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2.
Clin Exp Rheumatol ; 28(1): 91-3, 2010.
Article in English | MEDLINE | ID: mdl-20346246

ABSTRACT

Juvenile psoriatic arthritis was diagnosed in a girl of 15 and a half years old, who presented with severe poly-arthritis and psoriasis. Treatment with etanercept 25 mg by subcutaneous injections, twice a week was started. After 5 months of treatment, she developed microscopic hematuria, proteinuria and progressive acute renal failure with anaemia and hypertension. Renal histology, IF, and EM findings were consistent with severe extracapillary crescentic pauciimmune glomerulonephritis. The histology findings, the onset of renal symptoms after beginning treatment with etanercept, and the absence of any abnormality in the urine tests before administration of the drug, support the hypothesis of a rare case of secondary nephropathy due to treatment with an anti-TNF-alpha drug.


Subject(s)
Acute Kidney Injury/chemically induced , Antirheumatic Agents/adverse effects , Arthritis, Psoriatic/drug therapy , Glomerulonephritis/chemically induced , Immunoglobulin G/adverse effects , Acute Kidney Injury/pathology , Adolescent , Antirheumatic Agents/administration & dosage , Biopsy , Etanercept , Female , Glomerulonephritis/pathology , Humans , Immunoglobulin G/administration & dosage , Receptors, Tumor Necrosis Factor/administration & dosage , Tumor Necrosis Factor-alpha/antagonists & inhibitors
3.
Lupus ; 18(6): 567-9, 2009 May.
Article in English | MEDLINE | ID: mdl-19395461

ABSTRACT

Differential diagnosis in patients presenting with fever of unknown origin (FUO) is often difficult because infectious diseases, neoplasms, infective endocarditis or systemic autoimmune diseases may all be responsible for the condition. Furthermore, vasculitis may generate typical, atypical or limited syndromes depending on the extent of vascular involvement. Here, we report the case of a 73-year-old man with FUO and renal failure due to a rare variant of Wegener's granulomatosis, limited to the kidneys.


Subject(s)
Granulomatosis with Polyangiitis/complications , Nephritis/complications , Renal Insufficiency/etiology , Aged , Antibodies, Antineutrophil Cytoplasmic/analysis , Diagnosis, Differential , Enzyme-Linked Immunosorbent Assay , Follow-Up Studies , Granulomatosis with Polyangiitis/diagnosis , Granulomatosis with Polyangiitis/immunology , Humans , Immunohistochemistry , Kidney/pathology , Male , Nephritis/diagnosis , Nephritis/immunology , Renal Insufficiency/diagnosis
4.
Clin Exp Immunol ; 149(3): 504-12, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17590173

ABSTRACT

Expression of the autoimmune regulator gene (AIRE) and the presence of CD25(+)/forkhead box p3 (FoxP3)(+) T regulatory (T(reg)) cells were investigated in histologically normal adult thymi and in thymomas using immunohistochemistry and quantitative real-time polymerase chain reaction (PCR). In the normal thymus staining for AIRE was detected in the nucleus of some epithelial-like cells located in the medulla; in thymomas AIRE-positive cells were extremely rare and could be detected only in the areas of medullary differentiation of two B1 type, organoid thymomas. RNA was extracted from 36 cases of thymoma and 21 non-neoplastic thymi obtained from 11 myasthenic (MG(+)) and 10 non-myasthenic (MG(-)) patients. It was found that AIRE is 8.5-fold more expressed in non-neoplastic thymi than in thymomas (P = 0.01), and that the amount of AIRE transcripts present in the thymoma tissue are not influenced by the association with MG, nor by the histological type. A possible involvement of AIRE in the development of MG was suggested by the observation that medullary thymic epithelial cells isolated from AIRE-deficient mice contain low levels of RNA transcripts for CHRNA 1, a gene coding for acetylcholine receptor. Expression of human CHRNA 1 RNA was investigated in 34 human thymomas obtained from 20 MG(-) patients and 14 MG(+) patients. No significant difference was found in the two groups (thymoma MG(+), CHRNA1 = 0.013 +/- 0.03; thymoma MG-, CHRNA1 = 0.01 +/- 0.03). In normal and hyperplastic thymi CD25(+)/Foxp3(+) cells were located mainly in the medulla, and their number was not influenced by the presence of MG. Foxp3(+) and CD25(+) cells were significantly less numerous in thymomas. A quantitative estimate of T(reg) cells revealed that the levels of Foxp3 RNA detected in non-neoplastic thymi were significantly higher (P = 0.02) than those observed in 31 cases of thymomas. Our findings indicate that the tissue microenvironment of thymomas is defective in the expression of relevant functions that exert a crucial role in the negative selection of autoreactive lymphocytes.


