, the contribution of Vav1 GEF activity for allogeneic T cell activation has not been clarified yet. To address this question, we used knock-in mice bearing a mutated Vav1 with disrupted GEF activity but intact GEF-independent functions. T cells from these mice showed strongly reduced proliferation and activation in response to allogeneic stimulation. Furthermore, lack of Vav1 GEF activity strongly abrogated the in vivo expansion of T cells in a systemic graft-versus-host model. In a cardiac transplantation model, mice with disrupted Vav1 GEF activity show prolonged allograft survival. These findings demonstrate a strong requirement for Vav1 GEF activity for allogeneic T cell activation and graft rejection suggesting that disruption of Vav1 GEF activity alone is sufficient to induce significant immunosuppression. PMID:22456277
Hsu, Chih‐Kai; Lin, Chih‐Chung; Hsiao, Li‐Der
2015-01-01
Background and Purpose Sphingosine 1‐phosphate (S1P), an important inflammatory mediator, has been shown to regulate COX‐2 production and promote various cellular responses such as cell migration. Mevastatin, an inhibitor of 3‐hydroxy‐3‐methylglutaryl‐CoA reductase (HMG‐CoA), effectively inhibits inflammatory responses. However, the mechanisms underlying S1P‐evoked COX‐2‐dependent cell migration, which is modulated by mevastatin in human tracheal smooth muscle cells (HTSMCs) remain unclear. Experimental Approach The expression of COX‐2 was determined by Western blotting, real time‐PCR and promoter analyses. The signalling molecules were investigated by pretreatment with respective pharmacological inhibitors or transfection with siRNAs. The interaction between COX‐2 promoter and transcription factors was determined by chromatin immunoprecipitation assay. Finally, the effect of mevastatin on HTSMC migration and leukocyte counts in BAL fluid and COX‐2 expression induced by S1P was determined by a cell migration assay, cell counting and Western blot. Key Results S1P stimulated mTOR activation through the Nox2/ROS and PI3K/Akt pathways, which can further stimulate FoxO1 phosphorylation and translocation to the cytosol. We also found that S1P induced CREB activation and translocation via an mTOR‐independent signalling pathway. Finally, we showed that pretreatment with mevastatin markedly reduced S1P‐induced cell migration and COX‐2/PGE2 production via a PPARγ‐dependent signalling pathway. Conclusions and Implications Mevastatin attenuates the S1P‐induced increased expression of COX‐2 and cell migration via the regulation of FoxO1 and CREB phosphorylation and translocation by PPARγ in HTSMCs. Mevastatin could be beneficial for prevention of airway inflammation in the future. PMID:26359950
Vidal, Jorge E; Enríquez-Rincón, Fernando; Giono-Cerezo, Silvia; Ribas-Aparicio, Rosa María; Figueroa-Arredondo, Paula
2009-01-01
To investigate whether the HlyA-induced vacuolating effect is produced by V. cholerae O1 ElTor strains isolated from different geographic origins, including Mexico. Supernatant-induced haemolysis, vacuolating activity and cytotoxicity in Vero cells were recorded. PCR, RFLP analysis and molecular cloning were performed. All ElTor strains analyzed induced cellular vacuolation. Ribotype 2 strains isolates from the U.S. gulf coast yielded the highest titer of vacuolating activity. Eight of nine strains were haemolytic, while all strains were PCR positive for the hlyA gene. We cloned the hlyA gene from two ElTor strains, a toxigenic (2514-88, ctxAB+) and a non-toxigenic Mexican strain (CM 91-3, ctxAB-). Supernatant from those recombinant E. coli strains induced haemolysis, cell vacuolation and cytotoxicity. RFLP-PCR analysis revealed similarities in the hlyA gene from all strains tested. The HlyA-induced vacuolating effect is a widespread phenotype of epidemic V. cholerae O1 ElTor strains.
Osteoblasts Protect AML Cells from SDF-1-Induced Apoptosis
Kremer, Kimberly N.; Dudakovic, Amel; McGee-Lawrence, Meghan E.; Philips, Rachael L.; Hess, Allan D.; Smith, B. Douglas; van Wijnen, Andre J.; Karp, Judith E.; Kaufmann, Scott H.; Westendorf, Jennifer J.; Hedin, Karen E.
2014-01-01
The bone marrow provides a protective environment for acute myeloid leukemia (AML) cells that often allows leukemic stem cells to survive standard chemotherapeutic regimens. Targeting these leukemic stem cells within the bone marrow is critical for preventing relapse. We recently demonstrated that SDF-1, a chemokine abundant in the bone marrow, induces apoptosis in AML cell lines and in patient samples expressing high levels of its receptor, CXCR4. Here we show that a subset of osteoblast lineage cells within the bone marrow can protect AML cells from undergoing apoptosis in response to the SDF-1 naturally present in that location. In co-culture systems, osteoblasts at various stages of differentiation protected AML cell lines and patient isolates from SDF-1-induced apoptosis. The differentiation of the osteoblast cell lines, MC3T3 and W-20-17, mediated this protection via a cell contact-independent mechanism. In contrast, bone marrow-derived mesenchymal cells, the precursors of osteoblasts, induced apoptosis in AML cells via a CXCR4-dependent mechanism and failed to protect AML cells from exogenously added SDF-1. These results indicate that osteoblasts in the process of differentiation potently inhibit the SDF-1-driven apoptotic pathway of CXCR4-expressing AML cells residing in the bone marrow. Drugs targeting this protective mechanism could potentially provide a new approach to treating AML by enhancing the SDF-1-induced apoptosis of AML cells residing within the bone marrow microenvironment. PMID:24851270
T-cell differentiation of multipotent hematopoietic cell line EML in the OP9-DL1 coculture system
Kutleša, Snježana; Zayas, Jennifer; Valle, Alexandra; Levy, Robert B.; Jurecic, Roland
2011-01-01
Objective Multipotent hematopoietic cell line EML can differentiate into myeloid, erythroid, megakaryocytic, and B-lymphoid lineages, but it remained unknown whether EML cells have T-cell developmental potential as well. The goal of this study was to determine whether the coculture with OP9 stromal cells expressing Notch ligand Delta-like 1 (OP9-DL1) could induce differentiation of EML cells into T-cell lineage. Materials and Methods EML cells were cocultured with control OP9 or OP9-DL1 stromal cells in the presence of cytokines (stem cell factor, interleukin-7, and Fms-like tyrosine kinase 3 ligand). Their T-cell lineage differentiation was assessed through flow cytometry and reverse transcription polymerase chain reaction expression analysis of cell surface markers and genes characterizing and associated with specific stages of T-cell development. Results The phenotypic, molecular, and functional analysis has revealed that in EML/OP9-DL1 cocultures with cytokines, but not in control EML/OP9 cocultures, EML cell line undergoes T-cell lineage commitment and differentiation. In OP9-DL1 cocultures, EML cell line has differentiated into cells that 1) resembled double-negative, double-positive, and single-positive stages of T-cell development; 2) initiated expression of GATA-3, Pre-Tα, RAG-1, and T-cell receptor – Vβ genes; and 3) produced interferon-γ in response to T-cell receptor stimulation. Conclusions These results support the notion that EML cell line has the capacity for T-cell differentiation. Remarkably, induction of T-lineage gene expression and differentiation of EML cells into distinct stages of T-cell development were very similar to previously described T-cell differentiation of adult hematopoietic stem cells and progenitors in OP9-DL1 cocultures. Thus, EML/OP9-DL1 coculture could be a useful experimental system to study the role of particular genes in T-cell lineage specification, commitment, and differentiation. PMID:19447159
Schulte, Ralf; Autenrieth, Ingo B.
1998-01-01
In response to bacterial entry epithelial cells up-regulate expression and secretion of various proinflammatory cytokines, including interleukin-8 (IL-8). We studied Yersinia enterocolitica O:8-induced IL-8 secretion by intestinal epithelial cells as a function of cell differentiation. For this purpose, human T84 intestinal epithelial cells were grown on permeable supports, which led to the formation of tight monolayers of polarized intestinal epithelial cells. To analyze IL-8 secretion as a function of cell differentiation, T84 monolayers were infected from the apical or basolateral side at different stages of differentiation. Both virulent (plasmid-carrying) and nonvirulent (plasmid-cured) Y. enterocolitica strains invaded nondifferentiated T84 cells from the apical side. Yersinia invasion into T84 cells was followed by secretion of IL-8. After polarized differentiation of T84 cells Y. enterocolitica was no longer able to invade from the apical side or to induce IL-8 secretion by T84 cells. However, Y. enterocolitica invaded and induced IL-8 secretion by polarized T84 cells after infection from the basolateral side. Basolateral invasion required the presence of the Yersinia invasion locus, inv, suggesting β1 integrin-mediated cell invasion. After basolateral infection, Yersinia-induced IL-8 secretion was not strictly dependent on cell invasion. Thus, although the plasmid-carrying Y. enterocolitica strain did not significantly invade T84 cells, it induced significant IL-8 secretion. Taken together, these data show that Yersinia-triggered IL-8 secretion by intestinal epithelial cells depends on cell differentiation and might be induced by invasion as well as by basolateral adhesion, suggesting that invasion is not essential for triggering IL-8 production. Whether IL-8 secretion is involved in the pathogenesis of Yersinia-induced abscess formation in Peyer’s patch tissue remains to be shown. PMID:9488416
Dobos, Karen M.; Lucas, Megan; Spencer, John S.; Fang, Sunan; McDonald, Melissa A.; Pohl, Jan; Birkness, Kristin; Chamcha, Venkateswarlu; Ramirez, Melissa V.; Plikaytis, Bonnie B.; Posey, James E.; Amara, Rama Rao
2013-01-01
Glycosylation is the most abundant post-translational polypeptide chain modification in nature. Although carbohydrate modification of protein antigens from many microbial pathogens constitutes important components of B cell epitopes, the role in T cell immunity is not completely understood. Here, using ELISPOT and polychromatic flow cytometry, we show that O-mannosylation of the adhesin, Apa, of Mycobacterium tuberculosis (Mtb) is crucial for its T cell antigenicity in humans and mice after infection. However, subunit vaccination with both mannosylated and non-mannosylated Apa induced a comparable magnitude and quality of T cell response and imparted similar levels of protection against Mtb challenge in mice. Both forms equally improved waning BCG vaccine-induced protection in elderly mice after subunit boosting. Thus, O-mannosylation of Apa is required for antigenicity but appears to be dispensable for its immunogenicity and protective efficacy in mice. These results have implications for the development of subunit vaccines using post-translationally modified proteins such as glycoproteins against infectious diseases like tuberculosis. PMID:24130497
Nandakumar, Subhadra; Kannanganat, Sunil; Dobos, Karen M; Lucas, Megan; Spencer, John S; Fang, Sunan; McDonald, Melissa A; Pohl, Jan; Birkness, Kristin; Chamcha, Venkateswarlu; Ramirez, Melissa V; Plikaytis, Bonnie B; Posey, James E; Amara, Rama Rao; Sable, Suraj B
2013-01-01
Glycosylation is the most abundant post-translational polypeptide chain modification in nature. Although carbohydrate modification of protein antigens from many microbial pathogens constitutes important components of B cell epitopes, the role in T cell immunity is not completely understood. Here, using ELISPOT and polychromatic flow cytometry, we show that O-mannosylation of the adhesin, Apa, of Mycobacterium tuberculosis (Mtb) is crucial for its T cell antigenicity in humans and mice after infection. However, subunit vaccination with both mannosylated and non-mannosylated Apa induced a comparable magnitude and quality of T cell response and imparted similar levels of protection against Mtb challenge in mice. Both forms equally improved waning BCG vaccine-induced protection in elderly mice after subunit boosting. Thus, O-mannosylation of Apa is required for antigenicity but appears to be dispensable for its immunogenicity and protective efficacy in mice. These results have implications for the development of subunit vaccines using post-translationally modified proteins such as glycoproteins against infectious diseases like tuberculosis.
Jin, Xiaolu; Wang, Kai; Liu, Hongyun; Hu, Fuliang; Zhao, Fengqi; Liu, Jianxin
2016-01-01
The mammary epithelial cells (MECs) of high-producing dairy cows are likely to be subject to oxidative stress (OS) due to the intensive cell metabolism. The objectives of this study were to investigate the cytoprotective effects of resveratrol against hydrogen peroxide- (H2O2-) induced OS in cultured bovine MECs (MAC-T). Pretreatment of MAC-T cells with resveratrol could rescue the decrease in cell viability and resulted in lower intracellular reactive oxygen species (ROS) accumulation after H2O2 exposure. Resveratrol helped MAC-T cells to prevent H2O2-induced endoplasmic reticulum stress and mitochondria-related cell apoptosis. Moreover, resveratrol induced mRNA expression of multiple antioxidant defense genes in MAC-T cells under normal/oxidative conditions. Nuclear factor erythroid 2-related factor 2 (Nrf2) was required for the cytoprotective effects on MAC-T cells by resveratrol, as knockdown of Nrf2 significantly abolished resveratrol-induced cytoprotective effects against OS. In addition, by using selective inhibitors, we further confirmed that the induction of Nrf2 by resveratrol was mediated through the prolonged activation of PI3K/Akt and ERK/MAPK pathways but negatively regulated by p38/MAPK pathway. Overall, resveratrol has beneficial effects on bovine MECs redox balance and may be potentially used as a therapeutic medicine against oxidative insult in lactating animals.
Chao, Chen; Li, Feng; Tan, Zhiping; Zhang, Weizhi; Yang, Yifeng; Luo, Cheng
2018-01-15
Recent reports have demonstrated that RAF-1 L613V (a mutant of RAF-1) mutant mice show bone deformities similar to Noonan syndrome. It has been suggested that RAF-1 L613V might abnormally activate osteoblast differentiation of MC3T3-E1 cells. To demonstrate that RAF-1 is associated with bone deformity and that RAF-1 L613V dependent bone deformity could be inhibited by microRNA-195 (miR-195), we first investigated the amplifying influence of wild-type RAF-1 (WT) or RAF-1 L613V (L613V) on the viability and differentiation of MC3T3-E1 cells induced by bone morphogenetic protein-2 (BMP-2) via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining, quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analysis. Subsequently, we investigated the blocking effect and its mechanism of miR-195 for abnormal activation of osteoblast differentiation of MC3T3-E1 cells via targeting RAF-1. RAF-1, especially RAF-1 L613V , abnormally activates osteoblast differentiation of MC3T3-E1 cells induced by BMP-2. Meanwhile, miR-195 could inhibit the cell viability and differentiation of MC3T3-E1 cells. Transfection of miR-195 largely suppressed the L613V-induced viability and osteoblast differentiation of MC3T3-E1 cells and attenuated the accelerative effect of L613V on runt-related transcription factor-2 (Runx2), Osterix (OSX), alkaline phosphatase (ALP), osteocalcin (OCN), and distal-less homeobox 5 (DLX5) osteogenic gene expressions. In addition, miR-195 decreased the expression of RAF-1 mRNA and protein by directly targeting the 3'-untranslated regions (3'-UTR) of RAF-1 mRNA in MC3T3-E1 cells. Our findings indicated that miR-195 inhibited WT and L613V RAF-1 induced hyperactive osteoblast differentiation in MC3T3-E1 cells by targeting RAF-1. miR-195 might be a novel therapeutic agent for the treatment of L613V-induced bone deformity in Noonan syndrome. Copyright © 2017. Published by
Choi, Ra-Yeong; Ham, Ju Ri; Yeo, Jiyoung; Hur, Jae-Seoun; Park, Seok-Kyu; Kim, Myung-Joo; Lee, Mi-Kyung
2017-01-01
Thamnolia vermicularis (TV) is an edible lichen that is prevalent in the alpine zone of East Asia. This study evaluated the feasibility of using TV acetone extracts as a functional food based on experiments using cell line and obese mice. The cellular triglyceride levels and Oil red O staining of 3T3-L1 cells indicated that TV extracts (5 and 10 μg/mL) dose-dependently suppressed adipocyte differentiation and lipid accumulation compared with the control. The TV extract (0.4%, w/w) in a high-fat diet (HFD) was supplemented to C57BL/6N mice for 12 weeks, and TV extract supplement significantly reduced visceral fat mass and body weight compared with HFD feeding alone. The TV extract also induced significant decreases in serum and hepatic lipids, whereas it increased the serum high-density lipoproteins-cholesterol/total cholesterol ratio and fecal lipids levels. Moreover, the TV extract led to significantly lower homeostasis model assessment of insulin resistance in diet-induced obese mice. Taken together, these results suggest that the TV extract may have anti-obesity effects, including lipid-lowering, and it is a natural resource with the potential for use in obesity management. PMID:29333380
Cao, Jiang; Feng, Hao; Ding, Ning-Ning; Wu, Qing-Yun; Chen, Chong; Niu, Ming-Shan; Chen, Wei; Qiu, Ting-Ting; Zhu, Hong-Hu; Xu, Kai-Lin
2016-11-01
Homoharringtonine combined with aclarubicin and cytarabine (HAA) is a highly effective treatment for acute myeloid leukemia (AML), especially for t(8;21) AML. However, the underlying mechanisms by which HAA kills t(8;21) AML cells remain unclear. In this study, SKNO-1 and Kasumi-1 cells with t(8;21) were used. Compared with individual or pairwise administration of homoharringtonine, aclarubicin, or cytarabine, HAA showed the strongest inhibition of growth and induction of apoptosis in SKNO-1 and Kasumi-1 cells. HAA caused cleavage of the AML1-ETO (AE) oncoprotein to form truncated AE (ΔAE). Pretreatment with the caspase-3 inhibitor caspase-3 inhibitor Q-DEVD-OPh (QDO) not only suppressed HAA-induced apoptosis but also abrogated the cleavage of AE and generation of ΔAE. These results suggest that HAA synergistically induces apoptosis in t(8;21) leukemia cells and triggers caspase-3-mediated cleavage of the AML1-ETO oncoprotein, thus providing direct evidence for the strong activity of HAA toward t(8;21) AML. © 2016 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.
Lee, Jee H; Elly, Chris; Park, Yoon; Liu, Yun-Cai
2015-06-16
Foxp3(+) regulatory T (Treg) cells play a critical role in immune homeostasis; however, the mechanisms to maintain their function remain unclear. Here, we report that the E3 ubiquitin ligase VHL is essential for Treg cell function. Mice with Foxp3-restricted VHL deletion displayed massive inflammation associated with excessive Treg cell interferon-γ (IFN-γ) production. VHL-deficient Treg cells failed to prevent colitis induction, but converted into Th1-like effector T cells. VHL intrinsically orchestrated such conversion under both steady and inflammatory conditions followed by Foxp3 downregulation, which was reversed by IFN-γ deficiency. Augmented hypoxia-inducible factor 1α (HIF-1α)-induced glycolytic reprogramming was required for IFN-γ production. Furthermore, HIF-1α bound directly to the Ifng promoter. HIF-1α knockdown or knockout could reverse the increased IFN-γ by VHL-deficient Treg cells and restore their suppressive function in vivo. These findings indicate that regulation of HIF-1α pathway by VHL is crucial to maintain the stability and suppressive function of Foxp3(+) T cells. Copyright © 2015 Elsevier Inc. All rights reserved.
Iraolagoitia, Ximena L Raffo; Spallanzani, Raul G; Torres, Nicolás I; Araya, Romina E; Ziblat, Andrea; Domaica, Carolina I; Sierra, Jessica M; Nuñez, Sol Y; Secchiari, Florencia; Gajewski, Thomas F; Zwirner, Norberto W; Fuertes, Mercedes B
2016-08-01
Despite the classical function of NK cells in the elimination of tumor and of virus-infected cells, evidence for a regulatory role for NK cells has been emerging in different models of autoimmunity, transplantation, and viral infections. However, this role has not been fully explored in the context of a growing tumor. In this article, we show that NK cells can limit spontaneous cross-priming of tumor Ag-specific CD8(+) T cells, leading to reduced memory responses. After challenge with MC57 cells transduced to express the model Ag SIY (MC57.SIY), NK cell-depleted mice exhibited a significantly higher frequency of SIY-specific CD8(+) T cells, with enhanced IFN-γ production and cytotoxic capability. Depletion of NK cells resulted in a CD8(+) T cell population skewed toward an effector memory T phenotype that was associated with enhanced recall responses and delayed tumor growth after a secondary tumor challenge with B16.SIY cells. Dendritic cells (DCs) from NK cell-depleted tumor-bearing mice exhibited a more mature phenotype. Interestingly, tumor-infiltrating and tumor-draining lymph node NK cells displayed an upregulated expression of the inhibitory molecule programmed death ligand 1 that, through interaction with programmed death-1 expressed on DCs, limited DC activation, explaining their reduced ability to induce tumor-specific CD8(+) T cell priming. Our results suggest that NK cells can, in certain contexts, have an inhibitory effect on antitumor immunity, a finding with implications for immunotherapy in the clinic. Copyright © 2016 by The American Association of Immunologists, Inc.
Pusic, Kae M.; Hashimoto, Caryn N.; Lehrer, Axel; Aniya, Charmaine; Clements, David E.; Hui, George S.
2011-01-01
The C-terminal 42 kDa fragments of the P. falciparum Merozoite Surface Protein 1, MSP1-42 is a leading malaria vaccine candidate. MSP1-33, the N-terminal processed fragment of MSP1-42, is rich in T cell epitopes and it is hypothesized that they enhance antibody response toward MSP1-19. Here, we gave in vivo evidence that T cell epitope regions of MSP1-33 provide functional help in inducing anti-MSP1-19 antibodies. Eleven truncated MSP1-33 segments were expressed in tandem with MSP1-19, and immunogenicity was evaluated in Swiss Webster mice and New Zealand White rabbits. Analyses of anti-MSP1-19 antibody responses revealed striking differences in these segments' helper function despite that they all possess T cell epitopes. Only a few fragments induced a generalized response (100%) in outbred mice. These were comparable to or surpassed the responses observed with the full length MSP1-42. In rabbits, only a subset of truncated antigens induced potent parasite growth inhibitory antibodies. Notably, two constructs were more efficacious than MSP1-42, with one containing only conserved T cell epitopes. Moreover, another T cell epitope region induced high titers of non-inhibitory antibodies and they interfered with the inhibitory activities of anti-MSP1-42 antibodies. In mice, this region also induced a skewed TH2 cellular response. This is the first demonstration that T cell epitope regions of MSP1-33 positively or negatively influenced antibody responses. Differential recognition of these regions by humans may play critical roles in vaccine induced and/or natural immunity to MSP1-42. This study provides the rational basis to re-engineer more efficacious MSP1-42 vaccines by selective inclusion and exclusion of MSP1-33 specific T cell epitopes. PMID:21931852
βig-h3 Represses T-Cell Activation in Type 1 Diabetes.
Patry, Maeva; Teinturier, Romain; Goehrig, Delphine; Zetu, Cornelia; Ripoche, Doriane; Kim, In-San; Bertolino, Philippe; Hennino, Ana
2015-12-01
βig-h3/TGF-βi is a secreted protein capable of binding to both extracellular matrix and cells. Human genetic studies recently revealed that in the tgfbi gene encoding for βig-h3, three single nucleotide polymorphisms were significantly associated with type 1 diabetes (T1D) risk. Pancreatic islets express βig-h3 in physiological conditions, but this expression is reduced in β-cell insult in T1D. Since the integrity of islets is destroyed by autoimmune T lymphocytes, we thought to investigate the impact of βig-h3 on T-cell activation. We show here that βig-h3 inhibits T-cell activation markers as well as cytotoxic molecule production as granzyme B and IFN-γ. Furthermore, βig-h3 inhibits early T-cell receptor signaling by repressing the activation of the early kinase protein Lck. Moreover, βig-h3-treated T cells are unable to induce T1D upon transfer in Rag2 knockout mice. Our study demonstrates for the first time that T-cell activation is modulated by βig-h3, an islet extracellular protein, in order to efficiently avoid autoimmune response. © 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.
Differential requirement of RasGRP1 for γδ T cell development and activation
Chen, Yong; Ci, Xinxin; Gorentla, Balachandra; Sullivan, Sarah A.; Stone, James C.; Zhang, Weiguo; Pereira, Pablo; Lu, Jianxin; Zhong, Xiao-Ping
2012-01-01
γδ T cells (γδT) belong to a distinct T cell lineage that performs immune functions different from αβ T cells (αβT). Previous studies have established that Erk1/2 MAPKs are critical for positive selection of αβT cells. Additional evidence also suggests that elevated Erk1/2 activity promotes γδT cell generation. RasGRP1, a guanine nucleotide releasing factor for Ras, plays an important role in positive selection of αβT cells by activating the Ras-Erk1/2 pathway. In this report, we demonstrate that RasGRP1 is critical for TCR-induced Erk1/2 activation in γδT cells but exerts different roles for γδT cell generation and activation. Deficiency of RasGRP1 does not obviously affect γδT cell numbers in the thymus but leads to increased γδT cells, particularly CD4−CD8+ γδT cells, in the peripheral lymphoid organs. The virtually unhindered γδT cell development in the RasGRP1−/− thymus proved to be cell intrinsic, while the increase in CD8+ γδT cells is caused by non-cell-intrinsic mechanisms. Our data provides genetic evidence that decreased Erk1/2 activation in the absence of RasGRP1 is compatible for γδT cell generation. Although RasGRP1 is dispensable for γδT cell generation, RasGRP1-deficient γδT cells are defective in proliferation following TCR stimulation. Additionally, RasGRP1-deficient γδT cells are impaired to produce IL-17 but not IFNγ. Together, these observations have revealed that RasGRP1 plays differential roles for γδ and αβ T cell development but is critical for γδT cell proliferation and production of IL-17. PMID:22623331
Morris, John Henry; Nguyen, Tran; Nwadike, Abuoma; Geels, Mackenzie L; Kamp, Derrick L; Kim, Bo Ram; Boyer, Jean D; Shen, Anding
2017-02-01
In vitro, it is difficult to infect resting CD4 + T cells with human immunodeficiency virus type 1 (HIV), but infections readily occur in vivo. Endothelial cells (ECs) interact with resting CD4 + T cells in vivo, and we found previously that EC stimulation leads to productive and latent HIV infection of resting CD4 + T cells. In this study, we further characterize the interactions between EC and resting T cells. We found that resting CD4 + T cells did not require direct contact with EC for productive and/or latent infection to occur, indicating the involvement of soluble factors. Among 30 cytokines tested in a multiplex enzyme-linked immunosorbent assay (ELISA), we found that expressions for IL-6, IL-8, and CCL2 were much higher in EC-stimulated resting T cells than resting T cells cultured alone. IL-6 was found to be the soluble factor responsible for inducing productive infection of resting T cells, although direct contact with EC had an added effect. However, none of the cytokines tested, IL-6, IL-8, or CCL2, induced additional latent infection in resting T cells, suggesting that unidentified cytokines were involved. Intracellular molecules MURR1, c-Jun N-terminal kinase (JNK), and glucose transporter-1 (GLUT1) were previously shown in blocking HIV infection of resting CD4 + T cells. We found that the concentrations of these proteins were not significantly different in resting T cells before and after stimulation by EC; therefore, they are not likely involved in EC stimulation of resting CD4 + T cells, and a new mechanism is yet to be identified.
Morris, John Henry; Nguyen, Tran; Nwadike, Abuoma; Geels, Mackenzie L.; Kamp, Derrick L.; Kim, Bo Ram; Boyer, Jean D.
2017-01-01
Abstract In vitro, it is difficult to infect resting CD4+ T cells with human immunodeficiency virus type 1 (HIV), but infections readily occur in vivo. Endothelial cells (ECs) interact with resting CD4+ T cells in vivo, and we found previously that EC stimulation leads to productive and latent HIV infection of resting CD4+ T cells. In this study, we further characterize the interactions between EC and resting T cells. We found that resting CD4+ T cells did not require direct contact with EC for productive and/or latent infection to occur, indicating the involvement of soluble factors. Among 30 cytokines tested in a multiplex enzyme-linked immunosorbent assay (ELISA), we found that expressions for IL-6, IL-8, and CCL2 were much higher in EC-stimulated resting T cells than resting T cells cultured alone. IL-6 was found to be the soluble factor responsible for inducing productive infection of resting T cells, although direct contact with EC had an added effect. However, none of the cytokines tested, IL-6, IL-8, or CCL2, induced additional latent infection in resting T cells, suggesting that unidentified cytokines were involved. Intracellular molecules MURR1, c-Jun N-terminal kinase (JNK), and glucose transporter-1 (GLUT1) were previously shown in blocking HIV infection of resting CD4+ T cells. We found that the concentrations of these proteins were not significantly different in resting T cells before and after stimulation by EC; therefore, they are not likely involved in EC stimulation of resting CD4+ T cells, and a new mechanism is yet to be identified. PMID:27599784
Phung, Thuy Thi Bich; Sugamata, Ryuichi; Uno, Kazuko; Aratani, Yasuaki; Ozato, Keiko; Kawachi, Shoji; Thanh Nguyen, Liem; Nakayama, Toshinori; Suzuki, Kazuo
2011-12-01
Influenza virus infection causes severe respiratory disease such as that due to avian influenza (H5N1). Influenza A viruses proliferate in human epithelial cells, which produce inflammatory cytokines/chemokines as a "cytokine storm" attenuated with the viral nonstructural protein 1 (NS1). Cytokine/chemokine production in A549 epithelial cells infected with influenza A/H1N1 virus (PR-8) or nonstructural protein 1 (NS1) plasmid was examined in vitro. Because tumor necrosis factor-α (TNF-α) and regulated upon activation normal T-cell expressed and secreted (RANTES) are predominantly produced from cells infected with PR-8 virus, the effects of mRNA knockdown of these cytokines were investigated. Small interfering (si)TNF-α down-regulated RANTES expression and secretion of RANTES, interleukin (IL)-8, and monocyte chemotactic protein-1 (MCP-1). In addition, siRANTES suppressed interferon (IFN)-γ expression and secretion of RANTES, IL-8, and MCP-1, suggesting that TNF-α stimulates production of RANTES, IL-8, MCP-1, and IFN-γ, and RANTES also increased IL-8, MCP-1, and IFN-γ. Furthermore, administration of TNF-α promoted increased secretion of RANTES, IL-8, and MCP-1. Administration of RANTES enhanced IL-6, IL-8, and MCP-1 production without PR-8 infection. These results strongly suggest that, as an initial step, TNF-α regulates RANTES production, followed by increase of IL-6, IL-8, and MCP-1 and IFNs concentrations. At a later stage, cells transfected with viral NS1 plasmid showed production of a large amount of IL-8 and MCP-1 in the presence of the H(2)O(2)-myeloperoxidse (MPO) system, suggesting that NS1 of PR-8 may induce a "cytokine storm" from epithelial cells in the presence of an H(2)O(2)-MPO system. © 2011 The Societies and Blackwell Publishing Asia Pty Ltd.
T cell-dependent antibody production by Ly-1 B cells.
Taki, S; Schmitt, M; Tarlinton, D; Förster, I; Rajewsky, K
1992-05-04
Through the use of a SCID transfer system, we have demonstrated that under certain conditions, the production of Ig by Ly-1 B cells can be modulated by T cells. This modulation can take the form of enhanced isotype production or isotype-switch induction and to some extent appears to be dependent on the activation state of the T cells. Furthermore we have shown that Ly-1 B cells can mount an idiotypically restricted T cell-dependent immune response to the antigen PC-KLH. This result suggests that the previous failure to observe T cell-dependent responses by Ly-1 B cells has been due to these B cells being "blind" to the antigens used and is not due to some inherent property of these B cells. When one considers the previous reports of the substantial contribution of Ly-1 B cells to the natural serum immunoglobulin levels and the ability of T cells to affect Ig production by Ly-1 B cells documented in this report, it is clear that the interaction of T cells with the Ly-1 B-cell population is important in determining the "natural" serum Ig repertoire of the mouse.
Kassem, Sahar; Gaud, Guillaume; Bernard, Isabelle; Benamar, Mehdi; Dejean, Anne S; Liblau, Roland; Fournié, Gilbert J; Colacios, Céline; Malissen, Bernard; Saoudi, Abdelhadi
2016-07-01
The guanine nucleotide exchange factor Vav1 is essential for transducing T cell antigen receptor signals and therefore plays an important role in T cell development and activation. Our previous genetic studies identified a locus on rat chromosome 9 that controls the susceptibility to neuroinflammation and contains a non-synonymous polymorphism in the major candidate gene Vav1. To formally demonstrate the causal implication of this polymorphism, we generated a knock-in mouse bearing this polymorphism (Vav1R63W). Using this model, we show that Vav1R63W mice display reduced susceptibility to experimental autoimmune encephalomyelitis (EAE) induced by MOG35-55 peptide immunization. This is associated with a lower production of effector cytokines (IFN-γ, IL-17 and GM-CSF) by autoreactive CD4 T cells. Despite increased proportion of Foxp3+ regulatory T cells in Vav1R63W mice, we show that this lowered cytokine production is intrinsic to effector CD4 T cells and that Treg depletion has no impact on EAE development. Finally, we provide a mechanism for the above phenotype by showing that the Vav1R63W variant has normal enzymatic activity but reduced adaptor functions. Together, these data highlight the importance of Vav1 adaptor functions in the production of inflammatory cytokines by effector T cells and in the susceptibility to neuroinflammation.
Kassem, Sahar; Bernard, Isabelle; Dejean, Anne S.; Liblau, Roland; Fournié, Gilbert J.; Colacios, Céline
2016-01-01
The guanine nucleotide exchange factor Vav1 is essential for transducing T cell antigen receptor signals and therefore plays an important role in T cell development and activation. Our previous genetic studies identified a locus on rat chromosome 9 that controls the susceptibility to neuroinflammation and contains a non-synonymous polymorphism in the major candidate gene Vav1. To formally demonstrate the causal implication of this polymorphism, we generated a knock-in mouse bearing this polymorphism (Vav1R63W). Using this model, we show that Vav1R63W mice display reduced susceptibility to experimental autoimmune encephalomyelitis (EAE) induced by MOG35-55 peptide immunization. This is associated with a lower production of effector cytokines (IFN-γ, IL-17 and GM-CSF) by autoreactive CD4 T cells. Despite increased proportion of Foxp3+ regulatory T cells in Vav1R63W mice, we show that this lowered cytokine production is intrinsic to effector CD4 T cells and that Treg depletion has no impact on EAE development. Finally, we provide a mechanism for the above phenotype by showing that the Vav1R63W variant has normal enzymatic activity but reduced adaptor functions. Together, these data highlight the importance of Vav1 adaptor functions in the production of inflammatory cytokines by effector T cells and in the susceptibility to neuroinflammation. PMID:27438086
Jung, T.W.; Lee, K.T.; Lee, M.W.; Ka, K.H.
2012-01-01
Endoplasmic reticulum (ER) stress has been implicated in the pathology of type 2 diabetes mellitus (T2DM). Although SIRT1 has a therapeutic effect on T2DM, the mechanisms by which SIRT1 ameliorates insulin resistance (IR) remain unclear. In this study, we investigated the impact of SIRT1 on palmitate-induced ER stress in HepG2 cells and its underlying signal pathway. Treatment with resveratrol, a SIRT1 activator significantly inhibited palmitate-induced ER stress, leading to the protection against palmitate-induced ER stress and insulin resistance. Resveratrol and SIRT1 overexpression induced the expression of oxygen-regulated protein (ORP) 150 in HepG2 cells. Forkhead box O1 (FOXO1) was involved in the regulation of ORP150 expression because suppression of FOXO1 inhibited the induction of ORP150 by SIRT1. Our results indicate a novel mechanism by which SIRT1 regulates ER stress by overexpression of ORP150, and suggest that SIRT1 ameliorates palmitate-induced insulin resistance in HepG2 cells via regulation of ER stress.
Chae, Wook-Jin; Park, Jong-Hyun; Henegariu, Octavian; Yilmaz, Saliha; Hao, Liming; Bothwell, Alfred L M
2017-10-01
Induction of tolerance is a key mechanism to maintain or to restore immunological homeostasis. Here we show that Foxp3 + regulatory T (Treg) cells use Dickkopf-1 (DKK-1) to regulate T-cell-mediated tolerance in the T-cell-mediated autoimmune colitis model. Treg cells from DKK-1 hypomorphic doubleridge mice failed to control CD4 + T-cell proliferation, resulting in CD4 T-cell-mediated autoimmune colitis. Thymus-derived Treg cells showed a robust expression of DKK-1 but not in naive or effector CD4 T cells. DKK-1 expression in Foxp3 + Treg cells was further increased upon T-cell receptor stimulation in vitro and in vivo. Interestingly, Foxp3 + Treg cells expressed DKK-1 in the cell membrane and the functional inhibition of DKK-1 using DKK-1 monoclonal antibody abrogated the suppressor function of Foxp3 + Treg cells. DKK-1 expression was dependent on de novo protein synthesis and regulated by the mitogen-activated protein kinase pathway but not by the canonical Wnt pathway. Taken together, our results highlight membrane-bound DKK-1 as a novel Treg-derived mediator to maintain immunological tolerance in T-cell-mediated autoimmune colitis. © 2017 The Authors. Immunology Published by John Wiley & Sons Ltd.
Anipindi, Varun C.; Dizzell, Sara E.; Nguyen, Philip V.; Shaler, Christopher R.; Chu, Derek K.; Jiménez-Saiz, Rodrigo; Liang, Hong; Swift, Stephanie; Nazli, Aisha; Kafka, Jessica K.; Bramson, Jonathan; Xing, Zhou; Jordana, Manel; Wan, Yonghong; Snider, Denis P.; Stampfli, Martin R.; Kaushic, Charu
2016-01-01
Clinical and experimental studies have shown that estradiol (E2) confers protection against HIV and other sexually transmitted infections. Here, we investigated the underlying mechanism. Better protection in E2-treated mice, immunized against genital HSV-2, coincided with earlier recruitment and higher proportions of Th1 and Th17 effector cells in the vagina post-challenge, compared to placebo-treated controls. Vaginal APCs isolated from E2-treated mice induced 10-fold higher Th17 and Th1 responses, compared to APCs from progesterone-treated, placebo-treated, and estradiol-receptor knockout mice in APC-T cell co-cultures. CD11c+ DCs in the vagina were the predominant APC population responsible for priming these Th17 responses, and a potent source of IL-6 and IL-1β, important factors for Th17 differentiation. Th17 responses were abrogated in APC-T cell co-cultures containing IL-1β KO, but not IL-6 KO vaginal DCs, showing that IL-1β is a critical factor for Th17 induction in the genital tract. E2 treatment in vivo directly induced high expression of IL-1β in vaginal DCs, and addition of IL-1β restored Th17 induction by IL-1β KO APCs in co-cultures. Finally, we examined the role of IL-17 in anti-HSV-2 memory T cell responses. IL-17 KO mice were more susceptible to intravaginal HSV-2 challenge, compared to WT controls, and vaginal DCs from these mice were defective at priming efficient Th1 responses in vitro, indicating that IL-17 is important for the generation of efficient anti-viral memory responses. We conclude that the genital mucosa has a unique microenvironment whereby E2 enhances CD4+ T cell anti-viral immunity by priming vaginal DCs to induce Th17 responses through an IL-1-dependent pathway. PMID:27148737
Simon, Priscilla S.; Bardhan, Kankana; Chen, May R.; Paschall, Amy V.; Lu, Chunwan; Bollag, Roni J.; Kong, Feng-Chong; Jin, JianYue; Kong, Feng-Ming; Waller, Jennifer L.; Pollock, Raphael E.; Liu, Kebin
2016-01-01
Radiation modulates both tumor cells and immune cells in the tumor microenvironment to exert its anti-tumor activity; however, the molecular connection between tumor cells and immune cells that mediates radiation-exerted tumor suppression activity in the tumor microenvironment is largely unknown. We report here that radiation induces rapid activation of the p65/p50 and p50/p50 NF-κB complexes in human soft tissue sarcoma (STS) cells. Radiation-activated p65/p50 and p50/p50 bind to the TNFα promoter to activate its transcription in STS cells. Radiation-induced TNFα induces tumor cell death in an autocrine manner. A sublethal dose of Smac mimetic BV6 induces cIAP1 and cIAP2 degradation to increase tumor cell sensitivity to radiation-induced cell death in vitro and to enhance radiation-mediated suppression of STS xenografts in vivo. Inhibition of caspases, RIP1, or RIP3 blocks radiation/TNFα-induced cell death, whereas inhibition of RIP1 blocks TNFα-induced caspase activation, suggesting that caspases and RIP1 act sequentially to mediate the non-compensatory cell death pathways. Furthermore, we determined in a syngeneic sarcoma mouse model that radiation up-regulates IRF3, IFNβ, and the T cell chemokines CCL2 and CCL5 in the tumor microenvironment, which are associated with activation and increased infiltration of Th1/Tc1 T cells in the tumor microenvironment. Moreover, tumor-infiltrating T cells are in their active form since both the perforin and FasL pathways are activated in irradiated tumor tissues. Consequently, combined BV6 and radiation completely suppressed tumor growth in vivo. Therefore, radiation-induced NF-κB functions as a molecular link between tumor cells and immune cells in the tumor microenvironment for radiation-mediated tumor suppression. PMID:27014915
1983-01-01
Glucocorticosteroids (GCS) added to otherwise unstimulated cultures of human peripheral blood mononuclear cells (PBMC) induce the synthesis and secretion of all classes of immunoglobulin. The magnitude of this response is similar to that seen with other polyclonal B cell activators such as pokeweed mitogen (PWM), and like that of PWM, the steroid effect is dependent on both T cells and monocytes. To determine the cellular target for GCS in these cultures, separated populations of T cells and non-T cells were preincubated with steroids and then recombined. No immunoglobulin was produced in any of these preincubation experiments. As a different approach to this question, supernatants were collected from various cell populations following stimulation with PWM, concanavalin A (Con A), phytohemagglutinin (PHA), alloantigens, or GCS. These supernatants were tested for their effects on GCS-induced Ig production by B cells. Supernatants from 3-d cultures of unstimulated, as well as GCS-treated, PBMC contained a T cell- replacing factor that permitted T-depleted PBMC to produce Ig upon steroid stimulation. This supernatant factor (TRF-S) could be produced in the absence of steroid stimulation, but both the factor and GCS were necessary for the induction of Ig synthesis. Production of the TRF-S required the presence of both T cells and adherent cells in culture and was found in the highest concentrations at 3-4 d of culture. Supernatants from cultures stimulated with PWM, PHA, Con A, and alloantigens did not contain detectable TRF-S activity, and TRF-S was unable to replace helper T cells for PWM-induced Ig production. TRF-S required the presence of adherent cells in the T cell-depleted responder population for its action. Further, it was effective in inducing Ig production along with GCS in the presence of a sufficient concentration of cyclosporin A to block all T cell helper activity for primary responses of PBMC to PWM or GCS. TRF-S was inactivated by trypsin treatment
Di Rosso, María Emilia; Barreiro Arcos, María Laura; Elingold, Igal; Sterle, Helena; Baptista Ferreira, Sabrina; Ferreira, Vitor Francisco; Galleano, Mónica; Cremaschi, Graciela; Dubin, Marta
2013-10-01
Novel β-lapachone analogs 2-phenyl-3,4-dihydro-2H-benzo[h]chromene-5,6-dione (NQ1), 2-p-tolyl-3,4-dihydro-2H-benzo[h]chromene-5,6-dione (NQ3) and 2-methyl-2-phenyl-3,4-dihydro-2H-benzo[h]chromene-5,6-dione (NQ7), which have trypanocidal activity, were assayed for cytotoxic effects on murine EL-4 T lymphoma cells. The NQs inhibited the proliferation of EL-4 cells at concentrations above 1μM. Nuclear staining of the EL-4 cells revealed chromatin condensation and a nuclear morphology compatible with the induction of apoptosis. Flow cytometry assays with annexin V-FITC and propidium iodide confirmed the cell death by apoptosis. Using electron paramagnetic resonance (EPR), a semiquinone radical was detected in EL-4 cells treated with NQs. In addition, a decrease in the GSH level in parallel with reactive oxygen species (ROS) production was observed. Preincubation with n-acetyl-l-cysteine (NAC) was able to reverse the inhibitory effects of the NQs on cell proliferation, indicating that ROS generation is involved in NQ-induced apoptosis. In addition, the NQs induced a decrease in the mitochondrial membrane potential and increased the proteolytic activation of caspases 9 and 3 and the cleavage of Poly (ADP-Ribose) Polymerase (PARP). In conclusion, these results indicate that redox cycling is induced by the NQs in the EL-4 cell line, with the generation of ROS and other free radicals that could inhibit cellular proliferation as a result of the induction of the intrinsic apoptosis pathway. Copyright © 2013 Elsevier Ltd. All rights reserved.
