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Neutrophil interactions with sialyl Lewis X on human nonsmall cell lung carcinoma cells regulate invasive behavior.

ABSTRACT: The carbohydrate sialyl Lewis X (sLeX) is expressed on leukocytes and carcinoma cells and binds to selectins during inflammatory processes and early metastasis. Synthesis of sLeX depends on activity of enzymes, including ?(1,3/1,4) fucosyltransferase (FucT-III). Tumor necrosis factor-? (TNF-?) up-regulates FucT-III, resulting in increased sLeX in the airways of patients with respiratory disease; however, the mechanisms that regulate sLeX in the inflammatory tumor microenvironment are not well understood.The authors stably transfected human lung carcinoma cell lines with the FucT-III gene and exposed them to TNF-? to investigate its role in regulation of sLeX expression and selectin-binding ability using semiquantitative real-time polymerase chain reaction and flow cytometry. Cytokine expression was examined in transfected cells using chemiluminescent arrays and enzyme-linked immunosorbent assays, and invasion was studied using Matrigel assays and alterations in morphology. Human lung tissue arrays were analyzed for immunohistochemical detection of sLeX and neutrophils.Stimulation of FucT-III-transfected cells with recombinant human (rh) TNF-? up-regulated sLeX expression and increased E-selectin binding. Transfected cells secreted high levels of interleukin 8, growth-regulated oncogene-?, and mast cell proteinase-1. Cells exposed to rhTNF-?, neutrophil-conditioned media, and cultures with a 5:1 ratio of neutrophils to cancer cells had significantly increased sLeX expression and invasiveness and underwent nonadherent morphologic changes. In lung carcinomas, but not in normal lung tissues, 71% of tumors were highly positive for sLeX expression in areas of increased neutrophil infiltration.The current results indicated that neutrophils may be recruited to areas of FucT-III activity and sLeX expression in lung carcinomas to enhance the invasive and metastatic potential of lung cancer cells.

SUBMITTER: St Hill CA

PROVIDER: S-EPMC3134589 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Neutrophil interactions with sialyl Lewis X on human nonsmall cell lung carcinoma cells regulate invasive behavior.

St Hill Catherine A CA Krieser Katherine K Farooqui Mariya M

Cancer 20110322 19

<h4>Background</h4>The carbohydrate sialyl Lewis X (sLeX) is expressed on leukocytes and carcinoma cells and binds to selectins during inflammatory processes and early metastasis. Synthesis of sLeX depends on activity of enzymes, including α(1,3/1,4) fucosyltransferase (FucT-III). Tumor necrosis factor-α (TNF-α) up-regulates FucT-III, resulting in increased sLeX in the airways of patients with respiratory disease; however, the mechanisms that regulate sLeX in the inflammatory tumor microenvironm ...[more]

PMID: 21437888

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Project description:BackgroundLewis antigens such as Sialyl Lewis A (sLeA), Sialyl Lewis X (sLeX), Lewis X (LeX), and Lewis Y (LeY) are a class of carbohydrate molecules that are known to mediate adhesion between tumor cells and endothelium by interacting with its selectin ligands. However, their potential role in miscarriage remains enigmatic. This study aims to analyze the expression pattern of sLeA, sLeX, LeX, and LeY in the placental villi tissue of patients with a medical history of unexplained miscarriages.MethodsParaffin-embedded slides originating from placental tissue were collected from patients experiencing a miscarriage early in their pregnancy (6-13 weeks). Tissues collected from spontaneous (n = 20) and recurrent (n = 15) miscarriages were analyzed using immunohistochemical and immunofluorescent staining. Specimens obtained from legally terminated normal pregnancies were considered as control group (n = 18). Assessment of villous vessel density was performed in another cohort (n = 10 each group) of gestation ages-paired placenta tissue. Protein expression was evaluated with Immunoreactive Score (IRS). Statistical analysis was performed by using Graphpad Prism 8.ResultsExpression of sLeA, sLeX, LeX, and LeY in the syncytiotrophoblast was significantly upregulated in the control group compared with spontaneous and recurrent miscarriage groups. However, no prominent differences between spontaneous and recurrent miscarriage groups were identified. Potential key modulators ST3GAL6 and NEU1 were found to be significantly downregulated in the recurrent miscarriage group and upregulated in the spontaneous group, respectively. Interestingly, LeX and LeY expression was also detected in the endothelial cells of villous vessels in the control group but no significant expression in miscarriage groups. Furthermore, assessment of villous vessel density using CD31 found significantly diminished vessels in all size groups of villi (small villi <200 µm, P = 0.0371; middle villi between 200 and 400 µm, P = 0.0010 and large villi >400 µm, P = 0.0003). Immunofluorescent double staining also indicated the co-localization of LeX/Y and CD31.ConclusionsThe expression of four mentioned carbohydrate Lewis antigens and their potential modulators, ST3GAL6 and NEU1, in the placenta of patients with miscarriages was significantly different from the normal pregnancy. For the first time, their expression pattern in the placenta was illustrated, which might shed light on a novel understanding of Lewis antigens' role in the pathogenesis of miscarriages.

Project description:In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosylate terminal lactosaminyl glycans yielding Lewis-X (LeX; CD15) and/or sialyl Lewis-X (sLeX; CD15s), structures that play key functions in cell migration, development, and immunity. Prior studies analyzing α(1,3)-FT specificities utilized either purified and/or recombinant enzymes to modify synthetic substrates under nonphysiological reaction conditions or molecular biology approaches wherein α(1,3)-FTs were expressed in mammalian cell lines, notably excluding investigations using primary human cells. Accordingly, although significant insights into α(1,3)-FT catalytic properties have been obtained, uncertainty persists regarding their human LeX/sLeX biosynthetic range across various glycoconjugates. Here, we undertook a comprehensive evaluation of the lactosaminyl product specificities of intracellularly expressed α(1,3)-FTs using a clinically relevant primary human cell type, mesenchymal stem cells. Cells were transfected with modified mRNA encoding each human α(1,3)-FT, and the resultant α(1,3)-fucosylated lactosaminyl glycoconjugates were analyzed using a combination of flow cytometry and MS. The data show that biosynthesis of sLeX is driven by FTs-3, -5, -6, and -7, with FT6 and FT7 having highest potency. FT4 and FT9 dominantly biosynthesize LeX, and, among all FTs, FT6 holds a unique capacity in creating sLeX and LeX determinants across protein and lipid glycoconjugates. Surprisingly, FT4 does not generate sLeX on glycolipids, and neither FT4, FT6, nor FT9 synthesizes the internally fucosylated sialyllactosamine VIM-2 (CD65s). These results unveil the relevant human lactosaminyl glycans created by human α(1,3)-FTs, providing novel insights on how these isoenzymes stereoselectively shape biosynthesis of vital glycoconjugates, thereby biochemically programming human cell migration and tuning human immunologic and developmental processes.

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Neutrophil interactions with sialyl Lewis X on human nonsmall cell lung carcinoma cells regulate invasive behavior.

Publications

Neutrophil interactions with sialyl Lewis X on human nonsmall cell lung carcinoma cells regulate invasive behavior.

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