Subject(s)
T-Lymphocytes, Regulatory/immunology , Thymoma/immunology , Thymus Neoplasms/immunology , Transcription Factors/metabolism , Adult , Aged , Female , Forkhead Transcription Factors/metabolism , Humans , Immunoenzyme Techniques , Male , Middle Aged , Myasthenia Gravis/immunology , Polymerase Chain Reaction/methods , Thymus Gland/immunology , Transcription Factors/genetics , AIRE Protein
5.
J Pathol ; 211(3): 351-61, 2007 Feb.
Article in English | MEDLINE | ID: mdl-17152050

ABSTRACT

Collagen type I serves as an abundant structural and signalling component of skin. It is also an established target gene of the transcription factor, c-Myb. When c-myb-/- embryos were examined it was observed that their skin was markedly thinner than normal. Importantly, immunohistochemical investigation showed complete absence of collagen type I. Although these homozygous knock-out embryos fail to develop beyond day 15, fibroblasts established from these embryos (mouse embryonic fibroblasts [MEFs]) show defective proliferative responses. Furthermore, in vitro scratch wound assays demonstrated that these c-myb-/- MEFs also exhibit slower closure than their wild-type counterparts. Embryonic lethality has meant that examination of the role of c-Myb in adult mouse skin has not been reported to date. However, in view of the abundance of collagen type I in normal skin, its role in skin integrity and the in vitro data showing proliferative and migration defects in c-myb-/- MEFs, we investigated the consequences of heterozygous c-myb loss in adult mice on the complex process of skin repair in response to injury. Our studies clearly demonstrate that heterozygous c-myb deficiency has a functional effect on wound repair, collagen type I levels and, in response to wounding, transforming growth factor-beta1 (an important collagen stimulating factor) induction expression is aberrantly high. Manipulation of c-Myb may therefore provide new therapeutic opportunities for improving wound repair while uncontrolled expression may underpin some fibrotic disorders.


Subject(s)
Collagen Type I/metabolism , Extracellular Matrix/metabolism , Genes, myb , Proto-Oncogene Proteins c-myb/metabolism , Skin/metabolism , Wound Healing , Animals , Cell Cycle , Cell Proliferation , Cells, Cultured , Collagen Type I/analysis , DNA Primers/genetics , Extracellular Matrix/chemistry , Fibroblasts/metabolism , Fibroblasts/pathology , Heterozygote , Immunohistochemistry/methods , Mice , Mice, Inbred BALB C , Microscopy, Fluorescence , Models, Animal , Proto-Oncogene Proteins c-myb/analysis , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain Reaction , Skin/pathology , Transforming Growth Factor beta/analysis , Transforming Growth Factor beta/metabolism
6.
Clin Exp Rheumatol ; 22(3 Suppl 33): S66-72, 2004.
Article in English | MEDLINE | ID: mdl-15344601

ABSTRACT

OBJECTIVE: PTX3 is a secreted molecule which consists of a C-terminal domain similar to classical pentraxins (e.g. C-reactive protein) and of an unrelated N-terminal domain. Unlike the classical pentraxins, PTX3 is expressed in response to IL-1beta and TNF-alpha but not to IL-6. The present study was designed to investigate the expression of PTX3 in normal and scleroderma fibroblasts. METHODS: Normal and SSc fibroblasts were cultured in the presence and absence of inflammatory cytokines. PTX3 mRNA expression in fibroblasts was evaluated by Northern analysis. PTX3 protein levels in fibroblast culture medium were estimated by ELISA. RESULTS: Normal fibroblasts were induced to express high levels of P7X3 mRNA by IL-1beta and TNF-alpha but not by other cytokines or growth factors. Scleroderma fibroblasts, unlike normal fibroblasts, constitutively expressed high levels of PTX3 in the absence of deliberate stimulation. The constitutive expression of PTX3 in SSc fibroblasts was not modified by anti-TNF-alpha antibodies or IL-1 receptor antagonist. In contrast, IFN-gamma and TGF-beta inhibited the constitutive but not the stimulated expression of PTX3 in SSc fibroblasts. CONCLUSIONS: PTX3 is a main feature of activated scleroderma fibroblasts.


