Jean Louis Ravetti

CD8+CD28− T Regulatory Lymphocytes Inhibiting T Cell Proliferative and Cytotoxic Functions Infiltrate Human Cancers

Tumor growth is allowed by its ability to escape immune system surveillance. An important role in determining tumor evasion from immune control might be played by tumor-infiltrating regulatory lymphocytes. This study was aimed at characterizing phenotype and function of CD8+CD28− T regulatory cells infiltrating human cancer. Lymphocytes infiltrating primitive tumor lesion and/or satellite lymph node from a series of 42 human cancers were phenotypically studied and functionally analyzed by suppressor assays. The unprecedented observation was made that CD8+CD28− T regulatory lymphocytes are almost constantly present and functional in human tumors, being able to inhibit both T cell proliferation and cytotoxicity. CD4+CD25+ T regulatory lymphocytes associate with CD8+CD28− T regulatory cells so that the immunosuppressive activity of tumor-infiltrating regulatory T cell subsets, altogether considered, may become predominant. The infiltration of regulatory T cells seems tumor related, being present in metastatic but not in metastasis-free satellite lymph nodes; it likely depends on both in situ generation (via cytokine production) and recruitment from the periphery (via chemokine secretion). Collectively, these results have pathogenic relevance and implication for immunotherapy of cancer.

Expression of CXC chemokine receptors 1-5 and their ligands in human glioma tissues : Role of CXCR4 and SDF1 in glioma cell proliferation and migration

Chemokines have been involved in cellular processes associated to malignant transformation such as proliferation, migration and angiogenesis. The expression of five CXC chemokine receptors and their main ligands was analysed by RT-PCR in 31 human astrocytic neoplasms. The mRNAs for all the receptors analysed were identified in a high percentage of tumours, while their ligands showed lower expression. CXCR4 and SDF1 were the most frequently mRNA identified (29/31 and 13/31 of the gliomas studied, respectively). Thus, we further analysed the cell localization of CXCR4 and SDF1 in immunohistochemistry experiments. We show a marked co-localization of CXCR4 and SDF1 in tumour cells, mainly evident in psudolpalisade and microcystic degeneration areas and in the vascular endothelium. In addition, hSDF1alpha induced a significant increase of DNA synthesis in primary human glioblastoma cell cultures and chemotaxis in a glioblastoma cell line. These results provide evidence of the expression of multiple CXC chemokines and their receptors in brain tumours and that in particular CXCR4 and SDF1 sustain proliferation and migration of glioma cells to promote malignant progression.

Expression of the chemokine receptor CXCR4 and its ligand stromal cell-derived factor 1 in human brain tumors and their involvement in glial proliferation in vitro

Abstract: Chemokines are a family of proteins that chemoattract and activate cells by interacting with specific receptors on the surface of their targets. They are grouped into four classes based on the position of key cysteine residues: C, CC, CXC, and CX3C. Stromal cell‐derived factor 1 (SDF1), the ligand of the CXCR4 receptor, is a CXC chemokine involved in chemotaxis and brain development that also acts as coreceptor for HIV‐1 infection. It has been proposed that CXCR4 is overexpressed and required for proliferation in human brain tumor cells. We previously demonstrated that CXCR4 and SDF1 are expressed in culture of cortical type I rat astrocytes, cortical neurons, and cerebellar granule cells. In this study, we analyzed the expression of CXCR4 and SDF1 in four human brain tumor tissues, showing that CXCR4 is expressed in all tumors analyzed, whereas SDF1 is expressed only in two tumor tissues. We also investigated the possible functions of CXCR4 expressed in rat type I cortical astrocytes, demonstrating that SDF1α stimulates the proliferation of these cells in vitro. Moreover, we studied by western blot the intracellular pathway involved in cell proliferation, demonstrating that SDF1α induces the ERK1/2 phosphorylation that is reduced by the PD98059 compound, an MEK inhibitor.

Overexpression of Stromal Cell–Derived Factor 1 and Its Receptor CXCR4 Induces Autocrine/Paracrine Cell Proliferation in Human Pituitary Adenomas

