Invasion, isolation and evolution shape population genetic structure in Campanula rotundifolia.

The distribution and genetic structure of most plant species in Britain and Ireland bear the imprint of the last ice age. These patterns were largely shaped by random processes during recolonization but, in angiosperms, whole-genome duplication may also have been important. We investigate the distribution of cytotypes of Campanula rotundifolia, considering DNA variation, postglacial colonization, environmental partitioning and reproductive barriers. Cytotypes and genome size variation from across the species' range were determined by flow cytometry and genetic variation was assessed using cpDNA markers. A common garden study examined growth and flowering phenology of tetraploid, pentaploid and hexaploid cytotypes and simulated a contact zone for investigation of reproductive barriers. Irish populations were entirely hexaploid. In Britain, hexaploids occurred mostly in western coastal populations which were allopatric with tetraploids, and in occasional sympatric inland populations. Chloroplast markers resolved distinct genetic groups, related to cytotype and geographically segregated; allopatric hexaploids were distinct from tetraploids, whereas sympatric hexaploids were not. Genome downsizing occurred between cytotypes. Progeny of open-pollinated clones from the contact zone showed that maternal tetraploids rarely produced progeny of other cytotypes, whereas the progeny of maternal hexaploids varied, with frequent pentaploids and aneuploids. The presence of distinctive hexaploid chloroplast types in Ireland, Scottish islands and western mainland Britain indicates that its establishment preceded separation of these land masses by sea-level rise c. 16 000 years BP. This group did not originate from British tetraploids and probably diverged before postglacial invasion from mainland Europe. The combination of cytotype, molecular, contact zone and common garden data shows an overall pattern reflecting postglacial colonization events, now maintained by geographic separation, together with more recent occasional local in situ polyploidisation. Reproductive barriers favour the persistence of the tetraploid to the detriment of the hexaploid.

From the Data on Many, Precision Medicine for "One": The Case for Widespread Genomic Data Sharing.

Within the decade, genome sequencing promises to become a routine part of healthcare around the globe. Many millions of genomes linked to health records will soon be available for researchers and clinicians to make use of to advance precision medicine. To realise the full impact of genomic medicine, genomic and clinical data must be interoperable across traditional geographic, jurisdictional, sectoral, and domain boundaries. Extremely large and diverse data sets are needed to provide a context for interpretation of genetic sequences. No single country or institution can achieve the necessary scale and diversity alone. Data must be shared within an internationally federated, learning health system.

α/ß-Peptide Foldamers Targeting Intracellular Protein-Protein Interactions with Activity in Living Cells.

Peptides can be developed as effective antagonists of protein-protein interactions, but conventional peptides (i.e., oligomers of l-α-amino acids) suffer from significant limitations in vivo. Short half-lives due to rapid proteolytic degradation and an inability to cross cell membranes often preclude biological applications of peptides. Oligomers that contain both α- and ß-amino acid residues ("α/ß-peptides") manifest decreased susceptibility to proteolytic degradation, and when properly designed these unnatural oligomers can mimic the protein-recognition properties of analogous "α-peptides". This report documents an extension of the α/ß-peptide approach to target intracellular protein-protein interactions. Specifically, we have generated α/ß-peptides based on a "stapled" Bim BH3 α-peptide, which contains a hydrocarbon cross-link to enhance α-helix stability. We show that a stapled α/ß-peptide can structurally and functionally mimic the parent stapled α-peptide in its ability to enter certain types of cells and block protein-protein interactions associated with apoptotic signaling. However, the α/ß-peptide is nearly 100-fold more resistant to proteolysis than is the parent stapled α-peptide. These results show that backbone modification, a strategy that has received relatively little attention in terms of peptide engineering for biomedical applications, can be combined with more commonly deployed peripheral modifications such as side chain cross-linking to produce synergistic benefits.

Rangewide ploidy variation and evolution in Acacia senegal: a north-south divide?