Licochalcone C induces apoptosis via B-cell lymphoma 2 family proteins in T24 cells.
Wang, Penglong; Yuan, Xuan; Wang, Yan; Zhao, Hong; Sun, Xiling; Zheng, Qiusheng
2015-11-01
The current study investigated the mechanisms by which licochalcone C induces apoptosis of T24 human malignant bladder cancer cells. Cell viability was evaluated using an MTT assay. Apoptosis was investigated using a morphological assay, flow cytometry and a caspase‑3 activity assay. Alterations in the gene expression levels of Bcl‑2 family members were measured by semi‑quantitative reverse transcription‑polymerase chain reaction assays. The protein levels of pro‑caspase‑3 and cleaved poly(ADP ribose) polymerase were measured using western blotting. The results indicated that licochalcone C induced T24 cell apoptosis in a concentration‑dependent manner. Licochalcone C treatment reduced the levels of the anti‑apoptotic mRNAs (Bcl‑2, Bcl‑w and Bcl‑XL) and increased expression of the pro‑apoptotic mRNAs (Bax and Bim). The Bcl‑2 family inhibitor (ABT‑737) reduced apoptosis induced by licochalcone C in T24 cells. The current study demonstrated that licochalcone C may be a potential adjuvant therapeutic agent for bladder cancer.
Luo, Wen-Hui; Yang, Ya-Wun
2016-04-01
The aim of this study was to investigate the induction of antigen-specific T cell activation and cell cycle modulation by a poly-DL-lactide/glycolide (PLGA) nanoparticle (NP)-primed CD11b(+)Gr-1(high) subset isolated from mouse bone marrow. PLGA NPs containing the ovalbumin (OVA) antigen were prepared using the double emulsion and solvent evaporation method, and protein release rate and cell viability were determined. The Lin2(¯)CD11b(+)Gr-1(high)Ly6c(low) (Gr-1(high)) subset was sorted from the bone marrow of C57BL/6 J mice by fluorescence-activated cell sorting (FACS) and co-cultured with OT-I CD8(+) splenic T cells. Proliferation of OT-I CD8(+) T cells was monitored, and cell cycles were determined by 5-bromo-2'-deoxyuridine (BrdU) labeling. Treatment of Gr-1(high) cells with PLGA/OVA NPs upregulated expression of the SIINFEKL-H2K(b) complex in the context of MHC I. Co-cultures of OT-I CD8(+) T cells with the PLGA/OVA NP-primed Gr-1(high) cells induced the proliferation of T cells in vitro and modulated cell division and morphology. Treatment of Gr-1(high) cells with PLGA/OVA NPs also induced cell apoptosis and necrosis. This study demonstrated the function of PLGA/OVA NPs in the activation of OT-I CD8(+) T cells and the capability of cross-presentation via the Gr-1(high) polymorphonuclear subset from mouse bone marrow.
Soares, Kevin C.; Rucki, Agnieszka A.; Wu, Annie A.; Olino, Kelly; Xiao, Qian; Chai, Yi; Wamwea, Anthony; Bigelow, Elaine; Lutz, Eric; Liu, Linda; Yao, Sheng; Anders, Robert A.; Laheru, Daniel; Wolfgang, Christopher L.; Edil, Barish H.; Schulick, Richard D.; Jaffee, Elizabeth M.; Zheng, Lei
2014-01-01
Pancreatic ductal adenocarcinoma (PDA) has a poor prognosis due to late detection and resistance to conventional therapies. Published studies show that the PDA tumor microenvironment (TME) is predominantly infiltrated with immune suppressive cells and signals that if altered, would allow effective immunotherapy. However, single-agent checkpoint inhibitors including agents that alter immune suppressive signals in other human cancers such as cytotoxic T lymphocyte antigen-4 (CTLA-4), programmed death 1 (PD-1) and its ligand PD-L1, have failed to demonstrate objective responses when given as single agents to PDA patients. We recently reported that inhibition of the CTLA-4 pathway when given together with a T cell inducing vaccine gives objective responses in metastatic PDA patients. In this study, we evaluated blockade of the PD-1/PD-L1 pathway. We found that PD-L1 is weakly expressed at a low frequency in untreated human and murine PDAs but treatment with a GM-CSF secreting PDA vaccine (GVAX) significantly upregulates PD-L1 membranous expression after treatment of tumor bearing mice. In addition, combination therapy with vaccine and PD-1 antibody blockade improved murine survival compared to PD-1 antibody monotherapy or GVAX therapy alone. Furthermore, PD-1 blockade increased effector CD8+ T lymphocytes and tumor-specific interferon-γ production of CD8+ T cells in the TME. Immunosuppressive pathways, including regulatory T cells (Tregs) and CTLA-4 expression on T cells were overcome by the addition of vaccine and low dose cyclophosphamide to PD-1 blockade. Collectively, our study supports combining PD-1 or PD-L1 antibody therapy with a T cell inducing agent for PDA treatment. PMID:25415283
DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhang, Han; Sonoda, Koh-Hei, E-mail: sonodak@med.kyushu-u.ac.jp; Hijioka, Kuniaki
2009-04-17
Ocular neovascularization (NV) is the primary cause of blindness in a wide range of ocular diseases. The exact mechanism underlying the pathogenesis of ocular NV is not yet well understood, and so there is no satisfactory therapy for ocular NV. Here, we describe a strategy targeting Flk-1, a self-antigen overexpressed on proliferating endothelial cells in ocular NV, by antiangiogenic immunotherapy-DNA vaccine and adoptive T cell therapy. An oral DNA vaccine encoding Flk-1 carried by attenuated Salmonella typhimurium markedly suppressed development of laser-induced choroidal NV. We further demonstrated that adoptive transfer of vaccine-induced CD8{sup +} T cells reduced pathological preretinal NV,more » with a concomitant facilitation of physiological revascularization after oxygen-induced retinal vessel obliteration. However, physiological retinal vascular development was unaffected in neonatal mice transferred with vaccine-induced CD8{sup +} T cells. These findings suggested that antiangiogenic immunotherapy targeting Flk-1 such as vaccination and adoptive immunotherapy may contribute to future therapies for ocular NV.« less
Mishra, Ameet K; Kadoishi, Tanya; Wang, Xiaoguang; Driver, Emily; Chen, Zhangguo; Wang, Xiao-Jing; Wang, Jing H
2016-12-06
Squamous cell carcinoma (SCC) is the second commonest type of skin cancer. Moreover, about 90% of head and neck cancers are SCCs. SCCs develop at a significantly higher rate under chronic immunosuppressive conditions, implicating a role of immune surveillance in controlling SCCs. It remains largely unknown how SCCs evade immune recognition. Here, we established a mouse model by injecting tumor cells derived from primary SCCs harboring KrasG12D mutation and Smad4 deletion into wild-type (wt) or CD8-/- recipients. We found comparable tumor growth between wt and CD8-/- recipients, indicating a complete escape of CD8+ T cell-mediated anti-tumor responses by these SCCs. Mechanistically, CD8+ T cells apparently were not defective in infiltrating tumors given their relatively increased percentage among tumor infiltrating lymphocytes (TILs). CD8+ TILs exhibited phenotypes of chronic activation and exhaustion, including overexpression of activation markers, co-expression of programmed cell death 1 (PD-1) and lymphocyte activation gene-3 (LAG-3), as well as TCRβ downregulation. Among CD4+ TILs, T regulatory cells (Tregs) were preferentially expanded. Contradictory to prior findings in melanoma, Treg expansion was independent of CD8+ T cells in our SCC model. Unexpectedly, CD8+ T cells were required for promoting NK cell infiltration within SCCs. Furthermore, we uncovered AKT-dependent lymphocyte-induced PD-L1 upregulation on SCCs, which was contributed greatly by combinatorial effects of CD8+ T and NK cells. Lastly, dual blockade of PD-1 and LAG-3 inhibited the tumor growth of SCCs. Thus, our findings identify novel immune evasion mechanisms of SCCs and suggest that immunosuppressive mechanisms operate in a cancer-type specific and context-dependent manner.
Takada, Honami; Imadome, Ken-Ichi; Shibayama, Haruna; Yoshimori, Mayumi; Wang, Ludan; Saitoh, Yasunori; Uota, Shin; Yamaoka, Shoji; Koyama, Takatoshi; Shimizu, Norio; Yamamoto, Kouhei; Fujiwara, Shigeyoshi; Miura, Osamu; Arai, Ayako
2017-01-01
Epstein-Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-κB activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-κB activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-κB activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-κB activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-κB that suppresses NF-κB activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-κB-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells.
Shibayama, Haruna; Yoshimori, Mayumi; Wang, Ludan; Saitoh, Yasunori; Uota, Shin; Yamaoka, Shoji; Koyama, Takatoshi; Shimizu, Norio; Yamamoto, Kouhei; Fujiwara, Shigeyoshi; Miura, Osamu
2017-01-01
Epstein–Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-κB activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-κB activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-κB activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-κB activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-κB that suppresses NF-κB activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-κB-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells. PMID:28346502
Shorey, Lyndsey E.; Hagman, Amanda M.; Williams, David E.; Ho, Emily; Dashwood, Roderick H.; Benninghoff, Abby D.
2012-01-01
Certain bioactive food components, including indole-3-carbinol (I3C) and 3,3′-diindolylmethane (DIM) from cruciferous vegetables, have been shown to target cellular pathways regulating carcinogenesis. Previously, our laboratory showed that dietary I3C is an effective transplacental chemopreventive agent in a dibenzo[def,p]chrysene (DBC)-dependent model of murine T-cell lymphoblastic lymphoma. The primary objective of the present study was to extend our chemoprevention studies in mice to an analogous human neoplasm in cell culture. Therefore, we tested the hypothesis that I3C or DIM may be chemotherapeutic in human T-cell acute lymphoblastic leukemia (T-ALL) cells. Treatment of the T-ALL cell lines CCRF-CEM, CCRF-HSB2, SUP-T1 and Jurkat with DIM in vitro significantly reduced cell proliferation and viability at concentrations 8- to 25-fold lower than the parent compound I3C. DIM (7.5 µM) arrested CEM and HSB2 cells at the G1 phase of the cell cycle and 15 µM DIM significantly increased the percentage of apoptotic cells in all T-ALL lines. In CEM cells, DIM reduced protein expression of cyclin dependent kinases 4 and 6 (CDK4, CDK6) and D-type cyclin 3 (CCND3); DIM also significantly altered expression of eight transcripts related to human apoptosis (BCL2L10, CD40LG, HRK, TNF, TNFRSF1A, TNFRSF25, TNFSF8, TRAF4). Similar anticancer effects of DIM were observed in vivo. Dietary exposure to 100 ppm DIM significantly decreased the rate of growth of human CEM xenografts in immunodeficient SCID mice, reduced final tumor size by 44% and increased the apoptotic index compared to control-fed mice. Taken together, our results demonstrate a potential for therapeutic application of DIM in T-ALL. PMID:22514694
Cheung, Ka-Wai; Wu, Tongjin; Ho, Sai Fan; Wong, Yik Chun; Liu, Li; Wang, Hui; Chen, Zhiwei
2018-04-15
HIV-1 transmission occurs mainly through mucosal tissues. During mucosal transmission, HIV-1 preferentially infects α 4 β 7 + gut-homing CCR7 - CD4 + effector/effector memory T cells (T EM ) and results in massive depletion of these cells and other subsets of T EM in gut-associated lymphoid tissues. However, besides being eliminated by HIV-1, the role of T EM during the early stage of infection remains inconclusive. Here, using in vitro -induced α 4 β 7 + gut-homing T EM (α 4 β 7 + T EM ), we found that α 4 β 7 + T EM differentiated into CCR7 + CD4 + central memory T cells (T CM ). This differentiation was HIV-1 independent but was inhibited by SB431542, a specific transforming growth factor β (TGF-β) receptor I kinase inhibitor. Consistently, T EM -to-T CM differentiation was observed in α 4 β 7 + T EM stimulated with TGF-β1 (TGF-β). The T CM properties of the TGF-β-induced T EM -derived T CM (α 4 β 7 + T CM ) were confirmed by their enhanced CCL19 chemotaxis and the downregulation of surface CCR7 upon T cell activation in vitro Importantly, the effect of TGF-β on T CM differentiation also held in T EM directly isolated from peripheral blood. To investigate the significance of the TGF-β-dependent T EM -to-T CM differentiation in HIV/AIDS pathogenesis, we observed that both productively and latently infected α 4 β 7 + T CM could differentiate from α 4 β 7 + T EM in the presence of TGF-β during HIV-1 infection. Collectively, this study not only provides a new insight for the plasticity of T EM but also suggests that the TGF-β-dependent T EM -to-T CM differentiation is a previously unrecognized mechanism for the formation of latently infected T CM after HIV-1 infection. IMPORTANCE HIV-1 is the causative agent of HIV/AIDS, which has led to millions of deaths in the past 30 years. Although the implementation of highly active antiretroviral therapy has remarkably reduced the HIV-1-related morbidity and mortality, HIV-1 is not eradicated in
Fonseca, A M; Porto, G; Uchida, K; Arosa, F A
2001-05-15
Red blood cells (RBCs) are known to perform one prominent function: to carry and deliver oxygen to the tissues. Earlier studies, however, suggested a role for RBCs in potentiating T-cell proliferation in vitro. Here it is shown that the presence of RBCs in cultures of stimulated human peripheral blood lymphocytes strengthens T-cell proliferation and survival. Analysis of phosphatidylserine externalization and DNA fragmentation showed that RBCs inhibit T-cell apoptosis. This inhibition correlated with a reduction in CD71 but not CD95 expression. RBCs enhanced T-cell proliferation and survival upon activation with phytohemagglutinin and with OKT3 antibodies. Studies aimed at characterizing the cellular and molecular basis of the protection afforded to T cells by RBCs showed that (1) optimal protection required intact RBCs and red cell/T-cell contact but not monocytes; (2) RBCs markedly reduced the level of intracellular reactive oxygen species; and (3) RBCs inhibited the formation of protein-bound acrolein, a peroxidation adduct in biologic systems. Overall, these data indicate that human RBCs protect T cells from activation-induced cell death, at least in part by reducing the pro-oxidant state, and suggest a role for RBCs as conceivable modulators of T-cell homeostasis.
Characterization of the T-cell response to Dau c 1, the Bet v 1-homolog in carrot.
Zulehner, N; Nagl, B; Briza, P; Roulias, A; Ballmer-Weber, B; Zlabinger, G J; Ferreira, F; Bohle, B
2017-02-01
In contrast to other Bet v 1-related food allergens, the major carrot allergen, Dau c 1, has been suggested to induce food allergy independently from Bet v 1. As T cells are crucial in the sensitization process, we sought to characterize the T-cell response to Dau c 1 and its cross-reactivity with Bet v 1. Dau c 1-specific T-cell lines (TCL) and clones (TCC) established from PBMC of birch pollen-allergic patients with carrot allergy were analyzed for reactivity to Bet v 1, epitope specificity, allergen-induced cytokine secretion, and expression of integrins α4β7 and α4β1, critical for gut and lung homing, respectively. mRNA expression of GATA3 and Tbet was analyzed in sorted CD3 + CD4 + CFSE low cells proliferating upon stimulation of PBMC with Dau c 1 or Bet v 1. Dau c 1 was incubated with endolysosomal proteases, and the resulting fragments were identified by mass spectrometry. Among 14 distinct T-cell-activating regions, Dau c 1 139-153 was recognized by 55% of the patients. Only 6 of 15 (40%) Dau c 1-specific TCL and 9 of 21 (43%) TCC reacted with Bet v 1. Bet v 1-nonreactive TCC were mainly Th1-like and showed a higher expression of the integrin β7 and a significantly lower expression of the integrin β1 than Bet v 1-positive TCC. A Th1-like response was also detected in Dau c 1-reactive CD3 + CD4 + CFSE low cells. Full-length Dau c 1 was still detectable after 48 h of endolysosomal degradation. Proteolytic fragments of Dau c 1 matched its T-cell-activating regions. Dau c 1 displays several characteristics of sensitizing allergens, namely a major T-cell-activating region, low susceptibility to endolysosomal degradation, and induction of a Bet v 1-independent T-cell response. These cellular insights confirm that the major carrot allergen has a special status among Bet v 1-related food allergens. © 2016 The Authors. Allergy Published by John Wiley & Sons Ltd.
Biburger, Markus; Theiner, Gabi; Schädle, Mirjam; Schuler, Gerold; Tiegs, Gisa
2010-02-01
HO-1 is the only inducible one of three isoenzymes that catalyzes the oxidative degradation of heme. HO-1 is inducible by various cellular stress factors and exerts cytoprotective and immunomodulatory effects. Recent publications demonstrated that HO-1 is constitutively expressed by CD4(+)CD25(+) T(regs) and induced in CD4(+)CD25(-) T cells upon FoxP3 transfection. Here, we investigated whether HO-1 was essential and sufficient for human T(regs) to exert immunosuppression in vitro. PGJ(2) induced pronounced expression of HO-1 in CD4(+)CD25(-) T cells without accompanying FoxP3 induction. Treatment of CD4(+)CD25(-) T cells with PGJ(2) decreased their proliferation, whereas the HO-1 inhibitor SnPP enhanced the proliferation of HO-1-expressing T(regs), suggesting that HO-1 may modulate the proliferative capacity of T lymphocytes. HO-1 modulation by SnPP treatment of T(regs) or PGJ(2) treatment of CD4(+)CD25(-) T cells neither suppressed nor induced immune-modulatory function in these cells, respectively, as measured by responder-cell proliferation and/or IL-2 production. In summary, these data suggest that HO-1 expression by T(regs) might contribute to their typical reluctance to proliferate but does not account independently for their suppressive functions.
Stephen, Tom L; Rutkowski, Melanie R; Allegrezza, Michael J; Perales-Puchalt, Alfredo; Tesone, Amelia J; Svoronos, Nikolaos; Nguyen, Jenny M; Sarmin, Fahmida; Borowsky, Mark E; Tchou, Julia; Conejo-Garcia, Jose R
2014-09-18
Tumor-reactive T cells become unresponsive in advanced tumors. Here we have characterized a common mechanism of T cell unresponsiveness in cancer driven by the upregulation of the transcription factor Forkhead box protein P1 (Foxp1), which prevents CD8⁺ T cells from proliferating and upregulating Granzyme-B and interferon-γ in response to tumor antigens. Accordingly, Foxp1-deficient lymphocytes induced rejection of incurable tumors and promoted protection against tumor rechallenge. Mechanistically, Foxp1 interacted with the transcription factors Smad2 and Smad3 in preactivated CD8⁺ T cells in response to microenvironmental transforming growth factor-β (TGF-β), and was essential for its suppressive activity. Therefore, Smad2 and Smad3-mediated c-Myc repression requires Foxp1 expression in T cells. Furthermore, Foxp1 directly mediated TGF-β-induced c-Jun transcriptional repression, which abrogated T cell activity. Our results unveil a fundamental mechanism of T cell unresponsiveness different from anergy or exhaustion, driven by TGF-β signaling on tumor-associated lymphocytes undergoing Foxp1-dependent transcriptional regulation. Copyright © 2014 Elsevier Inc. All rights reserved.
Datta, Antara; Silverman, Lee; Phipps, Andrew J; Hiraragi, Hajime; Ratner, Lee; Lairmore, Michael D
2007-01-01
Background Human T-lymphotropic virus type-1 (HTLV-1) causes adult T-cell leukemia/lymphoma and is linked to a number of lymphocyte-mediated disorders. HTLV-1 contains both regulatory and accessory genes in four pX open reading frames. pX ORF-II encodes two proteins, p13 and p30, whose roles are still being defined in the virus life cycle and in HTLV-1 virus-host cell interactions. Proviral clones of HTLV-1 with pX ORF-II mutations diminish the ability of the virus to maintain viral loads in vivo. p30 expressed exogenously differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein/p300 and while acting as a repressor of many genes including Tax, in part by blocking tax/rex RNA nuclear export, selectively enhances key gene pathways involved in T-cell signaling/activation. Results Herein, we analyzed the role of p30 in cell cycle regulation. Jurkat T-cells transduced with a p30 expressing lentivirus vector accumulated in the G2-M phase of cell cycle. We then analyzed key proteins involved in G2-M checkpoint activation. p30 expression in Jurkat T-cells resulted in an increase in phosphorylation at serine 216 of nuclear cell division cycle 25C (Cdc25C), had enhanced checkpoint kinase 1 (Chk1) serine 345 phosphorylation, reduced expression of polo-like kinase 1 (PLK1), diminished phosphorylation of PLK1 at tyrosine 210 and reduced phosphorylation of Cdc25C at serine 198. Finally, primary human lymphocyte derived cell lines immortalized by a HTLV-1 proviral clone defective in p30 expression were more susceptible to camptothecin induced apoptosis. Collectively these data are consistent with a cell survival role of p30 against genotoxic insults to HTLV-1 infected lymphocytes. Conclusion Collectively, our data are the first to indicate that HTLV-1 p30 expression results in activation of the G2-M cell cycle checkpoint, events that would promote early viral spread and T-cell survival. PMID:17634129
Early T Cell Signalling Is Reversibly Altered in PD-1+ T Lymphocytes Infiltrating Human Tumors
Wang, Shu-Fang; Fouquet, Stéphane; Chapon, Maxime; Salmon, Hélène; Regnier, Fabienne; Labroquère, Karine; Badoual, Cécile; Damotte, Diane; Validire, Pierre; Maubec, Eve; Delongchamps, Nicolas B.; Cazes, Aurélie; Gibault, Laure; Garcette, Marylène; Dieu-Nosjean, Marie-Caroline; Zerbib, Marc; Avril, Marie-Françoise; Prévost-Blondel, Armelle; Randriamampita, Clotilde; Trautmann, Alain; Bercovici, Nadège
2011-01-01
To improve cancer immunotherapy, a better understanding of the weak efficiency of tumor-infiltrating T lymphocytes (TIL) is necessary. We have analyzed the functional state of human TIL immediately after resection of three types of tumors (NSCLC, melanoma and RCC). Several signalling pathways (calcium, phosphorylation of ERK and Akt) and cytokine secretion are affected to different extents in TIL, and show a partial spontaneous recovery within a few hours in culture. The global result is an anergy that is quite distinct from clonal anergy induced in vitro, and closer to adaptive tolerance in mice. PD-1 (programmed death -1) is systematically expressed by TIL and may contribute to their anergy by its mere expression, and not only when it interacts with its ligands PD-L1 or PD-L2, which are not expressed by every tumor. Indeed, the TCR-induced calcium and ERK responses were reduced in peripheral blood T cells transfected with PD-1. Inhibition by sodium stibogluconate of the SHP-1 and SHP-2 phosphatases that associate with several inhibitory receptors including PD-1, relieves part of the anergy apparent in TIL or in PD-1-transfected T cells. This work highlights some of the molecular modifications contributing to functional defects of human TIL. PMID:21408177
B cells and TCR avidity determine distinct functions of CD4+ T cells in retroviral infection1
Ploquin, Mickaël J-Y; Eksmond, Urszula; Kassiotis, George
2011-01-01
The T-cell-dependent B-cell response relies on cognate interaction between B cells and CD4+ Th cells. However, the consequences of this interaction for CD4+ T cells are not entirely known. B cells generally promote CD4+ T-cell responses to pathogens, albeit to a variable degree. In contrast, CD4+ T-cell responses to self or tumor antigens are often suppressed by B cells. Here we demonstrated that interaction with B cells dramatically inhibited the function of virus-specific CD4+ T cells in retroviral infection. We have used Friend virus (FV) infection of mice as a model for retroviral infection, in which the behavior of virus-specific CD4+ T cells was monitored according to their TCR avidity. We report that avidity for antigen and interaction with B cells determine distinct aspects of the primary CD4+ T-cell response to FV infection. Virus-specific CD4+ T cells followed exclusive Th1 and T follicular helper (Tfh) differentiation. High avidity for antigen facilitated expansion during priming and enhanced the capacity for IFN-γ and IL-21 production. In contrast, Tfh differentiation was not affected by avidity for antigen. By reducing or preventing B-cell interaction we found that B cells promoted Tfh differentiation, induced programmed death 1 (PD-1) expression and inhibited IFN-γ production by virus-specific CD4+ T cells. Ultimately, B cells protected hosts from CD4+ T-cell-mediated immune pathology, at the detriment of CD4+ T-cell-mediated protective immunity. Our results suggest that B-cell presentation of vaccine antigens could be manipulated to direct the appropriate CD4+ T-cell response. PMID:21841129
DOE Office of Scientific and Technical Information (OSTI.GOV)
Nozaki, Hirofumi; Itonori, Saki; Sugita, Mutsumi
2008-08-29
Interferon (IFN)-{gamma} and interleukin (IL)-4 regulate many types of immune responses. Here we report that acidic glycosphingolipids (AGLs) of Hypsizigus marmoreus and Pleurotus eryngii induced secretion of IFN- {gamma} and IL-4 from T cells in a CD11c-positive cell-dependent manner similar to that of {alpha}-galactosylceramide ({alpha}-GalCer) and isoglobotriaosylceramide (iGb3), although activated T cells by AGLs showed less secretion of cytokine than those activated by {alpha}-GalCer. In addition, stimulation of these mushroom AGLs induced proliferation of NK1.1 {alpha}/{beta} TCR-double positive cells in splenocytes. Administration of a mixture of {alpha}-GalCer and AGLs affected the stimulation of {alpha}-GalCer and generally induced a subtle Th1more » bias for splenocytes but induced an extreme Th2 bias for thymocytes. These results suggested that edible mushroom AGLs contribute to immunomodulation.« less
Hawse, William F; Sheehan, Robert P; Miskov-Zivanov, Natasa; Menk, Ashley V; Kane, Lawrence P; Faeder, James R; Morel, Penelope A
2015-05-15
Signaling via the Akt/mammalian target of rapamycin pathway influences CD4(+) T cell differentiation; low levels favor regulatory T cell induction and high levels favor Th induction. Although the lipid phosphatase phosphatase and tensin homolog (PTEN) suppresses Akt activity, the control of PTEN activity is poorly studied in T cells. In this study, we identify multiple mechanisms that regulate PTEN expression. During Th induction, PTEN function is suppressed via lower mRNA levels, lower protein levels, and an increase in C-terminal phosphorylation. Conversely, during regulatory T cell induction, PTEN function is maintained through the stabilization of PTEN mRNA transcription and sustained protein levels. We demonstrate that differential Akt/mammalian target of rapamycin signaling regulates PTEN transcription via the FoxO1 transcription factor. A mathematical model that includes multiple modes of PTEN regulation recapitulates our experimental findings and demonstrates how several feedback loops determine differentiation outcomes. Collectively, this work provides novel mechanistic insights into how differential regulation of PTEN controls alternate CD4(+) T cell fate outcomes. Copyright © 2015 by The American Association of Immunologists, Inc.
Thorburn, Alison N; Foster, Paul S; Gibson, Peter G; Hansbro, Philip M
2012-05-01
Asthma is an allergic airways disease (AAD) caused by dysregulated immune responses and characterized by eosinophilic inflammation, mucus hypersecretion, and airway hyperresponsiveness (AHR). NKT cells have been shown to contribute to AHR in some mouse models. Conversely, regulatory T cells (Tregs) control aberrant immune responses and maintain homeostasis. Recent evidence suggests that Streptococcus pneumoniae induces Tregs that have potential to be harnessed therapeutically for asthma. In this study, mouse models of AAD were used to identify the S. pneumoniae components that have suppressive properties, and the mechanisms underlying suppression were investigated. We tested the suppressive capacity of type-3-polysaccharide (T3P), isolated cell walls, pneumolysoid (Ply) and CpG. When coadministered, T3P + Ply suppressed the development of: eosinophilic inflammation, Th2 cytokine release, mucus hypersecretion, and AHR. Importantly, T3P + Ply also attenuated features of AAD when administered during established disease. We show that NKT cells contributed to the development of AAD and also were suppressed by T3P + Ply treatment. Furthermore, adoptive transfer of NKT cells induced AHR, which also could be reversed by T3P + Ply. T3P + Ply-induced Tregs were essential for the suppression of NKT cells and AAD, which was demonstrated by Treg depletion. Collectively, our results show that the S. pneumoniae components T3P + Ply suppress AAD through the induction of Tregs that blocked the activity of NKT cells. These data suggest that S. pneumoniae components may have potential as a therapeutic strategy for the suppression of allergic asthma through the induction of Tregs and suppression of NKT cells.
Hao, Feng; Kang, Jinsen; Cao, Yajun; Fan, Shengjun; Yang, Haopeng; An, Yu; Pan, Yan; Tie, Lu; Li, Xuejun
2015-11-01
Lipotoxicity plays a vital role in development and progression of type 2 diabetes. Prolonged elevation of free fatty acids especially the palmitate leads to pancreatic β-cell dysfunction and apoptosis. Curcumin (diferuloylmethane), a polyphenol from the curry spice turmeric, is considered to be a broadly cytoprotective agent. The present study was designed to determine the protective effect of curcumin on palmitate-induced apoptosis in β-cells and investigate underlying mechanisms. Our results showed that curcumin improved cell viability and enhanced glucose-induced insulin secretory function in MIN6 pancreatic β-cells. Palmitate incubation evoked chromatin condensation, DNA nick end labeling and activation of caspase-3 and -9. Curcumin treatment inhibited palmitate-induced apoptosis, relieved mitochondrial depolarization and up-regulated Bcl-2/Bax ratio. Palmitate induced the generation of reactive oxygen species and inhibited activities of antioxidant enzymes, which could be neutralized by curcumin treatment. Moreover, curcumin could promote rapid phosphorylation of Akt and nuclear exclusion of FoxO1 in MIN6 cells under lipotoxic condition. Phosphatidylinositol 3-kinase and Akt specific inhibitors abolished the anti-lipotoxic effect of curcumin and stimulated FoxO1 nuclear translocation. These findings suggested that curcumin protected MIN6 pancreatic β-Cells against apoptosis through activation of Akt, inhibition of nuclear translocation of FoxO1 and mitochondrial survival pathway.
Miao, Yong; Bhushan, Jaya; Dani, Adish; Vig, Monika
2017-05-11
T cell effector functions require sustained calcium influx. However, the signaling and phenotypic consequences of non-specific sodium permeation via calcium channels remain unknown. α-SNAP is a crucial component of Orai1 channels, and its depletion disrupts the functional assembly of Orai1 multimers. Here we show that α-SNAP hypomorph, hydrocephalus with hopping gait, Napa hyh/hyh mice harbor significant defects in CD4 T cell gene expression and Foxp3 regulatory T cell (Treg) differentiation. Mechanistically, TCR stimulation induced rapid sodium influx in Napa hyh/hyh CD4 T cells, which reduced intracellular ATP, [ATP] i . Depletion of [ATP] i inhibited mTORC2 dependent NFκB activation in Napa hyh/hyh cells but ablation of Orai1 restored it. Remarkably, TCR stimulation in the presence of monensin phenocopied the defects in Napa hyh/hyh signaling and Treg differentiation, but not IL-2 expression. Thus, non-specific sodium influx via bonafide calcium channels disrupts unexpected signaling nodes and may provide mechanistic insights into some divergent phenotypes associated with Orai1 function.
Miao, Yong; Bhushan, Jaya; Dani, Adish; Vig, Monika
2017-01-01
T cell effector functions require sustained calcium influx. However, the signaling and phenotypic consequences of non-specific sodium permeation via calcium channels remain unknown. α-SNAP is a crucial component of Orai1 channels, and its depletion disrupts the functional assembly of Orai1 multimers. Here we show that α-SNAP hypomorph, hydrocephalus with hopping gait, Napahyh/hyh mice harbor significant defects in CD4 T cell gene expression and Foxp3 regulatory T cell (Treg) differentiation. Mechanistically, TCR stimulation induced rapid sodium influx in Napahyh/hyh CD4 T cells, which reduced intracellular ATP, [ATP]i. Depletion of [ATP]i inhibited mTORC2 dependent NFκB activation in Napahyh/hyh cells but ablation of Orai1 restored it. Remarkably, TCR stimulation in the presence of monensin phenocopied the defects in Napahyh/hyh signaling and Treg differentiation, but not IL-2 expression. Thus, non-specific sodium influx via bonafide calcium channels disrupts unexpected signaling nodes and may provide mechanistic insights into some divergent phenotypes associated with Orai1 function. DOI: http://dx.doi.org/10.7554/eLife.25155.001 PMID:28492364
DOE Office of Scientific and Technical Information (OSTI.GOV)
Jeon, Youngsic; Jung, Yujung; Kim, Min Cheol
2014-08-08
Highlights: • SHQA increases PPARα/γ transactivation and inhibits MMP-2/-9 expression. • SHQA inhibits TNFα-induced AP-1 and MAPK signaling. • SHQA inhibits TNFα-induced p65 translocation and IκBα phosphorylation. • SHQA inhibits TNFα-induced AP-1 and NF-κB signaling via PPARα. - Abstract: Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor superfamily of ligand-activated transcription factors and expressed in various cell types in the skin, including keratinocytes, fibroblasts and infiltrating immune cells. Thus, their ligands are targets for the treatment of various skin disorders, such as photo-aging and chronological aging of skin. Intensive studies have revealed that PPARα/γ functions in photo-agingmore » and age-related inflammation by regulating matrix metalloproteinases (MMPs) via activator protein-1 (AP-1) and nuclear factor kappa B (NF-κB). However, the detailed mechanism of PPARα/γ’s role in skin aging has not yet been elucidated. In this study, we confirmed that sargahydroquinoic acid (SHQA) as a PPARα/γ ligand significantly decreased Tumor Necrosis Factor-alpha (TNFα)-induced MMP-2/-9 expression by downregulating TNFα-induced transcription factors, subsequently reducing IκBα degradation and blocking NF-κB p65 nuclear translocation in HaCaT human epidermal keratinocyte cells. Treatment of cells with SHQA and GW6471 (PPARα antagonist) not bisphenol A diglycidyl ether (PPARγ antagonists), reversed the effect on TNFα-induced inflammatory signaling pathway activation. Taken together, our data suggest that SHQA inhibit TNFα-induced MMP-2/-9 expression and age-related inflammation by suppressing AP-1 and NF-κB pathway via PPARα.« less
Kitazawa, Yusuke; Sawanobori, Yasushi; Ueno, Takamasa; Ueha, Satoshi; Matsushima, Kouji; Matsuno, Kenjiro
2018-01-01
Abstract Donor-specific blood transfusion is known to induce alloresponses and lead to immunosuppression. We examined their underlying mechanisms by employing fully allogeneic rat combinations. Transfused recipients efficiently produced alloantibodies of the IgM and IgG subclasses directed against donor class I MHC. The recipients exhibited active expansion of CD4+ T cells and CD4+FOXP3+ regulatory T cells (Treg cells), followed by CD45R+ B cells and IgM+ or IgG subclass+ antibody-forming cells mainly in the spleen. From 1.5 days, the resident MHCII+CD103+ dendritic cells (DCs) in the splenic T-cell area, periarterial lymphocyte sheath, formed clusters with recipient BrdU+ or 5-ethynyl-2′-deoxyuridine+ cells, from which the proliferative response of CD4+ T cells originated peaking at 3–4 days. Transfusion-induced antibodies had donor passenger cell-depleting activity in vitro and in vivo and could suppress acute GvH disease caused by donor T cells. Furthermore, Treg cells significantly suppressed mixed leukocyte reactions in a donor-specific manner. In conclusion, single blood transfusion efficiently induced a helper T-cell-dependent anti-donor class I MHC antibody-forming cell response with immunoglobulin class switching, and a donor-specific Treg cell response mainly in the spleen, probably by way of the indirect allorecognition via resident DCs. These antibodies and Treg cells may be involved, at least partly, in the donor-specific transfusion-induced suppression of allograft rejection. PMID:29361165
HLA Class I Binding 9mer Peptides from Influenza A Virus Induce CD4+ T Cell Responses
Wang, Mingjun; Larsen, Mette V.; Nielsen, Morten; Harndahl, Mikkel; Justesen, Sune; Dziegiel, Morten H.; Buus, Søren; Tang, Sheila T.; Lund, Ole; Claesson, Mogens H.
2010-01-01
Background Identification of human leukocyte antigen class I (HLA-I) restricted cytotoxic T cell (CTL) epitopes from influenza virus is of importance for the development of new effective peptide-based vaccines. Methodology/Principal Findings In the present work, bioinformatics was used to predict 9mer peptides derived from available influenza A viral proteins with binding affinity for at least one of the 12 HLA-I supertypes. The predicted peptides were then selected in a way that ensured maximal coverage of the available influenza A strains. One hundred and thirty one peptides were synthesized and their binding affinities for the HLA-I supertypes were measured in a biochemical assay. Influenza-specific T cell responses towards the peptides were quantified using IFNγ ELISPOT assays with peripheral blood mononuclear cells (PBMC) from adult healthy HLA-I typed donors as responder cells. Of the 131 peptides, 21 were found to induce T cell responses in 19 donors. In the ELISPOT assay, five peptides induced responses that could be totally blocked by the pan-specific anti-HLA-I antibody W6/32, whereas 15 peptides induced responses that could be completely blocked in the presence of the pan-specific anti-HLA class II (HLA-II) antibody IVA12. Blocking of HLA-II subtype reactivity revealed that 8 and 6 peptide responses were blocked by anti-HLA-DR and -DP antibodies, respectively. Peptide reactivity of PBMC depleted of CD4+ or CD8+ T cells prior to the ELISPOT culture revealed that effectors are either CD4+ (the majority of reactivities) or CD8+ T cells, never a mixture of these subsets. Three of the peptides, recognized by CD4+ T cells showed binding to recombinant DRA1*0101/DRB1*0401 or DRA1*0101/DRB5*0101 molecules in a recently developed biochemical assay. Conclusions/Significance HLA-I binding 9mer influenza virus-derived peptides induce in many cases CD4+ T cell responses restricted by HLA-II molecules. PMID:20479886
Raman spectrum reveals the cell cycle arrest of Triptolide-induced leukemic T-lymphocytes apoptosis
NASA Astrophysics Data System (ADS)
Zhang, Daosen; Feng, Yanyan; Zhang, Qinnan; Su, Xin; Lu, Xiaoxu; Liu, Shengde; Zhong, Liyun
2015-04-01
Triptolide (TPL), a traditional Chinese medicine extract, possesses anti-inflammatory and anti-tumor properties. Though some research results have implicated that Triptolide (TPL) can be utilized in the treatment of leukemia, it remains controversial about the mechanism of TPL-induced leukemic T-lymphocytes apoptosis. In this study, combining Raman spectroscopic data, principal component analysis (PCA) and atomic force microscopy (AFM) imaging, both the biochemical changes and morphological changes during TPL-induced cell apoptosis were presented. In contrast, the corresponding data during Daunorubicin (DNR)-induced cell apoptosis was also exhibited. The obtained results showed that Raman spectral changes during TPL-induced cell apoptosis were greatly different from DNR-induced cell apoptosis in the early stage of apoptosis but revealed the high similarity in the late stage of apoptosis. Moreover, above Raman spectral changes were respectively consistent with the morphological changes of different stages during TPL-induced apoptosis or DNR-induced apoptosis, including membrane shrinkage and blebbing, chromatin condensation and the formation of apoptotic bodies. Importantly, it was found that Raman spectral changes with TPL-induced apoptosis or DNR-induced apoptosis were respectively related with the cell cycle G1 phase arrest or G1 and S phase arrest.