Purpose: Hypothalamic or locally produced growth factors and cytokines control pituitary development, functioning, and cell division. We evaluated the expression of the chemokine stromal cell–derived factor 1 (SDF1) and its receptor CXCR4 in human pituitary adenomas and normal pituitary tissues and their role in cell proliferation. Experimental Design: The expression of SDF1 and CXCR4 in 65 human pituitary adenomas and 4 human normal pituitaries was determined by reverse transcription-PCR, immunohistochemistry, and confocal immunofluorescence. The proliferative effect of SDF1 was evaluated in eight fibroblast-free human pituitary adenoma cell cultures. Results: CXCR4 mRNA was expressed in 92% of growth hormone (GH)-secreting pituitary adenomas (GHoma) and 81% of nonfunctioning pituitary adenomas (NFPA), whereas SDF1 was identified in 63% and 78% of GHomas and NFPAs, respectively. Immunostaining for CXCR4 and SDF1 showed a strong homogenous labeling in all tumoral cells in both GHomas and NFPAs. In normal tissues, CXCR4 and SDF1 were expressed only in a subset of anterior pituitary cells, with a lower expression of SDF1 compared with its cognate receptor. CXCR4 and SDF1 were not confined to a specific cell population in the anterior pituitary but colocalized with discrete subpopulations of GH-, prolactin-, and adrenocorticorticotropic hormone–secreting cells. Conversely, most of the SDF1-containing cells expressed CXCR4. In six of eight pituitary adenoma primary cultures, SDF1 induced a statistically significant increase in DNA synthesis that was prevented by the treatment with the CXCR4 antagonist AMD3100 or somatostatin. Conclusions: CXCR4 and SDF1 are overexpressed in human pituitary adenomas and CXCR4 activation may contribute to pituitary cell proliferation and, possibly, to adenoma development `in humans.

Characterization of the intracellular mechanisms mediating somatostatin and lanreotide inhibition of DNA synthesis and growth hormone release from dispersed human GH‐secreting pituitary adenoma cells in vitro

objective Somatostatin is an endogenous inhibitor of hormone secretion and cell proliferation. Treatment with somatostatin analogues in humans causes a reduction in size and secretory activity of endocrine tumours, including GH‐secreting pituitary adenomas. This study was aimed to characterize the intracellular mechanisms mediating the in vitro antiproliferative and antisecretory effects of somatostatin and its analogue lanreotide, on primary cultures of GH‐secreting pituitary adenoma cells.

High ERp5/ADAM10 expression in lymph node microenvironment and impaired NKG2D ligands recognition in Hodgkin lymphomas

Herein we describe that in classic Hodgkin lymphomas (cHL, n = 25) the lymph node (LN) stroma displayed in situ high levels of transcription and expression of the disulfide-isomerase ERp5 and of the disintegrin-metalloproteinase ADAM10, able to shed the ligands for NKG2D (NKG2D-L) from the cell membrane. These enzymes were detected both in LN mesenchymal stromal cells (MSCs) and in Reed-Sternberg (RS) cells; in addition, MIC-A and ULBP3 were present in culture supernatants of LN MSCs or RS cells. NKG2D-L-negative RS cells could not be killed by CD8(+)αβT or γδT cells; tumor cell killing was partially restored by treating RS cells with valproic acid, which enhanced NKG2D-L surface expression. Upon coculture with LN MSCs, CD8(+)αβT and γδT cells strongly reduced their cytolytic activity against NKG2D-L(+) targets; this seems to be the result of TGF-β, present at the tumor site, produced in vitro by LN MSCs and able to down-regulate the expression of NKG2D on T lymphocytes. In addition, CD8(+)αβT and γδT cells from the lymph nodes of cHL patients, cocultured in vitro with LN MSCs, underwent TGF-β-mediated down regulation of NKG2D. Thus, in cHL the tumor microenvironment is prone to inhibit the development of an efficient antitumor response.

Testicular Sertoli Cell Tumours and Relative Sub-Types

Introduction: Sertoli cell tumours have a rare (0.4–1.5% of all testicular neoplasms) and heterogeneous pathology. The aim of this paper is to analyse the histological classification of Sertoli cell tumours, in order to assess if the three different histotypes – classic type, large cell calcifying Sertoli cell tumour (LCCSCT) and sclerosing Sertoli cell tumour (SSCT) – really present distinctive clinical and prognostic features. Materials and Methods: The current literature was reviewed; Sertoli cell tumour clinical series and single case reports were searched and analysed. Hence, more than 200 classic Sertoli cell tumours, 48 LCCSCTs and only 12 SSCTs were found. The thirteenth SSCT has been found by us in a 34-year-old man. Results: Every single sub-type presents clinical specific characteristics regarding age of onset, bilaterality, focality, abnormal hormone production, correlated systemic symptoms. Ultrasonographic findings, size and – above all – malignant potential. Conclusions: The precise classification of these tumours is not important only histologically: the currently recognised variants really differ in clinical presentation and course. Moreover, LCCSCTs can be further divided in two subgroups with very different clinical behaviour, those in older patients and those associated with well-known syndromes. These clinical and prognostic variables are of great importance when deciding on the therapeutical approach.

