XCL1 Aggravates Diabetic Nephropathy-Mediated Renal Glomerular Endothelial Cell Apoptosis and Inflammatory Response via Regulating p53/Nuclear Factor-Kappa B Pathway.

BACKGROUND: Glomerular endothelial cell damage plays an important role in the occurrence and development of diabetic nephropathy (DN). OBJECTIVES: This study aimed to clarify the role of XCL1 in DN-mediated glomerular endothelial cell apoptosis and whether the function was related to the activation of the p53/nuclear factor-kappa B (NF-κB) signaling pathway. METHODS: Candidate biomarkers were identified by least absolute shrinkage and selection operator (LASSO) regression model analysis. The area under the receiver operating characteristic curve value was calculated and used to evaluate the discriminating ability. Cell viability, apoptosis, and interleukin-1ß and tumor necrosis factor-α expression at messenger RNA and protein levels were detected by using the Cell Counting Kit-8, flow cytometry, ELISA, real-time polymerase chain reaction, and Western blotting assays. In vivo studies were conducted in the DN mice. RESULTS: The LASSO regression model displayed good discriminating performance, with a C-index of 0.803 and good calibration, and high XCL1 expression was identified as the predicting factor for DN in diabetes mellitus patients. XCL1 expression was upregulated in glomeruli of db/db mice, which was closely related to the expression of its receptor (XCR1). XCL1 overexpression played an important role in the apoptosis and inflammatory response of high glucose (HG)-treated human renal glomerular endothelial cells. Meanwhile, the expression of p53 and the levels of inflammatory cytokines were upregulated upon XCL1 overexpression. p53 silencing with its inhibitor blocked the apoptotic response and inflammatory response in XCL1-overexpressed cells exposed to HG. Besides, the XCL1 overexpression-induced downregulation of NF-κB was reversed by pifithrin-α pretreatment. CONCLUSIONS: Our findings in this work provided the mechanistic insights into the effects of XCL1 on the modulation of DN development, illustrating that XCL1 might serve as an essential prognostic indicator and therapeutic target for DN progression.

XCR1 promotes cell growth and migration and is correlated with bone metastasis in non-small cell lung cancer.

Bone metastasis occurs in approximately 30-40% patients with advanced non-small cell lung cancer (NSCLC), but the mechanism underlying this bone metastasis remains poorly understood. The chemokine super family is believed to play an important role in tumor metastasis in lung cancer. The chemokine receptor XCR1 has been identified to promote cell proliferation and migration in oral cancer and ovarian carcinoma, but the role of XCR1 in lung cancer has not been reported. In this study, we demonstrated for the first time that XCR1 was overexpressed in lung cancer bone metastasis as compared with that in patients with primary lung cancer. In addition, the XCR1 ligand XCL1 promoted the proliferation and migration of lung cancer cells markedly, and knockdown of XCR1 by siRNA abolished the effect of XCL1 in cell proliferation and migration. Furthermore, we identified JAK2/STAT3 as a novel downstream pathway of XCR1, while XCL1/XCR1 increased the mRNA level of the downstream of JAK2/STAT3 including PIM1, JunB, TTP, MMP2 and MMP9. These results indicate that XCR1 is a new potential therapeutic target for the treatment of lung cancer bone metastasis.

Aberrant tissue positioning of metallophilic macrophages in the thymus of XCL1-deficient mice.

Metallophilic macrophages hold a strategic position within the thymic tissue and play a considerable function in thymic physiology. The development and positioning of these cells within thymic tissue are regulated by complex molecular mechanisms involving different cytokine/chemokine axes. Herein, we studied the role of XCL1 signaling in these processes. We show that in the XCL1-deficient thymus numerous metallophilic macrophages are aberrantly positioned in the thymic cortex, instead of their normal location in the cortico-medullary zone. Still, these cells retain their normal appearance: very large size with prominent, ramifying cytoplasmic prolongations. This shows that XCL1 signaling is not involved in morphological development, but rather in correct positioning of metallophilic macrophages within the thymic tissue. In contrast to thymic metallophilic macrophages, the positioning of splenic marginal metallophilic macrophages is not affected by XCL1-deficiency.