Knowledge of rangewide variation in DNA content and ploidy level may be valuable in understanding the evolutionary history of a species. Recent studies of Acacia senegal report diploids and occasional tetraploids in the Sudano-Sahelian region of sub-Saharan Africa, but nothing is known about the overall extent of DNA ploidy variation within the species. In this study, we determine the DNA content and ploidy level of A. senegal across its native range, and explore whether the variation is related to its evolutionary and colonization history. We used propidium iodide flow cytometry (FCM) to estimate DNA content (2C value) and infer ploidy in 157 individuals from 54 populations on various tissues, using seeds, fresh leaves, dried leaves and twigs and herbarium specimens. The mean 2C DNA (pg ± s.d.) contents detected were 1.47 ± 0.09, 2.12 ± 0.02, 2.89 ± 0.12, and a single individual with 4.51 pg, corresponding to a polyploid series of diploid, triploid, tetraploid and hexaploid individuals. Diploids were confirmed by chromosome counts (2n = 2x = 26). Most populations (90.7 %) were of single ploidy level, while mixed ploidy populations (9.3 %) comprising mostly diploids (2x+3x, 2x+4x and 2x+6x) were restricted to the Sudano-Sahelian and Indian subcontinent regions, its northern range. The species is predominantly diploid, and no mixed ploidy populations were detected in east and southern Africa, its southern range. The geographic pattern of ploidy variation in conjunction with existing phylogeographic and phylogenetic data of the species suggests that polyploids have occurred multiple times in its evolutionary and recent colonization history, including contemporary ecological timescales. The successful use of external tissues of dried twigs in FCM is new, and presents the opportunity to study numerous other dryland woody species.

Aberrant regulation of argininosuccinate synthetase by TNF-alpha in human epithelial ovarian cancer.

The pro-inflammatory cytokine, tumour necrosis factor-alpha, TNF-alpha, is dysregulated in malignant compared with normal ovarian surface epithelium (OSE). Several epidemiological studies have associated inflammation with ovarian tumorigenesis, with TNF-alpha playing a key role in modulating invasion, angiogenesis and metastasis. Here, we show that TNF-alpha also induces expression of arate-limiting enzyme in arginine synthesis, argininosuccinate synthetase (AS), thereby linking inflammation with several arginine-dependent metabolic pathways, implicated in accelerated carcinogenesis and tumour progression. Having identified AS mRNA induction in TNF-alpha-treated IGROV-1 ovarian cancer cells, using RNA-arbitrarily primed-PCR, we then observed differential regulation of AS mRNA and protein in malignant, compared with normal, OSE cells. A cDNA cancer profiling array with matched normal ovarian and ovarian tumour samples revealed increased expression of AS mRNA in the latter. Moreover, AS protein co-localised with TNF-alpha in ovarian cancer cells, with significantly higher levels of AS in malignant compared with normal ovarian tissue. Increased co-expression of AS and TNF-alpha mRNA was also observed in 2 other epithelial tumours, non-small cell lung and stomach cancer, compared with normal corresponding tissues. In summary, high levels of AS expression, which may be required for several arginine-dependent processes in cancer, including the production of nitric oxide, proline, pyrimidines and polyamines, is regulated by TNF-alpha and may provide an important molecular pathway linking inflammation and metabolism to ovarian tumorigenesis.

The inflammatory cytokine tumor necrosis factor-alpha generates an autocrine tumor-promoting network in epithelial ovarian cancer cells.

