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21.01.2017 | Original Paper | Ausgabe 2/2017

Journal of Natural Medicines 2/2017

A novel lectin from Artocarpus lingnanensis induces proliferation and Th1/Th2 cytokine secretion through CD45 signaling pathway in human T lymphocytes

Zeitschrift:
Journal of Natural Medicines > Ausgabe 2/2017
Autoren:
Bo Cui, Lu Li, Qiyan Zeng, Faquan Lin, Lijun Yin, Liejun Liao, Min Huang, Jingping Wang
Wichtige Hinweise

Electronic supplementary material

The online version of this article (doi:10.​1007/​s11418-017-1073-x) contains supplementary material, which is available to authorized users.
B. Cui and L. Li contributed equally to this work.

Abstract

Lectins are carbohydrate-binding proteins and have been used for purification and characterization of glycoproteins. In this study, a novel 58.9-kDa tetrameric lectin from Artocarpus lingnanensis seeds was purified, characterized, and its mitogenic potential was evaluated. The hemagglutination inhibition assay indicated that Artocarpus lingnanensis lectin (ALL) showed specificity toward galactose. ALL was effectively purified in a single-step using affinity chromatography on a galactose-Sepharose column. ALL showed pH optima between 5.0 and 9.0, and optimal temperature between 20 and 40 °C. ALL triggered proliferation and activation of human T lymphocytes (e.g., CD4+ T lymphocytes). Flow cytometry and laser scanning confocal microscopy revealed binding of ALL to T cells and colocalized with CD45. Affinity chromatography and Western blot suggested that CD45 isolated from human T cell membrane fraction may be the major receptor of ALL. CD45 blocking antibody attenuated the binding and proliferation of T cells induced by ALL. CD45-PTPase inhibitor dephostatin reduced ALL-induced T cells proliferation and expression of CD25 and pZAP-70. Furthermore, secretion of ALL-induced Th1/Th2 cytokines was blocked with dephostatin. Also, dephostatin inhibited phosphorylation of ALL-mediated activation of ERK and p38MAPK. This study demonstrates the involvement of CD45-mediated signaling in ALL-induced T lymphocyte proliferation and Th1/Th2 cytokine secretion through activation of p38 and ERK.

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Zusatzmaterial
Effects of ALL on proliferation of T cells. T cells purified from healthy human PBL were stimulated with ALL at various concentrations and PHA as described above for 72 h. Data represent mean ± SD (n = 3). ∗p < 0.05, ∗∗p < 0.01, compared with untreated cells; #p < 0.05, compared with PHA treated cells. (TIFF 2158 kb)
11418_2017_1073_MOESM1_ESM.tif
ALL induce proliferation of CD4+ T lymphocytes. T cells were cultured for 24 h in the presence of lectins at indicated concentrations. Expression of CD4, CD8, CD8/CD4 in CD3+cells was analyzed by flow cytometry after labeling with the corresponding mAb. Data represent means + SD at least three experiments. (TIFF 1941 kb)
11418_2017_1073_MOESM2_ESM.tif
Effect of ALL on CD25 expression in T cells. T cells were incubated in ALL indicated concentrations for 24 h. Expression of CD25 in CD3+ T lymphocytes was analyzed by flow cytometry after labeling with the corresponding mAb. (TIFF 2049 kb)
11418_2017_1073_MOESM3_ESM.tif
T cells were pretreated with ERK inhibitor (PD98059) or p38 inhibitor (SB202190) for 45 min at 37 °C before incubating with ALL, followed by analysis of the culture supernatants for IL-2 levels using the ELISA kit. ∗p < 0.01, compared with ALL treatment alone. Data represent means + SD at least three experiments. (TIFF 2421 kb)
11418_2017_1073_MOESM4_ESM.tif
T cells were incubated in ALL indicated concentrations in the absence or presence of ERK inhibitor (PD98059) or p38 inhibitor (SB202190) for 24 h. Expression of CD25 in CD3+ T lymphocytes was analyzed by flow cytometry after labeling with the corresponding mAb. (TIFF 2677 kb)
11418_2017_1073_MOESM5_ESM.tif
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