Yang, Yin; Wu, Songfang; Wang, Yu; Pan, Shuang; Lan, Bei; Liu, Yaohui; Zhang, Liming; Leng, Qianli; Chen, Da; Zhang, Cuizhu; He, Bin; Cao, Youjia
2015-01-01
Herpes simplex virus 1 (HSV-1) is the most prevalent human virus and causes global morbidity because the virus is able to infect multiple cell types. Remarkably, HSV infection switches between lytic and latent cycles, where T cells play a critical role. However, the precise way of virus-host interactions is incompletely understood. Here we report that HSV-1 productively infected Jurkat T-cells and inhibited antigen-induced T cell receptor activation. We discovered that HSV-1-encoded Us3 protein interrupted TCR signaling and interleukin-2 production by inactivation of the linker for activation of T cells. This study unveils a mechanism by which HSV-1 intrudes into early events of TCR-mediated cell signaling and may provide novel insights into HSV infection, during which the virus escapes from host immune surveillance. PMID:25907557
Katz, Gil; Voss, Kelsey; Yan, Toria F; Kim, Yong Chan; Kortum, Robert L; Scott, David W; Snow, Andrew L
2018-05-01
Restimulation-induced cell death (RICD) is an apoptotic program that regulates effector T cell expansion, triggered by repeated stimulation through the T cell receptor (TCR) in the presence of interleukin-2 (IL-2). Although CD4 + regulatory T cells (Tregs) consume IL-2 and experience frequent TCR stimulation, they are highly resistant to RICD. Resistance in Tregs is dependent on the forkhead box P3 (FOXP3) transcription factor, although the mechanism remains unclear. T cells from patients with X-linked lymphoproliferative disease (XLP-1), that lack the adaptor molecule SLAM-associated protein (SAP), are also resistant to RICD. Here we demonstrate that normal Tregs express very low levels of SAP compared to conventional T cells. FOXP3 reduces SAP expression by directly binding to and repressing the SH2D1A (SAP) promoter. Indeed, ectopic SAP expression restores RICD sensitivity in human FOXP3 + Tregs. Our findings illuminate the mechanism behind FOXP3-mediated RICD resistance in Tregs, providing new insight into their long-term persistence. Published by Elsevier Inc.
Palma, Carla; Schiavoni, Giovanna; Abalsamo, Laura; Mattei, Fabrizio; Piccaro, Giovanni; Sanchez, Massimo; Fernandez, Carmen; Singh, Mahavir; Gabriele, Lucia
2013-09-01
The immunological mechanisms that modulate protection during Mycobacterium tuberculosis (Mtb) infection or vaccination are not fully understood. Secretion of IFN-γ and, to a lesser extent, of IL-17 by CD4(+) T cells plays a major role both in protection and immunopathology. Few Mtb Ags interacting with DCs affect priming, activation, and regulation of Ag-unrelated CD4(+) T-cell responses. Here we demonstrate that PstS1, a 38 kDa-lipoprotein of Mtb, promotes Ag-independent activation of memory T lymphocytes specific for Ag85B or Ag85A, two immunodominant protective Ags of Mtb. PstS1 expands CD4(+) and CD8(+) memory T cells, amplifies secretion of IFN-γ and IL-22 and induces IL-17 production by effector memory cells in an Ag-unrelated manner in vitro and in vivo. These effects were mediated through the stimulation of DCs, particularly of the CD8α(-) subtype, which respond to PstS1 by undergoing phenotypic maturation and by secreting IL-6, IL-1β and, to a lower extent, IL-23. IL-6 secretion by PstS1-stimulated DCs was required for IFN-γ, and to a lesser extent for IL-22 responses by Ag85B-specific memory T cells. These results may open new perspectives for immunotherapeutic strategies to control Th1/Th17 immune responses in Mtb infections and in vaccinations against tuberculosis. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Features of Effective T Cell-Inducing Vaccines against Chronic Viral Infections
Panagioti, Eleni; Klenerman, Paul; Lee, Lian N.; van der Burg, Sjoerd H.; Arens, Ramon
2018-01-01
For many years, the focus of prophylactic vaccines was to elicit neutralizing antibodies, but it has become increasingly evident that T cell-mediated immunity plays a central role in controlling persistent viral infections such as with human immunodeficiency virus, cytomegalovirus, and hepatitis C virus. Currently, various promising prophylactic vaccines, capable of inducing substantial vaccine-specific T cell responses, are investigated in preclinical and clinical studies. There is compelling evidence that protection by T cells is related to the magnitude and breadth of the T cell response, the type and homing properties of the memory T cell subsets, and their cytokine polyfunctionality and metabolic fitness. In this review, we evaluated these key factors that determine the qualitative and quantitative properties of CD4+ and CD8+ T cell responses in the context of chronic viral disease and prophylactic vaccine development. Elucidation of the mechanisms underlying T cell-mediated protection against chronic viral pathogens will facilitate the development of more potent, durable and safe prophylactic T cell-based vaccines. PMID:29503649
Zopun, Muhammet; Lieder, Barbara; Holik, Ann-Katrin; Ley, Jakop P; Hans, Joachim; Somoza, Veronika
2018-06-25
The role of sweet taste in energy intake and satiety regulation is still controversial. Noncaloric artificial sweeteners (NCSs) are thought to help reduce energy intake, although little is known about their impact on the satiating neurotransmitter serotonin (5-HT). In the gastrointestinal (GI) tract, 5-HT regulates gastric acid secretion and gastric motility, both part of the complex network of mechanisms regulating food intake and satiety. This study demonstrated a stimulating impact compared to controls (100%) on 5-HT release in human gastric tumor cells (HGT-1) by the NCSs cyclamate (50 mM, 157% ± 6.3%), acesulfame potassium (Ace K, 50 mM, 197% ± 8.6%), saccharin (50 mM, 147% ± 6.7%), sucralose (50 mM, 194% ± 11%), and neohesperidin dihydrochalcone (NHDC, 1 mM, 201% ± 13%). Although these effects were not associated with the sweet taste intensity of the NCSs tested, involvement of the sweet receptor subunit T1R3 in the NCS-evoked response was demonstrated by mRNA expression of TAS1R3, co-incubation experiments using the T1R3 receptor antagonist lactisole, and a TAS1R3 siRNA knockdown approach. Analysis of the downstream signaling revealed activation of the cAMP/ERK/Ca 2+ cascade. Co-treatment experiments with 10 mM glucose enhanced the 5-HT release induced by cyclamate, Ace K, saccharin, and sucralose, thereby supporting the enhancing effect of glucose on a NCS-mediated response. Overall, the results obtained identify NCSs as potent inducers of 5-HT release via T1R3 in human gastric parietal cells in culture and warrant in vivo studies to demonstrate their efficacy.
Chen, Yonglin; Peng, Hui; Chen, Yongyan; Wei, Haiming; Sun, Rui; Tian, Zhigang
2014-01-01
It is becoming increasingly clear that the T-cell-mediated immune response is important in many diseases. In this study, we used concanavalin A (Con A) -induced hepatitis to investigate the role of CD49a in the molecular and cellular mechanism of the T-cell-mediated immune response. We found that CD49a−/− mice had significantly reduced levels of serum alanine aminotransferase and were protected from Con A-induced hepatitis. CD49a deficiency led to decreased production of interferon-γ (IFN-γ) and interleukin-17A (IL-17A) after Con A injection. Furthermore, we found that hepatic CD4+ T cells and invariant natural killer T cells up-regulated CD49a expression, along with enhanced activation after Con A injection, leading to production of inflammatory cytokines by these T cells. Blockade of CD49a in vivo ameliorated Con A-induced hepatitis with reduced production of IFN-γ and IL-17A. Hence, CD49a promoted Con A-induced hepatitis through enhancing inflammatory cytokine production (IFN-γ and IL-17A) by CD4+ T and invariant natural killer T cells. The protective effect of CD49a blockade antibody suggested a new target therapeutic molecule for intervention of T-cell-mediated liver injury. PMID:24164540
Dual role of SLP-76 in mediating T cell receptor-induced activation of phospholipase C-gamma1.
Beach, Dvora; Gonen, Ronnie; Bogin, Yaron; Reischl, Ilona G; Yablonski, Deborah
2007-02-02
Phospholipase C-gamma1 (PLC-gamma1) activation depends on a heterotrimeric complex of adaptor proteins composed of LAT, Gads, and SLP-76. Upon T cell receptor stimulation, a portion of PLC-gamma1 is recruited to a detergent-resistant membrane fraction known as the glycosphingolipid-enriched membrane microdomains (GEMs), or lipid rafts, to which LAT is constitutively localized. In addition to LAT, PLC-gamma1 GEM recruitment depended on SLP-76, and, in particular, required the Gads-binding domain of SLP-76. The N-terminal tyrosine phosphorylation sites and P-I region of SLP-76 were not required for PLC-gamma1 GEM recruitment, but were required for PLC-gamma1 phosphorylation at Tyr(783). Thus, GEM recruitment can be insufficient for full activation of PLC-gamma1 in the absence of a second SLP-76-mediated event. Indeed, a GEM-targeted derivative of PLC-gamma1 depended on SLP-76 for T cell receptor-induced phosphorylation at Tyr783 and subsequent NFAT activation. On a biochemical level, SLP-76 inducibly associated with both Vav and catalytically active ITK, which efficiently phosphorylated a PLC-gamma1 fragment at Tyr783 in vitro. Both associations were disrupted upon mutation of the N-terminal tyrosine phosphorylation sites of SLP-76. The P-I region deletion disrupted Vav association and reduced SLP-76-associated kinase activity. A smaller deletion within the P-I region, which does not impair PLC-gamma1 activation, did not impair the association with Vav, but reduced SLP-76-associated kinase activity. These results provide new insight into the multiple roles of SLP-76 and the functional importance of its interactions with other signaling proteins.
Furlong, Suzanne J; Ridgway, Neale D; Hoskin, David W
2008-03-01
Bovine lactoferricin (LfcinB) is a cationic antimicrobial peptide that selectively induces apoptosis in several different types of human cancer cells. However, the potential use of LfcinB as an anticancer agent is presently limited by the need for relatively high concentrations of the peptide to trigger apoptosis. Ceramide is a membrane sphingolipid that is believed to function as a second messenger during apoptosis. In this study, we investigated the role of ceramide in LfcinB-induced apoptosis in CCRF-CEM and Jurkat T-leukemia cell lines. Exposure to LfcinB caused nuclear condensation and fragmentation, poly(ADP-ribose) polymerase (PARP) cleavage, and DNA fragmentation in CCRF-CEM and Jurkat T-cell acute lymphoblastic leukemia cell lines. Treatment with C6 ceramide, a cell-permeable, short-chain ceramide analog, also induced apoptotic nuclear morphology, PARP cleavage, and DNA fragmentation in T-leukemia cells. Although LfcinB treatment did not cause ceramide to accumulate in CCRF-CEM or Jurkat cells, the addition of C6 ceramide to LfcinB-treated T-leukemia cells resulted in increased DNA fragmentation. Furthermore, modulation of cellular ceramide metabolism either by inhibiting ceramidases with D-erythro-2-(N-myristoylamino)-1-phenyl-1-propanol or N-oleoylethanolamine, or by blocking glucosylceramide synthase activity with 1-phenyl-2-palmitoylamino-3-morpholino-1-propanol, enhanced the ability of LfcinB to trigger apoptosis in both Jurkat and CCRF-CEM cells. In addition, LfcinB-induced apoptosis of T-leukemia cells was enhanced in the presence of the antiestrogen tamoxifen, which has multiple effects on cancer cells, including inhibition of glucosylceramide synthase activity. We conclude that manipulation of cellular ceramide levels in combination with LfcinB therapy warrants further investigation as a novel strategy for the treatment of T cell-derived leukemias.
Vaid, Mudit; Prasad, Ram; Singh, Tripti; Elmets, Craig A.; Xu, Hui; Katiyar, Santosh K.
2013-01-01
Silymarin inhibits UVB-induced immunosuppression in mouse skin. To identify the molecular mechanisms underlying this effect, we used an adoptive transfer approach in which dendritic cells (DCs) from the draining lymph nodes of donor mice that had been UVB-exposed and sensitized to 2,4,-dinitrofluorobenzene (DNFB) were transferred into naïve recipient mice. The contact hypersensitivity (CHS) response of the recipient mice to DNFB was then measured. When DCs were obtained from UVB-exposed donor mice that were not treated with silymarin, the CHS response was suppressed confirming the role of DCs in the UVB-induced immunosuppression. Silymarin treatment of UVB-exposed donor mice relieved this suppression of the CHS response in the recipients. Silymarin treatment was associated with rapid repair of UVB-induced cyclobutane pyrimidine dimers (CPDs) in DCs and silymarin treatment did not prevent UV-induced immunosuppression in XPA-deficient mice which are unable to repair UV-induced DNA damage. The CHS response in mice receiving DCs from silymarin-treated UV-exposed donor mice also was associated with enhanced secretion of Th1-type cytokines and stimulation of T cells. Adoptive transfer of T cells revealed that transfer of either CD8+ or CD4+ cells from silymarin-treated, UVB-exposed donors resulted in enhancement of the CHS response. Cell culture study showed enhanced secretion of IL-2 and IFNγ by CD8+ T cells, and reduced secretion of Th2 cytokines by CD4+ cells, obtained from silymarin-treated UVB-exposed mice. These data suggest that DNA repair-dependent functional activation of DCs, a reduction in CD4+ regulatory T-cell activity, and stimulation of CD8+ effector T cells contribute to silymarin-mediated inhibition of UVB-induced immunosuppression. PMID:23395695
Li, Wenpeng; Guo, Linjie; Rathi, Purva; Marinova, Ekaterina; Gao, Xiuhua; Wu, Meng-Feng; Liu, Hao; Dotti, Gianpietro; Gottschalk, Stephen; Metelitsa, Leonid S.; Heczey, Andras
2017-01-01
T cells engineered to express CD19-specific chimeric antigen receptors (CARs) have shown breakthrough clinical successes in patients with B-cell lymphoid malignancies. However, similar therapeutic efficacy of CAR T cells in solid tumors is yet to be achieved. In this study we systematically evaluated a series of CAR constructs targeting glypican-3 (GPC3), which is selectively expressed on several solid tumors. We compared GPC3-specific CARs that encoded CD3ζ (Gz) alone or with costimulatory domains derived from CD28 (G28z), 4-1BB (GBBz), or CD28 and 4-1BB (G28BBz). All GPC3-CARs rendered T cells highly cytotoxic to GPC3-positive hepatocellular carcinoma, hepatoblastoma, and malignant rhabdoid tumor cell lines in vitro. GBBz induced the preferential production of Th1 cytokines (interferon γ/granulocyte macrophage colony-stimulating factor) while G28z preferentially induced Th2 cytokines (interleukin-4/interleukin-10). Inclusion of 4-1BB in G28BBz could only partially ameliorate the Th2-polarizing effect of CD28. 4-1BB induced superior expansion of CAR T cells in vitro and in vivo. T cells expressing GPC3-CARs incorporating CD28, 4-1BB, or both induced sustained tumor regressions in two xenogeneic tumor models. Thus, GBBz CAR endows T cells with superior proliferative potential, potent antitumor activity, and a Th1-biased cytokine profile, justifying further clinical development of GBBz CAR for immunotherapy of GPC3-positive solid tumors. PMID:27530312
Banga, Riddhima; Procopio, Francesco A.; Ruggiero, Alessandra; Noto, Alessandra; Ohmiti, Khalid; Cavassini, Matthias; Corpataux, Jean-Marc; Paxton, William A.; Pollakis, Georgios; Perreau, Matthieu
2018-01-01
We recently demonstrated that lymph nodes (LNs) PD-1+/T follicular helper (Tfh) cells from antiretroviral therapy (ART)-treated HIV-infected individuals were enriched in cells containing replication competent virus. However, the distribution of cells containing inducible replication competent virus has been only partially elucidated in blood memory CD4 T-cell populations including the Tfh cell counterpart circulating in blood (cTfh). In this context, we have investigated the distribution of (1) total HIV-infected cells and (2) cells containing replication competent and infectious virus within various blood and LN memory CD4 T-cell populations of conventional antiretroviral therapy (cART)-treated HIV-infected individuals. In the present study, we show that blood CXCR3-expressing memory CD4 T cells are enriched in cells containing inducible replication competent virus and contributed the most to the total pool of cells containing replication competent and infectious virus in blood. Interestingly, subsequent proviral sequence analysis did not indicate virus compartmentalization between blood and LN CD4 T-cell populations, suggesting dynamic interchanges between the two compartments. We then investigated whether the composition of blood HIV reservoir may reflect the polarization of LN CD4 T cells at the time of reservoir seeding and showed that LN PD-1+ CD4 T cells of viremic untreated HIV-infected individuals expressed significantly higher levels of CXCR3 as compared to CCR4 and/or CCR6, suggesting that blood CXCR3-expressing CD4 T cells may originate from LN PD-1+ CD4 T cells. Taken together, these results indicate that blood CXCR3-expressing CD4 T cells represent the major blood compartment containing inducible replication competent virus in treated aviremic HIV-infected individuals. PMID:29459864
Banga, Riddhima; Procopio, Francesco A; Ruggiero, Alessandra; Noto, Alessandra; Ohmiti, Khalid; Cavassini, Matthias; Corpataux, Jean-Marc; Paxton, William A; Pollakis, Georgios; Perreau, Matthieu
2018-01-01
We recently demonstrated that lymph nodes (LNs) PD-1 + /T follicular helper (Tfh) cells from antiretroviral therapy (ART)-treated HIV-infected individuals were enriched in cells containing replication competent virus. However, the distribution of cells containing inducible replication competent virus has been only partially elucidated in blood memory CD4 T-cell populations including the Tfh cell counterpart circulating in blood (cTfh). In this context, we have investigated the distribution of (1) total HIV-infected cells and (2) cells containing replication competent and infectious virus within various blood and LN memory CD4 T-cell populations of conventional antiretroviral therapy (cART)-treated HIV-infected individuals. In the present study, we show that blood CXCR3-expressing memory CD4 T cells are enriched in cells containing inducible replication competent virus and contributed the most to the total pool of cells containing replication competent and infectious virus in blood. Interestingly, subsequent proviral sequence analysis did not indicate virus compartmentalization between blood and LN CD4 T-cell populations, suggesting dynamic interchanges between the two compartments. We then investigated whether the composition of blood HIV reservoir may reflect the polarization of LN CD4 T cells at the time of reservoir seeding and showed that LN PD-1 + CD4 T cells of viremic untreated HIV-infected individuals expressed significantly higher levels of CXCR3 as compared to CCR4 and/or CCR6, suggesting that blood CXCR3-expressing CD4 T cells may originate from LN PD-1 + CD4 T cells. Taken together, these results indicate that blood CXCR3-expressing CD4 T cells represent the major blood compartment containing inducible replication competent virus in treated aviremic HIV-infected individuals.
Acute Viral Respiratory Infection Rapidly Induces a CD8+ T Cell Exhaustion-like Phenotype.
Erickson, John J; Lu, Pengcheng; Wen, Sherry; Hastings, Andrew K; Gilchuk, Pavlo; Joyce, Sebastian; Shyr, Yu; Williams, John V
2015-11-01
Acute viral infections typically generate functional effector CD8(+) T cells (TCD8) that aid in pathogen clearance. However, during acute viral lower respiratory infection, lung TCD8 are functionally impaired and do not optimally control viral replication. T cells also become unresponsive to Ag during chronic infections and cancer via signaling by inhibitory receptors such as programmed cell death-1 (PD-1). PD-1 also contributes to TCD8 impairment during viral lower respiratory infection, but how it regulates TCD8 impairment and the connection between this state and T cell exhaustion during chronic infections are unknown. In this study, we show that PD-1 operates in a cell-intrinsic manner to impair lung TCD8. In light of this, we compared global gene expression profiles of impaired epitope-specific lung TCD8 to functional spleen TCD8 in the same human metapneumovirus-infected mice. These two populations differentially regulate hundreds of genes, including the upregulation of numerous inhibitory receptors by lung TCD8. We then compared the gene expression of TCD8 during human metapneumovirus infection to those in acute or chronic lymphocytic choriomeningitis virus infection. We find that the immunophenotype of lung TCD8 more closely resembles T cell exhaustion late into chronic infection than do functional effector T cells arising early in acute infection. Finally, we demonstrate that trafficking to the infected lung alone is insufficient for TCD8 impairment or inhibitory receptor upregulation, but that viral Ag-induced TCR signaling is also required. Our results indicate that viral Ag in infected lungs rapidly induces an exhaustion-like state in lung TCD8 characterized by progressive functional impairment and upregulation of numerous inhibitory receptors. Copyright © 2015 by The American Association of Immunologists, Inc.
TIF-IA and Ebp1 regulate RNA synthesis in T cells.
Saudemont, Aurore
2015-04-16
In this issue of Blood, Nguyen et al show that mycophenolic acid (MPA) induces GTP depletion, which inhibits the function of transcription initiation factor I (TIF-IA) and impacts the interaction of TIF-IA with ErbB3-binding protein 1 (Ebp1), a key in regulating proliferating cell nuclear antigen (PCNA) expression and ribosomal RNA (rRNA) synthesis in T cells during activation.
Konkel, Joanne E; Frommer, Friederike; Leech, Melanie D; Yagita, Hideo; Waisman, Ari; Anderton, Stephen M
2010-01-01
The ultimate outcome of T-cell recognition of peptide–major histocompatibility complex (MHC) complexes is determined by the molecular context in which antigen presentation is provided. The paradigm is that, after exposure to peptides presented by steady-state dendritic cells (DCs), inhibitory signals dominate, leading to the deletion and/or functional inactivation of antigen-reactive T cells. This has been utilized in a variety of models providing peptide antigen in soluble form in the absence of adjuvant. A co-inhibitory molecule of considerable current interest is PD-1. Here we show that there is the opportunity for the PD-1/PD-L1 interaction to function in inhibiting the T-cell response during tolerance induction. Using traceable CD4+ T-cell receptor (TCR) transgenic cells, together with a blocking antibody to disrupt PD-1 signalling, we explored the roles of PD-1 in the induction of tolerance versus a productive immune response. Intact PD-1 signalling played a role in limiting the extent of CD4+ T-cell accumulation in response to an immunogenic stimulus. However, PD-1 signalling was not required for either the induction, or the maintenance, of peptide-induced tolerance; a conclusion underlined by successful tolerance induction in TCR transgenic cells genetically deficient for PD-1. These observations contrast with the reported requirement for PD-1 signals in CD8+ T-cell tolerance. PMID:20113370
Kamphorst, Alice O; Pillai, Rathi N; Yang, Shu; Nasti, Tahseen H; Akondy, Rama S; Wieland, Andreas; Sica, Gabriel L; Yu, Ke; Koenig, Lydia; Patel, Nikita T; Behera, Madhusmita; Wu, Hong; McCausland, Megan; Chen, Zhengjia; Zhang, Chao; Khuri, Fadlo R; Owonikoko, Taofeek K; Ahmed, Rafi; Ramalingam, Suresh S
2017-05-09
Exhausted T cells in chronic infections and cancer have sustained expression of the inhibitory receptor programmed cell death 1 (PD-1). Therapies that block the PD-1 pathway have shown promising clinical results in a significant number of advanced-stage cancer patients. Nonetheless, a better understanding of the immunological responses induced by PD-1 blockade in cancer patients is lacking. Identification of predictive biomarkers is a priority in the field, but whether peripheral blood analysis can provide biomarkers to monitor or predict patients' responses to treatment remains to be resolved. In this study, we analyzed longitudinal blood samples from advanced stage non-small cell lung cancer (NSCLC) patients ( n = 29) receiving PD-1-targeted therapies. We detected an increase in Ki-67+ PD-1+ CD8 T cells following therapy in ∼70% of patients, and most responses were induced after the first or second treatment cycle. This T-cell activation was not indiscriminate because we observed only minimal effects on EBV-specific CD8 T cells, suggesting that responding cells may be tumor specific. These proliferating CD8 T cells had an effector-like phenotype (HLA-DR + , CD38 + , Bcl-2 lo ), expressed costimulatory molecules (CD28, CD27, ICOS), and had high levels of PD-1 and coexpression of CTLA-4. We found that 70% of patients with disease progression had either a delayed or absent PD-1+ CD8 T-cell response, whereas 80% of patients with clinical benefit exhibited PD-1+ CD8 T-cell responses within 4 wk of treatment initiation. Our results suggest that peripheral blood analysis may provide valuable insights into NSCLC patients' responses to PD-1-targeted therapies.
Fu, Shin-Huei; Yeh, Li-Tzu; Chu, Chin-Chen; Yen, B Lin-Ju; Sytwu, Huey-Kang
2017-07-21
B lymphocyte-induced maturation protein-1 (Blimp-1) serves as a master regulator of the development and function of antibody-producing B cells. Given that its function in T lymphocytes has been identified within the past decade, we review recent findings with emphasis on its role in coordinated control of gene expression during the development, differentiation, and function of T cells. Expression of Blimp-1 is mainly confined to activated T cells and is essential for the production of interleukin (IL)-10 by a subset of forkhead box (Fox)p3 + regulatory T cells with an effector phenotype. Blimp-1 is also required to induce cell elimination in the thymus and critically modulates peripheral T cell activation and proliferation. In addition, Blimp-1 promotes T helper (Th) 2 lineage commitment and limits Th1, Th17 and follicular helper T cell differentiation. Furthermore, Blimp-1 coordinates with other transcription factors to regulate expression of IL-2, IL-21 and IL-10 in effector T lymphocytes. In CD8 + T cells, Blimp-1 expression is distinct in heterogeneous populations at the stages of clonal expansion, differentiation, contraction and memory formation when they encounter antigens. Moreover, Blimp-1 plays a fundamental role in coordinating cytokine receptor signaling networks and transcriptional programs to regulate diverse aspects of the formation and function of effector and memory CD8 + T cells and their exhaustion. Blimp-1 also functions as a gatekeeper of T cell activation and suppression to prevent or dampen autoimmune disease, antiviral responses and antitumor immunity. In this review, we discuss the emerging roles of Blimp-1 in the complex regulation of gene networks that regulate the destiny and effector function of T cells and provide a Blimp-1-dominated transcriptional framework for T lymphocyte homeostasis.
Mohanram, Venkatramanan; Johansson, Ulrika; Sköld, Annette E; Fink, Joshua; Kumar Pathak, Sushil; Mäkitalo, Barbro; Walther-Jallow, Lilian; Spetz, Anna-Lena
2011-01-01
Dendritic cells (DCs) are activated by signaling via pathogen-specific receptors or exposure to inflammatory mediators. Here we show that co-culturing DCs with apoptotic HIV-infected activated CD4(+) T cells (ApoInf) or apoptotic uninfected activated CD4(+) T cells (ApoAct) induced expression of co-stimulatory molecules and cytokine release. In addition, we measured a reduced HIV infection rate in DCs after co-culture with ApoAct. A prerequisite for reduced HIV infection in DCs was activation of CD4(+) T cells before apoptosis induction. DCs exposed to ApoAct or ApoInf secreted MIP-1α, MIP-1β, MCP-1, and TNF-α; this effect was retained in the presence of exogenous HIV. The ApoAct-mediated induction of co-stimulatory CD86 molecules and reduction of HIV infection in DCs were partially abrogated after blocking TNF-α using monoclonal antibodies. APOBEC3G expression in DCs was increased in co-cultures of DCs and ApoAct but not by apoptotic resting CD4(+) T cells (ApoRest). Silencing of APOBEC3G in DC abrogated the HIV inhibitory effect mediated by ApoAct. Sequence analyses of an env region revealed significant induction of G-to-A hypermutations in the context of GG or GA dinucleotides in DNA isolated from DCs exposed to HIV and ApoAct. Thus, ApoAct-mediated DC maturation resulted in induction of APOBEC3G that was important for inhibition of HIV-infection in DCs. These findings underscore the complexity of differential DC responses evoked upon interaction with resting as compared with activated dying cells during HIV infection.
CD47-ligation induced cell death in T-acute lymphoblastic leukemia.
Leclair, Pascal; Liu, Chi-Chao; Monajemi, Mahdis; Reid, Gregor S; Sly, Laura M; Lim, Chinten James
2018-05-10
CD47 is a cell-surface marker well recognized for its anti-phagocytic functions. As such, an emerging avenue for targeted cancer therapies involves neutralizing the anti-phagocytic function using monoclonal antibodies (mAbs) to enhance tumour cell immunogenicity. A lesser known consequence of CD47 receptor ligation is the direct induction of tumour cell death. While several mAbs and their derivatives with this property have been studied, the best characterized is the commercially available mAb B6H12, which requires immobilization for induction of cell death. Here, we describe a commercially available mAb, CC2C6, which induces T-cell acute lymphoblastic leukemia (ALL) cell death in soluble form. Soluble CC2C6 induces CD47-dependent cell death in a manner consistent with immobilized B6H12, which is characterized by mitochondrial deficiencies but is independent of caspase activation. Titration studies indicated that CC2C6 shares a common CD47-epitope with B6H12. Importantly, CC2C6 retains the anti-phagocytic neutralizing function, thus possessing dual anti-tumour properties. Although CD47-ligation induced cell death occurs in a caspase-independent manner, CC2C6 was found to stimulate increases in Mcl-1 and NOXA levels, two Bcl-2 family proteins that govern the intrinsic apoptosis pathway. Further analysis revealed that the ratio of Mcl-1:NOXA were minimally altered for cells treated with CC2C6, in comparison to cells treated with agents that induced caspase-dependent apoptosis which alter this ratio in favour of NOXA. Finally, we found that CC2C6 can synergize with low dose chemotherapeutic agents that induce classical apoptosis, giving rise to the possibility of an effective combination treatment with reduced long-term sequelae associated with high-dose chemotherapies in childhood ALL.
Wang, Yichuan; Solaymani-Mohammadi, Shahram; Frey, Blake; Kulkarni, Shweta; Andersen, Peter; Agger, Else Marie; Sui, Yongjun
2017-01-01
T cells with high functional avidity can sense and respond to low levels of cognate Ag, a characteristic that is associated with more potent responses against tumors and many infections, including HIV. Although an important determinant of T cell efficacy, it has proven difficult to selectively induce T cells of high functional avidity through vaccination. Attempts to induce high-avidity T cells by low-dose in vivo vaccination failed because this strategy simply gave no response. Instead, selective induction of high-avidity T cells has required in vitro culturing of specific T cells with low Ag concentrations. In this study, we combined low vaccine Ag doses with a novel potent cationic liposomal adjuvant, cationic adjuvant formulation 09, consisting of dimethyldioctadecylammonium liposomes incorporating two immunomodulators (monomycolyl glycerol analog and polyinosinic-polycytidylic acid) that efficiently induces CD4 Th cells, as well as cross-primes CD8 CTL responses. We show that vaccination with low Ag dose selectively primes CD4 T cells of higher functional avidity, whereas CD8 T cell functional avidity was unrelated to vaccine dose in mice. Importantly, CD4 T cells of higher functional avidity induced by low-dose vaccinations showed higher cytokine release per cell and lower inhibitory receptor expression (PD-1, CTLA-4, and the apoptosis-inducing Fas death receptor) compared with their lower-avidity CD4 counterparts. Notably, increased functional CD4 T cell avidity improved antiviral efficacy of CD8 T cells. These data suggest that potent adjuvants, such as cationic adjuvant formulation 09, render low-dose vaccination a feasible and promising approach for generating high-avidity T cells through vaccination. PMID:28348274
Chen, Weirong; Wan, Xiaoxiao; Ukah, Tobechukwu K; Miller, Mindy M; Barik, Subhasis; Cattin-Roy, Alexis N; Zaghouani, Habib
2016-11-01
To contain autoimmunity, pathogenic T cells must be eliminated or diverted from reaching the target organ. Recently, we defined a novel form of T cell tolerance whereby treatment with Ag downregulates expression of the chemokine receptor CXCR3 and prevents diabetogenic Th1 cells from reaching the pancreas, leading to suppression of type 1 diabetes (T1D). This report defines the signaling events underlying Ag-induced chemokine receptor-mediated tolerance. Specifically, we show that the mammalian target of rapamycin complex 1 (mTORC1) is a major target for induction of CXCR3 downregulation and crippling of Th1 cells. Indeed, Ag administration induces upregulation of programmed death-ligand 1 on dendritic cells in a T cell-dependent manner. In return, programmed death-ligand 1 interacts with the constitutively expressed programmed death-1 on the target T cells and stimulates docking of Src homology 2 domain-containing tyrosine phosphatase 2 phosphatase to the cytoplasmic tail of programmed death-1. Active Src homology 2 domain-containing tyrosine phosphatase 2 impairs the signaling function of the PI3K/protein kinase B (AKT) pathway, leading to functional defect of mTORC1, downregulation of CXCR3 expression, and suppression of T1D. Thus, mTORC1 component of the metabolic pathway serves as a target for chemokine receptor-mediated T cell tolerance and suppression of T1D. Copyright © 2016 by The American Association of Immunologists, Inc.
Spriggs, M K; Lioubin, P J; Slack, J; Dower, S K; Jonas, U; Cosman, D; Sims, J E; Bauer, J
1990-12-25
Primary human monocytes and the human monocytic cell line THP-1 were induced to express receptors for interleukin-1 alpha (IL-1 alpha) and IL-1 beta. Treatment of primary monocytes with dexamethasone resulted in a 10-fold increase in receptor number over untreated cells, to approximately 2,000 receptors/cell. Treatment of THP-1 cells with phorbol ester followed by prostaglandin E2 and dexamethasone resulted in the expression of approximately 30,000 receptors/cell. Competitive binding assays on THP-1 cells showed that both IL-1 alpha and IL-1 beta bind to the same receptor. The monocyte IL-1R is significantly smaller (63 kDa) than the T cell IL-1R (80 kDa) and is immunologically distinct. However, induction of monocytes and monocytic cell lines leads to the appearance of an abundant mRNA of approximately 5,000 bases which hybridizes to a cDNA probe from the T cell-type IL-1R. Sequence data obtained from a cDNA clone of this mRNA indicate that the message is identical to the T cell IL-1R mRNA throughout the coding region. A smaller mRNA, also homologous to the T cell IL-1R mRNA, accumulated in induced THP-1 cells and has a shorter 3'-untranslated region than the larger. Data are presented which suggest that neither form of this message encodes the 63-kDa IL-1R, but rather that this protein is the product of a separate nonhomologous mRNA.
Hypercholesterolemia induces T cell expansion in humanized immune mice.
Proto, Jonathan D; Doran, Amanda C; Subramanian, Manikandan; Wang, Hui; Zhang, Mingyou; Sozen, Erdi; Rymond, Christina C; Kuriakose, George; D'Agati, Vivette; Winchester, Robert; Sykes, Megan; Yang, Yong-Guang; Tabas, Ira
2018-06-01
Emerging data suggest that hypercholesterolemia has stimulatory effects on adaptive immunity and that these effects can promote atherosclerosis and perhaps other inflammatory diseases. However, research in this area has relied primarily on inbred strains of mice whose adaptive immune system can differ substantially from that of humans. Moreover, the genetically induced hypercholesterolemia in these models typically results in plasma cholesterol levels that are much higher than those in most humans. To overcome these obstacles, we studied human immune system-reconstituted mice (hu-mice) rendered hypercholesterolemic by treatment with adeno-associated virus 8-proprotein convertase subtilisin/kexin type 9 (AAV8-PCSK9) and a high-fat/high-cholesterol Western-type diet (WD). These mice had a high percentage of human T cells and moderate hypercholesterolemia. Compared with hu-mice that had lower plasma cholesterol, the PCSK9-WD mice developed a T cell-mediated inflammatory response in the lung and liver. Human CD4+ and CD8+ T cells bearing an effector memory phenotype were significantly elevated in the blood, spleen, and lungs of PCSK9-WD hu-mice, whereas splenic and circulating regulatory T cells were reduced. These data show that moderately high plasma cholesterol can disrupt human T cell homeostasis in vivo. This process may not only exacerbate atherosclerosis, but also contribute to T cell-mediated inflammatory diseases in the hypercholesterolemia setting.
Li, Guangming; Cheng, Menglan; Nunoya, Jun-ichi; Cheng, Liang; Guo, Haitao; Yu, Haisheng; Liu, Yong-jun; Su, Lishan; Zhang, Liguo
2014-01-01
The role of plasmacytoid dendritic cells (pDC) in human immunodeficiency virus type 1 (HIV-1) infection and pathogenesis remains unclear. HIV-1 infection in the humanized mouse model leads to persistent HIV-1 infection and immunopathogenesis, including type I interferons (IFN-I) induction, immune-activation and depletion of human leukocytes, including CD4 T cells. We developed a monoclonal antibody that specifically depletes human pDC in all lymphoid organs in humanized mice. When pDC were depleted prior to HIV-1 infection, the induction of IFN-I and interferon-stimulated genes (ISGs) were abolished during acute HIV-1 infection with either a highly pathogenic CCR5/CXCR4-dual tropic HIV-1 or a standard CCR5-tropic HIV-1 isolate. Consistent with the anti-viral role of IFN-I, HIV-1 replication was significantly up-regulated in pDC-depleted mice. Interestingly, the cell death induced by the highly pathogenic HIV-1 isolate was severely reduced in pDC-depleted mice. During chronic HIV-1 infection, depletion of pDC also severely reduced the induction of IFN-I and ISGs, associated with elevated HIV-1 replication. Surprisingly, HIV-1 induced depletion of human immune cells including T cells in lymphoid organs, but not the blood, was reduced in spite of the increased viral replication. The increased cell number in lymphoid organs was associated with a reduced level of HIV-induced cell death in human leukocytes including CD4 T cells. We conclude that pDC play opposing roles in suppressing HIV-1 replication and in promoting HIV-1 induced immunopathogenesis. These findings suggest that pDC-depletion and IFN-I blockade will provide novel strategies for treating those HIV-1 immune non-responsive patients with persistent immune activation despite effective anti-retrovirus treatment. PMID:25077616
Romo1 expression contributes to oxidative stress-induced death of lung epithelial cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Shin, Jung Ar; Chung, Jin Sil; Cho, Sang-Ho
Highlights: •Romo1 mediates oxidative stress-induced mitochondrial ROS production. •Romo1 induction by oxidative stress plays an important role in oxidative stress-induced apoptosis. •Romo1 overexpression correlates with epithelial cell death in patients with IPF. -- Abstract: Oxidant-mediated death of lung epithelial cells due to cigarette smoking plays an important role in pathogenesis in lung diseases such as idiopathic pulmonary fibrosis (IPF). However, the exact mechanism by which oxidants induce epithelial cell death is not fully understood. Reactive oxygen species (ROS) modulator 1 (Romo1) is localized in the mitochondria and mediates mitochondrial ROS production through complex III of the mitochondrial electron transport chain.more » Here, we show that Romo1 mediates mitochondrial ROS production and apoptosis induced by oxidative stress in lung epithelial cells. Hydrogen peroxide (H{sub 2}O{sub 2}) treatment increased Romo1 expression, and Romo1 knockdown suppressed the cellular ROS levels and cell death triggered by H{sub 2}O{sub 2} treatment. In immunohistochemical staining of lung tissues from patients with IPF, Romo1 was mainly localized in hyperplastic alveolar and bronchial epithelial cells. Romo1 overexpression was detected in 14 of 18 patients with IPF. TUNEL-positive alveolar epithelial cells were also detected in most patients with IPF but not in normal controls. These findings suggest that Romo1 mediates apoptosis induced by oxidative stress in lung epithelial cells.« less
Core/shell Fe3O4/Gd2O3 nanocubes as T1-T2 dual modal MRI contrast agents
NASA Astrophysics Data System (ADS)
Li, Fenfen; Zhi, Debo; Luo, Yufeng; Zhang, Jiqian; Nan, Xiang; Zhang, Yunjiao; Zhou, Wei; Qiu, Bensheng; Wen, Longping; Liang, Gaolin
2016-06-01
T1-T2 dual modal magnetic resonance imaging (MRI) has attracted considerable interest because it offers complementary diagnostic information, leading to more precise diagnosis. To date, a number of nanostructures have been reported as T1-T2 dual modal MR contrast agents (CAs). However, hybrids of nanocubes with both iron and gadolinium (Gd) elements as T1-T2 dual modal CAs have not been reported. Herein, we report the synthesis of novel core/shell Fe3O4/Gd2O3 nanocubes as T1-T2 dual-modal CAs and their application for enhanced T1-T2 MR imaging of rat livers. A relaxivity study at 1.5 T indicated that our Fe3O4/Gd2O3 nanocubes have an r1 value of 45.24 mM-1 s-1 and an r2 value of 186.51 mM-1 s-1, which were about two folds of those of Gd2O3 nanoparticles and Fe3O4 nanocubes, respectively. In vivo MR imaging of rats showed both T1-positive and T2-negative contrast enhancements in the livers. We envision that our Fe3O4/Gd2O3 nanocubes could be applied as T1-T2 dual modal MR CAs for a wide range of theranostic applications in the near future.T1-T2 dual modal magnetic resonance imaging (MRI) has attracted considerable interest because it offers complementary diagnostic information, leading to more precise diagnosis. To date, a number of nanostructures have been reported as T1-T2 dual modal MR contrast agents (CAs). However, hybrids of nanocubes with both iron and gadolinium (Gd) elements as T1-T2 dual modal CAs have not been reported. Herein, we report the synthesis of novel core/shell Fe3O4/Gd2O3 nanocubes as T1-T2 dual-modal CAs and their application for enhanced T1-T2 MR imaging of rat livers. A relaxivity study at 1.5 T indicated that our Fe3O4/Gd2O3 nanocubes have an r1 value of 45.24 mM-1 s-1 and an r2 value of 186.51 mM-1 s-1, which were about two folds of those of Gd2O3 nanoparticles and Fe3O4 nanocubes, respectively. In vivo MR imaging of rats showed both T1-positive and T2-negative contrast enhancements in the livers. We envision that our Fe3O4/Gd2O3 nanocubes
Chiasson, Valorie L; Pakanati, Abhinandan R; Hernandez, Marcos; Young, Kristina J; Bounds, Kelsey R; Mitchell, Brett M
2017-07-01
The immunosuppressive calcineurin inhibitors cyclosporine A and tacrolimus alter T-cell subsets and can cause hypertension, vascular dysfunction, and renal toxicity. We and others have reported that cyclosporine A and tacrolimus decrease anti-inflammatory regulatory T cells and increase proinflammatory interleukin-17-producing T cells; therefore, we hypothesized that inhibition of these effects using noncellular therapies would prevent the hypertension, endothelial dysfunction, and renal glomerular injury induced by calcineurin inhibitor therapy. Daily treatment of mice with cyclosporine A or tacrolimus for 1 week significantly decreased CD4 + /FoxP3 + regulatory T cells in the spleen and lymph nodes, as well as induced hypertension, vascular injury and dysfunction, and glomerular mesangial expansion in mice. Daily cotreatment with all-trans retinoic acid reported to increase regulatory T cells and decrease interleukin-17-producing T cells, prevented all of the detrimental effects of cyclosporine A and tacrolimus. All-trans retinoic acid also increased regulatory T cells and prevented the hypertension, endothelial dysfunction, and glomerular injury in genetically modified mice that phenocopy calcineurin inhibitor-treated mice (FKBP12-Tie2 knockout). Treatment with an interleukin-17-neutralizing antibody also increased regulatory T-cell levels and prevented the hypertension, endothelial dysfunction, and glomerular injury in cyclosporine A-treated and tacrolimus-treated mice and FKBP12-Tie2 knockout mice, whereas an isotype control had no effect. Augmenting regulatory T cells and inhibiting interleukin-17 signaling using noncellular therapies prevents the cardiovascular and renal toxicity of calcineurin inhibitors in mice. © 2017 American Heart Association, Inc.