The chemokines CXCL10 and XCL1 recruit human annulus fibrosus cells.

STUDY DESIGN: Human annulus fibrosus tissue and cells were analyzed for the presence of chemokine receptors and the migratory effect of selected chemokines. OBJECTIVE: To investigate spontaneous repair mechanisms and underlying cell recruitment in response to annular tears and degenerative defects. SUMMARY OF BACKGROUND DATA: Resorption of herniated disc tissue and the attempt to close annulus tears with repair tissue occur spontaneously. Although chemokines are suggested to play a role in resorption of herniated disc tissue, the role of chemokines in annulus fibrosus homeostasis and repair remains unclear. METHODS: Cells were isolated from annulus fibrosus tissue and expanded in the presence of human serum. Multiwell chemotaxis assays were used to analyze the migratory effect of human serum and 0 to 1000 nM concentrations of the chemokines CXCL7, CXCL10, CXCL12, CCL25, and XCL1 on annulus fibrosus cells (AFCs) (n = 9 per chemokine and dose). Presence of corresponding chemokine receptors in AFCs was determined by real-time polymerase chain reaction analysis and immunohistochemistry. RESULTS: Serum (0.1%-10%) significantly (P < 0.01) stimulates the migration of AFCs. Compared with untreated cells, the migration of cells was significantly (P < 0.01) enhanced upon stimulation with 100 to 1000 nM CXCL10 and 1000 nM XCL1. Chemokine receptors showed low expression levels in expanded AFCs as assessed by polymerase chain reaction. Immunohistochemical staining of the CXCL10 receptor CXCR3 and the XCL1 receptor XCR1 showed that the presence of the particular receptors in AFCs expanded under conventional cell culture conditions. In native annulus fibrosus tissue, CXCR3 was evident, whereas XCR1 could not be detected. CONCLUSION: The findings suggest that chemokines, in particular CXCL10, effectively recruit isolated AFCs. This suggests that chemokines are involved in annulus fibrosus homeostasis and potentially in spontaneous annulus repair attempts. This might have important implications for biological annulus-sealing strategies.

Glycosylated recombinant human XCL1/lymphotactin exhibits enhanced biologic activity.

Chemokines are a family of small, secreted chemoattractant cytokines that regulate distribution and function of leukocytes during immune responses. While most chemokines are members of the CC or CXC subgroups, XCL1, also known as lymphotactin, is the sole member of the C subgroup. XCL1 is produced by activated CD8(+) T cells, NK cells, gammadelta T cells, and mast cells. XCL1 differs from other chemokines in that it contains only a single disulfide bond and a mucin-like domain at its carboxy terminus that is glycosylated. Understanding the biologic functions of chemokines has largely depended upon expression of these recombinant molecules in E. coli. To examine the effects of glycosylation on the biologic activity of XCL1, we designed constructs for expression of human XCL1 in insect S2 cells. Comparison of this material with that expressed in E. coli reveals that glycosylation significantly increases the biologic activity of XCL1.

C and CX3C chemokines: cell sources and physiopathological implications.

Within the fascinating world of chemokines, C and CX3C chemokines have long been regarded as two minor components, even though they present unique features and show less redundancy than the other chemokine families. Nevertheless, the body of data on their expression and role in various inflammatory disorders has grown in the past few years. The C chemokine family is represented by two chemokines, XCL1/lymphotactin-alpha and XCL2/lymphotactin-beta, whereas the CX3C chemokine family contains only one member, called CX3CL1/ fractalkine. In this review, we present an overview on the structure, expression and signaling properties of these chemokines and their respective receptors and examine how they contribute to inflammation and the regulation of leukocyte trafficking, as well as their potential role in the pathophysiology of human inflammatory diseases. Taken together, these data expand the biological importance of C and CX3C chemokines from that of simple immune modulators to a much broader biological role, even though their precise commitment within the framework of immune responses has still to be determined.