Constitutive expression of the inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is characteristic of malignant ovarian surface epithelium. We investigated the hypothesis that this autocrine action of TNF-alpha generates and sustains a network of other mediators that promote peritoneal cancer growth and spread. When compared with two ovarian cancer cell lines that did not make TNF-alpha, constitutive production of TNF-alpha was associated with greater release of the chemokines CCL2 and CXCL12, the cytokines interleukin-6 (IL-6) and macrophage migration-inhibitory factor (MIF), and the angiogenic factor vascular endothelial growth factor (VEGF). TNF-alpha production was associated also with increased peritoneal dissemination when the ovarian cancer cells were xenografted. We next used RNA interference to generate stable knockdown of TNF-alpha in ovarian cancer cells. Production of CCL2, CXCL12, VEGF, IL-6, and MIF was decreased significantly in these cells compared with wild-type or mock-transfected cells, but in vitro growth rates were unaltered. Tumor growth and dissemination in vivo were significantly reduced when stable knockdown of TNF-alpha was achieved. Tumors derived from TNF-alpha knockdown cells were noninvasive and well circumscribed and showed high levels of apoptosis, even in the smallest deposits. This was reflected in reduced vascularization of TNF-alpha knockdown tumors. Furthermore, culture supernatants from such cells failed to stimulate endothelial cell growth in vitro. We conclude that autocrine production of TNF-alpha by ovarian cancer cells stimulates a constitutive network of other cytokines, angiogenic factors, and chemokines that may act in an autocrine/paracrine manner to promote colonization of the peritoneum and neovascularization of developing tumor deposits.

Endothelins induce CCR7 expression by breast tumor cells via endothelin receptor A and hypoxia-inducible factor-1.

Endothelin expression is increased in breast tumors and is associated with invasion and metastasis, whereas CCR7 expression by breast tumor cells may have a role in the organ specificity of breast cancer spread. In this article, we have analyzed whether endothelins influence breast tumor cell expression of the chemokine receptor CCR7. Stimulation of human breast tumor cell lines with endothelins increased cell surface expression of CCR7 via endothelin receptor A. The iron chelators desferrioxamine and cobalt chloride, which induce hypoxia-inducible factor (HIF)-mediated transcription, also increased CCR7 expression; transfection of a dominant-negative version of the HIF regulatory subunit, HIF-1alpha, into MCF-7 cells abolished CCR7 induction by endothelins, indicating that increased expression is due to HIF-1 stabilization. Endothelin stimulation promoted invasion toward the CCR7 ligands CCL19 and CCL21. Endothelin-mediated chemokine-independent invasion itself is dependent on CCR7 activity and could be abolished using a CCR7-neutralizing monoclonal antibody. In human breast carcinomas, mRNA expression of endothelins correlated with the level of CCR7 expression, both of which were associated with the presence of lymph node metastases. Expression of the CCR7 ligands CCL19 and CCL21 was also higher in breast cancer patients with lymph node involvement compared with those without, but expression of these chemokines did not correlate with endothelin expression. These data show that CCR7 may be regulated by the breast tumor microenvironment and further support the use of endothelin receptor antagonists in the treatment of invasive and metastatic breast cancer.

The chemokine network: a target in cancer biology?

Chemokine gradients are central to the movement of cells in both homeostatic and pathological processes. Most cancers express a complex array of chemokines that influence the local microenvironment through recruitment of stromal cells and by stimulating angiogenesis. Recently, the discovery of chemokine receptors on tumor cells has led to speculation that the chemokine system may be involved in cancer cell growth and survival, and possibly the development of site-specific spread. Understanding the networks of chemokines and their receptors in cancer will enable manipulation of this system. Both chemokines and their receptors represent targets for therapeutic intervention either with antibodies or small molecule antagonists. However, due to the complexity of the system, and the number of chemokines and receptors that are also expressed by normal cells, issues remain concerning whether systemic or local drug delivery are preferable and whether the redundancy of the system will compensate if one chemokine or receptor is targeted. Nevertheless, efficacy has been demonstrated in a number of experimental models. By targeting this network, it may be possible to generate anti-tumor immune responses by altering the chemokine and/or leukocyte balance in tumors; alternatively, chemokine/chemokine receptor-expressing cancer cells could be directly targeted.

Ovarian cancer cells polarize macrophages toward a tumor-associated phenotype.