Properties of HTLV-I transformed CD8+ T-cells in response to HIV-1 infection.
Gulzar, N; Shroff, A; Buberoglu, B; Klonowska, D; Kim, J E; Copeland, K F T
2010-10-25
HIV-1 infection studies of primary CD8(+) T-cells are hampered by difficulty in obtaining a significant number of targets for infection and low levels of productive infection. Further, there exists a paucity of CD8-expressing T-cell lines to address questions pertaining to the study of CD8(+) T-cells in the context of HIV-1 infection. In this study, a set of CD8(+) T-cell clones were originated through HTLV-I transformation in vitro, and the properties of these cells were examined. The clones were susceptible to T-cell tropic strains of the virus and exhibited HIV-1 production 20-fold greater than primary CD4(+) T-cells. Productive infection resulted in a decrease in expression of CD8 and CXCR4 molecules on the surface of the CD8(+) T-cell clones and antibodies to these molecules abrogated viral binding and replication. These transformed cells provide an important tool in the study of CD8(+) T-cells and may provide important insights into the mechanism(s) behind HIV-1 induced CD8(+) T-cell dysfunction. Copyright © 2010 Elsevier Inc. All rights reserved.
Krenger, W; Snyder, K M; Byon, J C; Falzarano, G; Ferrara, J L
1995-07-15
Acute graft-vs-host disease (GVHD) is thought to be mediated by alloreactive T cells with a type 1 cytokine phenotype. To prevent the development of acute GVHD, we have successfully polarized mature donor T cells toward a type 2 cytokine phenotype ex-vivo by incubating them with murine rIL-4 in a primary MLC. Polarized type 2 T cells were then transplanted with T cell-depleted bone marrow cells into irradiated recipients across either MHC class II (bm12-->C57BL/6) or class I (bm1-->C57BL/6) barriers, and the intensity of GVHD was measured by assessment of several in vitro and in vivo parameters. The injection of polarized type 2 T cells abrogated the mitogen-induced production of IFN-gamma by splenocytes from transplanted hosts on day 13 after bone marrow transplantation (BMT). Injection of polarized type 2 T cells failed to induce secretion of the effector phase cytokine TNF-alpha by splenocytes stimulated with LPS both in vitro and in vivo, and survival of transplanted mice after i.v. injection with LPS was significantly improved. Furthermore, cell-mixing experiments revealed that polarized type 2 T cells were able to inhibit type 1 cytokine responses induced by naive T cells after BMT. These data demonstrate that both polarized CD4+ and CD8+ type 2 alloreactive donor T cells can be generated in vitro from mature T cell populations. These cells function in vivo to inhibit type 1 T cell responses, and such inhibition attenuates the systemic morbidity of GVHD after BMT across both MHC class II or class I barriers in mice.
Inhibition of the K+ channel KCa3.1 ameliorates T cell-mediated colitis.
Di, Lie; Srivastava, Shekhar; Zhdanova, Olga; Ding, Yi; Li, Zhai; Wulff, Heike; Lafaille, Maria; Skolnik, Edward Y
2010-01-26
The calcium-activated K(+) channel KCa3.1 plays an important role in T lymphocyte Ca(2+) signaling by helping to maintain a negative membrane potential, which provides an electrochemical gradient to drive Ca(2+) influx. To assess the role of KCa3.1 channels in lymphocyte activation in vivo, we studied T cell function in KCa3.1(-/-) mice. CD4 T helper (i.e., Th0) cells isolated from KCa3.1(-/-) mice lacked KCa3.1 channel activity, which resulted in decreased T cell receptor-stimulated Ca(2+) influx and IL-2 production. Although loss of KCa3.1 did not interfere with CD4 T cell differentiation, both Ca(2+) influx and cytokine production were impaired in KCa3.1(-/-) Th1 and Th2 CD4 T cells, whereas T-regulatory and Th17 function were normal. We found that inhibition of KCa3.1(-/-) protected mice from developing severe colitis in two mouse models of inflammatory bowel disease, which were induced by (i) the adoptive transfer of mouse naïve CD4 T cells into rag2(-/-) recipients and (ii) trinitrobenzene sulfonic acid. Pharmacologic inhibitors of KCa3.1 have already been shown to be safe in humans. Thus, if these preclinical studies continue to show efficacy, it may be possible to rapidly test whether KCa3.1 inhibitors are efficacious in patients with inflammatory bowel diseases such as Crohn's disease and ulcerative colitis.
2017-01-01
Herpes simplex virus type 1 (HSV-1) latency in sensory ganglia such as trigeminal ganglia (TG) is associated with a persistent immune infiltrate that includes effector memory CD8+ T cells that can influence HSV-1 reactivation. In C57BL/6 mice, HSV-1 induces a highly skewed CD8+ T cell repertoire, in which half of CD8+ T cells (gB-CD8s) recognize a single epitope on glycoprotein B (gB498-505), while the remainder (non-gB-CD8s) recognize, in varying proportions, 19 subdominant epitopes on 12 viral proteins. The gB-CD8s remain functional in TG throughout latency, while non-gB-CD8s exhibit varying degrees of functional compromise. To understand how dominance hierarchies relate to CD8+ T cell function during latency, we characterized the TG-associated CD8+ T cells following corneal infection with a recombinant HSV-1 lacking the immunodominant gB498-505 epitope (S1L). S1L induced a numerically equivalent CD8+ T cell infiltrate in the TG that was HSV-specific, but lacked specificity for gB498-505. Instead, there was a general increase of non-gB-CD8s with specific subdominant epitopes arising to codominance. In a latent S1L infection, non-gB-CD8s in the TG showed a hierarchy targeting different epitopes at latency compared to at acute times, and these cells retained an increased functionality at latency. In a latent S1L infection, these non-gB-CD8s also display an equivalent ability to block HSV reactivation in ex vivo ganglionic cultures compared to TG infected with wild type HSV-1. These data indicate that loss of the immunodominant gB498-505 epitope alters the dominance hierarchy and reduces functional compromise of CD8+ T cells specific for subdominant HSV-1 epitopes during viral latency. PMID:29206240
DOE Office of Scientific and Technical Information (OSTI.GOV)
Hasegawa, Kazuhiro; Wakino, Shu; Yoshioka, Kyoko
2008-07-18
NAD{sup +}-dependent protein deacetylase Sirt1 regulates cellular apoptosis. We examined the role of Sirt1 in renal tubular cell apoptosis by using HK-2 cells, proximal tubular cell lines with or without reactive oxygen species (ROS), H{sub 2}O{sub 2}. Without any ROS, Sirt1 inhibitors enhanced apoptosis and the expression of ROS scavenger, catalase, and Sirt1 overexpression downregulated catalase. When apoptosis was induced with H{sub 2}O{sub 2}, Sirt1 was upregulated with the concomitant increase in catalase expression. Sirt1 overexpression rescued H{sub 2}O{sub 2}-induced apoptosis through the upregulation of catalase. H{sub 2}O{sub 2} induced the nuclear accumulation of forkhead transcription factor, FoxO3a and themore » gene silencing of FoxO3a enhanced H{sub 2}O{sub 2}-induced apoptosis. In conclusion, endogenous Sirt1 maintains cell survival by regulating catalase expression and by preventing the depletion of ROS required for cell survival. In contrast, excess ROS upregulates Sirt1, which activates FoxO3a and catalase leading to rescuing apoptosis. Thus, Sirt1 constitutes a determinant of renal tubular cell apoptosis by regulating cellular ROS levels.« less
γδ T cell homeostasis is established in competition with αβ T cells and NK cells
French, Jena D.; Roark, Christina L.; Born, Willi K.; O'Brien, Rebecca L.
2005-01-01
γδ T cells are a diverse population of lymphocytes that play an important role in immune regulation. The size of the γδ T cell pool is tightly regulated, comprising only 1-10% of total lymphoid T cells in mice and humans. We examined the homeostatic regulation of γδ T cells using a model of lymphopenia-induced homeostatic expansion. We found that IL-15 and, to a lesser extent, IL-7 play an important role in lymphoid γδ T cell homeostasis. Moreover, γδ T cell homeostatic expansion was limited not only by γδ T cells themselves but also by natural killer cells and αβ T cells. Our results suggest that CD8+ αβ T cells are the most potent inhibitors of γδ T cell homeostasis and exert their effect by competing for IL-15. PMID:16203967
Schindler, Tobias; Kagina, Benjamin M.; Zhang, Jitao David; Lukindo, Tedson; Mpina, Maxmillian; Bang, Peter; Kromann, Ingrid; Hoff, Søren T.; Andersen, Peter; Reither, Klaus; Churchyard, Gavin J.; Certa, Ulrich
2015-01-01
Tuberculosis (TB) remains a global health problem, with vaccination being a necessary strategy for disease containment and elimination. A TB vaccine should be safe and immunogenic as well as efficacious in all affected populations, including HIV-infected individuals. We investigated the induction and maintenance of vaccine-induced memory CD4+ T cells following vaccination with the subunit vaccine H1/IC31. H1/IC31 was inoculated twice on study days 0 and 56 among HIV-infected adults with CD4+ lymphocyte counts of >350 cells/mm3. Whole venous blood stimulation was conducted with the H1 protein, and memory CD4+ T cells were analyzed using intracellular cytokine staining and polychromatic flow cytometry. We identified high responders, intermediate responders, and nonresponders based on detection of interleukin-2 (IL-2), tumor necrosis factor alpha (TNF-α), and gamma interferon (IFN-γ) expressing central (TCM) and effector memory CD4+ T cells (TEM) 182 days after the first immunization. Amplicon-based transcript quantification using next-generation sequencing was performed to identify differentially expressed genes that correlated with vaccine-induced immune responses. Genes implicated in resolution of inflammation discriminated the responders from the nonresponders 3 days after the first inoculation. The volunteers with higher expression levels of genes involved in antiviral innate immunity at baseline showed impaired H1-specific TCM and TEM maintenance 6 months after vaccination. Our study showed that in HIV-infected volunteers, expression levels of genes involved in the antiviral innate immune response affected long-term maintenance of H1/IC31 vaccine-induced cellular immunity. (The clinical trial was registered in the Pan African Clinical Trials Registry [PACTR] with the identifier PACTR201105000289276.) PMID:25924764
Lopez, Pamela Aranda; Denny, Mark; Hartmann, Ann-Kathrin; Alflen, Astrid; Probst, Hans Christian; von Stebut, Esther; Tenzer, Stefan; Schild, Hansjörg; Stassen, Michael; Langguth, Peter; Radsak, Markus P
2017-09-01
Transcutaneous immunization (TCI) is a novel vaccination strategy utilizing the skin associated lymphatic tissue to induce immune responses. TCI using a cytotoxic T lymphocyte (CTL) epitope and the Toll-like receptor 7 (TLR7) agonist imiquimod mounts strong CTL responses by activation and maturation of skin-derived dendritic cells (DCs) and their migration to lymph nodes. However, TCI based on the commercial formulation Aldara only induces transient CTL responses that needs further improvement for the induction of durable therapeutic immune responses. Therefore we aimed to develop a novel imiquimod solid nanoemulsion (IMI-Sol) for TCI with superior vaccination properties suited to induce high quality T cell responses for enhanced protection against infections. TCI was performed by applying a MHC class I or II restricted epitope along with IMI-Sol or Aldara (each containing 5% Imiquimod) on the shaved dorsum of C57BL/6, IL-1R, Myd88, Tlr7 or Ccr7 deficient mice. T cell responses as well as DC migration upon TCI were subsequently analyzed by flow cytometry. To determine in vivo efficacy of TCI induced immune responses, CTL responses and frequency of peptide specific T cells were evaluated on day 8 or 35 post vaccination and protection in a lymphocytic choriomeningitis virus (LCMV) infection model was assessed. TCI with the imiquimod formulation IMI-Sol displayed equal skin penetration of imiquimod compared to Aldara, but elicited superior CD8 + as well as CD4 + T cell responses. The induction of T-cell responses induced by IMI-Sol TCI was dependent on the TLR7/MyD88 pathway and independent of IL-1R. IMI-Sol TCI activated skin-derived DCs in skin-draining lymph nodes more efficiently compared to Aldara leading to enhanced protection in a LCMV infection model. Our data demonstrate that IMI-Sol TCI can overcome current limitations of previous imiquimod based TCI approaches opening new perspectives for transcutaneous vaccination strategies and allowing the use of this
Antibody induced CD4 down-modulation of T cells is site-specifically mediated by CD64+ cells
Vogel, Stephanie; Grabski, Elena; Buschjäger, Daniela; Klawonn, Frank; Döring, Marius; Wang, Junxi; Fletcher, Erika; Bechmann, Ingo; Witte, Torsten; Durisin, Martin; Schraven, Burkhart; Mangsbo, Sara M.; Schönfeld, Kurt; Czeloth, Niklas; Kalinke, Ulrich
2015-01-01
Treatment of PBMC with the CD4-specific mAb BT-061 induces CD4 down-modulation of T cells. Here we report that addition of BT-061 to purified T cells did not confer this effect, whereas incubation of T cells in BT-061 coated wells restored CD4 down-modulation. These results implied that Fcγ receptor mediated cell-cell interactions played a role. In consistence with this hypothesis PBMC depleted of CD64+ monocytes did not confer CD4 down-modulation of BT-061 decorated T cells. Strikingly, CD4 down-modulation was observed in BT-061 treated synovial fluid punctuated from patients’ inflamed joints that comprised enhanced numbers of CD64+ cells. In contrast, in a circulating whole blood system injection of BT-061 did not induce CD4 down-modulation, due to CD64 saturation by serum IgG. Similarly, tonsil derived mononuclear cells devoid of CD64+ cells did not show CD4 down-modulation, whereas addition of blood derived monocytes restored the effect. Thus, the interaction of BT-061 decorated T cells with CD64+ cells is needed for CD4 down-modulation, implying that in patients BT-061 would primarily induce CD4 down-modulation at inflammatory sites. These results highlight the need not only to examine the interaction of a given mAb with single FcγR, but also the immunological environment that is appropriate to support such interactions. PMID:26670584
DeTemple, Daphne E.; Oldhafer, Felix; Falk, Christine S.; Chen‐Wacker, Chen; Figueiredo, Constanca; Kleine, Moritz; Ramackers, Wolf; Timrott, Kai; Lehner, Frank; Klempnauer, Juergen; Bock, Michael
2018-01-01
Hepatocyte transplantation is a promising therapeutic approach for various liver diseases. Despite the liver's tolerogenic potential, early immune‐mediated loss of transplanted cells is observed, and longterm acceptance has not been achieved yet. Patients deemed tolerant after liver transplantation presented an increased frequency of regulatory T cells (Tregs), which therefore also might enable reduction of posttransplant cell loss and enhance longterm allograft acceptance. We hence characterized hepatocyte‐induced immune reactions and evaluated the immunomodulatory potential of Tregs applying mixed lymphocyte cultures and mixed lymphocyte hepatocyte cultures. These were set up using peripheral blood mononuclear cells and primary human hepatocytes, respectively. Polyclonally expanded CD4+CD25highCD127low Tregs were added to cocultures in single‐/trans‐well setups with/without supplementation of anti‐interferon γ (IFNγ) antibodies. Hepatocyte‐induced alloresponses were then analyzed by multicolor flow cytometry. Measurements indicated that T cell response upon stimulation was associated with IFNγ‐induced major histocompatibility complex (MHC) class II up‐regulation on hepatocytes and mediated by CD4+ T cells. An indirect route of antigen presentation could be ruled out by use of fragmented hepatocytes and culture supernatants of hepatocytes. Allospecific proliferation was accompanied by inflammatory cytokine secretion. CD8+ T cells showed early up‐regulation of CD69 despite lack of cell proliferation in the course of coculture. Supplementation of Tregs effectively abrogated hepatocyte‐induced alloresponses and was primarily cell contact dependent. In conclusion, human hepatocytes induce a CD4+ T cell alloresponse in vitro, which is associated with MHC class II up‐regulation on hepatocytes and is susceptible to suppression by Tregs. Liver Transplantation 24 407–419 2018 AASLD. PMID:29365365
Saito, Masako; Nagasawa, Masayuki; Takada, Hidetoshi; Hara, Toshiro; Tsuchiya, Shigeru; Agematsu, Kazunaga; Yamada, Masafumi; Kawamura, Nobuaki; Ariga, Tadashi; Tsuge, Ikuya; Nonoyama, Shigeaki; Karasuyama, Hajime
2011-01-01
Hyper-IgE syndrome (HIES) is a primary immunodeficiency characterized by recurrent staphylococcal infections and atopic dermatitis associated with elevated serum IgE levels. Although defective differentiation of IL-17–producing CD4+ T cells (Th17) partly accounts for the susceptibility to staphylococcal skin abscesses and pneumonia, the pathogenesis of atopic manifestations in HIES still remains an enigma. In this study, we examined the differentiation and function of Th1, Th2, regulatory T cells (Treg cells), and dendritic cells (DCs) in HIES patients carrying either STAT3 or TYK2 mutations. Although the in vitro differentiation of Th1 and Th2 cells and the number and function of Treg cells in the peripheral blood were normal in HIES patients with STAT3 mutations, primary and monocyte-derived DCs showed defective responses to IL-10 and thus failed to become tolerogenic. When treated with IL-10, patient DCs showed impaired up-regulation of inhibitory molecules on their surface, including PD-L1 and ILT-4, compared with control DCs. Moreover, IL-10–treated DCs from patients displayed impaired ability to induce the differentiation of naive CD4+ T cells to FOXP3+ induced Treg cells (iTreg cells). These results suggest that the defective generation of IL-10–induced tolerogenic DCs and iTreg cells may contribute to inflammatory changes in HIES. PMID:21300911
Neurotoxicity Induced by Bupivacaine via T-Type Calcium Channels in SH-SY5Y Cells
Wen, Xianjie; Xu, Shiyuan; Liu, Hongzhen; Zhang, Quinguo; Liang, Hua; Yang, Chenxiang; Wang, Hanbing
2013-01-01
There is concern regarding neurotoxicity induced by the use of local anesthetics. A previous study showed that an overload of intracellular calcium is involved in the neurotoxic effect of some anesthetics. T-type calcium channels, which lower the threshold of action potentials, can regulate the influx of calcium ions. We hypothesized that T-type calcium channels are involved in bupivacaine-induced neurotoxicity. In this study, we first investigated the effects of different concentrations of bupivacaine on SH-SY5Y cell viability, and established a cell injury model with 1 mM bupivacaine. The cell viability of SH-SY5Y cells was measured following treatment with 1 mM bupivacaine and/or different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride, an antagonist of T-type calcium channels for 24 h. In addition, we monitored the release of lactate dehydrogenase, cytosolic Ca2+ ([Ca2+]i), cell apoptosis and caspase-3 expression. SH-SY5Y cells pretreated with different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride improved cell viability, reduced lactate dehydrogenase release, inhibited apoptosis, and reduced caspase-3 expression following bupivacaine exposure. However, the protective effect of NNC 55-0396 dihydrochloride plateaued. Overall, our results suggest that T-type calcium channels may be involved in bupivacaine neurotoxicity. However, identification of the specific subtype of T calcium channels involved requires further investigation. PMID:23658789
The Role of B Cells for in Vivo T Cell Responses to a Friend Virus-Induced Leukemia
NASA Astrophysics Data System (ADS)
Schultz, Kirk R.; Klarnet, Jay P.; Gieni, Randall S.; Hayglass, Kent T.; Greenberg, Philip D.
1990-08-01
B cells can function as antigen-presenting cells and accessory cells for T cell responses. This study evaluated the role of B cells in the induction of protective T cell immunity to a Friend murine leukemia virus (F-MuLV)-induced leukemia (FBL). B cell-deficient mice exhibited significantly reduced tumor-specific CD4^+ helper and CD8^+ cytotoxic T cell responses after priming with FBL or a recombinant vaccinia virus containing F-MuLV antigens. Moreover, these mice had diminished T cell responses to the vaccinia viral antigens. Tumor-primed T cells transferred into B cell-deficient mice effectively eradicated disseminated FBL. Thus, B cells appear necessary for efficient priming but not expression of tumor and viral T cell immunity.
Sanuki, Rina; Shionome, Chieko; Kuwabara, Akiko; Mitsui, Narihiro; Koyama, Yuki; Suzuki, Naoto; Zhang, Fan; Shimizu, Noriyoshi; Maeno, Masao
2010-04-01
In orthodontic tooth movement, prostaglandin E(2) (PGE(2)) released from osteoblasts can alter the normal process of bone remodeling. We previously showed that compressive force (CF) controls bone formation by stimulating the production of PGE(2) and Ep2 and/or Ep4 receptors in osteoblasts. The present study was undertaken to examine the effect of CF on the production of PGE(2), cyclooxygenase-2 (COX-2), macrophage colony-stimulating factor (M-CSF), receptor activator of NF-kappaB ligand (RANKL), and osteoprotegerin (OPG) using osteoblastic MC3T3-E1 cells and to examine the indirect effect of CF on osteoclast differentiation using RAW264.7 cells as osteoclast precursors. MC3T3-E1 cells were cultured with or without continuous CF (1.0 or 3.0 g/cm(2)) for 24 hr, and PGE(2) production was determined using ELISA. The expression of COX-2, M-CSF, RANKL, and OPG genes and proteins was determined using real-time PCR and ELISA, respectively. Osteoclast differentiation was estimated using tartrate-resistant acid phosphatase (TRAP) staining of RAW 264.7 cells cultured for 10 days with conditioned medium from CF-treated MC3T3-E1 cells and soluble RANKL. As CF increased, PGE(2) production and the expression of COX-2, M-CSF, and RANKL increased, whereas OPG expression decreased. The number of TRAP-positive cells increased as CF increased. Celecoxib, a specific inhibitor of COX-2, blocked the stimulatory effect of CF on TRAP staining and the production of PGE(2), M-CSF, RANKL, and OPG. These results suggest that CF induces osteoclast differentiation by increasing M-CSF production and decreasing OPG production via PGE(2) in osteoblasts.
Bégin, Philippe; Schulze, Janika; Baron, Udo; Olek, Sven; Bauer, Rebecca N; Passerini, Laura; Baccheta, Rosa; Nadeau, Kari C
2015-01-01
The FOXP3 gene is the master regulator for T regulatory cells and is under tight DNA methylation control at the Treg specific demethylated region (TSDR) in its first intron. This said, methylation of its promoter region, the significance of which is unknown, has also been associated with various immune-related disease states such as asthma, food allergy, auto-immunity and cancer. Here, we used induced T regulatory cells (iTreg) as a target cell population to identify candidate hypomethylated CpG sites in the FOXP3 gene promoter to design a DNA methylation quantitative assay for this region. Three CpG sites at the promoter region showed clear demethylation pattern associated with high FOXP3 expression after activation in presence of TGFβ and were selected as primary targets to design methylation-dependent RT-PCR primers and probes. We then examined the methylation of this 'inducible-promoter-demethylated-region' (IPDR) in various FOXP3+ T cell subsets. Both naïve and memory thymic-derived Treg cells were found to be fully demethylated at both the IPDR and TSDR. Interestingly, in addition to iTregs, both CD25- and CD25(lo) conventional memory CD4+CD45RA- T cells displayed a high fraction of IPDR demethylated cells in absence of TSDR demethylation. This implies that the fraction of memory T cells should be taken in account when interpreting FOXP3 promoter methylation results from clinical studies. This approach, which is available for testing in clinical samples could have diagnostic and prognostic value in patients with immune or auto-inflammatory diseases.
Candolfi, Marianela; Curtin, James F; Yagiz, Kader; Assi, Hikmat; Wibowo, Mia K; Alzadeh, Gabrielle E; Foulad, David; Muhammad, A K M G; Salehi, Sofia; Keech, Naomi; Puntel, Mariana; Liu, Chunyan; Sanderson, Nicholas R; Kroeger, Kurt M; Dunn, Robert; Martins, Gislaine; Lowenstein, Pedro R; Castro, Maria G
2011-10-01
We have demonstrated that modifying the tumor microenvironment through intratumoral administration of adenoviral vectors (Ad) encoding the conditional cytotoxic molecule, i.e., HSV1-TK and the immune-stimulatory cytokine, i.e., fms-like tyrosine kinase 3 ligand (Flt3L) leads to T-cell-dependent tumor regression in rodent models of glioblastoma. We investigated the role of B cells during immune-mediated glioblastoma multiforme regression. Although treatment with Ad-TK+Ad-Flt3L induced tumor regression in 60% of wild-type (WT) mice, it completely failed in B-cell-deficient Igh6(-/-) mice. Tumor-specific T-cell precursors were detected in Ad-TK+Ad-Flt3L-treated WT mice but not in Igh6(-/-) mice. The treatment also failed in WT mice depleted of total B cells or marginal zone B cells. Because we could not detect circulating antibodies against tumor cells and the treatment was equally efficient in WT mice and in mice with B-cell-specific deletion of Prdm 1 (encoding Blimp-1), in which B cells are present but unable to fully differentiate into antibody-secreting plasma cells, tumor regression in this model is not dependent on B cells' production of tumor antigen-specific immunoglobulins. Instead, B cells seem to play a role as antigen-presenting cells (APCs). Treatment with Ad-TK+Ad-Flt3L led to an increase in the number of B cells in the cervical lymph nodes, which stimulated the proliferation of syngeneic T cells and induced clonal expansion of antitumor T cells. Our data show that B cells act as APCs, playing a critical role in clonal expansion of tumor antigen-specific T cells and brain tumor regression.
Wells, James W; Cowled, Chris J; Darling, David; Guinn, Barbara-Ann; Farzaneh, Farzin; Noble, Alistair; Galea-Lauri, Joanna
2007-12-01
Alloreactive T-cell responses are known to result in the production of large amounts of proinflammatory cytokines capable of activating and maturing dendritic cells (DC). However, it is unclear whether these allogeneic responses could also act as an adjuvant for concurrent antigen-specific responses. To examine effects of simultaneous alloreactive and antigen-specific T-cell responses induced by semi-allogeneic DC. Semi-allogeneic DC were generated from the F(1) progeny of inbred strains of mice (C57BL/6 and C3H, or C57BL/6 and DBA). We directly primed antigen-specific CD8(+) and CD4(+) T-cells from OT-I and OT-II mice, respectively, in the absence of allogeneic responses, in vitro, and in the presence or absence of alloreactivity in vivo. In vitro, semi-allogeneic DC cross-presented ovalbumin (OVA) to naïve CD8(+) OT-I transgenic T-cells, primed naïve CD4(+) OT-II transgenic T-cells and could stimulate strong alloreactive T-cell proliferation in a primary mixed lymphocyte reaction (MLR). In vivo, semi-allogeneic DC migrated efficiently to regional lymph nodes but did not survive there as long as autologous DC. In addition, they were not able to induce cytotoxic T-lymphocyte (CTL) activity to a target peptide, and only weakly stimulated adoptively transferred OT-II cells. The CD4(+) response was unchanged in allo-tolerized mice, indicating that alloreactive T-cell responses could not provide help for concurrently activated antigen-specific responses. In an EL4 tumour-treatment model, vaccination with semi-allogeneic DC/EL4 fusion hybrids, but not allogeneic DC/EL4 hybrids, significantly increased mouse survival. Expression of self-Major histocompatibility complex (MHC) by semi-allogeneic DC can cause the induction of antigen-specific immunity, however, concurrently activated allogeneic bystander responses do not provide helper or adjuvant effects.
TGF-β induces surface LAP expression on murine CD4 T cells independent of Foxp3 induction.
Oida, Takatoku; Weiner, Howard L
2010-11-24
It has been reported that human FOXP3(+) CD4 Tregs express GARP-anchored surface latency-associated peptide (LAP) after activation, based on the use of an anti-human LAP mAb. Murine CD4 Foxp3(+) Tregs have also been reported to express surface LAP, but these studies have been hampered by the lack of suitable anti-mouse LAP mAbs. We generated anti-mouse LAP mAbs by immunizing TGF-β(-/-) animals with a mouse Tgfb1-transduced P3U1 cell line. Using these antibodies, we demonstrated that murine Foxp3(+) CD4 Tregs express LAP on their surface. In addition, retroviral transduction of Foxp3 into mouse CD4(+)CD25(-) T cells induced surface LAP expression. We then examined surface LAP expression after treating CD4(+)CD25(-) T cells with TGF-β and found that TGF-β induced surface LAP not only on T cells that became Foxp3(+) but also on T cells that remained Foxp3(-) after TGF-β treatment. GARP expression correlated with the surface LAP expression, suggesting that surface LAP is GARP-anchored also in murine T cells. Unlike human CD4 T cells, surface LAP expression on mouse CD4 T cells is controlled by Foxp3 and TGF-β. Our newly described anti-mouse LAP mAbs will provide a useful tool for the investigation and functional analysis of T cells that express LAP on their surface.
Jarmin, Sarah J.; David, Rachel; Ma, Liang; Chai, Jan-Guo; Dewchand, Hamlata; Takesono, Aya; Ridley, Anne J.; Okkenhaug, Klaus; Marelli-Berg, Federica M.
2008-01-01
The establishment of T cell–mediated inflammation requires the migration of primed T lymphocytes from the blood stream and their retention in antigenic sites. While naive T lymphocyte recirculation in the lymph and blood is constitutively regulated and occurs in the absence of inflammation, the recruitment of primed T cells to nonlymphoid tissue and their retention at the site are enhanced by various inflammatory signals, including TCR engagement by antigen-displaying endothelium and resident antigen-presenting cells. In this study, we investigated whether signals downstream of TCR ligation mediated by the phosphoinositide-3-kinase (PI3K) subunit p110δ contributed to the regulation of these events. T lymphocytes from mice expressing catalytically inactive p110δ displayed normal constitutive trafficking and migratory responses to nonspecific stimuli. However, these cells lost susceptibility to TCR-induced migration and failed to localize efficiently to antigenic tissue. Importantly, we showed that antigen-induced T cell trafficking and subsequent inflammation was abrogated by selective pharmacological inhibition of PI3K p110δ activity. These observations suggest that pharmacological targeting of p110δ activity is a viable strategy for the therapy of T cell–mediated pathology. PMID:18259608
Divalent Metal Ions Induced Osteogenic Differentiation of MC3T3E1
NASA Astrophysics Data System (ADS)
Wang, Guoshou; Su, Wenta; Chen, Pohung; Huang, Teyang
2017-12-01
Biomaterial scaffolds blended with biochemical signal molecules with adequate osteoinductive and osteoconductive properties have attracted significant interest in bone tissue engineering regeneration. The divalent metal ions can gradually release from the scaffold into the culture medium and then induced osteoblastic differentiation of MC3T3E1. These MC3T3E1 cells expressed high activity of alkaline phosphatase, bone-related gene expression of collagen type I, Runx2, osteopontin, osteocalcin, and significantly enhanced deposited minerals on scaffold after 21 days of culture. This experiment provided a useful inducer for osteogenic differentiation in bone repair.
Luz-Crawford, Patricia; Kurte, Monica; Bravo-Alegría, Javiera; Contreras, Rafael; Nova-Lamperti, Estefania; Tejedor, Gautier; Noël, Danièle; Jorgensen, Christian; Figueroa, Fernando; Djouad, Farida; Carrión, Flavio
2013-06-04
Mesenchymal stem cells (MSCs) are adult, multipotent, stem cells with immunomodulatory properties. The mechanisms involved in the capacity of MSCs to inhibit the proliferation of proinflammatory T lymphocytes, which appear responsible for causing autoimmune disease, have yet to be fully elucidated. One of the underlying mechanisms studied recently is the ability of MSCs to generate T regulatory (Treg) cells in vitro and in vivo from activated peripheral blood mononuclear cells (PBMC), T-CD4+ and also T-CD8(+) cells. In the present work we investigated the capacity of MSCs to generate Treg cells using T-CD4(+) cells induced to differentiate toward the proinflammatory Th1 and Th17 lineages. MSCs were obtained from mouse bone marrow and characterized according to their surface antigen expression and their multilineage differentiation potential. CD4(+) T cells isolated from mouse spleens were induced to differentiate into Th1 or Th17 cells and co-cultured with MSCs added at day 0, 2 or 4 of the differentiation processes. After six days, CD25, Foxp3, IL-17 and IFN-γ expression was assessed by flow cytometry and helios and neuropilin 1 mRNA levels were assessed by RT-qPCR. For the functional assays, the 'conditioned' subpopulation generated in the presence of MSCs was cultured with concanavalin A-activated CD4(+) T cells labeled with carboxyfluorescein succinimidyl ester. Finally, we used the encephalomyelitis autoimmune diseases (EAE) mouse model, in which mice were injected with MSCs at day 18 and 30 after immunization. At day 50, the mice were euthanized and draining lymph nodes were extracted for Th1, Th17 and Treg detection by flow cytometry. MSCs were able to suppress the proliferation, activation and differentiation of CD4(+) T cells induced to differentiate into Th1 and Th17 cells. This substantial suppressive effect was associated with an increase of the percentage of functional induced CD4(+)CD25(+)Foxp3(+) regulatory T cells and IL-10 secretion. However
Interleukin-29 induces epithelial production of CXCR3A ligands and T-cell infiltration.
Witte, Ellen; Kokolakis, Georgios; Witte, Katrin; Warszawska, Katarzyna; Friedrich, Markus; Christou, Demetrios; Kirsch, Stefan; Sterry, Wolfram; Volk, Hans-Dieter; Sabat, Robert; Wolk, Kerstin
2016-04-01
Psoriasis is considered as a model for chronic immune-mediated disorders. Th17-cells are pivotal players in those diseases. Recently, we demonstrated that Th17-cells produce interleukin (IL)-29 and that IL-29 is highly present in psoriatic lesions. Whether IL-29, with its action on epithelial cells and melanocytes, contributes to psoriasis pathogenesis, was unknown so far. Analysis of IL-29-treated human keratinocytes revealed induction of the chemokines CXCL10, CXCL11, and, to a much lesser extent, CXCL9. Unlike these CXCR3A ligands, known to attract Th1-, CD8(+), NK-, and Th1/Th17 transient cells, no influence was found on chemokines attracting other immune cell populations or on molecules modulating the CXCR3A/CXCR3A ligand interaction. CXCR3A ligand expression was also induced by IL-29 in melanocytes and in epidermis models and explanted skin. Regarding other psoriasis-relevant cytokines, interferon-γ and, less potently, tumor necrosis factor-α and IL-1β shared and strengthened IL-29's capacity. Murine IL-29 counterpart injected into mouse skin provoked local CXCL10 and CXCL11 expression, T-cell infiltration, and, in consequence, skin swelling. The elevated IL-29 expression in psoriatic lesions was associated with upregulation of CXCR3A ligands compared to non-lesional skin of these patients and to the skin of healthy donors and atopic dermatitis patients, which lack IL-29 production. Importantly, neutralization of IL-29 reduced CXCR3A ligand levels in explant cultures of psoriatic lesions. Finally, elevated blood CXCL11 levels were found in psoriasis that might be useful for monitoring lesional activity of the IL-29 axis. In summary, the Th17-cytokine IL-29 induces specific chemokines and, in consequence, provokes skin infiltration of potentially pathogenic T-cells. IL-29 selectively induces CXCR3A-binding chemokines (CXCL9, CXCL10, CXCL11) in skin cells. Murine IL-29 counterpart induces skin T-cell infiltration and inflammation in mice. CXCR3A ligands are
Romeo, Megan; Hutchison, Tetiana; Malu, Aditi; White, Averi; Kim, Janice; Gardner, Rachel; Smith, Katie; Nelson, Katherine; Bergeson, Rachel; McKee, Ryan; Harrod, Carolyn; Ratner, Lee; Lüscher, Bernhard; Martinez, Ernest; Harrod, Robert
2018-05-01
In normal cells, aberrant oncogene expression leads to the accumulation of cytotoxic metabolites, including reactive oxygen species (ROS), which can cause oxidative DNA-damage and apoptosis as an intrinsic barrier against neoplastic disease. The c-Myc oncoprotein is overexpressed in many lymphoid cancers due to c-myc gene amplification and/or 8q24 chromosomal translocations. Intriguingly, p53 is a downstream target of c-Myc and hematological malignancies, such as adult T-cell leukemia/lymphoma (ATL), frequently contain wildtype p53 and c-Myc overexpression. We therefore hypothesized that p53-regulated pro-survival signals may thwart the cell's metabolic anticancer defenses to support oncogene-activation in lymphoid cancers. Here we show that the Tp53-induced glycolysis and apoptosis regulator (TIGAR) promotes c-myc oncogene-activation by the human T-cell leukemia virus type-1 (HTLV-1) latency-maintenance factor p30 II , associated with c-Myc deregulation in ATL clinical isolates. TIGAR prevents the intracellular accumulation of c-Myc-induced ROS and inhibits oncogene-induced cellular senescence in ATL, acute lymphoblastic leukemia, and multiple myeloma cells with elevated c-Myc expression. Our results allude to a pivotal role for p53-regulated antioxidant signals as mediators of c-Myc oncogenic functions in viral and non-viral lymphoid tumors. Copyright © 2018 Elsevier Inc. All rights reserved.
Bryant, A E; Stevens, D L
1996-01-01
Clostridium perfringens phospholipase C (PLC) and perfringolysin O (PFO) differentially induced human umbilical vein endothelial cell expression and synthesis of endothelial cell-leukocyte adherence molecule-1 (ELAM-1), intracellular leukocyte adherence molecule-1 (ICAM-1), and interleukin-8 (IL-8). PLC strongly induced expression of ELAM-1, ICAM-1, and IL-8, while PFO stimulated early ICAM-1 expression but did not promote ELAM-1 expression or IL-8 synthesis. PLC caused human umbilical vein endothelial cells to assume a fibroblastoid morphology, whereas PFO, in high concentrations or after prolonged low-dose toxin exposure, caused cell death. The toxin-induced expression of proadhesive and activational proteins and direct cytopathic effects may contribute to the leukostasis, vascular compromise, and capillary leak characteristics of C. perfringens gas gangrene. PMID:8557365
Zaborsky, Nadja; Gassner, Franz Josef; Asslaber, Daniela; Reinthaler, Petra; Denk, Ursula; Flenady, Sabine; Hofbauer, Josefina Piñón; Danner, Barbara; Rebhandl, Stefan; Harrer, Andrea; Geisberger, Roland; Greil, Richard; Egle, Alexander
2016-08-02
Chronic lymphocytic leukemia develops within a complex network driven by genetic mutations and microenvironmental interactions. Among the latter a complex interplay with the immune system is established by the clone. Next to a proposed recruitment of support from T and myeloid cells, potential anti-CLL immune reactions need to be subverted. By using TCL1 mice as a CLL model, we show that TCR-Vβ7+ NK1.1+ T cells are overrepresented in this disease model and constitute a main subset of peripheral CD3+ cells with biased TCR usage, showing that these cells account for a major part for T cell skewing in TCL1 mice. Moreover, we show that overrepresentation is dependent on CD1d expression in TCL1 mice, implicating that these cells belong to a NKT-like cell fraction which are restricted to antigen presented by the MHC-like surface marker CD1d. Accordingly, we observed a high fraction of CD161+ cells within overrepresented T cells in CLL patients and we found downregulation of CD1d on the surface of CLL cells, both in TCL1 mice and patients. Finally, we show that in TCL1 mice, CD1d deficiency resulted in shortened overall survival. Our results point to an interaction between CLL and CD161+ T cells that may represent a novel therapeutic target for immune modulation.