Subject(s)
C-Reactive Protein/biosynthesis , Fibroblasts/metabolism , Scleroderma, Systemic/metabolism , Serum Amyloid P-Component/biosynthesis , Cell Culture Techniques , Cytokines/metabolism , Humans , Interferon-gamma/metabolism , Interleukin-1/metabolism , RNA, Messenger/biosynthesis , Transforming Growth Factor beta , Tumor Necrosis Factor-alpha/metabolism
8.
Eur J Immunol ; 31(10): 3101-10, 2001 Oct.
Article in English | MEDLINE | ID: mdl-11592087

ABSTRACT

The glycolipid alpha-galactosylceramide (alpha-GalCer), ligand of NKT cells, has been recently shown to induce antitumor immunity in mice through the induction of IL-12 production by dendritic cells. In the present study we compared alpha-GalCer and rIL-12 antitumor activities in the treatment of hepatic metastases of the C-26 murine colon carcinoma. We show that in immunocompetent mice the two molecules display similar efficacy, whereas in mice knockout (KO) for beta2-microglobulin (beta2m), IFN-gamma or IL-12p40, alpha-GalCer antitumor activity is severely impaired. Conversely,in all such KO mice, rIL-12 retains its efficacy. In this context, the IL-12 effect relies on NK cell function since it is abrogated by antibodies to NK1.1, expressed by both NK and NKT cells, but not in beta2m KO mice that lack NKT and CD8 T cells, but have a perfectly functional NK cell population. Furthermore, in IFN-gamma and IL-12p40 double KO mice, exogenous rIL-12 completely loses antitumor efficacy, suggesting the existence of an IFN-gamma-independent IL-12 effect that does require the presence of endogenous IL-12p40 chain.


Subject(s)
Colonic Neoplasms/therapy , Galactosylceramides/therapeutic use , Interleukin-12/therapeutic use , Liver Neoplasms, Experimental/drug therapy , Animals , Female , Interferon-gamma/biosynthesis , Interleukin-12/biosynthesis , Liver Neoplasms, Experimental/secondary , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Recombinant Proteins/therapeutic use , beta 2-Microglobulin/physiology
9.
Cancer Gene Ther ; 8(6): 459-68, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11498766

ABSTRACT

Phosphorothioate c-myc antisense oligodeoxynucleotides [S]ODNs (free INX-6295) were encapsulated in a new liposome formulation and the antitumor activity was compared to the unencapsulated antisense in a human melanoma xenograft. The systemic administration of INX-6295 encapsulated in stabilized antisense lipid particles (SALP INX-6295) improved plasma AUC (area under the plasma concentration-time curve) and initial half-life of free INX-6295, resulting in a significant enhancement in tumor accumulation and improvement in tumor distribution of antisense oligodeoxynucleotides. Animals treated with SALP INX-6295 exhibited a prolonged reduction of c-myc expression, reduced tumor growth and increased mice survival. When administered in combination with cisplatin (DDP), SALP INX-6295 produced a complete tumor regression in approximately 30% of treated mice, which persisted for at least 60 days following the first cycle of treatment. Finally, the median survival of mice treated with DDP/SALP INX-6295 increased by 105% compared to 84% for animals treated with the combination DDP/free INX-6295. These data indicate that the biological activity and the therapeutic efficacy of c-myc antisense therapy may be improved when these agents are administered in lipid-based delivery systems.


Subject(s)
Antineoplastic Agents/therapeutic use , Genes, myc/genetics , Melanoma/drug therapy , Oligonucleotides, Antisense , Animals , Area Under Curve , Blotting, Western , Cisplatin/pharmacology , Down-Regulation , Humans , Lipid Metabolism , Liposomes/metabolism , Male , Mice , Mice, Nude , Microscopy, Confocal , Neoplasm Transplantation , Time Factors , Tumor Cells, Cultured
10.
Cancer Res ; 61(17): 6532-9, 2001 Sep 01.
Article in English | MEDLINE | ID: mdl-11522651