Tumor-associated macrophages (TAM) may have tumor-promoting activity, but it is not clear how their phenotype is achieved. In this study, we demonstrate that ovarian cancer cells switch cocultured macrophages to a phenotype similar to that found in ovarian tumors. Tumor cells caused dynamic changes in macrophage cytokine, chemokine, and matrix metalloprotease mRNA, and protein-inducing mediators that are found in human cancer. Macrophage mannose, mannose receptor, and scavenger receptors (SR-As) were also up-regulated by coculture, but not by conditioned medium. To further validate the model, we studied SR-A regulation on TAM in vitro and in vivo. Coculture of murine macrophages from mice deficient in TNF-alpha or its receptors revealed that TNF-alpha was key to SR-A induction via its p75 receptor. SR-A expression was also reduced in TAM from ovarian cancers treated with anti-TNF-alpha Abs or grown in TNF-alpha(-/-) mice. Chemical communication between tumor cells and macrophages may be important in regulating the cancer cytokine microenvironment.

Expression and regulation of tumor necrosis factor alpha in normal and malignant ovarian epithelium.

Epidemiologic studies implicate inflammatory stimuli in the development of ovarian cancer. The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) and both its receptors (TNFRI and TNFRII) are expressed in biopsies of this malignancy. Here, we tested the hypothesis that TNF-alpha is a regulator of the proinflammatory microenvironment of ovarian cancer. A cancer profiling array showed higher expression of TNF-alpha in ovarian tumors compared with normal ovarian tissue, and cultured ovarian cancer cells expressed up to 1,000 times more TNF-alpha mRNA than cultured normal ovarian surface epithelial cells; TNF-alpha protein was only detected in the supernatant of tumor cell cultures. Treatment with TNF-alpha induced TNF-alpha mRNA via TNFRI in both malignant and normal cells with evidence for enhanced TNF-alpha mRNA stability in tumor cells. TNF-alpha induced TNF-alpha protein in an autocrine fashion in tumor but not in normal ovarian surface epithelial cells. The TNF-alpha neutralizing antibody infliximab reduced the constitutive levels of TNF-alpha mRNA in tumor cell lines capable of autocrine TNF-alpha production. Apart from TNF-alpha mRNA expression, several other proinflammatory cytokines were constitutively expressed in malignant and normal ovarian surface epithelial cells, including interleukin (IL)-1alpha, IL-6, CCL2, CXCL8, and M-CSF. TNF-alpha treatment further induced these cytokines with de novo transcription of IL-6 mRNA contrasting with the increased stability of CCL2 mRNA. RNA interference directed against TNF-alpha was highly effective in abolishing constitutive IL-6 production by ovarian tumor cells. In summary, we show that TNF-alpha is differentially regulated in ovarian cancer cells compared with untransformed cells and modulates production of several cytokines that may promote ovarian tumorigenesis. Infliximab treatment may have a role in suppressing the TNF-alpha-driven inflammatory response associated with ovarian cancer.

The inflammatory cytokine tumor necrosis factor-alpha regulates chemokine receptor expression on ovarian cancer cells.

Epithelial ovarian cancer cells express the chemokine receptor, CXCR4, which may be associated with increased survival and metastatic potential, but the regulation of this receptor is not understood. The inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is found in ovarian cancer biopsies and is associated with increased tumor grade. In this report, we show that CXCR4 expression on human epithelial ovarian cancer cells is associated with, and can be modulated by, TNF-alpha. Ovarian cancer cells with high endogenous expression of TNF-alpha expressed higher levels of CXCR4 mRNA and protein than cells with low TNF-alpha expression. Stimulation of ovarian cancer cell lines and primary epithelial cancer cells with TNF-alpha resulted in increased CXCR4 mRNA and protein. The TNF-alpha-stimulated increase in CXCR4 mRNA was due partly to de novo synthesis, and up-regulation of CXCR4 cell surface protein increased migration to the CXCR4 ligand CXCL12. CXCR4 mRNA and protein was down-regulated by anti-TNF-alpha antibody or by targeting TNF-alpha mRNA using RNAi. TNF-alpha stimulation activated components of the nuclear factor kappaB pathway, and overexpression of the inhibitor of kappaB also reduced CXCR4 expression. Coculture of macrophages with ovarian cancer cells also resulted in cancer cell up-regulation of CXCR4 mRNA in a TNF-alpha-dependent manner. Finally, there was a correlation between the levels of TNF-alpha and CXCR4 mRNA in clinical biopsies of ovarian cancer, and TNF-alpha protein was expressed in CXCR4-positive tumor cells. TNF-alpha is a critical mediator of tumor promotion in a number of experimental cancers. Our data suggest that one mechanism may be through nuclear factor kappaB-dependent induction of CXCR4.