Aline, Fleur; Bout, Daniel; Amigorena, Sébastian; Roingeard, Philippe; Dimier-Poisson, Isabelle
2004-01-01
It was previously demonstrated that immunizing mice with spleen dendritic cells (DCs) that had been pulsed ex vivo with Toxoplasma gondii antigens triggers a systemic Th1-biased specific immune response and induces protection against infection. T. gondii can cause severe sequelae in the fetuses of mothers who acquire the infection during pregnancy, as well as life-threatening neuropathy in immunocompromised patients, in particular those with AIDS. Here, we investigate the efficacy of a novel cell-free vaccine composed of DC exosomes, which are secreted antigen-presenting vesicles that express functional major histocompatibility complex class I and II and T-cell-costimulatory molecules. They have already been shown to induce potent antitumor immune responses. We investigated the potential of DC2.4 cell line-derived exosomes to induce protective immunity against toxoplasmosis. Our data show that most adoptively transferred T. gondii-pulsed DC-derived exosomes were transferred to the spleen, elicited a strong systemic Th1-modulated Toxoplasma-specific immune response in vivo, and conferred good protection against infection. These findings support the possibility that DC-derived exosomes can be used for T. gondii immunoprophylaxis and for immunoprophylaxis against many other pathogens. PMID:15213158
Zhou, Jing; Kaiser, Alaina; Ng, Colin; Karcher, Rachel; McConnell, Tim; Paczkowski, Patrick; Fernandez, Cristina; Zhang, Min; Mackay, Sean; Tsuji, Moriya
2017-07-03
Malaria is a severe infectious disease with relatively high mortality, thus having been a scourge of humanity. There are a few candidate malaria vaccines that have shown a protective efficacy in humans against malaria. One of the candidate human malaria vaccines, which is based on human malaria sporozoites and called PfSPZ Vaccine, has been shown to protect a significant proportion of vaccine recipients from getting malaria. PfSPZ Vaccine elicits a potent response of hepatic CD8+ T cells that are specific for malaria antigens in non-human primates. To further characterize hepatic CD8+ T cells induced by the sporozoite-based malaria vaccine in a mouse model, we have used a cutting-edge Single-cell Barcode (SCBC) assay, a recently emerged approach/method for investigating the nature of T-cells responses during infection or cancer. Using the SCBC technology, we have identified a population of hepatic CD8+ T cells that are polyfunctional at a single cell level only in a group of vaccinated mice upon malaria challenge. The cytokines/chemokines secreted by these polyfunctional CD8+ T-cell subsets include MIP-1α, RANTES, IFN-γ, and/or IL-17A, which have shown to be associated with protective T-cell responses against certain pathogens. Therefore, a successful induction of such polyfunctional hepatic CD8+ T cells may be a key to the development of effective human malaria vaccine. In addition, the SCBC technology could provide a new level of diagnostic that will allow for a more accurate determination of vaccine efficacy.
γδ T Cells Support Pancreatic Oncogenesis by Restraining αβ T Cell Activation.
Daley, Donnele; Zambirinis, Constantinos Pantelis; Seifert, Lena; Akkad, Neha; Mohan, Navyatha; Werba, Gregor; Barilla, Rocky; Torres-Hernandez, Alejandro; Hundeyin, Mautin; Mani, Vishnu Raj Kumar; Avanzi, Antonina; Tippens, Daniel; Narayanan, Rajkishen; Jang, Jung-Eun; Newman, Elliot; Pillarisetty, Venu Gopal; Dustin, Michael Loran; Bar-Sagi, Dafna; Hajdu, Cristina; Miller, George
2016-09-08
Inflammation is paramount in pancreatic oncogenesis. We identified a uniquely activated γδT cell population, which constituted ∼40% of tumor-infiltrating T cells in human pancreatic ductal adenocarcinoma (PDA). Recruitment and activation of γδT cells was contingent on diverse chemokine signals. Deletion, depletion, or blockade of γδT cell recruitment was protective against PDA and resulted in increased infiltration, activation, and Th1 polarization of αβT cells. Although αβT cells were dispensable to outcome in PDA, they became indispensable mediators of tumor protection upon γδT cell ablation. PDA-infiltrating γδT cells expressed high levels of exhaustion ligands and thereby negated adaptive anti-tumor immunity. Blockade of PD-L1 in γδT cells enhanced CD4(+) and CD8(+) T cell infiltration and immunogenicity and induced tumor protection suggesting that γδT cells are critical sources of immune-suppressive checkpoint ligands in PDA. We describe γδT cells as central regulators of effector T cell activation in cancer via novel cross-talk. Copyright © 2016 Elsevier Inc. All rights reserved.
Zunino, Susan J; Storms, David H
2017-08-01
Resveratrol has been reported to inhibit or induce DNA damage, depending upon the type of cell and the experimental conditions. Dietary resveratrol is present in the body predominantly as metabolites and limited data is available concerning the activities of these metabolic products. In the present study, physiologically obtainable levels of the resveratrol metabolites resveratrol-3-O-glucuronide, resveratrol-4'-O-glucuronide and resveratrol-3-O-sulfate were evaluated for their ability to protect Jurkat T cells against DNA damage induced by the topoisomerase I inhibitors camptothecin and topotecan. The cells were pretreated for 24 h with 10 µM resveratrol aglycone or each resveratrol metabolite prior to the induction of DNA damage with camptothecin or topotecan. In separate experiments, the cells were co-treated with resveratrol or its metabolites, and a topoisomerase I inhibitor. The detection of histone 2AX phosphorylation and terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) were used to determine DNA damage, and apoptosis was measured using an antibody against cleaved poly ADP-ribose polymerase. It was identified that pretreatment of the cells with resveratrol-3-O-glucuronide and resveratrol-4'-O-glucuronide reduced the mean fluorescence intensity of staining for DNA strand breaks following treatment with camptothecin, while the percentage of cells undergoing apoptosis was unchanged. However, pretreatment of the cells with resveratrol aglycone increased the DNA damage and apoptosis induced by the drugs. These results suggest that the glucuronide metabolites of resveratrol partially protected the cells from DNA damage, but did not influence the induction of cell death by camptothecin and topotecan. These data suggest that resveratrol aglycone treatment may be beneficial for treating types of cancer that have direct contact with resveratrol prior to its metabolism, including gastrointestinal cancers, which are routinely treated with
DeTemple, Daphne E; Oldhafer, Felix; Falk, Christine S; Chen-Wacker, Chen; Figueiredo, Constanca; Kleine, Moritz; Ramackers, Wolf; Timrott, Kai; Lehner, Frank; Klempnauer, Juergen; Bock, Michael; Vondran, Florian W R
2018-03-01
Hepatocyte transplantation is a promising therapeutic approach for various liver diseases. Despite the liver's tolerogenic potential, early immune-mediated loss of transplanted cells is observed, and longterm acceptance has not been achieved yet. Patients deemed tolerant after liver transplantation presented an increased frequency of regulatory T cells (Tregs), which therefore also might enable reduction of posttransplant cell loss and enhance longterm allograft acceptance. We hence characterized hepatocyte-induced immune reactions and evaluated the immunomodulatory potential of Tregs applying mixed lymphocyte cultures and mixed lymphocyte hepatocyte cultures. These were set up using peripheral blood mononuclear cells and primary human hepatocytes, respectively. Polyclonally expanded CD4 + CD25 high CD127 low Tregs were added to cocultures in single-/trans-well setups with/without supplementation of anti-interferon γ (IFNγ) antibodies. Hepatocyte-induced alloresponses were then analyzed by multicolor flow cytometry. Measurements indicated that T cell response upon stimulation was associated with IFNγ-induced major histocompatibility complex (MHC) class II up-regulation on hepatocytes and mediated by CD4 + T cells. An indirect route of antigen presentation could be ruled out by use of fragmented hepatocytes and culture supernatants of hepatocytes. Allospecific proliferation was accompanied by inflammatory cytokine secretion. CD8 + T cells showed early up-regulation of CD69 despite lack of cell proliferation in the course of coculture. Supplementation of Tregs effectively abrogated hepatocyte-induced alloresponses and was primarily cell contact dependent. In conclusion, human hepatocytes induce a CD4 + T cell alloresponse in vitro, which is associated with MHC class II up-regulation on hepatocytes and is susceptible to suppression by Tregs. Liver Transplantation 24 407-419 2018 AASLD. © 2018 The Authors. Liver Transplantation published by Wiley Periodicals, Inc
Tršan, Tihana; Vuković, Kristina; Filipović, Petra; Brizić, Ana Lesac; Lemmermann, Niels A W; Schober, Kilian; Busch, Dirk H; Britt, William J; Messerle, Martin; Krmpotić, Astrid; Jonjić, Stipan
2017-08-01
Designing CD8 + T-cell vaccines, which would provide protection against tumors is still considered a great challenge in immunotherapy. Here we show the robust potential of cytomegalovirus (CMV) vector expressing the NKG2D ligand RAE-1γ as CD8 + T cell-based vaccine against malignant tumors. Immunization with the CMV vector expressing RAE-1γ, delayed tumor growth or even provided complete protection against tumor challenge in both prophylactic and therapeutic settings. Moreover, a potent tumor control in mice vaccinated with this vector can be further enhanced by blocking the immune checkpoints TIGIT and PD-1. CMV vector expressing RAE-1γ potentiated expansion of KLRG1 + CD8 + T cells with enhanced effector properties. This vaccination was even more efficient in neonatal mice, resulting in the expansion and long-term maintenance of epitope-specific CD8 + T cells conferring robust resistance against tumor challenge. Our data show that immunomodulation of CD8 + T-cell responses promoted by herpesvirus expressing a ligand for NKG2D receptor can provide a powerful platform for the prevention and treatment of CD8 + T-cell sensitive tumors. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Tischner, D; Wiegers, G J; Fiegl, H; Drach, M; Villunger, A
2012-01-01
Transforming growth factor beta (TGF-β)- and Interleukin-2 (IL-2)-mediated signaling enables the generation and expansion of induced regulatory T (iTreg) cells that carry high hopes for the treatment of chronic inflammatory and autoimmune diseases. Knowledge about factors stabilizing their lineage commitment and lifespan, however, is limited. Here, we investigated the behavior of iTreg cells, derived from apoptosis-defective mouse mutants, during activated cell autonomous cell death, triggered by cytokine-deprivation, or activation-induced cell death (AICD) after restimulation of the T-cell receptor, and compared these responses with those of effector T cells. We observed that iTreg cells were much more sensitive to IL-2-deprivation but poorly susceptible to AICD. In fact, when apoptosis was compromised, T-cell receptor (TCR)-religation resulted in methylation-independent, ERK- and PI3K/mTOR-mediated loss of Foxp3 expression, impaired suppressive capacity and effector cytokine production. Although iTreg cells prevented colitis induction they rapidly lost Foxp3-GFP expression and gained ability to produce effector cytokines thereby imposing Th1 cell fate on resident effector cells. Surprisingly, iTreg cell conversion itself was limited by TGF-β-mediated Bim/Bcl2L11-dependent apoptosis. Hence, the very same cytokine that drives the generation of iTreg cells can trigger their demise. Our results provide novel insights in iTreg cell biology that will assist optimization of iTreg-based therapy. PMID:22322859
Pyo, Suhkneung; Kang, Chung Hyo; Lee, Chong Ock; Lee, Heung Kyoung; Choi, Sang Un; Park, Chi Hoon
2018-01-01
Gastric cancer is a malignancy that has a high mortality rate. Although progress has been made in the treatment of gastric cancer, many patients experience cancer recurrence and metastasis. Folate receptor 1 (FOLR1) is overexpressed on the cell surface in over one-third of gastric cancer patients, but rarely is expressed in normal tissue. This makes FOLR1 a potential target for chimeric antigen receptor (CAR) T cell immunotherapy, although the function of FOLR1 has not been elucidated. CAR are engineered fusion receptor composed of an antigen recognition region and signaling domains. T cells expressing CAR have specific activation and cytotoxic effects against cancer cells containing the target antigen. In this study, we generated a CAR that targets FOLR1 composed of a single-chain variable fragment (scFv) of FOLR1 antibody and signaling domains consisting of CD28 and CD3ζ. Both FOLR1-CAR KHYG-1, a natural killer cell line, and FOLR1-CAR T cells recognized FOLR1-positive gastric cancer cells in a MHC-independent manner and induced secretion of various cytokines and caused cell death. Conclusively, this is the first study to demonstrate that CAR KHYG-1/T cells targeting FOLR1 are effective against FOLR1-positive gastric cancer cells. PMID:29874279
Choi, Dong Hoon; Kim, Kwang Soon; Yang, Se Hwan; Chung, Doo Hyun; Song, Boyeong; Sprent, Jonathan; Cho, Jae Ho; Sung, Young Chul
2011-12-15
Dendritic cells (DC) present α-galactosylceramide (αGalCer) to invariant T-cell receptor-expressing natural killer T cells (iNKT) activating these cells to secrete a variety of cytokines, which in turn results in DC maturation and activation of other cell types, including NK cells, B cells, and conventional T cells. In this study, we showed that αGalCer-pulsing of antigen-activated CD8 T cells before adoptive transfer to tumor-bearing mice caused a marked increase in donor T-cell proliferation, precursor frequency, and cytotoxic lymphocyte activity. This effect was interleukin (IL)-2 dependent and involved both natural killer T cells (NKT) and DCs, as mice lacking IL-2, NKTs, and DCs lacked any enhanced response to adoptively transferred αGalCer-loaded CD8 T cells. iNKT activation was mediated by transfer of αGalCer from the cell membrane of the donor CD8 T cells onto the αGalCer receptor CD1d which is present on host DCs. αGalCer transfer was increased by prior activation of the donor CD8 T cells and required AP-2-mediated endocytosis by host DCs. In addition, host iNKT cell activation led to strong IL-2 synthesis, thereby increasing expansion and differentiation of donor CD8 T cells. Transfer of these cells led to improved therapeutic efficacy against established solid tumors in mice. Thus, our findings illustrate how αGalCer loading of CD8 T cells after antigen activation in vitro may leverage the therapeutic potential of adoptive T-cell therapies.
Progesterone and HMOX-1 promote fetal growth by CD8+ T cell modulation
Solano, María Emilia; Kowal, Mirka Katharina; O’Rourke, Greta Eugenia; Horst, Andrea Kristina; Modest, Kathrin; Plösch, Torsten; Barikbin, Roja; Remus, Chressen Catharina; Berger, Robert G.; Jago, Caitlin; Ho, Hoang; Sass, Gabriele; Parker, Victoria J.; Lydon, John P.; DeMayo, Francesco J.; Hecher, Kurt; Karimi, Khalil; Arck, Petra Clara
2015-01-01
Intrauterine growth restriction (IUGR) affects up to 10% of pregnancies in Western societies. IUGR is a strong predictor of reduced short-term neonatal survival and impairs long-term health in children. Placental insufficiency is often associated with IUGR; however, the molecular mechanisms involved in the pathogenesis of placental insufficiency and IUGR are largely unknown. Here, we developed a mouse model of fetal-growth restriction and placental insufficiency that is induced by a midgestational stress challenge. Compared with control animals, pregnant dams subjected to gestational stress exhibited reduced progesterone levels and placental heme oxygenase 1 (Hmox1) expression and increased methylation at distinct regions of the placental Hmox1 promoter. These stress-triggered changes were accompanied by an altered CD8+ T cell response, as evidenced by a reduction of tolerogenic CD8+CD122+ T cells and an increase of cytotoxic CD8+ T cells. Using progesterone receptor– or Hmox1-deficient mice, we identified progesterone as an upstream modulator of placental Hmox1 expression. Supplementation of progesterone or depletion of CD8+ T cells revealed that progesterone suppresses CD8+ T cell cytotoxicity, whereas the generation of CD8+CD122+ T cells is supported by Hmox1 and ameliorates fetal-growth restriction in Hmox1 deficiency. These observations in mice could promote the identification of pregnancies at risk for IUGR and the generation of clinical interventional strategies. PMID:25774501
Dopamine, T cells and multiple sclerosis (MS).
Levite, Mia; Marino, Franca; Cosentino, Marco
2017-05-01
Dopamine is a key neurotransmitter that induces critical effects in the nervous system and in many peripheral organs, via 5 dopamine receptors (DRs): D1R-D5R. Dopamine also induces many direct and very potent effects on many DR-expressing immune cells, primarily T cells and dendritic cells. In this review, we focus only on dopamine receptors, effects and production in T cells. Dopamine by itself (at an optimal concentration of~0.1 nM) induces multiple function of resting normal human T cells, among them: T cell adhesion, chemotactic migration, homing, cytokine secretion and others. Interestingly, dopamine activates resting effector T cells (Teffs), but suppresses regulatory T cells (Tregs), and both effects lead eventually to Teff activation. Dopamine-induced effects on T cells are dynamic, context-sensitive and determined by the: T cell activation state, T cell type, DR type, and dopamine concentration. Dopamine itself, and also few dopaminergic molecules/ drugs that are in clinical use for cardiac, neurological and other non-immune indications, have direct effects on human T cells (summarized in this review). These dopaminergic drugs include: dopamine = intropin, L-DOPA, bromocriptine, pramipexole, pergolide, haloperidol, pimozide, and amantadine. Other dopaminergic drugs were not yet tested for their direct effects on T cells. Extensive evidence in multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE) show dopaminergic dysregulations in T cells in these diseases: D1-like DRs are decreased in Teffs of MS patients, and dopamine does not affect these cells. In contrast, D1-like DRs are increased in Tregs of MS patients, possibly causing functional Treg impairment in MS. Treatment of MS patients with interferon β (IFN-β) increases D1-like DRs and decreases D2-like DRs in Teffs, decreases D1-like DRs in Tregs, and most important: restores responsiveness of patient's Teffs to dopamine. DR agonists and antagonists confer some benefits in
Schoenlaub, Laura; Cherla, Rama; Zhang, Yan; Zhang, Guoquan
2016-12-01
Our recent study demonstrated that virulent Coxiella burnetii Nine Mile phase I (NMI) is capable of infecting and replicating within peritoneal B1a cells and that B1a cells play an important role in host defense against C. burnetii infection in mice. However, it remains unknown if avirulent Nine Mile phase II (NMII) can infect and replicate in B1a cells and whether NMI and NMII can differentially interact with B1a cells. In this study, we examined if NMI and NMII can differentially modulate host cell apoptotic signaling in B1a cells. The results showed that NMII induced dose-dependent cell death in murine peritoneal B1a cells but NMI did not, suggesting that NMI and NMII may differentially activate host cell apoptotic signaling in B1a cells. Western blotting indicated that NMII-induced B1a cell death was not dependent on either caspase-3 or PARP-1 cleavage, but cleavage of caspase-1 was detected in NMII-infected B1a cells. In addition, inhibition or deficiency of caspase-1 activity blocked NMII-induced B1a cell death. These results suggest that NMII induces a caspase-1-dependent pyroptosis in murine peritoneal B1a cells. We also found that heat-killed NMII and type 4 secretion system (T4SS) mutant NMII were unable to induce B1a cell death and that NMII infection did not induce cell death in peritoneal B1a cells from Toll-like receptor 2 (TLR-2)- or NLRP3 inflammasome-deficient mice. These data suggest that NMII-induced caspase-1-dependent pyroptosis may require its T4SS and activation of the TLR-2 and NLRP3 signaling pathways. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
Abernathy-Carver, K J; Sampson, H A; Picker, L J; Leung, D Y
1995-01-01
The extravasation of T cells at sites of inflammation is critically dependent on the activity of homing receptors (HR) involved in endothelial cell recognition and binding. Two such HR (the cutaneous lymphocyte antigen [CLA] and L-selectin) have been shown to be selectively involved in T cell migration to skin and peripheral lymph nodes, respectively. This study was designed to assess the relationship between the organ specificity of an allergic reaction to food and the expression of HR on T cells activated in vitro by the relevant food allergen. Peripheral blood mononuclear cells were isolated from seven milk allergic children with a history of eczema when exposed to milk. All patients had a positive prick skin test and double-blind placebo-controlled food challenge to milk. 10 children with either allergic eosinophilic gastroenteritis or milk-induced enterocolitis and 8 nonatopic adults served as controls. Five-parameter flow cytometry using monoclonal antibodies was used for detection of the specific HR on freshly isolated T cells versus T cell blasts induced by a 6-d incubation with casein, as compared with Candida albicans. After in vitro stimulation with casein, but not C. albicans, patients with milk allergy and atopic dermatitis had a significantly greater percentage of CLA+ T cells (P < 0.01) than controls with milk-induced enterocolitis, allergic eosinophilic gastroenteritis, or nonatopic healthy controls. In contrast, the percentage of L-selectin-expressing T cells did not differ significantly between these groups. These data suggest that after casein stimulation allergic patients with milk-induced skin disease have an expanded population of CLA+ T cells, as compared with nonatopics or allergic patients without skin involvement. We postulate that heterogeneity in the regulation of HR expression on antigen-specific T cells may play a role in determining sites of involvement in tissue-directed allergic responses. Images PMID:7532192
Abernathy-Carver, K J; Sampson, H A; Picker, L J; Leung, D Y
1995-02-01
The extravasation of T cells at sites of inflammation is critically dependent on the activity of homing receptors (HR) involved in endothelial cell recognition and binding. Two such HR (the cutaneous lymphocyte antigen [CLA] and L-selectin) have been shown to be selectively involved in T cell migration to skin and peripheral lymph nodes, respectively. This study was designed to assess the relationship between the organ specificity of an allergic reaction to food and the expression of HR on T cells activated in vitro by the relevant food allergen. Peripheral blood mononuclear cells were isolated from seven milk allergic children with a history of eczema when exposed to milk. All patients had a positive prick skin test and double-blind placebo-controlled food challenge to milk. 10 children with either allergic eosinophilic gastroenteritis or milk-induced enterocolitis and 8 nonatopic adults served as controls. Five-parameter flow cytometry using monoclonal antibodies was used for detection of the specific HR on freshly isolated T cells versus T cell blasts induced by a 6-d incubation with casein, as compared with Candida albicans. After in vitro stimulation with casein, but not C. albicans, patients with milk allergy and atopic dermatitis had a significantly greater percentage of CLA+ T cells (P < 0.01) than controls with milk-induced enterocolitis, allergic eosinophilic gastroenteritis, or nonatopic healthy controls. In contrast, the percentage of L-selectin-expressing T cells did not differ significantly between these groups. These data suggest that after casein stimulation allergic patients with milk-induced skin disease have an expanded population of CLA+ T cells, as compared with nonatopics or allergic patients without skin involvement. We postulate that heterogeneity in the regulation of HR expression on antigen-specific T cells may play a role in determining sites of involvement in tissue-directed allergic responses.
Attrition of memory CD8 T cells during sepsis requires LFA-1.
Serbanescu, Mara A; Ramonell, Kimberly M; Hadley, Annette; Margoles, Lindsay M; Mittal, Rohit; Lyons, John D; Liang, Zhe; Coopersmith, Craig M; Ford, Mandy L; McConnell, Kevin W
2016-11-01
CD8 T cell loss and dysfunction have been implicated in the increased susceptibility to opportunistic infections during the later immunosuppressive phase of sepsis, but CD8 T cell activation and attrition in early sepsis remain incompletely understood. With the use of a CLP model, we assessed CD8 T cell activation at 5 consecutive time points and found that activation after sepsis results in a distinct phenotype (CD69 + CD25 int CD62L HI ) independent of cognate antigen recognition and TCR engagement and likely through bystander-mediated cytokine effects. Additionally, we observed that sepsis concurrently results in the preferential depletion of a subset of memory-phenotype CD8 T cells that remain "unactivated" (i.e., fail to up-regulate activation markers) by apoptosis. Unactivated CD44 HI OT-I cells were spared from sepsis-induced attrition, as were memory-phenotype CD8 T cells of mice treated with anti-LFA-1 mAb, 1 h after CLP. Perhaps most importantly, we demonstrate that attrition of memory phenotype cells may have a pathologic significance, as elevated IL-6 levels were associated with decreased numbers of memory-phenotype CD8 T cells in septic mice, and preservation of this subset after administration of anti-LFA-1 mAb conferred improved survival at 7 d. Taken together, these data identify potentially modifiable responses of memory-phenotype CD8 T cells in early sepsis and may be particularly important in the application of immunomodulatory therapies in sepsis. © Society for Leukocyte Biology.
Attrition of memory CD8 T cells during sepsis requires LFA-1
Serbanescu, Mara A.; Ramonell, Kimberly M.; Hadley, Annette; Margoles, Lindsay M.; Mittal, Rohit; Lyons, John D.; Liang, Zhe; Coopersmith, Craig M.; Ford, Mandy L.; McConnell, Kevin W.
2016-01-01
CD8 T cell loss and dysfunction have been implicated in the increased susceptibility to opportunistic infections during the later immunosuppressive phase of sepsis, but CD8 T cell activation and attrition in early sepsis remain incompletely understood. With the use of a CLP model, we assessed CD8 T cell activation at 5 consecutive time points and found that activation after sepsis results in a distinct phenotype (CD69+CD25intCD62LHI) independent of cognate antigen recognition and TCR engagement and likely through bystander-mediated cytokine effects. Additionally, we observed that sepsis concurrently results in the preferential depletion of a subset of memory-phenotype CD8 T cells that remain “unactivated” (i.e., fail to up-regulate activation markers) by apoptosis. Unactivated CD44HI OT-I cells were spared from sepsis-induced attrition, as were memory-phenotype CD8 T cells of mice treated with anti-LFA-1 mAb, 1 h after CLP. Perhaps most importantly, we demonstrate that attrition of memory phenotype cells may have a pathologic significance, as elevated IL-6 levels were associated with decreased numbers of memory-phenotype CD8 T cells in septic mice, and preservation of this subset after administration of anti-LFA-1 mAb conferred improved survival at 7 d. Taken together, these data identify potentially modifiable responses of memory-phenotype CD8 T cells in early sepsis and may be particularly important in the application of immunomodulatory therapies in sepsis. PMID:27286793
Hirane, Miku; Araki, Mutsumi; Dong, Yan; Honoki, Kanya; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi
2013-11-08
Reactive oxygen species (ROS) are known to mediate a variety of biological responses, including cell motility. Recently, we indicated that lysophosphatidic acid (LPA) receptor-3 (LPA3) increased cell motile activity stimulated by hydrogen peroxide. In the present study, we assessed the role of LPA1 in the cell motile activity mediated by ROS in mouse fibroblast 3T3 cells. 3T3 cells were treated with hydrogen peroxide and 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) at concentrations of 0.1 and 1 μM for 48 h. In cell motility assays with Cell Culture Inserts, the cell motile activities of 3T3 cells treated with hydrogen peroxide and DMNQ were significantly higher than those of untreated cells. 3T3 cells treated with hydrogen peroxide and DMNQ showed elevated expression levels of the Lpar3 gene, but not the Lpar1 and Lpar2 genes. To investigate the effects of LPA1 on the cell motile activity induced by hydrogen peroxide and DMNQ, Lpar1-overexpressing (3T3-a1) cells were generated from 3T3 cells and treated with hydrogen peroxide and DMNQ. The cell motile activities stimulated by hydrogen peroxide and DMNQ were markedly suppressed in 3T3-a1 cells. These results suggest that LPA signaling via LPA1 inhibits the cell motile activities stimulated by hydrogen peroxide and DMNQ in 3T3 cells. Copyright © 2013 Elsevier Inc. All rights reserved.
Mangifera indica L. extract protects T cells from activation-induced cell death.
Hernández, Patricia; Delgado, Rene; Walczak, Henning
2006-09-01
The aqueous stem bark extract of Mangifera indica L. (Vimang) has been reported to have antioxidant properties. AIDS is characterized by up-regulation of CD95 ligand (CD95L) expression and enhancement of activation-induced cell death (AICD). Recent studies demonstrate oxidative signals combined with simultaneous calcium (Ca(2+)) influx into the cytosol are required for induction of CD95L expression. In this study we show that M. indica extract attenuated anti-CD3-induced accumulation of reactive oxygen species (ROS) and intracellular free Ca(2+) and consequently, downregulates CD95L mRNA expression and CD95-mediated AICD. In addition, TCR triggering caused an elevation in the antioxidant enzyme manganous superoxide dismutase (Mn-SOD) and the increase in c-Jun N-terminal kinase (JNK) phosphorylation, both effects being prevented by M. indica extract. We provide a number of evidences regarding how M. indica extract enhance T-cell survival by inhibiting AICD, a finding associated with a decrease in oxidative stress generated through the TCR signaling pathway in activated T cells.
Duewell, P; Steger, A; Lohr, H; Bourhis, H; Hoelz, H; Kirchleitner, S V; Stieg, M R; Grassmann, S; Kobold, S; Siveke, J T; Endres, S; Schnurr, M
2014-01-01
Pancreatic cancer is characterized by a microenvironment suppressing immune responses. RIG-I-like helicases (RLH) are immunoreceptors for viral RNA that induce an antiviral response program via the production of type I interferons (IFN) and apoptosis in susceptible cells. We recently identified RLH as therapeutic targets of pancreatic cancer for counteracting immunosuppressive mechanisms and apoptosis induction. Here, we investigated immunogenic consequences of RLH-induced tumor cell death. Treatment of murine pancreatic cancer cell lines with RLH ligands induced production of type I IFN and proinflammatory cytokines. In addition, tumor cells died via intrinsic apoptosis and displayed features of immunogenic cell death, such as release of HMGB1 and translocation of calreticulin to the outer cell membrane. RLH-activated tumor cells led to activation of dendritic cells (DCs), which was mediated by tumor-derived type I IFN, whereas TLR, RAGE or inflammasome signaling was dispensable. Importantly, CD8α+ DCs effectively engulfed apoptotic tumor material and cross-presented tumor-associated antigen to naive CD8+ T cells. In comparison, tumor cell death mediated by oxaliplatin, staurosporine or mechanical disruption failed to induce DC activation and antigen presentation. Tumor cells treated with sublethal doses of RLH ligands upregulated Fas and MHC-I expression and were effectively sensitized towards Fas-mediated apoptosis and cytotoxic T lymphocyte (CTL)-mediated lysis. Vaccination of mice with RLH-activated tumor cells induced protective antitumor immunity in vivo. In addition, MDA5-based immunotherapy led to effective tumor control of established pancreatic tumors. In summary, RLH ligands induce a highly immunogenic form of tumor cell death linking innate and adaptive immunity. PMID:25012502
Manzar, Gohar S; Kim, Eun-Mi; Zavazava, Nicholas
2017-08-25
Type 1 diabetes (T1D) can be managed by transplanting either the whole pancreas or isolated pancreatic islets. However, cadaveric pancreas is scarcely available for clinical use, limiting this approach. As such, there is a great need to identify alternative sources of clinically usable pancreatic tissues. Here, we used induced pluripotent stem (iPS) cells derived from patients with T1D to generate glucose-responsive, insulin-producing cells (IPCs) via 3D culture. Initially, T1D iPS cells were resistant to differentiation, but transient demethylation treatment significantly enhanced IPC yield. The cells responded to high-glucose stimulation by secreting insulin in vitro The shape, size, and number of their granules, as observed by transmission electron microscopy, were identical to those found in cadaveric β cells. When the IPCs were transplanted into immunodeficient mice that had developed streptozotocin-induced diabetes, they promoted a dramatic decrease in hyperglycemia, causing the mice to become normoglycemic within 28 days. None of the mice died or developed teratomas. Because the cells are derived from "self," immunosuppression is not required, providing a much safer and reliable treatment option for T1D patients. Moreover, these cells can be used for drug screening, thereby accelerating drug discovery. In conclusion, our approach eliminates the need for cadaveric pancreatic tissue.
Singh, Kailash; Kadesjö, Erik; Lindroos, Julia; Hjort, Marcus; Lundberg, Marcus; Espes, Daniel; Carlsson, Per-Ola; Sandler, Stellan; Thorvaldson, Lina
2015-07-30
The anti-inflammatory cytokine IL-35 is produced by regulatory T (Treg) cells to suppress autoimmune and inflammatory responses. The role of IL-35 in type 1 diabetes (T1D) remains to be answered. To elucidate this, we investigated the kinetics of Treg cell response in the multiple low dose streptozotocin induced (MLDSTZ) T1D model and measured the levels of IL-35 in human T1D patients. We found that Treg cells were increased in MLDSTZ mice. However, the Treg cells showed a decreased production of anti-inflammatory (IL-10, IL-35, TGF-β) and increased pro-inflammatory (IFN-γ, IL-2, IL-17) cytokines, indicating a phenotypic shift of Treg cells under T1D condition. IL-35 administration effectively both prevented development of, and counteracted established MLDSTZ T1D, seemingly by induction of Eos expression and IL-35 production in Treg cells, thus reversing the phenotypic shift of the Treg cells. IL-35 administration reversed established hyperglycemia in NOD mouse model of T1D. Moreover, circulating IL-35 levels were decreased in human T1D patients compared to healthy controls. These findings suggest that insufficient IL-35 levels play a pivotal role in the development of T1D and that treatment with IL-35 should be investigated in treatment of T1D and other autoimmune diseases.
Engelmann, Robby; Sellmann, Tina; Köhling, Rüdiger; Müller-Hilke, Brigitte
2017-01-01
Growing clinical and laboratory evidence corroborates a role for the immune system in the pathophysiology of epilepsy. In order to delineate the immune response following pilocarpine-induced status epilepticus (SE) in the mouse, we monitored the kinetics of leukocyte presence in the hippocampus over the period of four weeks. SE was induced following a ramping protocol of pilocarpine injection into 4–5 weeks old C57BL/6 mice. Brains were removed at days 1–4, 14 or 28 after SE, and the hippocampi were analyzed via flow cytometry, via quantitative reverse transcriptase PCR (qRT-PCR) and via immunohistochemistry. Epileptogenesis was confirmed by Timm staining of mossy fiber sprouting in the inner molecular layer of the dentate gyrus. The flow cytometry data revealed a biphasic immune response following pilocarpine-induced SE with a transient increase in activated CD11b+ and F4/80+ macrophages within the first four days replaced by an increase in CD3+ T-lymphocytes around day 28. This delayed T cell response was confirmed via qRT-PCR and via immunohistochemistry. In addition, qRT-PCR data could show that the delayed T cell response was associated with an increased CD8/CD4 ratio indicating a cytotoxic T cell response after SE. Intriguingly, early intervention with mycophenolate mofetil administration on days 0–3 after SE prevented this delayed T cell response. These results show an orchestrated immunological sequela and provide evidence that the delayed T cell response is sensitive to early immunomodulatory intervention. PMID:29182639
Magazin, Marilyn; Poszepczynska-Guigné, Ewa; Bagot, Martine; Boumsell, Laurence; Pruvost, Christelle; Chalon, Pascale; Culouscou, Jean-Michel; Ferrara, Pascual; Bensussan, Armand
2004-01-01
Circulating malignant Sezary cells are a clonal proliferation of CD4+CD45RO+ T lymphocytes primarily involving the skin. To study the biology of these malignant T lymphocytes, we tested their ability to migrate in chemotaxis assays. Previously, we had shown that the neuropeptide neurotensin (NT) binds to freshly isolated Sezary malignant cells and induces through NT1 receptors the cell migration of the cutaneous T cell lymphoma cell line Cou-L. Here, we report that peripheral blood Sezary cells as well as the Sezary cell line Pno fail to migrate in response to neurotensin although they are capable of migrating to the chemokine stromal-cell-derived factor 1 alpha. This is in contrast with normal circulating CD4+ or CD8+ lymphocytes, which respond to both types of chemoattractants except after ex vivo short-time anti-CD3 monoclonal antibody activation, which abrogates the neurotensin-induced lymphocyte migration. Furthermore, we demonstrate that neurotensin-responsive T lymphocytes express the functional NT1 receptor responsible for chemotaxis. In these cells, but not in Sezary cells, neurotensin induces recruitment of phosphatidylinositol-3 kinase, and redistribution of phosphorylated cytoplasmic tyrosine kinase focal adhesion kinase and filamentous actin. Taken together, these results, which show functional distinctions between normal circulating lymphocytes and Sezary syndrome cells, contribute to further understanding of the physiopathology of these atypical cells.
Han, Ihn; Choi, Eun Ha
2017-05-30
Non-thermal atmospheric pressure plasma is ionized matter, composed of highly reactive species that include positive ions, negative ions, free radicals, neutral atoms, and molecules. Recent reports have suggested that non-thermal biocompatible plasma (NBP) can selectively kill a variety of cancer cells, and promote stem cell differentiation. However as of yet, the regulation of proliferation and differentiation potential of NBP has been poorly understood.Here, we investigated the effects of NBP on the osteogenic differentiation of precursor cell lines of osteoblasts, MC3T3 E1 and SaOS-2. For in vitro osteogenic differentiation, precursor cell lines were treated with NBP, and cultured with osteogenic induction medium. After 10 days of treatment, the NBP was shown to be effective in osteogenic differentiation in MC3T3 E1 cells by von Kossa and Alizarin Red S staining assay. Real-time PCR was then performed to investigate the expression of osteogenic specific genes, Runx2, OCN, COL1, ALP and osterix in MC3T3 E1 cells after treatment with NBP for 4 days. Furthermore, analysis of the protein expression showed that NBP treatment significantly reduced PI3K/AKT signaling and MAPK family signaling. However, p38 controlled phosphorylation of transcription factor forkhead box O1 (FoxO1) that related to cell differentiation with increased phosphorylated p38. These results suggest that non-thermal atmospheric pressure plasma can induce osteogenic differentiation, and enhance bone formation.
2013-01-01
Background The naphthoquinone pigment, shikonin, is a major component of Lithospermum erythrorhizon and has been shown to have various biological functions, including antimicrobial, anti-inflammatory, and antitumor effects. In this study, we investigated the effect of shikonin on adipocyte differentiation and its mechanism of action in 3T3-L1 cells. Methods To investigate the effects of shikonin on adipocyte differentiation, 3T3-L1 cells were induced to differentiate using 3-isobutyl-1-methylzanthine, dexamethasone, and insulin (MDI) for 8 days in the presence of 0–2 μM shikonin. Oil Red O staining was performed to determine the lipid accumulation in 3T3-L1 cells. To elucidate the anti-adipogenic mechanism of shikonin, adipogenic transcription factors, the phosphorylation levels of ERK, and adipogenic gene expression were analyzed by Western blotting and quantitative real-time PCR. To further confirm that shikonin inhibits adipogenic differentiation through downregulation of ERK 1/2 activity, 3T3-L1 cells were treated with shikonin in the presence of FGF-2, an activator, or PD98059, an inhibitor, of the ERK1/2 signaling pathway. Results Shikonin effectively suppressed adipogenesis and downregulated the protein levels of 2 major transcription factors, PPARγ and C/EBPα, as well as the adipocyte specific gene aP2 in a dose-dependent manner. qRT-PCR analysis revealed that shikonin inhibited mRNA expression of adipogenesis-related genes, such as PPARγ, C/EBPα, and aP2. Adipocyte differentiation was mediated by ERK 1/2 phosphorylation, which was confirmed by pretreatment with PD98059 (an ERK 1/2 inhibitor) or FGF-2 (an ERK 1/2 activator). The phosphorylation of ERK1/2 during the early stages of adipogenesis in 3T3-L1 cells was inhibited by shikonin. We also confirmed that FGF-2-stimulated ERK 1/2 activity was attenuated by shikonin. Conclusions These results demonstrate that shikonin inhibits adipogenic differentiation via suppression of the ERK signaling pathway
Gwon, So Young; Ahn, Ji Yun; Jung, Chang Hwa; Moon, Bo Kyung; Ha, Tae Youl
2013-08-06
The naphthoquinone pigment, shikonin, is a major component of Lithospermum erythrorhizon and has been shown to have various biological functions, including antimicrobial, anti-inflammatory, and antitumor effects. In this study, we investigated the effect of shikonin on adipocyte differentiation and its mechanism of action in 3T3-L1 cells. To investigate the effects of shikonin on adipocyte differentiation, 3T3-L1 cells were induced to differentiate using 3-isobutyl-1-methylzanthine, dexamethasone, and insulin (MDI) for 8 days in the presence of 0-2 μM shikonin. Oil Red O staining was performed to determine the lipid accumulation in 3T3-L1 cells. To elucidate the anti-adipogenic mechanism of shikonin, adipogenic transcription factors, the phosphorylation levels of ERK, and adipogenic gene expression were analyzed by Western blotting and quantitative real-time PCR. To further confirm that shikonin inhibits adipogenic differentiation through downregulation of ERK 1/2 activity, 3T3-L1 cells were treated with shikonin in the presence of FGF-2, an activator, or PD98059, an inhibitor, of the ERK1/2 signaling pathway. Shikonin effectively suppressed adipogenesis and downregulated the protein levels of 2 major transcription factors, PPARγ and C/EBPα, as well as the adipocyte specific gene aP2 in a dose-dependent manner. qRT-PCR analysis revealed that shikonin inhibited mRNA expression of adipogenesis-related genes, such as PPARγ, C/EBPα, and aP2. Adipocyte differentiation was mediated by ERK 1/2 phosphorylation, which was confirmed by pretreatment with PD98059 (an ERK 1/2 inhibitor) or FGF-2 (an ERK 1/2 activator). The phosphorylation of ERK1/2 during the early stages of adipogenesis in 3T3-L1 cells was inhibited by shikonin. We also confirmed that FGF-2-stimulated ERK 1/2 activity was attenuated by shikonin. These results demonstrate that shikonin inhibits adipogenic differentiation via suppression of the ERK signaling pathway during the early stages of adipogenesis.
Girardin, S E; Benjannet, S; Barale, J C; Chrétien, M; Seidah, N G
1998-07-24
mLIM3, a member of the LIM homeobox family, was recently demonstrated to be critical for proliferation and differentiation of the pituitary cell lineage. Using a pool of degenerate oligonucleotides we determined the DNA sequence ANNAGGAAA(T/C)GA(CIG)AA as the set preferentially recognized by mLIM3. A nearly identical sequence is found in the prolactin (PRL) promoter, within a 15-mer stretch from nucleotides (nts) -218 to -204 which is highly conserved between human, rat, and bovine. In order to test the hypothesis of a transcriptional effect of mLIM3 on the prolactin promoter, stable transfectants of mLIM3 cDNA in AtT20 tumor cells revealed that PRL mRNA expression was induced in 3 separate stable clones. Gel retardation experiments performed using nuclear extracts isolated from one of the AtT20/mLIM3 stable transfectants revealed affinity towards the 15-mer element of the PRL promoter. From these results, we propose that the PRL promoter element (nts -218 to -204) could be functional in vivo. Finally, we demonstrate that in AtT20 cells prolactin mRNA expression is not induced by the Pit-1/GHF-1 pathway and that growth hormone mRNA is not detected concomitantly with prolactin. We conclude that mLIM3 may play a key role in inducing PRL gene expression in lactotrophs by binding to a conserved motif close to a Pit-1/GHF-1 site within the proximal PRL promoter.