CXCR4 and SDF1 expression in human meningiomas: A proliferative role in tumoral meningothelial cells in vitro

Chemokines participate in cellular processes associated with tumor proliferation, migration, and angiogenesis. We previously demonstrated that stromal cell-derived factor 1 (SDF1) exerts a mitogenic activity in glioblastomas through the activation of its receptor CXCR4. Here we studied the expression of this chemokine in human meningiomas and its possible role in cell proliferation. Reverse transcriptase-PCR analysis for CXCR4 and SDF1 was performed on 55 human meningiomas (47 WHO grade I, 5 WHO II, and 3 WHO III). Immunolabeling for CXCR4 and SDF1 was performed on paraffin-embedded sections of these tumors. [(3)H]Thymidine uptake and Western blot analyses were performed on primary meningeal cell cultures of tumors to evaluate the proliferative activity of human SDF1alpha (hSDF1alpha) in vitro and the involvement of extracellular signal-regulated kinase 1/2 (ERK1/2) activation in this process. CXCR4 mRNA was expressed by 78% of the tumor specimens and SDF1 mRNA by 53%. CXCR4 and SDF1 were often detected in the same tumor tissues and colocalized with epithelial membrane antigen immunostaining. In 9 of 12 primary cultures from meningiomas, hSDF1alpha induced significant cell proliferation that was strongly reduced by the mitogen-activated protein kinase kinase inhibitor PD98059, involving ERK1/2 activation in the proliferative signal of hSDF1alpha. In fact, CXCR4 stimulation led to ERK1/2 phosphorylation/activation. In addition, the hSDF1alpha-induced cell proliferation was significantly correlated with the MIB1 staining index in the corresponding surgical specimen. In conclusion, we found that human meningiomas express CXCR4 and SDF1 and that hSDF1alpha induces proliferation in primary meningioma cell cultures through the activation of ERK1/2.

CXC receptor and chemokine expression in human meningioma: SDF1/CXCR4 signaling activates ERK1/2 and stimulates meningioma cell proliferation

Abstract:u2002 Recent evidence indicates that cancer cells express chemokine (CK) receptors and that their signaling is crucial for tumor proliferation, migration, and angiogenesis. The profiles of expression of CXC CK receptors (CXCR1‐5) and their main ligands (growth‐related oncogene, GRO1‐2‐3/CXCL1‐2‐3; interleukin 8, IL‐8/CXCL8; monokine‐induced γ‐interferon MIG/CXCL9; γ‐interferon‐inducible‐protein‐10, IP‐10/CXCL10; stromal cell‐derived factor‐1, SDF1/CXCL12; B‐cell activating CK‐1, BCA‐1/CXCL13) were analyzed by reverse transcription polymerase chain reaction (RT‐PCR) in surgical samples of human meningiomas. All the five receptors displayed high percentages of positive cases: 92% CXCR1, 89% CXCR2, 83% CXCR3, 78% CXCR4, and 94% CXCR5. Conversely, their ligands showed a lower pattern of expression: 40% IL‐8, 42% GRO1‐3, 42% IP‐10, 28% MIG, 53% SDF1, and 3% BCA‐1. SDF1/CXCR4 interaction plays a pivotal role in cancer proliferation. Thus, the signaling mechanisms activated by the exclusive binding between SDF1 and CXCR4 was investigated in 12 primary cultures from meningioma tissues. CXCR4 was functionally coupled as demonstrated by the significant increase of DNA synthesis in meningioma cells in response to SDF1, measured by [3H]‐thymidine uptake. In three primary cultures, the SDF1‐dependent mitogenic activity was associated with a marked phosphorylation of extracellular signal‐regulated kinase (ERK1/2) as evaluated by Western blots. PD98059 (a MEK inhibitor) significantly reduced ERK1/2 activation, thus linking the SDF1/CXCR4 pathway to meningioma cell proliferation via ERK1/2 signal transduction. We demonstrate, for the first time in human meningiomas, the simultaneous expression of CXCR1‐5 and their CKs and the mitogenic activity of SDF1/CXCR4, suggesting a pivotal role of these receptor–ligand pairs in meningeal tumors.

Balance between somatostatin and D2 receptor expression drives TSH-secreting adenoma response to somatostatin analogues and dopastatins

Contextu2002 First‐line therapy for thyrotropin‐secreting pituitary adenomas (TSHomas) is neurosurgery, while medical treatment rests mainly on somatostatin analogues. Clinically available sst2‐preferring analogues, octreotide and lanreotide, induce normalization of hormone levels in approximately 90% of patients and tumour shrinkage in 45%.