Macrophages induce invasiveness of epithelial cancer cells via NF-kappa B and JNK.

Tumor-associated macrophages may influence tumor progression, angiogenesis and invasion. To investigate mechanisms by which macrophages interact with tumor cells, we developed an in vitro coculture model. Previously we reported that coculture enhanced invasiveness of the tumor cells in a TNF-alpha- and matrix metalloprotease-dependent manner. In this report, we studied intracellular signaling pathways and induction of inflammatory genes in malignant cells under the influence of macrophage coculture. We report that coculture of macrophages with ovarian or breast cancer cell lines led to TNF-alpha-dependent activation of JNK and NF-kappaB pathways in tumor cells, but not in benign immortalized epithelial cells. Tumor cells with increased JNK and NF-kappaB activity exhibited enhanced invasiveness. Inhibition of the NF-kappaB pathway by TNF-alpha neutralizing Abs, an NF-kappaB inhibitor, RNAi to RelA, or overexpression of IkappaB inhibited tumor cell invasiveness. Blockade of JNK also significantly reduced invasiveness, but blockade of p38 MAPK or p42 MAPK had no effect. Cocultured tumor cells were screened for the expression of 22 genes associated with inflammation and invasion that also contained an AP-1 and NF-kappaB binding site. EMMPRIN and MIF were up-regulated in cocultured tumor cells in a JNK- and NF-kappaB-dependent manner. Knocking down either MIF or EMMPRIN by RNAi in the tumor cells significantly reduced tumor cell invasiveness and matrix metalloprotease activity in the coculture supernatant. We conclude that TNF-alpha, via NF-kappaB, and JNK induces MIF and EMMPRIN in macrophage to tumor cell cocultures and this leads to increased invasive capacity of the tumor cells.

The chemokine network in cancer--much more than directing cell movement.

Cytokine and chemokine gradients are central to the directed movement of cells in both homeostatic and pathological processes. Most cancers have a complex chemokine network which can influence immune responses to the tumor, direct the extent and cellular composition of the leukocyte infiltrate and also play a role in angiogenesis. Tumor cells can also hijack the chemokine system and gain expression of certain chemokine receptors and respond to specific chemokine gradients. Chemokine receptor expression and activation on malignant cells may be central to the growth, survival and migration of cancer cells from the primary tumor. Chemokine receptors, both CC and CXC have been detected on malignant cells and the relevant ligands are sometimes expressed at the tumor site and at sites of tumor spread, suggesting a role for the chemokine family in malignant growth and metastasis.

A chemokine receptor antagonist inhibits experimental breast tumor growth.

The leukocyte infiltrate of human and murine epithelial cancers is regulated by chemokine production in the tumor microenvironment. In this article, we tested the hypothesis that chemokine receptor antagonists may have anticancer activity by inhibiting this infiltrate. We first characterized CC chemokines, chemokine receptors, and the leukocyte infiltrate in the 410.4 murine model of breast cancer. We found that CCL5 (RANTES) was produced by the tumor cells, and its receptors, CCR1 and CCR5, were expressed by the leukocyte infiltrate. As Met-CCL5 is an antagonist of CCR1 and CCR5 with activity in models of inflammatory disease, we tested its activity against 410.4 tumors. After 5 weeks of daily treatment with Met-CCL5, the volume and weight of 410.4 tumors was significantly decreased compared with control-treated tumors. Met-CCL5 was also active against established tumors. The total cell number obtained after collagenase digestion was decreased in Met-CCL5-treated tumors as was the proportion of infiltrating macrophages. Furthermore, chemokine antagonist treatment increased stromal development and necrosis. Our results provide direct evidence that macrophages contribute to tumor development and are the first indication that chemokine receptor antagonists may provide novel strategies in cancer prevention and treatment.