Cendron, Delphine; Ingoure, Sophie; Martino, Angelo; Casetti, Rita; Horand, Françoise; Romagné, François; Sicard, Hélène; Fournié, Jean-Jacques; Poccia, Fabrizio
2007-02-01
Phosphoantigens are mycobacterial non-peptide antigens that might enhance the immunogenicity of current subunit candidate vaccines for tuberculosis. However, their testing requires monkeys, the only animal models suitable for gammadelta T cell responses to mycobacteria. Thus here, the immunogenicity of 6-kDa early secretory antigenic target-mycolyl transferase complex antigen 85B (ESAT-6-Ag85B) (H-1 hybrid) fusion protein associated or not to a synthetic phosphoantigen was compared by a prime-boost regimen of two groups of eight cynomolgus. Although phosphoantigen activated immediately a strong release of systemic Th1 cytokines (IL-2, IL-6, IFN-gamma, TNF-alpha), it further anergized blood gammadelta T lymphocytes selectively. By contrast, the hybrid H-1 induced only memory alphabeta T cell responses, regardless of phosphoantigen. These latter essentially comprised cytotoxic T lymphocytes specific for Ag85B (on average + 430 cells/million PBMC) and few IFN-gamma-secreting cells (+ 40 cells/million PBMC, equally specific for ESAT-6 and for Ag85B). Hence, in macaques, a prime-boost with the H-1/phosphoantigen subunit combination induces two waves of immune responses, successively by gammadelta T and alphabeta T lymphocytes.
4-1BB regulates NKG2D costimulation in human cord blood CD8+ T cells.
Kim, Young-June; Han, Myung-Kwan; Broxmeyer, Hal E
2008-02-01
Ligation of NKG2D, a potent costimulatory receptor, can be either beneficial or detrimental to CD8(+) cytotoxic T cell (CTL) responses. Factors for these diverse NKG2D effects remain elusive. In this study, we demonstrate that 4-1BB, another costimulatory receptor, is an essential regulator of NKG2D in CD8(+) T cells. Costimulation of NKG2D caused down-modulation of NKG2D, but induced 4-1BB expression on the cell surface, even in the presence of TGF-beta1, which inhibits 4-1BB expression. Resulting NKG2D(-)4-1BB(+) cells were activated but still in an immature state with low cytotoxic activity. However, subsequent 4-1BB costimulation induced cytotoxic activity and restored down-modulated NKG2D. The cytotoxic activity and NKG2D expression induced by 4-1BB on NKG2D(+)4-1BB(+) cells were refractory to TGF-beta1 down-modulation. Such 4-1BB effects were enhanced by IL-12. In contrast, in the presence of IL-4, 4-1BB effects were abolished because IL-4 down-modulated NKG2D and 4-1BB expression in cooperation with TGF-beta1, generating another CD8(+) T-cell type lacking both NKG2D and 4-1BB. These NKG2D(-)4-1BB(-) cells were inert and unable to gain cytotoxic activity. Our results suggest that 4-1BB plays a critical role in protecting NKG2D from TGF-beta1-mediated down-modulation. Co-expression of NKG2D and 4-1BB may represent an important biomarker for defining competency of tumor infiltrating CD8(+) T cells.
Gao, Jian-mei; Li, Ran; Zhang, Lei; Jia, Li-long; Ying, Xi-xiang; Dou, De-qiang; Li, Jian-chun; Li, Hai-bo
2013-07-09
Cuscuta chinensis (C. chinensis) is a well-known traditional Chinese herb that has been used to treat heart disease, diabetes, liver injury, cancer, and aging. Murine osteoblastic MC3T3-E1 cells were treated with various concentrations of C. chinensis water extraction at different time intervals. The antioxidant effect of C. chinensis on MC3T3-E1 cells was evaluated using MTT and TUNEL assays. The effect of C. chinensis on cell cycle was analyzed by flow cytometry with propidium iodide. Lipid peroxidation was measured by the HPLC method. The cellular redox status was determined from the reduced glutathione to oxidized glutathione ratio (GSH/GSSG) and the enzymes involved in glutathione metabolism, including glutathione reductase (GR), Glutathione S-transferase (GST), and Glucose-6-phosphate dehydrogenase (G6PD). The changes in relative mitochondrial transmembrane potential (ΔΨm) in the MC3T3-E1 cells were analyzed with rhodamine 123 staining. Western blot analysis was used to evaluate the levels of cytochrome c (cyto c), Bax, Bcl-2, caspase 3, Sirt3, and IDH2 expressions. The C. chinensis water extraction protects tertiary butyl hydroperoxide (TBHP)-treated MC3T3-E1 cells from death in a dose-dependent manner. C. chinensis treatment significantly inhibited the reactive oxygen species (ROS) generation, malondialdehyde (MDA) production, and increased the activity of superoxide dismutase (SOD), GR, GST, and G6PD. The release of cyto c from mitochondria was reduced by C. chinensis, which increased the expression of antiapoptotic IDH2, Sirt3, and Bcl-2 and decreased the expression of Bax, cyto c, and caspase 3. C. chinensis modulated the oxidative stress-induced apoptosis in MC3T3-E1 cells, probably due to its antioxidant activity and functioning via mitochondria-dependent pathways. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.
Sakane, T; Honda, M; Taniguchi, Y; Kotani, H
1981-08-01
Very few normal human peripheral blood T cells are capable of binding autologous erythrocytes to form rosettes, whereas in the T cell population activated by concanavalin A (Con A) the autorosette levels are markedly enhanced. Fractionation of the Con A-activated T cells with autologous erythrocytes into autorosetting and nonrosetting cells demonstrates that suppressor, but not helper, activity resides in the autorosetting population, whereas the reverse is true of the nonrosetting population. Both these activities are found to be Con A dependent. The Con A-induced human suppressor cells can be identified and separated from the Con A-induced human helper cells by the autorosette technique. Studies on the surface properties of autorosetting and nonrosetting T cells indicate that there is little correlation between the activated suppressor and helper T cell subsets defined by autorosette technique and either those defined by monoclonal antibodies (which are able to distinguish these subsets in the resting but not activated T cells) or those defined by Fc receptors. Since the autorosetting T cell population (which acts as suppressor cells) bears receptors for peanut agglutinin, the nature of Con A-induced human suppressor cells appears to be analogous to that of Con A-induced murine suppressor cells.
Liu, Xiaobin; Xavier, Christy; Jann, Jamieson; Wu, Hongli
2016-01-01
Protein glutathionylation, defined as the formation of protein mixed disulfides (PSSG) between cysteine residues and glutathione (GSH), can lead to cell death. Glutaredoxin 1 (Grx1) is a thiol repair enzyme which catalyzes the reduction of PSSG. Therefore, Grx1 exerts strong anti-apoptotic effects by improving the redox state, especially in times of oxidative stress. However, there is currently no compound that is identified as a Grx1 activator. In this study, we identified and characterized Salvianolic acid B (Sal B), a natural compound, as a Grx1 inducer, which potently protected retinal pigment epithelial (RPE) cells from oxidative injury. Our results showed that treatment with Sal B protected primary human RPE cells from H2O2-induced cell damage. Interestingly, we found Sal B pretreatment upregulated Grx1 expression in RPE cells in a time- and dose-dependent manner. Furthermore, NF-E2-related factor 2 (Nrf2), the key transcription factor that regulates the expression of Grx1, was activated in Sal B treated RPE cells. Further investigation showed that knockdown of Grx1 by small interfering RNA (siRNA) significantly reduced the protective effects of Sal B. We conclude that Sal B protects RPE cells against H2O2-induced cell injury through Grx1 induction by activating Nrf2 pathway, thus preventing lethal accumulation of PSSG and reversing oxidative damage. PMID:27827892
Khouri, Ricardo; Silva-Santos, Gilvanéia; Dierckx, Tim; Menezes, Soraya Maria; Decanine, Daniele; Theys, Kristof; Silva, Aline Clara; Farré, Lourdes; Bittencourt, Achiléa; Mangino, Massimo; Roederer, Mario; Vandamme, Anne-Mieke; Van Weyenbergh, Johan
2018-01-01
Adult T-cell leukemia (ATL) is an aggressive, chemotherapy-resistant CD4 + CD25 + leukemia caused by HTLV-1 infection, which usually develops in a minority of patients several decades after infection. IFN + AZT combination therapy has shown clinical benefit in ATL, although its mechanism of action remains unclear. We have previously shown that an IFN-responsive FAS promoter polymorphism in a STAT1 binding site (rs1800682) is associated to ATL susceptibility and survival. Recently, CD4 T stem cell memory (T SCM ) Fas hi cells have been identified as the hierarchical cellular apex of ATL, but a possible link between FAS, apoptosis, proliferation and IFN response in ATL has not been studied. In this study, we found significant ex vivo antiproliferative, antiviral and immunomodulatory effects of IFN-α treatment in short-term culture of primary mononuclear cells from ATL patients (n = 25). Bayesian Network analysis allowed us to integrate ex vivo IFN-α response with clinical, genetic and immunological data from ATL patients, thereby revealing a central role for FAS -670 polymorphism and apoptosis in the coordinated mechanism of action of IFN-α. FAS genotype-dependence of IFN-induced apoptosis was experimentally validated in an independent cohort of healthy controls (n = 20). The same FAS -670 polymorphism also determined CD4 T SCM levels in a genome-wide twin study (p = 7 × 10 -11 , n = 460), confirming a genetic link between apoptosis and T SCM levels. Transcriptomic analysis and cell type deconvolution confirmed the FAS genotype/T SCM link and IFN-α-induced downregulation of CD4 T SCM -specific genes in ATL patient cells. In conclusion, ex vivo IFN-α treatment exerts a pleiotropic effect on primary ATL cells, with a genetic IFN/STAT1/Fas axis determining apoptosis vs. proliferation and underscoring the CD4 T SCM model of ATL leukemogenesis.
Butyric acid induces apoptosis via oxidative stress in Jurkat T-cells.
Kurita-Ochiai, T; Ochiai, K
2010-07-01
Reactive oxygen species (ROS) are essential for the induction of T-cell apoptosis by butyric acid, an extracellular metabolite of periodontopathic bacteria. To determine the involvement of oxidative stress in apoptosis pathways, we investigated the contribution of ROS in mitochondrial signaling pathways, death-receptor-initiated signaling pathway, and endoplasmic reticulum stress in butyric-acid-induced T-cell apoptosis. N-acetyl-L-Cysteine (NAC) abrogated mitochondrial injury, cytochrome c, AIF, and Smac release, and Bcl-2 and Bcl-xL suppression and Bax and Bad activation induced by butyric acid. However, the decrease in cFLIP expression by butyric acid was not restored by treatment with NAC; increases in caspase-4 and -10 activities by butyric acid were completely abrogated by NAC. NAC also affected the elevation of GRP78 and CHOP/GADD153 expression by butyric acid. These results suggest that butyric acid is involved in mitochondrial-dysfunction- and endoplasmic reticulum stress-mediated apoptosis in human Jurkat T-cells via a ROS-dependent mechanism.
Monti, Paolo; Scirpoli, Miriam; Maffi, Paola; Ghidoli, Nadia; De Taddeo, Francesca; Bertuzzi, Federico; Piemonti, Lorenzo; Falcone, Marika; Secchi, Antonio; Bonifacio, Ezio
2008-01-01
Successful transplantation requires the prevention of allograft rejection and, in the case of transplantation to treat autoimmune disease, the suppression of autoimmune responses. The standard immunosuppressive treatment regimen given to patients with autoimmune type 1 diabetes who have received an islet transplant results in the loss of T cells. In many other situations, the immune system responds to T cell loss through cytokine-dependant homeostatic proliferation of any remaining T cells. Here we show that T cell loss after islet transplantation in patients with autoimmune type 1 diabetes was associated with both increased serum concentrations of IL-7 and IL-15 and in vivo proliferation of memory CD45RO+ T cells, highly enriched in autoreactive glutamic acid decarboxylase 65–specific T cell clones. Immunosuppression with FK506 and rapamycin after transplantation resulted in a chronic homeostatic expansion of T cells, which acquired effector function after immunosuppression was removed. In contrast, the cytostatic drug mycophenolate mofetil efficiently blocked homeostatic T cell expansion. We propose that the increased production of cytokines that induce homeostatic expansion could contribute to recurrent autoimmunity in transplanted patients with autoimmune disease and that therapy that prevents the expansion of autoreactive T cells will improve the outcome of islet transplantation. PMID:18431516
Kamphorst, Alice O.; Pillai, Rathi N.; Yang, Shu; Nasti, Tahseen H.; Sica, Gabriel L.; Yu, Ke; Koenig, Lydia; Patel, Nikita T.; Behera, Madhusmita; Wu, Hong; McCausland, Megan; Chen, Zhengjia; Zhang, Chao; Khuri, Fadlo R.; Owonikoko, Taofeek K.; Ahmed, Rafi; Ramalingam, Suresh S.
2017-01-01
Exhausted T cells in chronic infections and cancer have sustained expression of the inhibitory receptor programmed cell death 1 (PD-1). Therapies that block the PD-1 pathway have shown promising clinical results in a significant number of advanced-stage cancer patients. Nonetheless, a better understanding of the immunological responses induced by PD-1 blockade in cancer patients is lacking. Identification of predictive biomarkers is a priority in the field, but whether peripheral blood analysis can provide biomarkers to monitor or predict patients’ responses to treatment remains to be resolved. In this study, we analyzed longitudinal blood samples from advanced stage non–small cell lung cancer (NSCLC) patients (n = 29) receiving PD-1–targeted therapies. We detected an increase in Ki-67+ PD-1+ CD8 T cells following therapy in ∼70% of patients, and most responses were induced after the first or second treatment cycle. This T-cell activation was not indiscriminate because we observed only minimal effects on EBV-specific CD8 T cells, suggesting that responding cells may be tumor specific. These proliferating CD8 T cells had an effector-like phenotype (HLA-DR+, CD38+, Bcl-2lo), expressed costimulatory molecules (CD28, CD27, ICOS), and had high levels of PD-1 and coexpression of CTLA-4. We found that 70% of patients with disease progression had either a delayed or absent PD-1+ CD8 T-cell response, whereas 80% of patients with clinical benefit exhibited PD-1+ CD8 T-cell responses within 4 wk of treatment initiation. Our results suggest that peripheral blood analysis may provide valuable insights into NSCLC patients’ responses to PD-1–targeted therapies. PMID:28446615
Type 1 Responses of Human Vγ9Vδ2 T Cells to Influenza A Viruses▿
Qin, Gang; Liu, Yinping; Zheng, Jian; Ng, Iris H. Y.; Xiang, Zheng; Lam, Kwok-Tai; Mao, Huawei; Li, Hong; Peiris, J. S. Malik; Lau, Yu-Lung; Tu, Wenwei
2011-01-01
γδ T cells are essential constituents of antimicrobial and antitumor defenses. We have recently reported that phosphoantigen isopentenyl pyrophosphate (IPP)-expanded human Vγ9Vδ2 T cells participated in anti-influenza virus immunity by efficiently killing both human and avian influenza virus-infected monocyte-derived macrophages (MDMs) in vitro. However, little is known about the noncytolytic responses and trafficking program of γδ T cells to influenza virus. In this study, we found that Vγ9Vδ2 T cells expressed both type 1 cytokines and chemokine receptors during influenza virus infection, and IPP-expanded cells had a higher capacity to produce gamma interferon (IFN-γ). Besides their potent cytolytic activity against pandemic H1N1 virus-infected cells, IPP-activated γδ T cells also had noncytolytic inhibitory effects on seasonal and pandemic H1N1 viruses via IFN-γ but had no such effects on avian H5N1 or H9N2 virus. Avian H5N1 and H9N2 viruses induced significantly higher CCL3, CCL4, and CCL5 production in Vγ9Vδ2 T cells than human seasonal H1N1 virus. CCR5 mediated the migration of Vγ9Vδ2 T cells toward influenza virus-infected cells. Our findings suggest a novel therapeutic strategy of using phosphoantigens to boost the antiviral activities of human Vγ9Vδ2 T cells against influenza virus infection. PMID:21752902
Ebola Virus Binding to Tim-1 on T Lymphocytes Induces a Cytokine Storm.
Younan, Patrick; Iampietro, Mathieu; Nishida, Andrew; Ramanathan, Palaniappan; Santos, Rodrigo I; Dutta, Mukta; Lubaki, Ndongala Michel; Koup, Richard A; Katze, Michael G; Bukreyev, Alexander
2017-09-26
consistently linked with fatal disease outcome. Previous findings have demonstrated that specific T-cell subsets are key contributors to the onset of a cytokine storm. In this study, we investigated the role of Tim-1, a T-cell-receptor-independent trigger of T-cell activation. We first demonstrated that Tim-1-knockout (KO) mice survive lethal Ebola virus challenge. We then used a series of in vitro assays to demonstrate that Ebola virus directly binds primary T cells in a Tim-1-phosphatidylserine-dependent manner. We noted that binding induces a cytokine storm-like phenomenon and that blocking Tim-1-phosphatidylserine interactions reduces viral binding, T-cell activation, and cytokine production. These findings highlight a previously unknown role of Tim-1 in the development of a cytokine storm and "immune paralysis." Copyright © 2017 Younan et al.
Radiation induces premature chromatid separation via the miR-142-3p/Bod1 pathway in carcinoma cells.
Pan, Dong; Du, Yarong; Ren, Zhenxin; Chen, Yaxiong; Li, Xiaoman; Wang, Jufang; Hu, Burong
2016-09-13
Radiation-induced genomic instability plays a vital role in carcinogenesis. Bod1 is required for proper chromosome biorientation, and Bod1 depletion increases premature chromatid separation. MiR-142-3p influences cell cycle progression and inhibits proliferation and invasion in cervical carcinoma cells. We found that radiation induced premature chromatid separation and altered miR-142-3p and Bod1 expression in 786-O and A549 cells. Overexpression of miR-142-3p increased premature chromatid separation and G2/M cell cycle arrest in 786-O cells by suppressing Bod1 expression. We also found that either overexpression of miR-142-3p or knockdown of Bod1 sensitized 786-O and A549 cells to X-ray radiation. Overexpression of Bod1 inhibited radiation- and miR-142-3p-induced premature chromatid separation and increased resistance to radiation in 786-O and A549 cells. Taken together, these results suggest that radiation alters miR-142-3p and Bod1 expression in carcinoma cells, and thus contributes to early stages of radiation-induced genomic instability. Combining ionizing radiation with epigenetic regulation may help improve cancer therapies.
Identification of an inducible regulator of c-myb expression during T-cell activation.
Phan, S C; Feeley, B; Withers, D; Boxer, L M
1996-01-01
Resting T cells express very low levels of c-Myb protein. During T-cell activation, c-myb expression is induced and much of the increase in expression occurs at the transcriptional level. We identified a region of the c-myb 5' flanking sequence that increased c-myb expression during T-cell activation. In vivo footprinting by ligation-mediated PCR was performed to correlate in vivo protein binding with functional activity. A protein footprint was visible over this region of the c-myb 5' flanking sequence in activated T cells but not in unactivated T cells. An electrophoretic mobility shift assay (EMSA) with nuclear extract from activated T cells and an oligonucleotide of this binding site demonstrated a new protein-DNA complex, referred to as CMAT for c-myb in activated T cells; this complex was not present in unactivated T cells. Because the binding site showed some sequence similarity with the nuclear factor of activated T cells (NFAT) binding site, we compared the kinetics of induction of the two binding complexes and the molecular masses of the two proteins. Studies of the kinetics of induction showed that the NFAT EMSA binding complex appeared earlier than the CMAT complex. The NFAT protein migrated more slowly in a sodium dodecyl sulfate-polyacrylamide gel than the CMAT protein did. In addition, an antibody against NFAT did not cross-react with the CMAT protein. The appearance of the CMAT binding complex was inhibited by both cyclosporin A and rapamycin. The CMAT protein appears to be a novel inducible protein involved in the regulation of c-myb expression during T-cell activation. PMID:8628306
Quan, Juan-Hua; Kang, Byung-Hun; Cha, Guang-Ho; Zhou, Wei; Koh, Young-Bok; Yang, Jung-Bo; Yoo, Heon-Jong; Lee, Min-A; Ryu, Jae-Sook; Noh, Heung-Tae; Kwon, Jaeyul; Lee, Young-Ha
2014-01-01
To elucidate the roles of metalloproteinases and the Bcl-2 family of proteins in Trichovaginalis. vaginalis-induced apoptosis in human cervical cancer cells (SiHa cells) and vaginal epithelial cells (MS74 cells), SiHa cells and MS74 cells were incubated with live T. vaginalis, T. vaginalis excretory and secretory products (ESP), and T. vaginalis lysates, either with or without the specific metalloproteinase inhibitor 1,10-phenanthroline (1,10-PT), and examined apoptotic events and Bcl-2 signaling. The live T. vaginalis and the T. vaginalis ESP induced the release of cytochrome c into the cytosol, the activation of caspase-3 and caspase-9, and the cleavage of PARP. Additionally, the live T. vaginalis, but not the T. vaginalis lysate, induced the cleavage of the proapoptotic Bim protein. The live T. vaginalis and the T. vaginalis ESP, but not the T. vaginalis lysate, induced the dose-dependent cleavage of the antiapoptotic Bcl-xL and Mcl-1 proteins and decreased the association levels of Bcl-xL/Bim and Mcl-1/Bim complexes. We performed gelatin zymography and casein-hydrolysis assays on the live T. vaginalis and the T. vaginalis ESP to identify the apoptosis-inducing factor. Both the live T. vaginalis and the ESP contained high levels of metalloproteinases, of which activities were significantly inhibited by 1,10-PT treatment. Furthermore, the 1,10-PT blocked the cleavage of Bcl-xL, Mcl-1, PARP, caspase-3, and caspase-9, as well as the release of cytochrome c into the cytosol, and it significantly increased the association levels of the Bcl-xL/Bim and Mcl-1/Bim protein complexes, returning them to normal levels. Our results demonstrate that T. vaginalis induces mitochondria-dependent apoptosis in SiHa cells through the dissociation of Bcl-xL/Bim and Mcl-1/Bim complexes and that the apoptosis is blocked by the metalloproteinase inhibitor 1,10-PT. These results expand our understanding of the role of metalloproteinases in T. vaginalis-induced apoptosis and the signaling
MAO, JINGJIE; LI, ZUANFANG; LIN, RUHUI; ZHU, XIAOQIN; LIN, JIUMAO; PENG, JUN; CHEN, LIDIAN
2015-01-01
Spasticity is common in various central neurological conditions, including after a stroke. Such spasticity may cause additional problems, and often becomes a primary concern for afflicted individuals. A number of studies have identified nuclear factor (erythroid-derived 2)-like 2 (Nrf2) as a key regulator in the adaptive survival response to oxidative stress. Elevated expression of Nrf2, combined with heme oxygenase 1 (HO-1) resistance, in the central nervous system is known to elicit key internal and external oxidation protection. Gua Lou Gui Zhi decoction (GLGZD) is a popular traditional Chinese formula with a long history of clinical use in China for the treatment of muscular spasticity following a stroke, epilepsy or a spinal cord injury. However, the mechanism underlying the efficacy of the medicine remains unclear. In the present study, the antioxidative effects of GLGZD were evaluated and the underlying molecular mechanisms were investigated, using hydrogen peroxide (H2O2)-induced rat pheochromocytoma cells (PC12 cells) as an in vitro oxidative stress model of neural cells. Upon application of different concentrations of GLGZD, a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay and ATP measurement were conducted to assess the impact on PC12 cell proliferation. In addition, inverted microscopy observations, and the MTT and ATP assessments, revealed that GLGZD attenuated H2O2-induced oxidative damage and signaling repression in PC12 cells. Furthermore, the mRNA and protein expression levels of Nrf2 and HO-1, which are associated with oxidative stress, were analyzed using reverse transcription quantitative polymerase chain reaction (PCR) and confocal microscopy. Confocal microscopy observations, as well as the quantitative PCR assay, revealed that GLGZD exerted a neuroprotective function against H2O2-induced oxidative damage in PC12 cells. Therefore, the results demonstrated that GLGZD protected PC12 cells injured by H2O2, which may be
Driver, John P; Scheuplein, Felix; Chen, Yi-Guang; Grier, Alexandra E; Wilson, S Brian; Serreze, David V
2010-02-01
In part, activation of invariant natural killer T (iNKT)-cells with the superagonist alpha-galactosylceramide (alpha-GalCer) inhibits the development of T-cell-mediated autoimmune type 1 diabetes in NOD mice by inducing the downstream differentiation of antigen-presenting dendritic cells (DCs) to an immunotolerogenic state. However, in other systems iNKT-cell activation has an adjuvant-like effect that enhances rather than suppresses various immunological responses. Thus, we tested whether in some circumstances genetic variation would enable activated iNKT-cells to support rather than inhibit type 1 diabetes development. We tested whether iNKT-conditioned DCs in NOD mice and a major histocompatibility complex-matched C57BL/6 (B6) background congenic stock differed in capacity to inhibit type 1 diabetes induced by the adoptive transfer of pathogenic AI4 CD8 T-cells. Unlike those of NOD origin, iNKT-conditioned DCs in the B6 background stock matured to a state that actually supported rather than inhibited AI4 T-cell-induced type 1 diabetes. The induction of a differing activity pattern of T-cell costimulatory molecules varying in capacity to override programmed death-ligand-1 inhibitory effects contributes to the respective ability of iNKT-conditioned DCs in NOD and B6 background mice to inhibit or support type 1 diabetes development. Genetic differences inherent to both iNKT-cells and DCs contribute to their varying interactions in NOD and B6.H2(g7) mice. This great variability in the interactions between iNKT-cells and DCs in two inbred mouse strains should raise a cautionary note about considering manipulation of this axis as a potential type 1 diabetes prevention therapy in genetically heterogeneous humans.
Markopoulou, Soultana; Kontargiris, Evangelos; Batsi, Christina; Tzavaras, Theodore; Trougakos, Ioannis; Boothman, David A.; Gonos, Efstathios S.; Kolettas, Evangelos
2016-01-01
Vanadium exerts a variety of biological effects, including antiproliferative responses through activation of the respective signaling pathways and the generation of reactive oxygen species. As epidermal cells are exposed to environmental insults, human keratinocytes (HaCaT) were used to investigate the mechanism of the antiproliferative effects of vanadyl(IV) sulfate (VOSO4). Treatment of HaCaT cells with VOSO4 inhibited proliferation and induced apoptosis in a dose-dependent manner. Inhibition of proliferation was associated with downregulation of cyclins D1 and E, E2F1, and the cyclin-dependent kinase inhibitors p21Cip1/Waf1 and p27Kip1. Induction of apoptosis correlated with upregulation of the c-fos oncoprotein, changes in the expression of clusterin (CLU), an altered ratio of antiapoptotic to proapoptotic Bcl-2 protein family members, and poly(ADP-ribose) poly-merase-1 cleavage. Forced overexpression of c-fos induced apoptosis in HaCaT cells that correlated with secretory CLU downregulation and upregulation of nuclear CLU (nCLU), a pro-death protein. Overexpression of Bcl-2 protected HaCaT cells from vanadium-induced apoptosis, whereas secretory CLU overexpression offered no cytoprotection. In contrast, nCLU sensitized HaCaT cells to apoptosis. Our data suggest that vanadium-mediated apoptosis was promoted by c-fos, leading to alterations in CLU isoform processing and induction of the pro-death nCLU protein. PMID:19531052
van der Heiden, Marieke; Duizendstra, Aafke; Berbers, Guy A M; Boots, Annemieke M H; Buisman, Anne-Marie
2017-10-09
Vaccines frequently induce suboptimal immune responses in the elderly, due to immunological ageing. Timely vaccination may be a strategy to overcome this problem, which classifies middle-aged adults asan interesting target group for future vaccine interventions. However, the immunological fitness of the middle-aged population is ill-defined. It is currently unknown whether effective T-cell help towards B-cells is initiated by conjugate-carrier vaccines at middle-age. We characterized systemic Tetanus Toxoid (TT) specific T-helper cell responses in the circulation of middle-aged adults (50-65years of age, n=31) having received the MenACWY-TT vaccination. Blood samples were taken pre- as well as 7days, 28days, and 1year post-vaccination. TT-specific T-cell responses were determined by IFNγ Elispot and by the secretion of IFNγ, IL13, IL10, IL17, and IL21 in cell culture supernatants. Circulating CD4+CXCR5+ICOS+IL21+ cells were analyzed by flow cytometry, and meningococcal and TT-specific IgG responses by bead-based immunoassays. The correlation between the T-cell help and humoral responses was evaluated. Vaccination with a TT-carrier protein induced a mixed TT-specific Th1 (IFNγ), Th2 (IL13, IL10), and Th17 (IL17) response in most participants. Additionally, circulating CD4+CXCR5+ICOS+IL21+ cells were significantly increased 7days post-vaccination. Pre-vaccination TT-specific cytokine production and post-vaccination Th2 responses correlated positively with the increase of CD4+CXCR5+ICOS+IL21+ cells. No correlation between T-cell help and antibody responses was found. The characteristics of the T-cell response upon a TT-carrier vaccination suggests effective T-cell help towards B-cells in response to meningococcal polysaccharides, although the absence of a correlation with the antibody responses warrants further clarification. However, the robust T-helper cell response in middle-aged adults, decades after previous TT vaccinations, strengthens the classification of
Yang, Jie; Liu, Lidong; Yang, Yiming; Kong, Ning; Jiang, Xueyu; Sun, Juan; Xie, Rufeng
2017-01-01
Tolerogenic dendritic cells (tDCs) can expand TGF- β -induced regulatory T cells (iTregs); however, the therapeutic utility of these expanded iTregs in autoimmune diseases remains unknown. We sought to determine the properties of iTregs expanded by mature tolerogenic dendritic cells (iTreg mtDC ) in vitro and explore their potential to ameliorate collagen-induced arthritis (CIA) in a mouse model. After induction by TGF- β and expansion by mature tDCs (mtDCs), the phenotype and proliferation of iTreg mtDC were assessed by flow cytometry. The ability of iTregs and iTreg mtDC to inhibit CD4 + T cell proliferation and suppress Th17 cell differentiation was compared. Following adoptive transfer of iTregs and iTreg mtDC to mice with CIA, the clinical and histopathologic scores, serum levels of IFN- γ , TNF- α , IL-17, IL-6, IL-10, TGF- β and anti-CII antibodies, and the distribution of the CD4 + Th subset were assessed. Compared with iTregs, iTreg mtDC expressed higher levels of Foxp3 and suppressed CD4 + T cell proliferation and Th17 cell differentiation to a greater extent. In vivo, iTreg mtDC reduced the severity and progression of CIA more significantly than iTregs, which was associated with a modulated inflammatory cytokine profile, reduced anti-CII IgG levels, and polarized Treg/Th17 balance. This study highlights the potential therapeutic utility of iTreg mtDC in autoimmune arthritis and should facilitate the future design of iTreg immunotherapeutic strategies.
Liu, Lidong; Kong, Ning; Jiang, Xueyu; Sun, Juan; Xie, Rufeng
2017-01-01
Objective Tolerogenic dendritic cells (tDCs) can expand TGF-β-induced regulatory T cells (iTregs); however, the therapeutic utility of these expanded iTregs in autoimmune diseases remains unknown. We sought to determine the properties of iTregs expanded by mature tolerogenic dendritic cells (iTregmtDC) in vitro and explore their potential to ameliorate collagen-induced arthritis (CIA) in a mouse model. Methods After induction by TGF-β and expansion by mature tDCs (mtDCs), the phenotype and proliferation of iTregmtDC were assessed by flow cytometry. The ability of iTregs and iTregmtDC to inhibit CD4+ T cell proliferation and suppress Th17 cell differentiation was compared. Following adoptive transfer of iTregs and iTregmtDC to mice with CIA, the clinical and histopathologic scores, serum levels of IFN-γ, TNF-α, IL-17, IL-6, IL-10, TGF-β and anti-CII antibodies, and the distribution of the CD4+ Th subset were assessed. Results Compared with iTregs, iTregmtDC expressed higher levels of Foxp3 and suppressed CD4+ T cell proliferation and Th17 cell differentiation to a greater extent. In vivo, iTregmtDC reduced the severity and progression of CIA more significantly than iTregs, which was associated with a modulated inflammatory cytokine profile, reduced anti-CII IgG levels, and polarized Treg/Th17 balance. Conclusion This study highlights the potential therapeutic utility of iTregmtDC in autoimmune arthritis and should facilitate the future design of iTreg immunotherapeutic strategies. PMID:28702462
Yu, Ya-qiong; Guo, Jia-jie; Qiu, Li-hong; Lv, You; Jia, Ge; Guo, Yan
2013-08-01
To investigate the effect of NF-κB signaling on the expression of interleukin-6(IL-6) induced by lipopolysaccharides(LPS) extracted from Porphyromonas endodontalis(P.e) in MC3T3-El cells. MC3T3-E1 cells were pretreated with BAY-117082 for 1 h, and then were treated with 10 mg/L P.e-LPS for different times. The translocation of NF-κB was observed by immunofluorescence. The expression of IL-6 was detected by reverse transcription polymerse chain reaction (RT-PCR) and enzyme-linked immuno sorbent assay (ELISA). Statistical analysis was performed using multi-way ANOVA and Dunnett's t test with SPSS 13.0 software package. The staining of NF-κB was mostly in cytoplasm in untreated cells. Rapid translocation of NF-κB into nucleus was observed in the cells stimulated for 30 min and mostly relocalization of NF-κB from nucleus to cytoplasm was observed after 60 min. Pretreatment with 10 μmol/L BAY-117082 for 1h significantly inhibited P.e-LPS-induced translocation of NF-κB .The mRNA and proteins of IL-6 decreased significantly after pretreatment with 10 μmol/L BAY-117082 and the expression of IL-6 proteins was reduced from (774.983±6.585) ng/L to (377.384±14.620) ng/L (P<0.01). The group of treatment with BAY-117082 alone had no significant difference from the blank control group. P.e-LPS can induce translocation of NF-κB in mouse osteoblast MC3T3-El, and P.e-LPS may induce the expression of IL-6 in mouse osteoblast through the signaling of NF-κB.
Muri, Jonathan; Heer, Sebastian; Matsushita, Mai; Pohlmeier, Lea; Tortola, Luigi; Fuhrer, Tobias; Conrad, Marcus; Zamboni, Nicola; Kisielow, Jan; Kopf, Manfred
2018-05-10
The thioredoxin-1 (Trx1) system is an important contributor to cellular redox balance and is a sensor of energy and glucose metabolism. Here we show critical c-Myc-dependent activation of the Trx1 system during thymocyte and peripheral T-cell proliferation, but repression during T-cell quiescence. Deletion of thioredoxin reductase-1 (Txnrd1) prevents expansion the CD4 - CD8 - thymocyte population, whereas Txnrd1 deletion in CD4 + CD8 + thymocytes does not affect further maturation and peripheral homeostasis of αβT cells. However, Txnrd1 is critical for expansion of the activated T-cell population during viral and parasite infection. Metabolomics show that TrxR1 is essential for the last step of nucleotide biosynthesis by donating reducing equivalents to ribonucleotide reductase. Impaired availability of 2'-deoxyribonucleotides induces the DNA damage response and cell cycle arrest of Txnrd1-deficient T cells. These results uncover a pivotal function of the Trx1 system in metabolic reprogramming of thymic and peripheral T cells and provide a rationale for targeting Txnrd1 in T-cell leukemia.
Choi, Seung-Chul; Hutchinson, Tarun E.; Titov, Anton A.; Seay, Howard R.; Li, Shiwu; Brusko, Todd M.; Croker, Byron P.; Salek-Ardakani, Shahram; Morel, Laurence
2016-01-01
Pbx1 controls chromatin accessibility to a large number of genes and is entirely conserved between mice and humans. The Pbx1-d dominant negative isoform is more frequent in the CD4+ T cells from lupus patients than from healthy controls. Pbx1-d is associated with the production of autoreactive T cells in mice carrying the Sle1a1 lupus susceptibility locus. Transgenic expression of Pbx1-d in CD4+ T cells reproduced the phenotypes of Sle1a1 mice, with increased inflammatory functions of CD4+ T cells and impaired regulatory T cell homeostasis. Pbx1-d Tg also expanded the number of follicular helper T cells in a cell-intrinsic and antigen-specific manner that was enhanced in recall responses, and resulted in TH1-biased antibodies. Moreover, Pbx1-d Tg CD4+ T cells upregulated the expression of miR-10a, miR-21 and miR-155, which have been implicated in Treg and TFH cell homeostasis. Our results suggest that Pbx1-d impacts lupus development by regulating effector T cell differentiation and promoting TFH cells at the expense of Treg cells. In addition, our results identify Pbx1 as a novel regulator of CD4+ T cell effector function. PMID:27296664
Zunino, Susan J.; Storms, David H.
2017-01-01
Resveratrol has been reported to inhibit or induce DNA damage, depending upon the type of cell and the experimental conditions. Dietary resveratrol is present in the body predominantly as metabolites and limited data is available concerning the activities of these metabolic products. In the present study, physiologically obtainable levels of the resveratrol metabolites resveratrol-3-O-glucuronide, resveratrol-4′-O-glucuronide and resveratrol-3-O-sulfate were evaluated for their ability to protect Jurkat T cells against DNA damage induced by the topoisomerase I inhibitors camptothecin and topotecan. The cells were pretreated for 24 h with 10 µM resveratrol aglycone or each resveratrol metabolite prior to the induction of DNA damage with camptothecin or topotecan. In separate experiments, the cells were co-treated with resveratrol or its metabolites, and a topoisomerase I inhibitor. The detection of histone 2AX phosphorylation and terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) were used to determine DNA damage, and apoptosis was measured using an antibody against cleaved poly ADP-ribose polymerase. It was identified that pretreatment of the cells with resveratrol-3-O-glucuronide and resveratrol-4′-O-glucuronide reduced the mean fluorescence intensity of staining for DNA strand breaks following treatment with camptothecin, while the percentage of cells undergoing apoptosis was unchanged. However, pretreatment of the cells with resveratrol aglycone increased the DNA damage and apoptosis induced by the drugs. These results suggest that the glucuronide metabolites of resveratrol partially protected the cells from DNA damage, but did not influence the induction of cell death by camptothecin and topotecan. These data suggest that resveratrol aglycone treatment may be beneficial for treating types of cancer that have direct contact with resveratrol prior to its metabolism, including gastrointestinal cancers, which are routinely treated with
Function of the Nucleotide Exchange Activity of Vav1 in T cell Development and Activation*
Saveliev, Alexander; Vanes, Lesley; Ksionda, Olga; Rapley, Jonathan; Smerdon, Stephen J.; Rittinger, Katrin; Tybulewicz, Victor L. J.
2012-01-01
The guanine nucleotide exchange factor (GEF) Vav1 is essential for transducing T cell antigen receptor (TCR) signals and therefore plays a critical role in the development and activation of T cells. It has been presumed that the GEF activity of Vav1 is important for its function; however, there has been no direct demonstration of this. Here, we generated mice expressing enzymatically inactive, but normally folded, Vav1 protein. Analysis of these mice showed that the GEF activity of Vav1 was necessary for the selection of thymocytes and for the optimal activation of T cells, including signal transduction to Rac1, Akt, and integrins. In contrast, the GEF activity of Vav1 was not required for TCR-induced calcium flux, activation of extracellular signal–regulated kinase (ERK) and protein kinase D1 (PKD1), and cell polarization. Thus, in T cells, the GEF activity of Vav1 is essential for some, but not all, of its functions. PMID:20009105
The cathepsin B inhibitor z-FA-CMK induces cell death in leukemic T cells via oxidative stress.
Liow, K Y; Chow, Sek C
2018-01-01
The cathepsin B inhibitor benzyloxycarbonyl-phenylalanine-alanine-chloromethyl ketone (z-FA-CMK) was recently found to induce apoptosis at low concentrations in Jurkat T cells, while at higher concentrations, the cells die of necrosis. In the present study, we showed that z-FA-CMK readily depletes intracellular glutathione (GSH) with a concomitant increase in reactive oxygen species (ROS) generation. The toxicity of z-FA-CMK in Jurkat T cells was completely abrogated by N-acetylcysteine (NAC), suggesting that the toxicity mediated by z-FA-CMK is due to oxidative stress. We found that L-buthionine sulfoximine (BSO) which depletes intracellular GSH through the inhibition of GSH biosynthesis in Jurkat T cells did not promote ROS increase or induce cell death. However, NAC was still able to block z-FA-CMK toxicity in Jurkat T cells in the presence of BSO, indicating that the protective effect of NAC does not involve GSH biosynthesis. This is further corroborated by the protective effect of the non-metabolically active D-cysteine on z-FA-CMK toxicity. Furthermore, in BSO-treated cells, z-FA-CMK-induced ROS increased which remains unchanged, suggesting that the depletion of GSH and increase in ROS generation mediated by z-FA-CMK may be two separate events. Collectively, our results demonstrated that z-FA-CMK toxicity is mediated by oxidative stress through the increase in ROS generation.
Hettich, Michael; Lahoti, Jayashree; Prasad, Shruthi; Niedermann, Gabriele
2016-08-15
T cell-recruiting bispecific antibodies (bsAb) show promise in hematologic malignancies and are also being evaluated in solid tumors. In this study, we investigated whether T cell-recruiting bsAbs synergize with hypofractionated tumor radiotherapy (hRT) and/or blockade of the programmed death-1 (PD-1) immune checkpoint, both of which can increase tumor-infiltrating lymphocyte (TIL) numbers. Unexpectedly, large melanomas treated with hRT plus bsAb (AC133×CD3) relapsed faster than those treated with hRT alone, accompanied by massive TIL apoptosis. This fast relapse was delayed by the further addition of anti-PD-1. Mechanistic investigations revealed restimulation-induced cell death mediated by BIM and FAS as an additional cause of bsAb-mediated TIL depletion. In contrast, the double combination of hRT and anti-PD-1 strongly increased TIL numbers, and even very large tumors were completely eradicated. Our study reveals the risk that CD3-engaging bsAbs can induce apoptotic TIL depletion followed by rapid tumor regrowth, reminiscent of tolerance induction by CD3 mAb-mediated T-cell depletion, warranting caution in their use for the treatment of solid tumors. Our findings also argue that combining radiotherapy and anti-PD-1 can be quite potent, including against very large tumors. Cancer Res; 76(16); 4673-83. ©2016 AACR. ©2016 American Association for Cancer Research.