ABSTRACT

We had demonstrated previously a functional bridge between altered homebox (HOX) gene expression and tumor progression through HOXB7 transactivation of basic fibroblast growth factor. Here, we have studied whether HOXB7, in addition to basic fibroblast growth factor, may induce other genes directly or indirectly related to neoangiogenesis and tumor invasion. Parental, beta-galactosidase-transduced, and HOXB7-transduced SkBr3 cell lines were examined for the expression of several growth factors and growth factor receptors involved in the proliferative and angiogenic processes. Vascular endothelial growth factor, melanoma growth-stimulatory activity/growth-related oncogenene alpha, interleukin-8, and angiopoietin-2 were up-regulated by HOXB7 transduction. The exception was angiopoietin-1 expression that was abrogated. Additional analyses included the expression levels of enzymes such as matrix metalloprotease (MMP)-2 and MMP-9 and heparanase, capable of proteolytic degradation of extracellular matrix and basement membranes. Results showed an induction of only MMP-9. The functional implication of such a finding was tested using an in vitro coculture assay in a three-dimensional matrix. A delay of differentiation with persistent nests of proliferating cells was found in endothelial cells cocultured with HOXB7-transduced SkBr3 cells. Tumorigenicity of these cells has been evaluated in vivo. Xenograft into athymic nude mice showed that SkBr3/HOXB7 cells developed tumors in mice, either irradiated or not, whereas parental SkBr3 cells did not show any tumor take unless mice were sublethally irradiated. Comparison of tumor nodules for vascularization by CD-31 and CD-34 immunostaining revealed an increased number of blood vessels in tumors expressing HOXB7. Together, the results indicate HOXB7 as a key factor up-regulating a variety of proangiogenic stimuli. Thus, HOXB7 gene or protein is a target to aim at to inhibit tumor-associated neoangiogenesis, considering the number and the redundancy of proangiogenic molecules that should be targeted one by one to theoretically achieve the same effect.


Subject(s)
Adenocarcinoma/blood supply , Breast Neoplasms/blood supply , Gene Expression Regulation, Neoplastic , Homeodomain Proteins/genetics , Neovascularization, Pathologic/genetics , Adenocarcinoma/genetics , Adenocarcinoma/metabolism , Angiopoietin-1 , Animals , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Coculture Techniques , Endothelial Growth Factors/biosynthesis , Endothelial Growth Factors/genetics , Endothelium, Vascular/cytology , Gene Expression Regulation, Enzymologic , Glucuronidase/biosynthesis , Glucuronidase/genetics , Humans , Lymphokines/biosynthesis , Lymphokines/genetics , Matrix Metalloproteinase 2/biosynthesis , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 9/biosynthesis , Matrix Metalloproteinase 9/genetics , Membrane Glycoproteins/biosynthesis , Membrane Glycoproteins/genetics , Mice , Mice, Nude , Neoplasm Transplantation , Neovascularization, Pathologic/metabolism , Protein Isoforms , Receptors, Growth Factor/biosynthesis , Receptors, Growth Factor/genetics , Reverse Transcriptase Polymerase Chain Reaction , Transduction, Genetic , Transplantation, Heterologous , Tumor Cells, Cultured , Up-Regulation , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth Factors
11.
J Clin Invest ; 107(9): 1173-81, 2001 May.
Article in English | MEDLINE | ID: mdl-11342581

ABSTRACT

Fractalkine (FKN, CX3CL1) is a membrane-bound CX3C chemokine induced by primary proinflammatory signals in vascular endothelial cells (ECs). Here we examined the role of FKN in polarized Th1 or Th2 responses. Proinflammatory signals, including LPS, IL-1, TNF, and CD40 ligand, induced FKN, as did IFN-gamma, which had synergistic activity with TNF. IL-4 and IL-13 did not stimulate the expression of FKN and markedly reduced induction by TNF and IFN-gamma. TNF alone or combined with IFN-gamma also induced release of soluble FKN, which was inhibited by IL-4 and IL-13. In light of this differential regulation of FKN by the master cytokines that control polarized responses, we analyzed the interaction of FKN with natural killer (NK) cells and polarized T-cell populations. NK cells expressed high levels of the FKN receptor CX3CR1 and responded to FKN. CX3CR1 was preferentially expressed in Th1 compared with Th2 cells. Th1 but not Th2 cells responded to FKN. By immunohistochemistry, FKN was expressed on ECs in psoriasis, a Th1-dominated skin disorder, but not in Th2-driven atopic dermatitis. Similarly, ECs in Mycobacterium tuberculosis granulomatous lymphadenitis, but not those in reactive lymph node hyperplasia or in Castelman's disease, showed immunoreactive FKN. These results indicate that regulated expression of FKN in ECs participates in an amplification circuit of polarized type I responses.