Lymphotactin expression by engineered myeloma cells drives tumor regression: mediation by CD4+ and CD8+ T cells and neutrophils expressing XCR1 receptor.

The C chemokine lymphotactin has been characterized as a T cell chemoattractant both in vitro and in vivo. To determine whether lymphotactin expression within tumors could influence tumor growth, we transfected an expression vector for lymphotactin into SP2/0 myeloma cells and tested their ability to form tumors in BALB/c and nude mice. Transfection did not alter cell growth in vitro. Whereas SP2/0 cells gave rise to a 100% tumor incidence, lymphotactin-expressing SP2/0-Lptn tumors invariably regressed in BALB/c mice and became infiltrated with CD4(+) and CD8(+) T cells and neutrophils. Regression of the SP2/0-Lptn tumors was associated with a type 1 cytokine response and dependent on both CD4(+) and CD8(+) T cells, but not NK cells. Both SP2/0 and SP2/0-Lptn tumors grew in nude mice, but growth of the latter tumors was retarded and associated with heavy neutrophil responses; this retardation of SP2/0-Lptn tumor growth was reversed by neutrophil depletion of the mice. Our data also indicate that mouse neutrophils express the lymphotactin receptor XCR1 and that lymphotactin specifically chemoattracts these cells in vitro. Thus, lymphotactin has natural adjuvant activities that may augment antitumor responses via effects on both T cells and neutrophils and thereby could be important in gene transfer immunotherapies for some cancers.

Gene disruption through homologous recombination in Spiroplasma citri: an scm1-disrupted motility mutant is pathogenic.

To determine whether homologous recombination could be used to inactivate selected genes in Spiroplasma citri, plasmid constructs were designed to disrupt the motility gene scm1. An internal scm1 gene fragment was inserted into plasmid pKT1, which replicates in Escherichia coli but not in S. citri, and into the S. citri oriC plasmid pBOT1, which replicates in spiroplasma cells as well as in E. coli. Electrotransformation of S. citri with the nonreplicative, recombinant plasmid pKTM1 yielded no transformants. In contrast, spiroplasmal transformants were obtained with the replicative, pBOT1-derived plasmid pCJ32. During passaging of the transformants, the plasmid was found to integrate into the chromosome by homologous recombination either at the oriC region or at the scm1 gene. In the latter case, plasmid integration by a single crossover between the scm1 gene fragment carried by the plasmid and the full-length scm1 gene carried by the chromosome led to a nonmotile phenotype. Transmission of the scm1-disrupted mutant to periwinkle (Catharanthus roseus) plants through injection into the leafhopper vector (Circulifer haematoceps) showed that the motility mutant multiplied in the insects and was efficiently transmitted to plants, in which it induced symptoms similarly to the wild-type S. citri strain. These results suggest that the spiroplasmal motility may not be essential for pathogenicity and that, more broadly, the S. citri oriC plasmids can be considered promising tools for specific gene disruption by promoting homologous recombination in S. citri, a mollicute which probably lacks a functional RecA protein.

Identification of a chicken "C" chemokine related to lymphotactin.

In this study we describe the isolation and characterization of a new chicken (Gallus gallus) chemokine. This molecule belongs to the C or gamma-chemokine family and is related to the mouse and human lymphotactin (Lptn). Mouse and human Lptn are distinguished from alpha and beta chemokines by the absence of two cysteines (Cys 1 and 3) that form a disulfide bridge; the novel chicken chemokine shows the same cysteine pattern, but replaces a long carboxy-terminal tail found in the other Lptn proteins with a short extension rich in Arg residues. The 1-kb mRNA is mainly expressed in spleen, although weaker signals have been detected in liver and colon. It is interesting to note that the chicken chemokine seems to preferentially induce the migration of spleen B cells over T cells or B cells from the bursa of Fabricius.