Multiple actions of the chemokine CXCL12 on epithelial tumor cells in human ovarian cancer.

Of 14 chemokine receptors investigated, only CXCR4 was expressed on ovarian cancer cells [C. J. Scotton et al., Cancer Res., 61: 4961-4965, 2001]. To further understand the role of this chemokine receptor in ovarian tumor biology, we studied the action of its ligand, CXCL12 (stromal cell-derived factor 1), on the CXCR4-expressing ovarian cancer cell lines IGROV. Ligand stimulation of the CXCR4 receptor resulted in sustained activation of Akt/protein kinase B and biphasic phosphorylation of p44/42 mitogen-activated protein kinase in IGROV. When IGROV cells were cultured under suboptimal conditions, CXCL12 stimulated their in vitro growth, an effect that was abrogated by neutralizing antibodies to CXCR4. This increase in cell number was attributable to stimulation of DNA synthesis, not protection from apoptosis. CXCL12 treatment of IGROV cells also induced mRNA and protein for tumor necrosis factor alpha, a cytokine that is expressed by tumor cells in ovarian cancer biopsies. IGROV cells invaded through Matrigel toward a CXCL12 gradient. Invasion was abrogated by the broad spectrum matrix metalloproteinase and TNFalpha converting enzyme inhibitor Marimastat and was partially inhibited by neutralizing antitumor necrosis factor alpha antibodies. These effects were not limited to the IGROV cell line. They could also be demonstrated in the CAOV-3 ovarian cancer cell line and primary ovarian tumor cells isolated from ovarian ascites. These biological effects of CXCL12 on IGROV cells were also inhibited by the small molecular weight CXCR4 antagonist AMD3100. Finally, we found abundant intracellular CXCL12 protein in tumor cells in 15 of 18 ovarian cancer biopsies but not in epithelial cells from normal ovary or borderline disease. The chemokine CXCL12 may have multiple biological effects in ovarian cancer, stimulating cell migration and invasion through extracellular matrix, as well as DNA synthesis and establishment of a cytokine network in situations that are suboptimal for tumor cell growth.

Natural killer and dendritic cell contact in lesional atopic dermatitis skin--Malassezia-influenced cell interaction.

The regulation of dendritic cells is far from fully understood. Interestingly, several recent reports have suggested a role for natural killer cells in affecting dendritic cell maturation and function upon direct contact between the cells. It is not known if this interaction takes place also in vivo, or if a potential interaction of natural killer cells and dendritic cells would be affected by allergen exposure of the dendritic cells. The yeast Malassezia can act as an allergen in atopic eczema/dermatitis syndrome, and induce maturation of dendritic cells. Our aims were to study the distribution of natural killer cells in the skin from atopic eczema/dermatitis syndrome patients with the emphasis on possible natural killer cell-dendritic cell interaction, and to assess whether the interaction of Malassezia with dendritic cells would affect subsequent interaction between dendritic cells and natural killer cells. A few scattered natural killer (CD56+/CD3-) cells were found in the dermis of healthy individuals and in nonlesional skin from atopic eczema/dermatitis syndrome patients. In lesional skin and in biopsies from Malassezia atopy-patch-test-positive skin, however, natural killer cells were differentially distributed and for the first time we could show close contact between natural killer cells and CD1a+ dendritic cells. Dendritic cells preincubated with Malassezia became less susceptible to natural-killer-cell-induced cell death, suggesting a direct effect imposed by Malassezia upon interaction of dendritic cells with natural killer cells. These findings indicate that natural killer cells and dendritic cells can interact in the skin and that Malassezia affects the interaction between natural killer cells and dendritic cells. Our data suggest that natural killer cells may play a role in regulating dendritic cells in atopic eczema/dermatitis syndrome.

Endothelin-2 is a macrophage chemoattractant: implications for macrophage distribution in tumors.