Subject(s)
Chemokines, CX3C/biosynthesis , Endothelium, Vascular/immunology , Killer Cells, Natural/immunology , Membrane Proteins/biosynthesis , Th1 Cells/immunology , Adult , CD40 Ligand/metabolism , CX3C Chemokine Receptor 1 , Castleman Disease/immunology , Chemokine CX3CL1 , Chemotaxis, Leukocyte , Dermatitis, Atopic/immunology , Endothelium, Vascular/drug effects , Humans , Infant, Newborn , Interferon-gamma/metabolism , Interleukin-13/metabolism , Interleukin-4/metabolism , Lymphadenitis/immunology , Psoriasis/immunology , Receptors, Cytokine/metabolism , Receptors, HIV/metabolism , Th2 Cells/immunology
12.
J Pathol ; 194(1): 4-8, 2001 May.
Article in English | MEDLINE | ID: mdl-11329134

ABSTRACT

In the last 10 years, evidence has accumulated that overexpression of Met protein is a distinguishing feature of almost every case of well-differentiated papillary carcinoma. Increased expression of the protein is probably due to enhanced transcription of the MET gene and/or to post-transcriptional mechanisms. So far, alterations of the MET gene have not been recognized, but evidence has been provided that activated RAS and RET can cause accumulation of MET RNA. Thus, the possibility exists that dysregulation of MET is the final result of different molecular pathways capable of inducing thyroid cell transformation; RET rearrangements might account for some of the cases, but the demonstration that the majority of papillary carcinomas do not have recognized alterations of the RET gene strongly suggests that MET gene dysregulation can also be achieved through other molecular pathways. Dysregulation of MET causes marked accumulation of Met protein in tumour cells that is promptly detected by immunohistochemistry. Thus, overexpression of Met protein might represent an immunohistochemical marker of papillary carcinoma, potentially helpful in problematic cases, but caution is required; moderate expression of Met protein is observed in non-neoplastic thyroid diseases, such as Graves' and Hashimoto's thyroiditis, and reagents active on paraffin sections may have a low affinity and/or low specificity for Met protein, leading to artifactual staining. Met protein-positive papillary carcinoma cells may produce hepatocyte growth factor (HGF) and may activate HGF through the urokinase-type plasminogen activator (uPA) bound to urokinase-type plasminogen activator receptor (uPA-R). Thus, papillary carcinoma cells possess the molecular machinery necessary for a productive HGF/Met interaction. In vitro studies have demonstrated that HGF enhances the motility and invasiveness of tumour cells and induces the synthesis and release of chemokines active in the recruitment of dendritic cells. These observations provide a rational basis for the understanding of two distinguishing features of papillary carcinoma. First, the tumour is often characterized by early metastatic spread to regional lymph nodes and by multifocal involvement of the gland, which suggests highly invasive behaviour. Second, a prominent peritumoural inflammatory reaction is often observed, which suggests cross-talk between tumour cells and the immune system.


Subject(s)
Biomarkers, Tumor/metabolism , Carcinoma, Papillary/metabolism , Hepatocyte Growth Factor/metabolism , Proto-Oncogene Proteins , Receptors, Growth Factor , Thyroid Neoplasms/metabolism , Trans-Activators/metabolism , Carcinoma, Papillary/pathology , Cell Transformation, Neoplastic/metabolism , Humans , Neoplasm Invasiveness , Neoplasm Proteins/metabolism , Proto-Oncogene Proteins c-met , Thyroid Neoplasms/pathology
13.
Eur Cytokine Netw ; 12(1): 177-86, 2001 Mar.
Article in English | MEDLINE | ID: mdl-11282563

ABSTRACT

The role of endogenous IL-1beta in regulating spontaneous and Fas-triggered apoptosis of human PMN has been studied in relation to the activity of the IL-1beta-generating enzyme ICE (caspase-1), an enzyme also involved in the mechanism of cell death. Upon in vitro culture, PMN undergo spontaneous apoptosis and express increasing levels of IL-1beta, caspase-1- and caspase-3-like enzymes. Endogenous IL-1beta protects PMN from apoptosis, since inhibition of either IL-1beta or caspase-1 activity can accelerate PMN apoptotic death. Thus, in spontaneous PMN apoptosis caspase-1 essentially plays an anti-apoptotic role by inducing maturation of protective IL-1beta, whereas other molecules are responsible of driving apoptosis. Upon Fas triggering, PMN apoptosis is greatly accelerated, in correlation with increased caspase activity, whereas IL-1beta production is not augmented. Inhibition of IL-1beta activity can increase Fas-induced apoptosis, whereas caspase-1 inhibitors are without significant effect. It is hypothesized that in Fas-induced PMN apoptosis caspase-1 has a double role: it can protect from apoptosis through generation of protective IL-1beta, as in spontaneous apoptosis, and it can also exert pro-apoptotic activity which counterbalances the protective effect and allows accelerated apoptosis.