Torres, AnnMarie; Luke, Joanna D; Kullas, Amy L; Kapilashrami, Kanishk; Botbol, Yair; Koller, Antonius; Tonge, Peter J; Chen, Emily I; Macian, Fernando; van der Velden, Adrianus W M
2016-02-01
Salmonellae are pathogenic bacteria that induce immunosuppression by mechanisms that remain largely unknown. Previously, we showed that a putative type II l-asparaginase produced by Salmonella Typhimurium inhibits T cell responses and mediates virulence in a murine model of infection. Here, we report that this putative L-asparaginase exhibits L-asparagine hydrolase activity required for Salmonella Typhimurium to inhibit T cells. We show that L-asparagine is a nutrient important for T cell activation and that L-asparagine deprivation, such as that mediated by the Salmonella Typhimurium L-asparaginase, causes suppression of activation-induced mammalian target of rapamycin signaling, autophagy, Myc expression, and L-lactate secretion. We also show that L-asparagine deprivation mediated by the Salmonella Typhimurium L-asparaginase causes suppression of cellular processes and pathways involved in protein synthesis, metabolism, and immune response. Our results advance knowledge of a mechanism used by Salmonella Typhimurium to inhibit T cell responses and mediate virulence, and provide new insights into the prerequisites of T cell activation. We propose a model in which l-asparagine deprivation inhibits T cell exit from quiescence by causing suppression of activation-induced metabolic reprogramming. © Society for Leukocyte Biology.
Shikonin induces apoptosis of HaCaT cells via the mitochondrial, Erk and Akt pathways
JING, HUILING; SUN, WENYAN; FAN, JINGHUA; ZHANG, YANMIN; YANG, JIAO; JIA, JINJING; LI, JICHANG; GUO, JIAQI; LUO, SUJU; ZHENG, YAN
2016-01-01
Shikonin, which is a major ingredient of the traditional Chinese herb Lithospermum erythrorhizon, possesses various biological functions, including antimicrobial, anti-inflammatory, and antitumor activities. The present study aimed to determine the molecular mechanisms underlying the effects of shikonin on HaCaT cell apoptosis. Treatment with shikonin significantly inhibited the viability of HaCaT cells in a dose- and time-dependent manner, and promoted cell cycle arrest at G0/G1 phase and apoptosis. In addition, shikonin treatment reduced the mitochondrial membrane potential and induced reactive oxygen species generation. The results of a western blot analysis demonstrated that shikonin significantly activated caspase 3 expression, downregulated B-cell lymphoma 2 (Bcl-2) expression, and upregulated Bcl-2-associated X protein and Bcl-2 homologous antagonist killer expression in a dose-dependent manner in HaCaT cells. Furthermore, shikonin decreased extracellular signal-regulated kinase (Erk) and Akt phosphorylation. These results indicated that shikonin may exert its anti-proliferative effects by inducing apoptosis via activation of the mitochondrial signaling pathway and inactivation of the Akt and Erk pathways in HaCaT cells. Therefore, the present study suggested that shikonin may have potential as a component of therapeutic strategies for the treatment of skin diseases. PMID:26935874
Nonhuman TRIM5 Variants Enhance Recognition of HIV-1-Infected Cells by CD8+ T Cells
Jimenez-Moyano, Esther; Ruiz, Alba; Kløverpris, Henrik N.; Rodriguez-Plata, Maria T.; Peña, Ruth; Blondeau, Caroline; Selwood, David L.; Izquierdo-Useros, Nuria; Moris, Arnaud; Clotet, Bonaventura; Goulder, Philip; Towers, Greg J.
2016-01-01
ABSTRACT Tripartite motif-containing protein 5 (TRIM5) restricts human immunodeficiency virus type 1 (HIV-1) in a species-specific manner by uncoating viral particles while activating early innate responses. Although the contribution of TRIM5 proteins to cellular immunity has not yet been studied, their interactions with the incoming viral capsid and the cellular proteasome led us to hypothesize a role for them. Here, we investigate whether the expression of two nonhuman TRIM5 orthologs, rhesus TRIM5α (RhT5) and TRIM-cyclophilin A (TCyp), both of which are potent restrictors of HIV-1, could enhance immune recognition of infected cells by CD8+ T cells. We illustrate how TRIM5 restriction improves CD8+ T-cell-mediated HIV-1 inhibition. Moreover, when TRIM5 activity was blocked by the nonimmunosuppressive analog of cyclosporine (CsA), sarcosine-3(4-methylbenzoate)–CsA (SmBz-CsA), we found a significant reduction in CD107a/MIP-1β expression in HIV-1-specific CD8+ T cells. This finding underscores the direct link between TRIM5 restriction and activation of CD8+ T-cell responses. Interestingly, cells expressing RhT5 induced stronger CD8+ T-cell responses through the specific recognition of the HIV-1 capsid by the immune system. The underlying mechanism of this process may involve TRIM5-specific capsid recruitment to cellular proteasomes and increase peptide availability for loading and presentation of HLA class I antigens. In summary, we identified a novel function for nonhuman TRIM5 variants in cellular immunity. We hypothesize that TRIM5 can couple innate viral sensing and CD8+ T-cell activation to increase species barriers against retrovirus infection. IMPORTANCE New therapeutics to tackle HIV-1 infection should aim to combine rapid innate viral sensing and cellular immune recognition. Such strategies could prevent seeding of the viral reservoir and the immune damage that occurs during acute infection. The nonhuman TRIM5 variants, rhesus TRIM5α (RhT5) and TRIM
Function of the nucleotide exchange activity of vav1 in T cell development and activation.
Saveliev, Alexander; Vanes, Lesley; Ksionda, Olga; Rapley, Jonathan; Smerdon, Stephen J; Rittinger, Katrin; Tybulewicz, Victor L J
2009-12-15
The guanine nucleotide exchange factor (GEF) Vav1 is essential for transducing T cell antigen receptor (TCR) signals and therefore plays a critical role in the development and activation of T cells. It has been presumed that the GEF activity of Vav1 is important for its function; however, there has been no direct demonstration of this. Here, we generated mice expressing enzymatically inactive, but normally folded, Vav1 protein. Analysis of these mice showed that the GEF activity of Vav1 was necessary for the selection of thymocytes and for the optimal activation of T cells, including signal transduction to Rac1, Akt, and integrins. In contrast, the GEF activity of Vav1 was not required for TCR-induced calcium flux, activation of extracellular signal-regulated kinase and protein kinase D1, and cell polarization. Thus, in T cells, the GEF activity of Vav1 is essential for some, but not all, of its functions.
Rai, Priyamvada
2012-01-01
Oncogenic RAS-induced reactive oxygen species (ROS) trigger barriers to cell transformation and cancer progression through tumor-suppressive responses such as cellular senescence or cell death. We have recently shown that oncogenic RAS-induced DNA damage and attendant premature senescence can be prevented by overexpressing human MutT Homolog 1 (MTH1), the major mammalian detoxifier of the oxidized DNA precursor, 8-oxo-dGTP. Paradoxically, RAS-induced ROS are also able to participate in tumor progression via transformative processes such as mitogenic signaling, the epithelial-mesenchymal transition (EMT), anoikis inhibition, and PI3K/Akt-mediated survival signaling. Here we provide a preliminary insight into the influence of MTH1 levels on the EMT phenotype and Akt activation in RAS-transformed HMLE breast epithelial cells. Within this context, we will discuss the implications of MTH1 upregulation in oncogenic RAS-sustaining cells as a beneficial adaptive change that inhibits ROS-mediated cell senescence and participates in the maintenance of ROS-associated tumor-promoting mechanisms. Accordingly, targeting MTH1 in RAS-transformed tumor cells will not only induce proliferative defects but also potentially enhance therapeutic cytotoxicity by shifting cellular response away from pro-survival mechanisms.
Cornejo, Melanie G.; Scholl, Claudia; Liu, Jianing; Leeman, Dena S.; Haydu, J. Erika; Fröhling, Stefan; Lee, Benjamin H.; Gilliland, D. Gary
2008-01-01
To study the impact of oncogenic K-Ras on T-cell leukemia/lymphoma development and progression, we made use of a conditional K-RasG12D murine knockin model, in which oncogenic K-Ras is expressed from its endogenous promoter. Transplantation of whole bone marrow cells that express oncogenic K-Ras into wild-type recipient mice resulted in a highly penetrant, aggressive T-cell leukemia/lymphoma. The lymphoblasts were composed of a CD4/CD8 double-positive population that aberrantly expressed CD44. Thymi of primary donor mice showed reduced cellularity, and immunophenotypic analysis demonstrated a block in differentiation at the double-negative 1 stage. With progression of disease, approximately 50% of mice acquired Notch1 mutations within the PEST domain. Of note, primary lymphoblasts were hypersensitive to γ-secretase inhibitor treatment, which is known to impair Notch signaling. This inhibition was Notch-specific as assessed by down-regulation of Notch1 target genes and intracellular cleaved Notch. We also observed that the oncogenic K-Ras-induced T-cell disease was responsive to rapamycin and inhibitors of the RAS/MAPK pathway. These data indicate that patients with T-cell leukemia with K-Ras mutations may benefit from therapies that target the NOTCH pathway alone or in combination with inhibition of the PI3K/AKT/MTOR and RAS/MAPK pathways. PMID:18663146
Candida albicans morphology and dendritic cell subsets determine T helper cell differentiation
Gerami-Nejad, Maryam; Kumamoto, Yosuke; Mohammed, Javed A.; Jarrett, Elizabeth; Drummond, Rebecca A.; Zurawski, Sandra M.; Zurawski, Gerard; Berman, Judith; Iwasaki, Akiko; Brown, Gordon D.; Kaplan, Daniel H.
2015-01-01
Summary Candida albicans is a dimorphic fungus responsible for chronic mucocutaneous and systemic infections. Mucocutaneous immunity to C. albicans requires T helper-17 (Th17) cell differentiation that is thought to depend on recognition of filamentous C. albicans. Systemic immunity is considered T cell independent. Using a murine skin infection model, we compared T helper cell responses to yeast and filamentous C. albicans, We found that only yeast induced Th17 cell responses through a mechanism that required Dectin-1 mediated expression of interleukin-6 (IL-6) by Langerhans cells. Filamentous forms induced Th1 without Th17 cell responses due to the absence of Dectin-1 ligation. Notably, Th17 cell responses provided protection against cutaneous infection while Th1 cell responses provided protection against systemic infection. Thus, C. albicans morphology drives distinct T helper cell responses that provide tissue specific protection. These findings provide insight into compartmentalization of Th responses, C. albicans pathogenesis and have critical implications for vaccine strategies. PMID:25680275
Lyng, Fiona M; Desplanques, Maxime; Jella, Kishore Kumar; Garcia, Amaya; McClean, Brendan
2012-10-01
The aim of this study was to investigate the importance of serum serotonin levels in the measurement of bystander cell death. The study was undertaken as part of an intercomparison exercise involving seven European laboratories funded under the European Union Sixth Framework Programme (FP6) Non-Targeted Effects (NOTE) integrated project. Three batches of foetal bovine serum were tested; serum with high and low serotonin content from the intercomparison exercise as well as serum from the home laboratory. Three sets of human keratinocytes (HaCaT cell line) were cultured in DMEM:F12 medium supplemented with serum with high or low serotonin content or serum from the home laboratory and both donor and recipient HaCaT cells were plated. The donor HaCaT cells were irradiated (0.5 Gy) using a cobalt 60 teletherapy unit, the medium was harvested 1 hour post irradiation and transferred to the recipient HaCaT cells. Bystander induced cell death was measured by the clonogenic survival assay and the Alamar blue viability assay. A significant reduction in cell survival, as measured by the clonogenic assay, and in cell viability, as measured by the Alamar blue assay, was observed in the recipient HaCaT cells treated with medium from irradiated cells compared to the cells treated with medium from unirradiated cells. No significant difference was found between the three batches of serum. The data suggest that in our cell system and with our endpoints (clonogenic assay and Alamar blue assay), serum serotonin levels do not play a role in bystander-induced cell death.
Dexamethasone attenuates echinococcosis-induced allergic reactions via regulatory T cells in mice.
Zhang, Qin; Ye, Jianrong; Zheng, Hong
2016-03-11
Cystic echinococcosis (CE), caused by infection with Echinococcus granulosus larvae, is a potentially life-threatening disease in humans. Anaphylactic shock caused by CE is very dangerous, and is highly prevalent during surgery. Dexamethasone (DEX) is used clinically before operations to prevent allergic reactions; Regulatory T cells (Treg cells) are believed to be associated with negative immune response, which play an important role in alleviating allergic reactions. However, the association of Treg cells with DEX remains unknown. In this study, C57BL/6 mice were divided into uninfected group, untreated group and DEX group which were inoculated with protoscoleces from E. granulosus and sensitized using a cyst fluid suspension to induce anaphylactic shock. In addition, the mice in DEX group were treated with 10 mg/kg DEX by intraperitoneal injection 30 min before being sensitized. It was found that 93.75 % of all sensitized mice experienced allergic symptoms. The levels of IgE, IgE/IgG, and IgE/IgG1 were significantly higher in both untreated group and DEX group. The proportion of CD4 + CD25 + FOXP3 + Treg cells relative to CD4+ Treg cells, and the levels of interleukin-10 (IL-10) and tumor growth factor-β (TGF-β1) were significantly higher in DEX group. The level of IL-13 was significantly higher in the sensitized mice than in the other groups. These cells may play a key role in alleviating the immune response in CE-induced anaphylactic shock. The protective effect of DEX may be due to Treg cell upregulating IL-10 and TGF-β1 levels, and inhibiting helper T cell 2 cytokines.
Zheng, Bin; van Bergenhenegouwen, Jeroen; Overbeek, Saskia; van de Kant, Hendrik J G; Garssen, Johan; Folkerts, Gert; Vos, Paul; Morgan, Mary E; Kraneveld, Aletta D
2014-01-01
While some probiotics have shown beneficial effects on preventing or treating colitis development, others have shown no effects. In this study, we have assessed the immunomodulating effects of two probiotic strains, Lactobacillus rhamnosus (L. rhamnosus) and Bifidobacterium breve (B. breve) on T cell polarization in vitro, using human peripheral blood mononuclear cells (PBMC), and in vivo, using murine dextran sodium sulfate (DSS) colitis model. With respect to the latter, the mRNA expression of T cell subset-associated transcription factors and cytokines in the colon was measured and the T helper type (Th) 17 and regulatory T cell (Treg) subsets were determined in the Peyer's patches. Both L. rhamnosus and B. breve incubations in vitro reduced Th17 and increased Th2 cell subsets in human PBMCs. In addition, B. breve incubation was also able to reduce Th1 and increase Treg cell subsets in contrast to L. rhamnosus. In vivo intervention with B. breve, but not L. rhamnosus, significantly attenuated the severity of DSS-induced colitis. In DSS-treated C57BL/6 mice, intervention with B. breve increased the expression of mRNA encoding for Th2- and Treg-associated cytokines in the distal colon. In addition, intervention with B. breve led to increases of Treg and decreases of Th17 cell subsets in Peyer's patches of DSS-treated mice. B. breve modulates T cell polarization towards Th2 and Treg cell-associated responses in vitro and in vivo. In vivo B. breve intervention ameliorates DSS-induced colitis symptoms and this protective effect may mediated by its effects on the T-cell composition.
Zheng, Bin; van Bergenhenegouwen, Jeroen; Overbeek, Saskia; van de Kant, Hendrik J. G.; Garssen, Johan; Folkerts, Gert; Vos, Paul; Morgan, Mary E.; Kraneveld, Aletta D.
2014-01-01
While some probiotics have shown beneficial effects on preventing or treating colitis development, others have shown no effects. In this study, we have assessed the immunomodulating effects of two probiotic strains, Lactobacillus rhamnosus (L. rhamnosus) and Bifidobacterium breve (B. breve) on T cell polarization in vitro, using human peripheral blood mononuclear cells (PBMC), and in vivo, using murine dextran sodium sulfate (DSS) colitis model. With respect to the latter, the mRNA expression of T cell subset-associated transcription factors and cytokines in the colon was measured and the T helper type (Th) 17 and regulatory T cell (Treg) subsets were determined in the Peyer's patches. Both L. rhamnosus and B. breve incubations in vitro reduced Th17 and increased Th2 cell subsets in human PBMCs. In addition, B. breve incubation was also able to reduce Th1 and increase Treg cell subsets in contrast to L. rhamnosus. In vivo intervention with B. breve, but not L. rhamnosus, significantly attenuated the severity of DSS-induced colitis. In DSS-treated C57BL/6 mice, intervention with B. breve increased the expression of mRNA encoding for Th2- and Treg-associated cytokines in the distal colon. In addition, intervention with B. breve led to increases of Treg and decreases of Th17 cell subsets in Peyer's patches of DSS-treated mice. B. breve modulates T cell polarization towards Th2 and Treg cell-associated responses in vitro and in vivo. In vivo B. breve intervention ameliorates DSS-induced colitis symptoms and this protective effect may mediated by its effects on the T-cell composition. PMID:24787575
The Interaction of CD154 with the α5β1 Integrin Inhibits Fas-Induced T Cell Death
Yacoub, Daniel; Salti, Suzanne; Alaaeddine, Nada; Aoudjit, Fawzi; Hassan, Ghada S.; Mourad, Walid
2016-01-01
CD154, a critical regulator of the immune response, is usually associated with chronic inflammatory, autoimmune diseases as well as malignant disorders. In addition to its classical receptor CD40, CD154 is capable of binding other receptors, members of the integrin family, the αIIbβ3, αMβ2 and α5β1. Given the role attributed to integrins and particularly the β1 integrins in inhibiting apoptotic events in normal as well as malignant T cells, we were highly interested in investigating the role of the CD154/α5β1 interaction in promoting survival of malignant T cells contributing as such to tumor development and/or propagation. To support our hypothesis, we first show that soluble CD154 binds to the T-cell acute lymphoblastic leukemia cell line, Jurkat E6.1 in a α5β1-dependent manner. Binding of soluble CD154 to α5β1 integrin of Jurkat cells leads to the activation of key survival proteins, including the p38 and ERK1/2 mitogen-activated protein kinases (MAPKs), phosphoinositide 3 kinase (PI-3K), and Akt. Interestingly, soluble CD154 significantly inhibits Fas-mediated apoptosis in T cell leukemia-lymphoma cell lines, Jurkat E6.1 and HUT78 cells, an important hallmark of T cell survival during malignancy progression. These anti-apoptotic effects were mainly mediated by the activation of the PI-3K/Akt pathway but also involved the p38 and the ERK1/2 MAPKs cascades. Our data also demonstrated that the CD154-triggered inhibition of the Fas-mediated cell death response was dependent on a suppression of caspase-8 cleavage, but independent of de novo protein synthesis or alterations in Fas expression on cell surface. Together, our results highlight the impact of the CD154/α5β1 interaction in T cell function/survival and identify novel targets for the treatment of malignant disorders, particularly of T cell origin. PMID:27391025
The Interaction of CD154 with the α5β1 Integrin Inhibits Fas-Induced T Cell Death.
Bachsais, Meriem; Naddaf, Nadim; Yacoub, Daniel; Salti, Suzanne; Alaaeddine, Nada; Aoudjit, Fawzi; Hassan, Ghada S; Mourad, Walid
2016-01-01
CD154, a critical regulator of the immune response, is usually associated with chronic inflammatory, autoimmune diseases as well as malignant disorders. In addition to its classical receptor CD40, CD154 is capable of binding other receptors, members of the integrin family, the αIIbβ3, αMβ2 and α5β1. Given the role attributed to integrins and particularly the β1 integrins in inhibiting apoptotic events in normal as well as malignant T cells, we were highly interested in investigating the role of the CD154/α5β1 interaction in promoting survival of malignant T cells contributing as such to tumor development and/or propagation. To support our hypothesis, we first show that soluble CD154 binds to the T-cell acute lymphoblastic leukemia cell line, Jurkat E6.1 in a α5β1-dependent manner. Binding of soluble CD154 to α5β1 integrin of Jurkat cells leads to the activation of key survival proteins, including the p38 and ERK1/2 mitogen-activated protein kinases (MAPKs), phosphoinositide 3 kinase (PI-3K), and Akt. Interestingly, soluble CD154 significantly inhibits Fas-mediated apoptosis in T cell leukemia-lymphoma cell lines, Jurkat E6.1 and HUT78 cells, an important hallmark of T cell survival during malignancy progression. These anti-apoptotic effects were mainly mediated by the activation of the PI-3K/Akt pathway but also involved the p38 and the ERK1/2 MAPKs cascades. Our data also demonstrated that the CD154-triggered inhibition of the Fas-mediated cell death response was dependent on a suppression of caspase-8 cleavage, but independent of de novo protein synthesis or alterations in Fas expression on cell surface. Together, our results highlight the impact of the CD154/α5β1 interaction in T cell function/survival and identify novel targets for the treatment of malignant disorders, particularly of T cell origin.
Oxaliplatin antagonizes HIV-1 latency by activating NF-κB without causing global T cell activation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhu, Xiaoli; Liu, Sijie; Wang, Pengfei
Highlights: • The chemotherapeutic drug oxaliplatin reactivates latent HIV-1 in this cell line model of HIV-1 latency. • Reactivation is synergized when oxaliplatin is used in combination with valproic acid. • Oxaliplatin reactivates latent HIV-1 through activation of NF-kB and does not induce T cell activation. - Abstract: Reactivation of latent HIV-1 is a promising strategy for the clearance of the viral reservoirs. Because of the limitations of current agents, identification of new latency activators is urgently required. Using an established model of HIV-1 latency, we examined the effect of Oxaliplatin on latent HIV-1 reactivation. We showed that Oxaliplatin, alonemore » or in combination with valproic acid (VPA), was able to reactivate HIV-1 without inducing global T cell activation. We also provided evidence that Oxaliplatin reactivated HIV-1 expression by inducing nuclear factor kappa B (NF-κB) nuclear translocation. Our results indicated that Oxaliplatin could be a potential drug candidate for anti-latency therapies.« less
Schmidt, Jonas D; Ahlström, Malin G; Johansen, Jeanne D; Dyring-Andersen, Beatrice; Agerbeck, Christina; Nielsen, Morten M; Poulsen, Steen S; Woetmann, Anders; Ødum, Niels; Thomsen, Allan R; Geisler, Carsten; Bonefeld, Charlotte M
2017-04-01
Skin-resident memory T (T RM ) cells are associated with immunological memory in the skin. Whether immunological memory responses to allergens in the skin are solely localized to previously allergen-exposed sites or are present globally in the skin is not clear. Furthermore, the mechanisms whereby T RM cells induce rapid recall responses need further investigation. To study whether contact allergens induce local and/or global memory, and to determine the mechanisms involved in memory responses in the skin. To address these questions, we analysed responses to contact allergens in mice and humans sensitized to 2,4-dinitrofluorobenzene and nickel, respectively. Challenge responses in both mice and humans were dramatically increased at sites previously exposed to allergens as compared with previously unexposed sites. Importantly, the magnitude of the challenge response correlated with the epidermal accumulation of interleukin (IL)-17A-producing and interferon (IFN)-γ-producing T RM cells. Moreover, IL-17A and IFN-γ enhanced allergen-induced IL-1β production in keratinocytes. We show that sensitization with contact allergens induces a strong, long-lasting local memory and a weaker, temporary global immunological memory response to the allergen that is mediated by IL-17A-producing and IFN-γ-producing CD8 + T RM cells. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Lee, Jun Sik; Lee, Je Chul; Lee, Chang-Min; Jung, In Duk; Jeong, Young-Il; Seong, Eun-Young; Chung, Hae-Young; Park, Yeong-Min
2007-06-30
Acinetobacter baumannii is an increasing hospital-acquired pathogen that causes a various type of infections, but little is known about the protective immune response to this microorganism. Outer membrane protein A of A. baumannii (AbOmpA) is a major porin protein and plays an important role in pathogenesis. We analyzed interaction between AbOmpA and dendritic cells (DCs) to characterize the role of this protein in promoting innate and adaptive immune responses. AbOmpA functionally activates bone marrow-derived DCs by augmenting expression of the surface markers, CD40, CD54, B7 family (CD80 and CD86) and major histocompatibility complex class I and II. AbOmpA induces production of Th1-promoting interleukin-12 from DCs and augments the syngeneic and allogeneic immunostimulatory capacity of DCs. AbOmpA stimulates production of interferon-gamma from T cells in mixed lymphocyte reactions, which suggesting Th1-polarizing capacity. CD4(+) T cells stimulated by AbOmpA-stimulated DCs show a Th1-polarizing cytokine profile. The expression of surface markers on DCs is mediated by both mitogen-activated protein kinases and NF-kappaB pathways. Our findings suggest that AbOmpA induces maturation of DCs and drives Th1 polarization, which are important properties for determining the nature of immune response against A. baumannii.
Arumuggam, Niroshaathevi; Melong, Nicole; Too, Catherine Kl; Berman, Jason N; Rupasinghe, Hp Vasantha
2017-01-01
The overall clinical outcome in T-cell acute lymphoblastic leukemia (T-ALL) can be improved by minimizing risk for treatment failure using effective pharmacological adjuvants. Phloridzin (PZ), a flavonoid precursor found in apple peels, was acylated with docosahexaenoic acid (DHA) yielding a novel ester known as phloridzin docosahexaenoate (PZ-DHA). Here, we have studied the cytotoxic effects of PZ-DHA on human leukemia cells using in vitro and in vivo models. The inhibitory effects of PZ-DHA were tested on human Jurkat T-ALL cells in comparison to K562 chronic myeloid leukemia (CML) cells and non-malignant murine T-cells. PZ-DHA, not PZ or DHA alone, reduced cell viability and ATP levels, increased intracellular LDH release, and caused extensive morphological alterations in both Jurkat and K562 cells. PZ-DHA also inhibited cell proliferation, and selectively induced apoptosis in Jurkat and K562 cells while sparing normal murine T-cells. The cytotoxic effects of PZ-DHA on Jurkat cells were associated with caspase activation, DNA fragmentation, and selective down-regulation of STAT3 phosphorylation. PZ-DHA significantly inhibited Jurkat cell proliferation in zebrafish larvae; however, the proliferation of K562 cells was not affected in vivo . We propose that PZ-DHA-induced cytotoxic response is selective towards T-ALL in the presence of a tumor-stromal microenvironment. Prospective studies evaluating the combinatorial effects of PZ-DHA with conventional chemotherapy for T-ALL are underway.
Arumuggam, Niroshaathevi; Melong, Nicole; Too, Catherine KL; Berman, Jason N; Rupasinghe, HP Vasantha
2017-01-01
The overall clinical outcome in T-cell acute lymphoblastic leukemia (T-ALL) can be improved by minimizing risk for treatment failure using effective pharmacological adjuvants. Phloridzin (PZ), a flavonoid precursor found in apple peels, was acylated with docosahexaenoic acid (DHA) yielding a novel ester known as phloridzin docosahexaenoate (PZ-DHA). Here, we have studied the cytotoxic effects of PZ-DHA on human leukemia cells using in vitro and in vivo models. The inhibitory effects of PZ-DHA were tested on human Jurkat T-ALL cells in comparison to K562 chronic myeloid leukemia (CML) cells and non-malignant murine T-cells. PZ-DHA, not PZ or DHA alone, reduced cell viability and ATP levels, increased intracellular LDH release, and caused extensive morphological alterations in both Jurkat and K562 cells. PZ-DHA also inhibited cell proliferation, and selectively induced apoptosis in Jurkat and K562 cells while sparing normal murine T-cells. The cytotoxic effects of PZ-DHA on Jurkat cells were associated with caspase activation, DNA fragmentation, and selective down-regulation of STAT3 phosphorylation. PZ-DHA significantly inhibited Jurkat cell proliferation in zebrafish larvae; however, the proliferation of K562 cells was not affected in vivo. We propose that PZ-DHA-induced cytotoxic response is selective towards T-ALL in the presence of a tumor-stromal microenvironment. Prospective studies evaluating the combinatorial effects of PZ-DHA with conventional chemotherapy for T-ALL are underway. PMID:29312799
Kochetkova, Irina; Thornburg, Theresa; Callis, Gayle; Pascual, David W.
2011-01-01
Oral immunization with a Salmonella vaccine vector expressing enterotoxigenic E. coli colonization factor antigen I (CFA/I) can protect against collagen-induced arthritis (CIA) by dampening IL-17 and IFN-γ via enhanced IL-4, IL-10, and TGF-β. To identify the responsible regulatory CD4+ T cells making the host refractory to CIA, Salmonella-CFA/I induced CD39+CD4+ T cells with enhanced apyrase activity relative to Salmonella vector-immunized mice. Adoptive transfer of vaccine-induced CD39+CD4+ T cells into CIA mice conferred complete protection, while CD39−CD4+ T cells did not. Subsequent analysis of vaccinated FoxP3-GFP mice revealed the CD39+ T cells were composed of FoxP3-GFP− and FoxP3-GFP+ subpopulations. Although each adoptively transferred Salmonella-CFA/I-induced FoxP3− and FoxP3+CD39+CD4+ T cells could protect against CIA, each subset was not as efficacious as total CD39+CD4+ T cells, suggesting their interdependence for optimal protection. Cytokine analysis revealed FoxP3− CD39+CD4+ T cells produced TGF-β, and FoxP3+CD39+CD4+ T cells produced IL-10, showing a segregation of function. Moreover, donor FoxP3-GFP− CD4+ T cells converted to FoxP3-GFP+ CD39+CD4+ T cells in the recipients, showing plasticity of these regulatory T cells. TGF-β was found to be essential for protection since in vivo TGF-β neutralization reversed activation of cAMP-response element-binding protein (CREB) and reduced the development of CD39+CD4+ T cells. Thus, CD39 apyrase-expressing CD4+ T cells stimulated by Salmonella-CFA/I are composed of TGF-β-producing FoxP3− CD39+CD4+ T cells and support the stimulation of IL-10-producing FoxP3+ CD39+CD4+ T cells. PMID:21967895
Li, Ke-Ran; Yang, Su-Qing; Gong, Yi-Qing; Yang, Hong; Li, Xiu-Miao; Zhao, Yu-Xia; Yao, Jin; Jiang, Qin; Cao, Cong
2016-05-06
Excessive UV radiation and reactive oxygen species (ROS) cause retinal pigment epithelium (RPE) cell injuries. Nrf2 regulates transcriptional activation of many anti-oxidant genes. Here, we tested the potential role of 3H-1,2-dithiole-3-thione (D3T) against UV or ROS damages in cultured RPE cells (both primary cells and ARPE-19 line). We showed that D3T significantly inhibited UV-/H2O2-induced RPE cell death and apoptosis. UV-stimulated ROS production was dramatically inhibited by D3T pretreatment. D3T induced Nrf2 phosphorylation in cultured RPE cells, causing Nrf2 disassociation with KEAP1 and its subsequent nuclear accumulation. This led to expression of antioxidant response elements (ARE)-dependent gene heme oxygenase-1 (HO-1). Nrf2-HO-1 activation was required for D3T-mediated cytoprotective effect. Nrf2 shRNA knockdown or S40T dominant negative mutation as well as the HO-1 inhibitor Zinc protoporphyrin (ZnPP) largely inhibited D3T's RPE cytoprotective effects against UV radiation. Yet, exogenous overexpression Nrf2 enhanced D3T's activity in RPE cells. Further studies showed that D3T activated Akt/mTORC1 in cultured RPE cells. Akt-mTORC1 inhibitors, or Akt1 knockdown by shRNA, not only inhibited D3T-induced Nrf2-HO-1 activation, but also abolished the RPE cytoprotective effects. In vivo, D3T intravitreal injection protected from light-induced retinal dysfunctions in mice. Thus, D3T protects RPE cells from UV-induced damages via activation of Akt-mTORC1-Nrf2-HO-1 signaling axis.
Jin, Xinchun; Sun, Yanyun; Xu, Ji; Liu, Wenlan
2015-03-01
Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates blood-brain barrier injury and increases the risk of symptomatic cerebral hemorrhage. The mechanism through which tPA enhances MMP-9 activity is not well understood. Here we report an important role of caveolin-1 in mediating tPA-induced MMP-9 synthesis. Brain microvascular endothelial cell line bEnd3 cells were incubated with 5 or 20 μg/ml tPA for 24 hrs before analyzing MMP-9 levels in the conditioned media and cellular extracts by gelatin zymography. tPA at a dose of 20 μg/mL tPA, but not 5 μg/mL, significantly increased MMP-9 level in cultured media while decreasing it in cellular extracts. Concurrently, tPA treatment induced a 2.3-fold increase of caveolin-1 protein levels in endothelial cells. Interestingly, knockdown of Cav-1 with siRNA inhibited tPA-induced MMP-9 mRNA up-regulation and MMP-9 increase in the conditioned media, but did not affect MMP-9 decrease in cellular extracts. These results suggest that caveolin-1 critically contributes to tPA-mediated MMP-9 up-regulation, but may not facilitate MMP-9 secretion in endothelial cells. Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates ischemic blood brain barrier (BBB) injury and increases the risk of symptomatic cerebral hemorrhage. Our results suggest a novel mechanism underlying this tPA-MMP 9 axis. In response to tPA treatment, caveolin-1 protein levels increased in endothelial cells, which mediate MMP-9 mRNA up-regulation and its secretion into extracellular space. Caveolin-1 may, however, not facilitate MMP-9 secretion in endothelial cells. Our data suggest caveolin-1 as a novel therapeutic target for protecting the BBB against ischemic damage. The schematic outlines tPA-induced MMP-9 upreguation. © 2015 International Society for Neurochemistry.
CD22 Promotes B-1b Cell Responses to T Cell-Independent Type 2 Antigens.
Haas, Karen M; Johnson, Kristen L; Phipps, James P; Do, Cardinal
2018-03-01
CD22 (Siglec-2) is a critical regulator of B cell activation and survival. CD22 -/- mice generate significantly impaired Ab responses to T cell-independent type 2 (TI-2) Ags, including haptenated Ficoll and pneumococcal polysaccharides, Ags that elicit poor T cell help and activate BCR signaling via multivalent epitope crosslinking. This has been proposed to be due to impaired marginal zone (MZ) B cell development/maintenance in CD22 -/- mice. However, mice expressing a mutant form of CD22 unable to bind sialic acid ligands generated normal TI-2 Ab responses, despite significantly reduced MZ B cells. Moreover, mice treated with CD22 ligand-binding blocking mAbs, which deplete MZ B cells, had little effect on TI-2 Ab responses. We therefore investigated the effects of CD22 deficiency on B-1b cells, an innate-like B cell population that plays a key role in TI-2 Ab responses. B-1b cells from CD22 -/- mice had impaired BCR-induced proliferation and significantly increased intracellular Ca 2+ concentration responses following BCR crosslinking. Ag-specific B-1b cell expansion and plasmablast differentiation following TI-2 Ag immunization was significantly impaired in CD22 -/- mice, consistent with reduced TI-2 Ab responses. We generated CD22 -/- mice with reduced CD19 levels (CD22 -/- CD19 +/- ) to test the hypothesis that augmented B-1b cell BCR signaling in CD22 -/- mice contributes to impaired TI-2 Ab responses. BCR-induced proliferation and intracellular Ca 2+ concentration responses were normalized in CD22 -/- CD19 +/- B-1b cells. Consistent with this, TI-2 Ag-specific B-1b cell expansion, plasmablast differentiation, survival, and Ab responses were rescued in CD22 -/- CD19 +/- mice. Thus, CD22 plays a critical role in regulating TI-2 Ab responses through regulating B-1b cell signaling thresholds. Copyright © 2018 by The American Association of Immunologists, Inc.
Sojka, Dorothy K.; Fowell, Deborah J.
2011-01-01
CD4+CD25+Forkhead box P3 (Foxp3)+ regulatory T cells (Tregs) control immune responses to self and foreign antigens in secondary lymphoid organs and at tissue sites of inflammation. Tregs can modify the function of many immune cells and have been proposed to block early proliferation, differentiation, and effector function. Acute ablation of Tregs has revealed rapid cytokine production immediately after Treg removal, suggesting that Tregs may regulate effector function acutely rather than regulating the programming for immune function. We developed in vitro and in vivo models that enabled the direct test of Treg regulation of T-helper cell type 1 (Th1) differentiation. CD28 signaling is known to abrogate Treg suppression of IL-2 secretion and proliferation, but our studies show that Treg suppression of IFN-γ during Th1 priming proceeds despite enhanced CD28 signaling. Importantly, during Th1 differentiation, Tregs inhibited early IFN-γ transcription without disrupting expression of Th1-specific T-box transcription factor (Tbet) and Th1 programming. Acute shutoff of effector cytokine production by Tregs was selective for IFN-γ but not TNF-α and was independent of TGF-β and Epstein-Barr virus-induced gene 3. In vivo, Tregs potently controlled CD4 IFN-γ and CD4 effector cell expansion in the lymph node (four- to fivefold reduction) but not Th1 programming, independent of IL-10. Tregs additionally reduced CD4 IFN-γ in the inflamed dermis (twofold reduction) dependent on their production of IL-10. We propose a model for Treg inhibition of effector function based on acute cytokine regulation. Interestingly, Tregs used different regulatory mechanisms to regulate IFN-γ (IL-10–dependent or –independent) subject to the target T-cell stage of activation and its tissue location. PMID:22025707
Smith, Michelle J; Webber, Beau R; Mohtashami, Mahmood; Stefanski, Heather E; Zúñiga-Pflücker, Juan Carlos; Blazar, Bruce R
2015-11-01
T lymphocytes are critical mediators of the adaptive immune system and have the capacity to serve as therapeutic agents in the areas of transplant and cancer immunotherapy. While T cells can be isolated and expanded from patients, T cells derived in vitro from both hematopoietic stem/progenitor cells (HSPCs) and human pluripotent stem cells (hPSCs) offer great potential advantages in generating a self-renewing source of T cells that can be readily genetically modified. T-cell differentiation in vivo is a complex process requiring tightly regulated signals; providing the correct signals in vitro to induce T-cell lineage commitment followed by their development into mature, functional, single positive T cells, is similarly complex. In this review, we discuss current methods for the in vitro derivation of T cells from murine and human HSPCs and hPSCs that use feeder-cell and feeder-cell-free systems. Furthermore, we explore their potential for adoption for use in T-cell-based therapies. © 2015 AlphaMed Press.
Frommer, Friederike; Heinen, Tobias J A J; Wunderlich, F Thomas; Yogev, Nir; Buch, Thorsten; Roers, Axel; Bettelli, Estelle; Müller, Werner; Anderton, Stephen M; Waisman, Ari
2008-10-15
B cells have been shown in various animal models to induce immunological tolerance leading to reduced immune responses and protection from autoimmunity. We show that interaction of B cells with naive T cells results in T cell triggering accompanied by the expression of negative costimulatory molecules such as PD-1, CTLA-4, B and T lymphocyte attenuator, and CD5. Following interaction with B cells, T cells were not induced to proliferate, in a process that was dependent on their expression of PD-1 and CTLA-4, but not CD5. In contrast, the T cells became sensitive to Ag-induced cell death. Our results demonstrate that B cells participate in the homeostasis of the immune system by ablation of conventional self-reactive T cells.
CD95 co-stimulation blocks activation of naive T cells by inhibiting T cell receptor signaling
Lindquist, Jonathan A.; Arhel, Nathalie; Felder, Edward; Karl, Sabine; Haas, Tobias L.; Fulda, Simone; Walczak, Henning; Kirchhoff, Frank; Debatin, Klaus-Michael
2009-01-01
CD95 is a multifunctional receptor that induces cell death or proliferation depending on the signal, cell type, and cellular context. Here, we describe a thus far unknown function of CD95 as a silencer of T cell activation. Naive human T cells triggered by antigen-presenting cells expressing a membrane-bound form of CD95 ligand (CD95L) or stimulated by anti-CD3 and -CD28 antibodies in the presence of recombinant CD95L had reduced activation and proliferation, whereas preactivated, CD95-sensitive T cells underwent apoptosis. Triggering of CD95 during T cell priming interfered with proximal T cell receptor signaling by inhibiting the recruitment of ζ-chain–associated protein of 70 kD, phospholipase-γ, and protein kinase C-θ into lipid rafts, thereby preventing their mutual tyrosine protein phosphorylation. Subsequently, Ca2+ mobilization and nuclear translocation of transcription factors NFAT, AP1, and NF-κB were strongly reduced, leading to impaired cytokine secretion. CD95-mediated inhibition of proliferation in naive T cells could not be reverted by the addition of exogenous interleukin-2 and T cells primed by CD95 co-stimulation remained partially unresponsive upon secondary T cell stimulation. HIV infection induced CD95L expression in primary human antigeen-presenting cells, and thereby suppressed T cell activation, suggesting that CD95/CD95L-mediated silencing of T cell activation represents a novel mechanism of immune evasion. PMID:19487421
INHIBITION OF INDUCED DIFFERENTIATION OF C3H/1OT 1/2 CLONE 8 MOUSE EMBRYO CELLS BY TUMOR PROMOTERS
C3H/10T 1/2 cells were induced to differentiate into muscle cells by treatment with 5-azacytidine, and the effects of tumor promoters, nonpromoters, and inhibitors of tumor promotion on this induced differentiation were investigated. Cell morphology was dramatically changed withi...