Endothelins (ET-1, ET-2 and ET-3) are 21-amino acid vasoactive peptides that bind to G-protein-linked transmembrane receptors, ET-RA and ET-RB. As well as modulating vasoconstriction, endothelins regulate growth in several cell types and may also affect differentiation, inflammation and angiogenesis. Both macrophages and endothelins are found in areas of hypoxia in solid tumors and ET-2 expression may be modulated by hypoxia in some tumors. As the peptide structure of mature endothelins is similar to that of CXC chemokines, we asked if endothelins contribute to control of macrophage distribution in tumors. We found that ET-2 is a chemoattractant for macrophages and THP-1 monocytic cells, but not for freshly isolated monocytes. The chemotactic response to ET-2 shows a typical bell-shaped response curve. Experiments with endothelin receptor antagonists showed that migration to ET-2 is mediated via the ET-RB receptor. Moreover, monocytes do not express ET-RB. Chemotaxis towards ET-2 is via the MAPK pathway: p44 and p42 are phosphorylated when THP-1 cells are stimulated with ET-2, and the MAPKK inhibitor PD98059 stops chemotaxis. As with 'classical' chemokines, migration toET-2 is also inhibited by hypoxia and by pertussis toxin. As well as its chemotactic properties, ET-2 leads to activation of macrophages. In human breast tumors that express ET-2, endothelins and ET-RB expressing macrophages often co-localized. While shorter than 'classical' chemokines, ET-2 shares a similar peptide sequence with chemokines and may signal via a similar receptor and MAPK-mediated pathway. Furthermore, ET-2 expression by tumors may modulate the behavior of macrophages such that activated cells accumulate in areas of hypoxia.

The role of cytokines in the epithelial cancer microenvironment.

The epithelial tumour microenvironment is a complex tissue comprising variable numbers of tumour cells, fibroblasts, endothelial cells and infiltrating leucocytes. Cytokines are key molecules controlling autocrine or paracrine communications within and between these individual cell types. Under some circumstances, endogenous cytokines may orchestrate host responses against the tumour, but there is increasing evidence that the cytokine network contributes to tumour growth, progression and host immuno-suppression. In this review we outline some of the actions of endogenous cytokines in epithelial tumours with particular emphasis on tumour necrosis factor alpha, TNF, related inflammatory cytokines and the chemokine group of chemoattractant cytokines.

Analysis of chemokines and chemokine receptor expression in ovarian cancer ascites.

PURPOSE: Ascitic disease is a common occurrence in human ovarian cancer, but it is unclear how the cellular composition of ascitic fluid is determined. Because chemokines can determine host cell infiltration in solid ovarian cancer, we assessed CC chemokine protein and CC chemokine receptor expression in ovarian cancer ascites. EXPERIMENTAL DESIGN: We used reverse transcription-PCR and RNase protection assay to determine CC chemokine and chemokine receptor mRNA expression and ELISA to measure CC chemokine protein levels. Flow cytometry was used to identify cell populations and their chemokine receptor protein expression. RESULTS: mRNA for the CC chemokines CCL2, -3, -4, -5, -8, and -22 was expressed in cell isolates from ascites samples, and the corresponding proteins were detected in ascitic fluid. mRNA for CC chemokine receptors CCR1, -2a, -2b, -3, -4, -5, and -8 was detected in cells from ascites. Fluorescence-activated cell-sorting analysis showed variable numbers of macrophages and CD3(+) T lymphocytes (predominantly CD4(+)) within ovarian cancer ascites. CD14(+) macrophages within ascites consistently expressed protein for CCR1, -2, and -5. CCR1 was expressed by >60% of all T cells, but more CD4(+) than CD8(+) T cells expressed CCR2 and -5. A direct correlation was found between the CCL5 concentration and CD3(+) T-cell infiltration. CONCLUSIONS: We conclude that there is a complex chemokine/chemokine receptor network in ovarian cancer ascites. However, associations between chemokine receptor expression, chemokine levels, and cell counts were limited.