Subject(s)
Apoptosis , Caspase 1/metabolism , Cell Survival , Interleukin-1/metabolism , Neutrophils/cytology , Adult , Apoptosis/physiology , Caspase Inhibitors , Humans , fas Receptor/physiology
14.
Eur J Immunol ; 31(3): 812-22, 2001 Mar.
Article in English | MEDLINE | ID: mdl-11241286

ABSTRACT

Macrophage-derived chemokine (MDC)/CCL22 is a CC chemokine active on dendritic cells (DC), NK cells and Th2 lymphocytes. The present study was aimed at comprehensively investigating MDC production in vitro and in vivo. DC were the most potent producers of MDC among leukocytes tested. Endothelial cells did not produce MDC under a variety of conditions. Signals that induce maturation (lipopolysaccharide, IL-1, TNF, CD40 ligand, recognition of bacteria and yeast) dramatically augmented MDC production, and dexamethasone and vitamin D3 blocked it. Prostaglandin E(2), which blocked the acquisition of IL-12 production and the capacity to promote Th1 generation, did not affect MDC production. Using mass spectrometry-based techniques, DC supernatants were found to contain N-terminally truncated forms of MDC [MDC(3-69), MDC(5-69) and MD(C7-69)] as well as the full-length molecule. In vivo, CD1a(+), CD83(+), MDC(+) DC were found in reactive lymph nodes, and in Langerhans' cell histiocytosis. Skin lesions of atopic dermatitis patients showed that CD1a(+) or CD1b(+) DC, and DC with a CD83(+) phenotype were responsible for MDC production in this Th2-oriented disorder. Thus, DC are the predominant source of MDC in vitro and in vivo under a variety of experimental and clinical conditions. Processing of MDC to MDC(3-69) and shorter forms which do not recognize CCR4 is likely to represent a feedback mechanism of negative regulation.


Subject(s)
Chemokines, CC/genetics , Dendritic Cells/immunology , Cells, Cultured , Chemokine CCL22 , Chemokines, CC/biosynthesis , Cholecalciferol/pharmacology , Chromatography, High Pressure Liquid , Dendritic Cells/drug effects , Dendritic Cells/metabolism , Dermatitis/immunology , Dermatitis/metabolism , Dexamethasone/pharmacology , Dinoprostone/pharmacology , Endocytosis , Histiocytosis, Langerhans-Cell/immunology , Histiocytosis, Langerhans-Cell/metabolism , Humans , Leukocytes/immunology , Leukocytes/metabolism , Lipopolysaccharides/pharmacology , Lymphatic Diseases/immunology , Lymphatic Diseases/metabolism , Mass Spectrometry , Monocytes/immunology , Protein Isoforms/biosynthesis , RNA, Messenger/biosynthesis , Transcriptional Activation/drug effects
15.
Cancer Res ; 61(1): 309-17, 2001 Jan 01.
Article in English | MEDLINE | ID: mdl-11196179

ABSTRACT

Fibroblast growth factor-2 (FGF2) is a pleiotropic heparin-binding growth factor endowed with a potent angiogenic activity in vitro and in vivo. To investigate the impact of the modulation of FGF2 expression on the neovascularization at different stages of tumor growth, we generated stable transfectants (Tet-FGF2) from the human endometrial adenocarcinoma HEC-1-B cell line in which FGF2 expression is under the control of the tetracycline-responsive promoter (Tet-off system). After transfection, independent clones were obtained in which FGF2 mRNA and protein were up-regulated compared with parental cells. Also, the conditioned medium of Tet-FGF2 transfectants caused proliferation, urokinase-type plasminogen activator up-regulation, migration, and sprouting of cultured endothelial cells. A 3-day treatment of Tet-FGF2 cell cultures with tetracycline abolished FGF2 overexpression and the biological activity of the conditioned medium without affecting their proliferative capacity. Tet-FGF2 cells formed tumors when nude mice received s.c. injections. The administration of 2.0 mg/ml tetracycline in the drinking water before cell transplantation, continued throughout the whole experiment, inhibited FGF2 expression in Tet-FGF2 tumor lesions. This was paralleled by a significant decrease in the rate of tumor growth and vascularization to values similar to those observed in lesions generated by parental HEC-1-B cells. Tetracycline administration 20 days after tumor cell implant, although equally effective in reducing FGF2 expression and inhibiting tumor vascularity, only minimally impaired the growth of established Tet-FGF2 tumors. The results indicate that FGF2 expression deeply affects the initial tumor growth and neovascularization of HEC-1-B human endometrial adenocarcinoma in nude mice. On the contrary, the growth of established tumors appears to be independent of the inhibition of FGF2 expression and decreased vascular density. The possibility that a significant reduction of angiogenesis may not affect the progression of large tumors points to the use of antiangiogenic therapy in early tumor stage.