Huang, Chuanshu; Li, Jingxia; Zhang, Qi; Huang, Xi
2010-01-01
Activator protein-1 (AP-1) and nuclear factor of activated T cells (NFAT) are two important transcription factors responsible for the regulation of cytokines, which are involved in cell proliferation and inflammation. Coal workers’ pneumoconiosis (CWP) is an occupational lung disease that may be related to chronic inflammation caused by coal dust exposure. In the present study, we demonstrate that coal from the Pennsylvania (PA) coalmine region, which has a high prevalence of CWP, can activate both AP-1 and NFAT in JB6 mouse epidermal cells. In contrast, coal from the Utah (UT) coalmine region, which has a low prevalence of CWP, has no such effects. The PA coal stimulates mitogen-activated protein kinase (MAPK) family members of extracellular signal-regulated kinases (ERKs) and p38 MAPK but not c-Jun-NH2-terminal kinases, as determined by the phosphorylation assay. The increase in AP-1 by the PA coal was completely eliminated by the pretreatment of cells with PD98059, a specific MAPK kinase inhibitor, and SB202190, a p38 kinase inhibitor, further confirming that the PA coal-induced AP-1 activation is mediated through ERKs and p38 MAPK pathways. Deferoxamine (DFO), an iron chelator, synergistically enhanced the PA coal-induced AP-1 activity, but inhibited NFAT activity. For comparison, cells were treated with ferrous sulfate and/or DFO. We have found that iron transactivated both AP-1 and NFAT, and DFO further enhanced iron-induced AP-1 activation but inhibited NFAT. These results indicate that activation of AP-1 and NFAT by the PA coal is through bioavailable iron present in the coal. These data are in agreement with our previous findings that the prevalence of CWP correlates well with levels of bioavailable iron in coals from various mining regions. PMID:12397016
Lentiviral Delivery of HIV-1 Vpr Protein Induces Apoptosis in Transformed Cells
NASA Astrophysics Data System (ADS)
Stewart, Sheila A.; Poon, Betty; Jowett, Jeremy B. M.; Xie, Yiming; Chen, Irvin S. Y.
1999-10-01
Most current anticancer therapies act by inducing tumor cell stasis followed by apoptosis. HIV-1 Vpr effectively induces apoptosis of T cells after arrest of cells at a G2/M checkpoint. Here, we investigated whether this property of Vpr could be exploited for use as a potential anticancer agent. As a potentially safer alternative to transfer of genes encoding Vpr, we developed a method to efficiently introduce Vpr protein directly into cells. Vpr packaged into HIV-1 virions lacking a genome induced efficient cell cycle arrest and apoptosis. Introduction of Vpr into tumor cell lines of various tissue origin, including those bearing predisposing mutations in p53, XPA, and hMLH1, induced cell cycle arrest and apoptosis with high efficiency. Significantly, apoptosis mediated by virion-associated Vpr was more effective on rapidly dividing cells compared with slow-growing cells, thus, in concept, providing a potential differential effect between some types of tumor cells and surrounding normal cells. This model system provides a rationale and proof of concept for the development of potential cancer therapeutic agents based on the growth-arresting and apoptotic properties of Vpr.
Seo, Seung-Hee; Jeong, Gil-Saeng
2015-12-01
Oxidative skin damage and skin inflammation play key roles in the pathogenesis of skin-related diseases. Fisetin is a naturally occurring flavonoid abundantly found in several vegetables and fruits. Fisetin has been shown to exert various positive biological effects, such as anti-cancer, anti-proliferative, neuroprotective and anti-oxidative effects. In this study, we investigate the skin protective effects and anti-inflammatory properties of fisetin in hydrogen peroxide- and TNF-α-challenged human keratinocyte HaCaT cells. When HaCaT cells were treated with non-cytotoxic concentrations of fisetin (1-20μM), heme oxygenase (HO)-1 mRNA and protein expression increased in a dose-dependent manner. Furthermore, fisetin dose-dependently increased cell viability and reduced ROS production in hydrogen peroxide-treated HaCaT cells. Fisetin also inhibited the production of NO, PGE2 IL-1β, IL-6, expression of iNOS and COX-2, and activation of NF-κB in HaCaT cells treated with TNF-α. Fisetin induced Nrf2 translocation to the nuclei. HO-1 siRNA transient transfection reversed the effects of fisetin on cytoprotection, ROS reduction, NO, PGE2, IL-1β, IL-6, and TNF-α production, and NF-κB DNA-binding activity. Moreover, fisetin increased Akt phosphorylation and a PI3K pathway inhibitor (LY294002) abolished fisetin-induced cytoprotection and NO inhibition. Taken together, these results provide evidence for a beneficial role of fisetin in skin therapy. Copyright © 2015. Published by Elsevier B.V.
Thy-1+ dendritic epidermal cells express T3 antigen and the T-cell receptor gamma chain.
Stingl, G; Koning, F; Yamada, H; Yokoyama, W M; Tschachler, E; Bluestone, J A; Steiner, G; Samelson, L E; Lew, A M; Coligan, J E
1987-01-01
The murine epidermis is a heterogeneous epithelium composed of keratinocytes, melanocytes, Langerhans cells, and a recently described subpopulation (2-3%) of bone-marrow-derived leukocytes with a dendritic morphology and the cell surface phenotype Thy-1+, L3T4-, Lyt-2-. Previous studies have demonstrated that cell lines derived from freshly explanted Thy-1+ dendritic epidermal cells (DEC) have abundant mRNA for rearranged T-cell receptor (TCR) gamma-chain genes. Analysis of Thy-1+ DEC in situ, freshly isolated cell suspensions of Thy-1+ DEC, and long-term Thy-1+ DEC lines demonstrated that 100% of the Thy-1+ DEC reacted with a monoclonal antibody to the epsilon chain of the murine T3 complex and that 40-60% of resident Thy-1+ DEC were also reactive with an antiserum to the TCR gamma chain. Two Thy-1+ DEC lines expressed a disulfide-linked 70-kDa molecule that could be precipitated with an anti-gamma-chain antiserum and could be coprecipitated with an antiserum to the T3 delta chain; the molecule appeared as a single 34-kDa band under reducing conditions. The phenotype of Thy-1+ DEC (T3+, L3T4-, Lyt-2-, TCR gamma chain+) thus resembles that of the recently described subpopulation of murine and human lymphocytes that have been identified in the thymus, peripheral blood, and fetal blood. Images PMID:2885839
Antiadopogenic effects of rice hull smoke extract in 3T3-L1 cells
USDA-ARS?s Scientific Manuscript database
The present study investigates the inhibitory effects of a rice hull smoke extract (RHSE) against adipogenesis in 3T3-L1 pre-adipocyte cells. At concentrations of 0.1% and 0.5% RHSE, MDI-induced cells were shown to reduce their cellular lipid content by about 72% and 88%, respectively, compared to ...
Zeis, M; Uharek, L; Hartung, G; Glass, B; Steinmann, J; Schmitz, N
2001-01-01
The transfer of allogeneic lymphocytes contained in a hematopoietic stem cell graft confers an immune-mediated antileukemic effect, termed the graft-vs-leukemia (GVL) effect. Graft-vs-host disease (GVHD), the most detrimental complication of allogeneic BMT, largely resides within the same lymphocyte population. Therefore, separation of GVL- and GVH-reactions is a long-standing goal of experimental studies dealing with allogeneic transplantation of hematopoietic stem cells. The objective of the current study was to assess the potential of Th1- and Th2-type CD4+ T cells in mediating GVHD and GVL effects in a fully allogeneic murine transplant model. BALB/c (H-2d) mice were given a dose of A20 (H-2d, B-cell leukemia) cells two days prior to lethal total body irradiation (TBI) and transplantation of fully mismatched (C57BL/6, H-2b) T-cell depleted (anti-Thy1.2, CD90) bone marrow (TCD-BM) cells. Graded numbers of either unmanipulated, Th1- or Th2-polarized highly enriched CD4+ donor type T cells (10(6) or 10(7)) were administered 2 h posttransplant. Infusion of 10(6) of unmanipulated, Th1-, or Th2-primed CD4+ T cells resulted in moderate GVHD-related mortality (40%, 50%, 10%) and significantly improved long-term survival (50%, 45%, 46% surviving the observation period of 120 days) as compared to animals receiving TCD-BM alone (18%). The administration of 10(7) unmanipulated or Th1-type CD4+ T cells given shortly after transplantation led to death of all mice within 50 days due to fatal acute GVHD. In contrast, the adoptive transfer of 10(7) Th2-primed CD4+ T cells resulted in significant improvement of long-term survival (80%) compared to the TCD-BM group. This powerful GVL effect was associated with a substantially lower incidence of lethal acute GVHD (10%) if compared to the results of transplantation of Th1-type CD4+ T cells. These results demonstrate that allogeneic Th2-type CD4+ T cells given post BMT can induce GVL effects in a cell-dose-dependent manner without
Ma, Xuefei; Lin, Wenjian; Lin, Zhenyu; Hao, Ming; Gao, Xinyuan; Zhang, Yue; Kuang, Hongyu
2017-06-01
Retinal ganglion cells (RGCs), which exist in the inner retina, are the retinal neurons which can be damaged in the early stage of diabetic retinopathy (DR). Liraglutide, a glucagon-like peptide-1 (GLP-1) analog, exerts biological functions by binding the receptor (GLP-1R), the expression of which in RGC-5 cells was first shown by our team in 2012. It was reported that liraglutide prevented retinal neurodegeneration in diabetic subjects. However, the involvement of mechanisms such as autophagy and mitochondrial balance in liraglutide-induced retinal protection is unknown. Here, we aimed to investigate the protective effects of liraglutide and explore the potential mechanisms of liraglutide-induced retinal RGC protection. RGC-5 cells were treated with H 2 O 2 and/or liraglutide. Cell viability was detected with the CCK-8 kit. The axon marker GAP43, autophagy and mitophagy indicators LC3A/B, Beclin-1, p62, Parkin, BCL2/Adenovirus E1B 19kDa protein-interacting protein 3-like (BNIP3L) and the key regulator of mitochondrial biogenesis PGC-1α were examined via western blot analysis. Autophagy was also evaluated using the ImageXpress Micro XLS system and transmission electron microscopy (TEM). Reactive oxygen species (ROS), mitochondrial membrane potential and fluorescent staining for mitochondria were also measured using the ImageXpress Micro XLS system. Our results showed that pretreatment with liraglutide significantly prevented H 2 O 2 -induced cell viability decline, mitochondrial morphological deterioration and induction of autophagy, which appeared as increased expression of LC3 II/I and Beclin-1, along with p62 degradation. Moreover, liraglutide suppressed the H 2 O 2 -induced decline in GAP43 expression, thus protecting cells. However, rapamycin induced autophagy and blocked the protective process. Liraglutide also provided mitochondrial protection and appeared to alleviate H 2 O 2 -induced ROS overproduction and a decline in mitochondrial membrane potential
Luo, Jixian; Li, Dingyun; Wei, Dan; Wang, Xiaoguang; Wang, Lan; Zeng, Xianlu
2017-12-01
Stromal cell-derived factor-1 (SDF-1) signaling is important to the maintenance and progression of T-cell acute lymphoblastic leukemia by inducing chemotaxis migration. To identify the mechanism of SDF-1 signaling in the migration of T-ALL, Jurkat acute lymphoblastic leukemia cells were used. Results showed that SDF-1 induces Jurkat cell migration by F-actin redistribution and assembly, which is dependent on Rho activity. SDF-1 induced RhoA and RhoC activation, as well as reactive oxygen species (ROS) production, which was inhibited by Rho inhibitor. The Rho-dependent ROS production led to subsequent cytoskeleton redistribution and assembly in the process of migration. Additionally, RhoA and RhoC were involved in SDF-1-induced Jurkat cell migration. Taken together, we found a SDF-1/CXCR4-RhoA and RhoC-ROS-cytoskeleton pathway that regulates Jurkat cell migration in response to SDF-1. This work will contribute to a clearer insight into the migration mechanism of acute lymphoblastic leukemia.
Zhou, Wei; Koh, Young-Bok; Yang, Jung-Bo; Yoo, Heon-Jong; Lee, Min-A; Ryu, Jae-Sook; Noh, Heung-Tae; Kwon, Jaeyul; Lee, Young-Ha
2014-01-01
To elucidate the roles of metalloproteinases and the Bcl-2 family of proteins in Trichovaginalis. vaginalis-induced apoptosis in human cervical cancer cells (SiHa cells) and vaginal epithelial cells (MS74 cells), SiHa cells and MS74 cells were incubated with live T. vaginalis, T. vaginalis excretory and secretory products (ESP), and T. vaginalis lysates, either with or without the specific metalloproteinase inhibitor 1,10-phenanthroline (1,10-PT), and examined apoptotic events and Bcl-2 signaling. The live T. vaginalis and the T. vaginalis ESP induced the release of cytochrome c into the cytosol, the activation of caspase-3 and caspase-9, and the cleavage of PARP. Additionally, the live T. vaginalis, but not the T. vaginalis lysate, induced the cleavage of the proapoptotic Bim protein. The live T. vaginalis and the T. vaginalis ESP, but not the T. vaginalis lysate, induced the dose-dependent cleavage of the antiapoptotic Bcl-xL and Mcl-1 proteins and decreased the association levels of Bcl-xL/Bim and Mcl-1/Bim complexes. We performed gelatin zymography and casein-hydrolysis assays on the live T. vaginalis and the T. vaginalis ESP to identify the apoptosis-inducing factor. Both the live T. vaginalis and the ESP contained high levels of metalloproteinases, of which activities were significantly inhibited by 1,10-PT treatment. Furthermore, the 1,10-PT blocked the cleavage of Bcl-xL, Mcl-1, PARP, caspase-3, and caspase-9, as well as the release of cytochrome c into the cytosol, and it significantly increased the association levels of the Bcl-xL/Bim and Mcl-1/Bim protein complexes, returning them to normal levels. Our results demonstrate that T. vaginalis induces mitochondria-dependent apoptosis in SiHa cells through the dissociation of Bcl-xL/Bim and Mcl-1/Bim complexes and that the apoptosis is blocked by the metalloproteinase inhibitor 1,10-PT. These results expand our understanding of the role of metalloproteinases in T. vaginalis-induced apoptosis and the signaling
Innovative T Cell-Targeted Therapy for Ovarian Cancer
2012-10-01
from co-culture with EL4 -ROR1neg and EL4 -ROR1+ tumor targets. Ovarian cancer cell lines (A2780, EFO21, EFO27, IGROV1, OC314, and UPN251) were...profiled for ROR1 expression in normoxia (20% O2) and hypoxia (1% O2). Four-hour CRA was used to evaluate cytotoxicity against the OvCa and EL4 tumor...loaded aAPC for negative controls. EL4 is a murine T cell lymphoma cell line used to test specificity of CAR+ T cells with limited allo-reactivity
Role of gamma-delta T cells in host response against Staphylococcus aureus-induced pneumonia
2012-01-01
Background Staphylococcus aureus is the major cause of hospital-acquired and community-acquired pneumonia. Host defense to S.aureus infection is largely mediated by the innate immune system. γδ T cells play an important role in innate immunity to many infectious diseases. However, less is known about the role of these cells during S.aureus-induced pneumonia. In this study, we examined the response and the role of γδ T cells to pulmonary S.aureus infection. Results Mice infected with S. aureus intranasally showed rapid γδ T cells accumulation in the lung. Deficiency of γδ T cells led to attenuated bacterial clearance and less tissue damage in lung compared with WT mice. Moreover, TCR-δ−/− mice exhibited impaired neutrophil recruitment and reduced cytokine production at the site of infection. The γδ T cells in response to pulmonary S. aureus infection mainly secreted IL-17 and γδ T cells deficiency reduced IL-17 production, which might regulate the production of neutrophil-inducing cytokine/chemokine in the S. aureus-infected lungs. Conclusions Accumulation of γδ T cells in the lungs to S. aureus infection is beneficial for bacteria clearance and also contributes to the tissue damage. These cells were the primary source of IL-17, which might influence the recruitment of neutrophils at the early stage of infection. PMID:22776294
Positive MRI contrast enhancement in THP-1 cells with Gd2O3 nanoparticles.
Klasson, Anna; Ahrén, Maria; Hellqvist, Eva; Söderlind, Fredrik; Rosén, Anders; Käll, Per-Olov; Uvdal, Kajsa; Engström, Maria
2008-01-01
There is a demand for more efficient and tissue-specific MRI contrast agents and recent developments involve the design of substances useful as molecular markers and magnetic tracers. In this study, nanoparticles of gadolinium oxide (Gd2O3) have been investigated for cell labeling and capacity to generate a positive contrast. THP-1, a monocytic cell line that is phagocytic, was used and results were compared with relaxivity of particles in cell culture medium (RPMI 1640). The results showed that Gd2O3-labeled cells have shorter T1 and T2 relaxation times compared with untreated cells. A prominent difference in signal intensity was observed, indicating that Gd2O3 nanoparticles can be used as a positive contrast agent for cell labeling. The r1 for cell samples was 4.1 and 3.6 s(-1) mm(-1) for cell culture medium. The r2 was 17.4 and 12.9 s(-1) mm(-1), respectively. For r1, there was no significant difference in relaxivity between particles in cells compared to particles in cell culture medium, (p(r1) = 0.36), but r2 was significantly different for the two different series (p(r2) = 0.02). Viability results indicate that THP-1 cells endure treatment with Gd2O3 nanoparticles for an extended period of time and it is therefore concluded that results in this study are based on viable cells. Copyright 2008 John Wiley & Sons, Ltd.
Lee, Yueh-Lun; Lee, Lin-Wen; Su, Chen-Yao; Hsiao, George; Yang, Yi-Yuan; Leu, Sy-Jye; Shieh, Ying-Hua; Burnouf, Thierry
2013-09-01
Platelet concentrate lysates (PCLs) are increasingly used in regenerative medicine. We have developed a solvent/detergent (S/D)-treated PCL. The functional properties of this preparation should be unveiled. We hypothesized that, due to transforming growth factor-β1 (TGF-β1) content, PCLs may exert immunosuppressive and anti-inflammatory functions. PCL was prepared by S/D treatment, oil extraction, and hydrophobic interaction chromatography. The content of TGF-β in PCL was determined by enzyme-linked immunosorbent assay. Cultured CD4+ T cells were used to investigate the effects of PCL on expression of transcription factor forkhead box P3 (Foxp3), the inhibition of T-cell proliferation, and cytokine production. The regulatory function of PCL-converted CD4+ T cells was analyzed by suppressive assay. The BALB/c mice were given PCL-converted CD4+ T cells before ovalbumin (OVA) sensitization and challenge using an asthma model. Inflammatory parameters, such as the level of immunoglobulin E (IgE), airway hyperresponsiveness (AHR), bronchial lavage fluid eosinophils, and cytokines were assayed. Recombinant human (rHu) TGF-β1 was used as control. PCL significantly enhanced the development of CD4+Foxp3+-induced regulatory T cells (iTregs). Converted iTregs produced neither Th1 nor Th2 cytokines and inhibited normal T-cell proliferation. PCL- and rHuTGF-β-converted CD4+ T cells prevented OVA-induced asthma. PCL- and rHuTGF-β-modified T cells both significantly reduced expression levels of OVA-specific IgE and significantly inhibited the development of AHR, airway eosinophilia, and Th2 responses in mice. S/D-treated PCL promotes Foxp3+ iTregs and exerts immunosuppressive and anti-inflammatory properties. This finding may help to understand the clinical properties of platelet lysates. © 2013 American Association of Blood Banks.
Schuster, Cornelia; Brosi, Helen; Stifter, Katja; Boehm, Bernhard O.; Schirmbeck, Reinhold
2013-01-01
Coinhibitory PD-1/PD-L1 (B7-H1) interactions provide critical signals for the regulation of autoreactive T-cell responses. We established mouse models, expressing the costimulator molecule B7.1 (CD80) on pancreatic beta cells (RIP-B7.1 tg mice) or are deficient in coinhibitory PD-L1 or PD-1 molecules (PD-L1−/− and PD-1−/− mice), to study induction of preproinsulin (ppins)-specific CD8 T-cell responses and experimental autoimmune diabetes (EAD) by DNA-based immunization. RIP-B7.1 tg mice allowed us to identify two CD8 T-cell specificities: pCI/ppins DNA exclusively induced Kb/A12–21-specific CD8 T-cells and EAD, whereas pCI/ppinsΔA12–21 DNA (encoding ppins without the COOH-terminal A12–21 epitope) elicited Kb/B22–29-specific CD8 T-cells and EAD. Specific expression/processing of mutant ppinsΔA12–21 (but not ppins) in non-beta cells, targeted by intramuscular DNA-injection, thus facilitated induction of Kb/B22–29-specific CD8 T-cells. The A12–21 epitope binds Kb molecules with a very low avidity as compared with B22–29. Interestingly, immunization of coinhibition-deficient PD-L1−/− or PD-1−/− mice with pCI/ppins induced Kb/A12–21-monospecific CD8 T-cells and EAD but injections with pCI/ppinsΔA12–21 did neither recruit Kb/B22–29-specific CD8 T-cells into the pancreatic target tissue nor induce EAD. PpinsΔA12–21/(Kb/B22–29)-mediated EAD was efficiently restored in RIP-B7.1+/PD-L1−/− mice, differing from PD-L1−/− mice only in the tg B7.1 expression in beta cells. Alternatively, an ongoing beta cell destruction and tissue inflammation, initiated by ppins/(Kb/A12–21)-specific CD8 T-cells in pCI/ppins+pCI/ppinsΔA12–21 co-immunized PD-L1−/− mice, facilitated the expansion of ppinsΔA12–21/(Kb/B22–29)-specific CD8 T-cells. CD8 T-cells specific for the high-affinity Kb/B22–29- (but not the low-affinity Kb/A12–21)-epitope thus require stimulatory ´help from beta cells or inflamed islets to expand in PD-L1
Hu, Lifang; Su, Peihong; Li, Runzhi; Yan, Kun; Chen, Zhihao; Shang, Peng; Qian, Airong
2015-01-01
Microtubule actin crosslinking factor 1 (MACF1), a widely expressed cytoskeletal linker, plays important roles in various cells by regulating cytoskeleton dynamics. However, its role in osteoblastic cells is not well understood. Based on our previous findings that the association of MACF1 with F-actin and microtubules in osteoblast-like cells was altered under magnetic force conditions, here, by adopting a stable MACF1-knockdown MC3T3-E1 osteoblastic cell line, we found that MACF1 knockdown induced large cells with a binuclear/multinuclear structure. Further, immunofluorescence staining showed disorganization of F-actin and microtubules in MACF1-knockdown cells. Cell counting revealed significant decrease of cell proliferation and cell cycle analysis showed an S phase cell cycle arrest in MACF1-knockdown cells. Moreover and interestingly, MACF1 knockdown showed a potential effect on cellular MTT reduction activity and mitochondrial content, suggesting an impact on cellular metabolic activity. These results together indicate an important role of MACF1 in regulating osteoblastic cell morphology and function. [BMB Reports 2015; 48(10): 583-588] PMID:26277981
Hu, Lifang; Su, Peihong; Li, Runzhi; Yan, Kun; Chen, Zhihao; Shang, Peng; Qian, Airong
2015-10-01
Microtubule actin crosslinking factor 1 (MACF1), a widely expressed cytoskeletal linker, plays important roles in various cells by regulating cytoskeleton dynamics. However, its role in osteoblastic cells is not well understood. Based on our previous findings that the association of MACF1 with F-actin and microtubules in osteoblast-like cells was altered under magnetic force conditions, here, by adopting a stable MACF1-knockdown MC3T3-E1 osteoblastic cell line, we found that MACF1 knockdown induced large cells with a binuclear/multinuclear structure. Further, immunofluorescence staining showed disorganization of F-actin and microtubules in MACF1-knockdown cells. Cell counting revealed significant decrease of cell proliferation and cell cycle analysis showed an S phase cell cycle arrest in MACF1-knockdown cells. Moreover and interestingly, MACF1 knockdown showed a potential effect on cellular MTT reduction activity and mitochondrial content, suggesting an impact on cellular metabolic activity. These results together indicate an important role of MACF1 in regulating osteoblastic cell morphology and function.
Lactisole inhibits the glucose-sensing receptor T1R3 expressed in mouse pancreatic β-cells.
Hamano, Kunihisa; Nakagawa, Yuko; Ohtsu, Yoshiaki; Li, Longfei; Medina, Johan; Tanaka, Yuji; Masuda, Katsuyoshi; Komatsu, Mitsuhisa; Kojima, Itaru
2015-07-01
Glucose activates the glucose-sensing receptor T1R3 and facilitates its own metabolism in pancreatic β-cells. An inhibitor of this receptor would be helpful in elucidating the physiological function of the glucose-sensing receptor. The present study was conducted to examine whether or not lactisole can be used as an inhibitor of the glucose-sensing receptor. In MIN6 cells, in a dose-dependent manner, lactisole inhibited insulin secretion induced by sweeteners, acesulfame-K, sucralose and glycyrrhizin. The IC50 was ∼4 mmol/l. Lactisole attenuated the elevation of cytoplasmic Ca2+ concentration ([Ca2+]c) evoked by sucralose and acesulfame-K but did not affect the elevation of intracellular cAMP concentration ([cAMP]c) induced by these sweeteners. Lactisole also inhibited the action of glucose in MIN6 cells. Thus, lactisole significantly reduced elevations of intracellular [NADH] and intracellular [ATP] induced by glucose, and also inhibited glucose-induced insulin secretion. To further examine the effect of lactisole on T1R3, we prepared HEK293 cells stably expressing mouse T1R3. In these cells, sucralose elevated both [Ca2+]c and [cAMP]c. Lactisole attenuated the sucralose-induced increase in [Ca2+]c but did not affect the elevation of [cAMP]c. Finally, lactisole inhibited insulin secretion induced by a high concentration of glucose in mouse islets. These results indicate that the mouse glucose-sensing receptor was inhibited by lactisole. Lactisole may be useful in assessing the role of the glucose-sensing receptor in mouse pancreatic β-cells. © 2015 Society for Endocrinology.
Hertz, Tomer; Ahmed, Hasan; Friedrich, David P; Casimiro, Danilo R; Self, Steven G; Corey, Lawrence; McElrath, M Juliana; Buchbinder, Susan; Horton, Helen; Frahm, Nicole; Robertson, Michael N; Graham, Barney S; Gilbert, Peter
2013-01-01
Several recent large clinical trials evaluated HIV vaccine candidates that were based on recombinant adenovirus serotype 5 (rAd-5) vectors expressing HIV-derived antigens. These vaccines primarily elicited T-cell responses, which are known to be critical for controlling HIV infection. In the current study, we present a meta-analysis of epitope mapping data from 177 participants in three clinical trials that tested two different HIV vaccines: MRKAd-5 HIV and VRC-HIVAD014-00VP. We characterized the population-level epitope responses in these trials by generating population-based epitope maps, and also designed such maps using a large cohort of 372 naturally infected individuals. We used these maps to address several questions: (1) Are vaccine-induced responses randomly distributed across vaccine inserts, or do they cluster into immunodominant epitope hotspots? (2) Are the immunodominance patterns observed for these two vaccines in three vaccine trials different from one another? (3) Do vaccine-induced hotspots overlap with epitope hotspots induced by chronic natural infection with HIV-1? (4) Do immunodominant hotspots target evolutionarily conserved regions of the HIV genome? (5) Can epitope prediction methods be used to identify these hotspots? We found that vaccine responses clustered into epitope hotspots in all three vaccine trials and some of these hotspots were not observed in chronic natural infection. We also found significant differences between the immunodominance patterns generated in each trial, even comparing two trials that tested the same vaccine in different populations. Some of the vaccine-induced immunodominant hotspots were located in highly variable regions of the HIV genome, and this was more evident for the MRKAd-5 HIV vaccine. Finally, we found that epitope prediction methods can partially predict the location of vaccine-induced epitope hotspots. Our findings have implications for vaccine design and suggest a framework by which different
SOCS1 and SOCS3 Are Targeted by Hepatitis C Virus Core/gC1qR Ligation To Inhibit T-Cell Function
Yao, Zhi Qiang; Waggoner, Stephen N.; Cruise, Michael W.; Hall, Caroline; Xie, Xuefang; Oldach, David W.; Hahn, Young S.
2005-01-01
T cells play an important role in the control of hepatitis C virus (HCV) infection. We have previously demonstrated that the HCV core inhibits T-cell responses through interaction with gC1qR. We show here that core proteins from chronic and resolved HCV patients differ in sequence, gC1qR-binding ability, and T-cell inhibition. Specifically, chronic core isolates bind to gC1qR more efficiently and inhibit T-cell proliferation as well as gamma interferon (IFN-γ) production more profoundly than resolved core isolates. This inhibition is mediated by the disruption of STAT phosphorylation through the induction of SOCS molecules. Silencing either SOCS1 or SOCS3 by small interfering RNA dramatically augments the production of IFN-γ in T cells, thereby abrogating the inhibitory effect of core. Additionally, the ability of core proteins from patients with chronic infections to induce SOCS proteins and suppress STAT activation greatly exceeds that of core proteins from patients with resolved infections. These results suggest that the HCV core/gC1qR-induced T-cell dysfunction involves the induction of SOCS, a powerful inhibitor of cytokine signaling, which represents a novel mechanism by which a virus usurps the host machinery for persistence. PMID:16306613
Liu, Danya; Burd, Eileen M.; Coopersmith, Craig M.; Ford, Mandy L.
2016-01-01
Following T cell encounter with antigen, multiple signals are integrated to collectively induce distinct differentiation programs within antigen-specific CD8+ T cell populations. Several factors contribute to these cell fate decisions including the amount and duration of antigen, exposure to inflammatory cytokines, and degree of ligation of cosignaling molecules. The inducible costimulator (ICOS) is not expressed on resting T cells but is rapidly upregulated upon encounter with antigen. However, the impact of ICOS signaling on programmed differentiation is not well understood. In this study we therefore sought to determine the role of ICOS signaling on CD8+ T cell programmed differentiation. Through the creation of novel ICOS retrogenic antigen-specific TCR transgenic CD8+ T cells, we interrogated the phenotype, functionality, and recall potential of CD8+ T cells that receive early and sustained ICOS signaling during antigen exposure. Our results reveal that these ICOS signals critically impacted cell fate decisions of antigen-specific CD8+ T cells, resulting in increased frequencies of KLRG-1hiCD127lo cells, altered BLIMP-1, T-bet, and eomesodermin expression, and increased cytolytic capacity as compared to empty vector controls. Interestingly, however, ICOS retrogenic CD8+ T cells also preferentially homed to non-lymphoid organs, and exhibited reduced multi-cytokine functionality and reduced ability to mount secondary recall responses upon challenge in vivo. In sum, our results suggest that an altered differentiation program is induced following early and sustained ICOS expression, resulting in the generation of more cytolyticly potent, terminally differentiated effectors that possess limited capacity for recall response. PMID:26729800
Structural Elements Recognized by Abacavir-Induced T Cells.
Yerly, Daniel; Pompeu, Yuri Andreiw; Schutte, Ryan J; Eriksson, Klara K; Strhyn, Anette; Bracey, Austin W; Buus, Soren; Ostrov, David A
2017-07-07
Adverse drug reactions are one of the leading causes of morbidity and mortality in health care worldwide. Human leukocyte antigen (HLA) alleles have been strongly associated with drug hypersensitivities, and the causative drugs have been shown to stimulate specific T cells at the sites of autoimmune destruction. The structural elements recognized by drug-specific T cell receptors (TCRs) in vivo are poorly defined. Drug-stimulated T cells express TCRs specific for peptide/HLA complexes, but the characteristics of peptides (sequence, or endogenous or exogenous origin) presented in the context of small molecule drugs are not well studied. Using HLA-B*57:01 mediated hypersensitivity to abacavir as a model system, this study examines structural similarities of HLA presented peptides recognized by drug-specific TCRs. Using the crystal structure of HLA-B*57:01 complexed with abacavir and an immunogenic self peptide, VTTDIQVKV SPT5a 976-984, peptide side chains exhibiting flexibility and solvent exposure were identified as potential drug-specific T cell recognition motifs. Viral sequences with structural motifs similar to the immunogenic self peptide were identified. Abacavir-specific T cell clones were used to determine if virus peptides presented in the context of abacavir stimulate T cell responsiveness. An abacavir-specific T cell clone was stimulated by VTQQAQVRL, corresponding to HSV1/2 230-238, in the context of HLA-B*57:01. These data suggest the T cell polyclonal response to abacavir consists of multiple subsets, including T cells that recognize self peptide/HLA-B*57:01 complexes and crossreact with viral peptide/HLA-B*57:01 complexes due to similarity in TCR contact residues.
Structural Elements Recognized by Abacavir-Induced T Cells
Yerly, Daniel; Pompeu, Yuri Andreiw; Schutte, Ryan J.; Eriksson, Klara. K.; Strhyn, Anette; Bracey, Austin. W.; Buus, Soren; Ostrov, David A.
2017-01-01
Adverse drug reactions are one of the leading causes of morbidity and mortality in health care worldwide. Human leukocyte antigen (HLA) alleles have been strongly associated with drug hypersensitivities, and the causative drugs have been shown to stimulate specific T cells at the sites of autoimmune destruction. The structural elements recognized by drug-specific T cell receptors (TCRs) in vivo are poorly defined. Drug-stimulated T cells express TCRs specific for peptide/HLA complexes, but the characteristics of peptides (sequence, or endogenous or exogenous origin) presented in the context of small molecule drugs are not well studied. Using HLA-B*57:01 mediated hypersensitivity to abacavir as a model system, this study examines structural similarities of HLA presented peptides recognized by drug-specific TCRs. Using the crystal structure of HLA-B*57:01 complexed with abacavir and an immunogenic self peptide, VTTDIQVKV SPT5a 976–984, peptide side chains exhibiting flexibility and solvent exposure were identified as potential drug-specific T cell recognition motifs. Viral sequences with structural motifs similar to the immunogenic self peptide were identified. Abacavir-specific T cell clones were used to determine if virus peptides presented in the context of abacavir stimulate T cell responsiveness. An abacavir-specific T cell clone was stimulated by VTQQAQVRL, corresponding to HSV1/2 230–238, in the context of HLA-B*57:01. These data suggest the T cell polyclonal response to abacavir consists of multiple subsets, including T cells that recognize self peptide/HLA-B*57:01 complexes and crossreact with viral peptide/HLA-B*57:01 complexes due to similarity in TCR contact residues. PMID:28686208
Chiozzini, Chiara; Arenaccio, Claudia; Olivetta, Eleonora; Anticoli, Simona; Manfredi, Francesco; Ferrantelli, Flavia; d'Ettorre, Gabriella; Schietroma, Ivan; Andreotti, Mauro; Federico, Maurizio
2017-09-01
Intact HIV-1 and exosomes can be internalized by dendritic cells (DCs) through a common pathway leading to their transmission to CD4 + T lymphocytes by means of mechanisms defined as trans-infection and trans-dissemination, respectively. We previously reported that exosomes from HIV-1-infected cells activate both uninfected quiescent CD4 + T lymphocytes, which become permissive to HIV-1, and latently infected cells, with release of HIV-1 particles. However, nothing is known about the effects of trans-dissemination of exosomes produced by HIV-1-infected cells on uninfected or latently HIV-1-infected CD4 + T lymphocytes. Here, we report that trans-dissemination of exosomes from HIV-1-infected cells induces cell activation in resting CD4 + T lymphocytes, which appears stronger with mature than immature DCs. Using purified preparations of both HIV-1 and exosomes, we observed that mDC-mediated trans-dissemination of exosomes from HIV-1-infected cells to resting CD4 + T lymphocytes induces efficient trans-infection and HIV-1 expression in target cells. Most relevant, when both mDCs and CD4 + T lymphocytes were isolated from combination anti-retroviral therapy (ART)-treated HIV-1-infected patients, trans-dissemination of exosomes from HIV-1-infected cells led to HIV-1 reactivation from the viral reservoir. In sum, our data suggest a role of exosome trans-dissemination in both HIV-1 spread in the infected host and reactivation of the HIV-1 reservoir.
Takemiya, Takako; Takeuchi, Chisen; Kawakami, Marumi
2017-12-19
Microsomal prostaglandin synthetase-1 (mPGES-1) is an inducible terminal enzyme that produces prostaglandin E₂ (PGE₂). In our previous study, we investigated the role of mPGES-1 in the inflammation and demyelination observed in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, using mPGES - 1 -deficient ( mPGES-1 -/- ) and wild-type (wt) mice. We found that mPGES-1 facilitated inflammation, demyelination, and paralysis and was induced in vascular endothelial cells and macrophages and microglia around inflammatory foci. Here, we investigated the role of interleukin-1β (IL-1β) in the intercellular mechanism stimulated by mPGES-1 in EAE spinal cords in the presence of inflammation. We found that the area invaded by CD4-positive (CD4⁺) T cells was extensive, and that PGE₂ receptors EP1-4 were more induced in activated CD4⁺ T cells of wt mice than in those of mPGES - 1 -/- mice. Moreover, IL-1β and IL-1 receptor 1 (IL-1r1) were produced by 65% and 48% of CD4⁺ T cells in wt mice and by 44% and 27% of CD4⁺ T cells in mPGES-1 -/- mice. Furthermore, interleukin-17 (IL-17) was released from the activated CD4⁺ T cells. Therefore, mPGES-1 stimulates an intercellular interaction between CD4⁺ T cells by upregulating the autocrine function of IL-1β in activated CD4⁺ T cells, which release IL-17 to facilitate axonal and myelin damage in EAE mice.
Dudani, Renu; Murali-Krishna, Kaja; Krishnan, Lakshmi; Sad, Subash
2014-01-01
Memory T cells are critical for the control of intracellular pathogens and require few signals for maintenance; however, erosion of established preexisting memory CD8+ T cells has been shown to occur during infection with heterologous viral infections. We evaluated whether this also occurs during infection with various intracellular bacteria and what mechanisms may be involved. We demonstrate that erosion of established memory is also induced during infection of mice with various intracellular bacteria, such as Listeria monocytogenes, Salmonella typhimurium, and Mycobacterium bovis (bacillus Calmette-Guérin). The extent of erosion of established CD8+ T cell memory was dependent on the virulence of the heterologous pathogen, not persistence. Furthermore, when antibiotics were used to comprehensively eliminate the heterologous pathogen, the numbers of memory CD8+ T cells were not restored, indicating that erosion of preexisting memory CD8+ T cells was irreversible. Irrespective of the initial numbers of memory CD8+ T cells, challenge with the heterologous pathogen resulted in a similar extent of erosion of memory CD8+ T cells, suggesting that cellular competition was not responsible for erosion. After challenge with the heterologous pathogen, effector memory CD8+ T cells were rapidly eliminated. More importantly, erosion of preexisting memory CD8+ T cells was abrogated in the absence of IFN-γ. These studies help reveal the paradoxical role of IFN-γ. Although IFN-γ promotes the control of intracellular bacterial replication during primary infection, this comes at the expense of erosion of preexisting memory CD8+ T cells in the wake of infection with heterologous pathogens. PMID:18641306
Zhou, Ji-Hao; Yao, Yu-Shi; Li, Yong-Hui; Xu, Yi-Han; Li, Jing-Xin; Gao, Xiao-Ning; Zhou, Min-Hang; Jiang, Meng-Meng; Gao, Li; Ding, Yi; Lu, Xue-Chun; Shi, Jin-Long; Luo, Xu-Feng; Wang, Jia; Wang, Li-Li; Qu, Chunfeng; Bai, Xue-Feng; Yu, Li
2013-01-01
Lack of immunogenicity of cancer cells has been considered a major reason for their failure in induction of a tumor specific T cell response. In this paper, we present evidence that decitabine (DAC), a DNA methylation inhibitor that is currently used for the treatment of myelodysplastic syndrome (MDS), acute myeloid leukemia (AML) and other malignant neoplasms, is capable of eliciting an anti-tumor cytotoxic T lymphocyte (CTL) response in mouse EL4 tumor model. C57BL/6 mice with established EL4 tumors were treated with DAC (1.0 mg/kg body weight) once daily for 5 days. We found that DAC treatment resulted in infiltration of IFN-γ producing T lymphocytes into tumors and caused tumor rejection. Depletion of CD8+, but not CD4+ T cells resumed tumor growth. DAC-induced CTL response appeared to be elicited by the induction of CD80 expression on tumor cells. Epigenetic evidence suggests that DAC induces CD80 expression in EL4 cells via demethylation of CpG dinucleotide sites in the promoter of CD80 gene. In addition, we also showed that a transient, low-dose DAC treatment can induce CD80 gene expression in a variety of human cancer cells. This study provides the first evidence that epigenetic modulation can induce the expression of a major T cell co-stimulatory molecule on cancer cells, which can overcome immune tolerance, and induce an efficient anti-tumor CTL response. The results have important implications in designing DAC-based cancer immunotherapy. PMID:23671644
Wang, Li-Xin; Mei, Zhen-Yang; Zhou, Ji-Hao; Yao, Yu-Shi; Li, Yong-Hui; Xu, Yi-Han; Li, Jing-Xin; Gao, Xiao-Ning; Zhou, Min-Hang; Jiang, Meng-Meng; Gao, Li; Ding, Yi; Lu, Xue-Chun; Shi, Jin-Long; Luo, Xu-Feng; Wang, Jia; Wang, Li-Li; Qu, Chunfeng; Bai, Xue-Feng; Yu, Li
2013-01-01
Lack of immunogenicity of cancer cells has been considered a major reason for their failure in induction of a tumor specific T cell response. In this paper, we present evidence that decitabine (DAC), a DNA methylation inhibitor that is currently used for the treatment of myelodysplastic syndrome (MDS), acute myeloid leukemia (AML) and other malignant neoplasms, is capable of eliciting an anti-tumor cytotoxic T lymphocyte (CTL) response in mouse EL4 tumor model. C57BL/6 mice with established EL4 tumors were treated with DAC (1.0 mg/kg body weight) once daily for 5 days. We found that DAC treatment resulted in infiltration of IFN-γ producing T lymphocytes into tumors and caused tumor rejection. Depletion of CD8(+), but not CD4(+) T cells resumed tumor growth. DAC-induced CTL response appeared to be elicited by the induction of CD80 expression on tumor cells. Epigenetic evidence suggests that DAC induces CD80 expression in EL4 cells via demethylation of CpG dinucleotide sites in the promoter of CD80 gene. In addition, we also showed that a transient, low-dose DAC treatment can induce CD80 gene expression in a variety of human cancer cells. This study provides the first evidence that epigenetic modulation can induce the expression of a major T cell co-stimulatory molecule on cancer cells, which can overcome immune tolerance, and induce an efficient anti-tumor CTL response. The results have important implications in designing DAC-based cancer immunotherapy.