Subject(s)
Adenocarcinoma/blood supply , Endometrial Neoplasms/blood supply , Fibroblast Growth Factor 2/biosynthesis , Neovascularization, Pathologic/metabolism , Tetracycline/pharmacology , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Animals , Cell Division/drug effects , Cell Division/physiology , DNA, Complementary/genetics , Endometrial Neoplasms/genetics , Endometrial Neoplasms/pathology , Female , Fibroblast Growth Factor 2/genetics , Gene Expression/drug effects , Genetic Vectors , Humans , Mice , Mice, Nude , Promoter Regions, Genetic , Time Factors , Transfection , Tumor Cells, Cultured , Xenograft Model Antitumor Assays
18.
EMBO J ; 19(13): 3337-48, 2000 Jul 03.
Article in English | MEDLINE | ID: mdl-10880446

ABSTRACT

The multiplicity of Notch receptors raises the question of the contribution of specific isoforms to T-cell development. Notch3 is expressed in CD4(-)8(-) thymocytes and is down-regulated across the CD4(-)8(-) to CD4(+)8(+) transition, controlled by pre-T-cell receptor signaling. To determine the effects of Notch3 on thymocyte development, transgenic mice were generated, expressing lck promoter-driven intracellular Notch3. Thymuses of young transgenics showed an increased number of thymocytes, particularly late CD4(-)8(-) cells, a failure to down-regulate CD25 in post-CD4(-)8(-) subsets and sustained activity of NF-kappaB. Subsequently, aggressive multicentric T-cell lymphomas developed with high penetrance. Tumors sustained characteristics of immature thymocytes, including expression of CD25, pTalpha and activated NF-kappaB via IKKalpha-dependent degradation of IkappaBalpha and enhancement of NF-kappaB-dependent anti-apoptotic and proliferative pathways. Together, these data identify activated Notch3 as a link between signals leading to NF-kappaB activation and T-cell tumorigenesis. The phenotypes of pre-malignant thymocytes and of lymphomas indicate a novel and particular role for Notch3 in co-ordinating growth and differentiation of thymocytes, across the pre-T/T cell transition, consistent with the normal expression pattern of Notch3.


Subject(s)
I-kappa B Proteins , Leukemia, T-Cell/metabolism , Lymphoma, T-Cell/metabolism , NF-kappa B/metabolism , Proto-Oncogene Proteins/physiology , Receptors, Cell Surface/physiology , Animals , Apoptosis/physiology , Base Sequence , CD4-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/cytology , Cell Survival , DNA Primers , DNA-Binding Proteins/physiology , Leukemia, T-Cell/pathology , Lymphoma, T-Cell/pathology , Mice , Mice, Transgenic , NF-KappaB Inhibitor alpha , NF-kappa B/antagonists & inhibitors , Proto-Oncogene Proteins/genetics , Receptor, Notch3 , Receptor, Notch4 , Receptors, Cell Surface/genetics , Receptors, Notch , Thymus Gland/pathology
19.
J Immunol ; 164(11): 5998-6004, 2000 Jun 01.
Article in English | MEDLINE | ID: mdl-10820283

ABSTRACT

Members of the Toll-like receptor (TLR) family probably play a fundamental role in pathogen recognition and activation of innate immunity. The present study used a systematic approach to analyze how different human leukocyte populations express specific transcripts for the first five characterized TLR family members. TLR1 was expressed in all leukocytes examined, including monocytes, polymorphonuclear leukocytes, T and B cells, and NK cells. In contrast TLR2, TLR4, and TLR5 were expressed in myelomonocytic elements. Exposure to bacterial products, such as LPS or lipoarabinomannan, or to proinflammatory cytokines increased TLR4 expression in monocytes and polymorphonuclear leukocytes, whereas IL-10 blocked this effect. TLR3 was only expressed in human dendritic cells (DC) wherein maturation induced by bacterial products or cytokines was associated with reduced expression. TLR3 mRNA expression was detected by in situ hybridization in DC and lymph nodes. These results demonstrate that TLR1 through TLR5 mRNAs are differentially expressed and regulated in human leukocytes. In particular, expression of TLR3 transcripts is restricted to DC that are the only elements which express the full TLR repertoire. These data suggest that TLR can be classified based on expression pattern as ubiquitous (TLR1), restricted (TLR2, TLR4, and TLR5 in myelomonocytic cells), and specific (TLR3 in DC) molecules.


Subject(s)
Dendritic Cells/metabolism , Drosophila Proteins , Leukocytes/metabolism , Membrane Glycoproteins/biosynthesis , Receptors, Cell Surface/biosynthesis , Cells, Cultured , Dendritic Cells/immunology , Humans , Leukocytes/immunology , Membrane Glycoproteins/classification , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Monocytes/immunology , Monocytes/metabolism , Neutrophils/immunology , Neutrophils/metabolism , RNA, Messenger/biosynthesis , Receptors, Cell Surface/classification , Receptors, Cell Surface/genetics , Receptors, Cell Surface/metabolism , Toll-Like Receptor 1 , Toll-Like Receptor 2 , Toll-Like Receptor 3 , Toll-Like Receptor 4 , Toll-Like Receptor 5 , Toll-Like Receptors , Transcription, Genetic
20.
Cancer Res ; 60(8): 2095-100, 2000 Apr 15.
Article in English | MEDLINE | ID: mdl-10786666

ABSTRACT

Chromosomal rearrangements observed in T-cell prolymphocytic leukemia involve the translocation of one T-cell receptor gene to either chromosome 14q32 or Xq28, deregulating the expression of cellular protooncogenes of unknown function, such as TCL1 or its homologue, MTCP1. In the human hematopoietic system, TCL1 expression is predominantly observed in developing B lymphocytes, whereas its overexpression in T cells causes mature T-cell proliferation in transgenic mice. In this study, using a newly generated monoclonal antibody against recombinant TCL1 protein, we extended our analysis mainly by immunohistochemistry and also by fluorescence-activated cell sorting and Western blot to a large tumor lymphoma data bank including 194 cases of lymphoproliferative disorders of B- and T-cell origin as well as reactive lymphoid tissues. The results obtained show that in reactive lymphoid tissues, TCL1 is strongly expressed by a subset of mantle zone B lymphocytes and is expressed to a lesser extent by follicle center cells and by scattered interfollicular small lymphocytes. In B-cell neoplasia, TCL1 was expressed in the majority of the cases, including lymphoblastic lymphoma, chronic lymphocytic leukemia, mantle cell lymphoma, follicular lymphoma, Burkitt lymphoma, diffuse large B-cell lymphoma (60%), and primary cutaneous B cell lymphoma (55%). TCL1 expression was observed in both the cytoplasmic and nuclear compartments, as confirmed by Western blot analysis. Conversely, TCL1 was not expressed in Hodgkin/Reed-Sternberg cells, multiple myelomas, marginal zone B-cell lymphomas, CD30+ anaplastic large cell lymphoma, lymphoblastic T-cell lymphoma, peripheral T-cell lymphoma, and mycosis fungoides. These data indicate that TCL1 is expressed in more differentiated B cells, under both reactive and neoplastic conditions, from antigen committed B cells and in germinal center B cells and is down-regulated in the latest stage of B-cell differentiation.


Subject(s)
DNA-Binding Proteins/metabolism , Gene Expression Regulation, Neoplastic , Lymphoma, B-Cell/metabolism , Lymphoma, T-Cell/metabolism , Proto-Oncogene Proteins , Pseudolymphoma/metabolism , Transcription Factors/metabolism , Aged , Antibodies, Monoclonal/immunology , Antibody Specificity/immunology , Blotting, Western , Cell Differentiation , Cell Nucleus/metabolism , Cytoplasm/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/immunology , Flow Cytometry , Hodgkin Disease/metabolism , Hodgkin Disease/pathology , Humans , Immunohistochemistry , Leukemia, Lymphoid/metabolism , Leukemia, Lymphoid/pathology , Leukemia, Prolymphocytic/metabolism , Leukemia, Prolymphocytic/pathology , Leukocytes, Mononuclear/metabolism , Lymphoid Tissue/metabolism , Lymphoid Tissue/pathology , Lymphoma, B-Cell/genetics , Lymphoma, B-Cell/pathology , Lymphoma, Large-Cell, Anaplastic/metabolism , Lymphoma, Large-Cell, Anaplastic/pathology , Lymphoma, T-Cell/genetics , Lymphoma, T-Cell/pathology , Pseudolymphoma/genetics , Pseudolymphoma/pathology , Transcription Factors/genetics , Transcription Factors/immunology
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