Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 46
Filter
1.
Clin Exp Immunol ; 203(3): 400-408, 2021 03.
Article in English | MEDLINE | ID: mdl-33020923

ABSTRACT

Rheumatoid arthritis (RA) is a systemic inflammatory autoimmune disease that leads to joint destruction and disability. Despite a significant progress in administration of biological agents for RA patients, there is still a need for improved therapy. Intravenous immunoglobulins (IVIG), a pooled polyspecific immunoglobulin (Ig)G extracted from 5000 to 20 000 healthy subjects, showed beneficial therapeutic effect in patients with immune deficiency, sepsis and autoimmune diseases. The current study aimed to investigate the beneficial effect of treatment with IVIG in established collagen-induced arthritis in DBA/1j mice. Murine arthritis was induced in DBA/1j mice. Treatment with IVIG began when the disease was established. The clinical score was followed twice a week until day 48. The mice were bled for plasma and the paws were hematoxylin and eosin (H&E)-stained. Cytokine profile in the plasma was analyzed by Luminex technology and titers of circulating anti-collagen antibodies in the plasma was tested by enzyme-linked immunosorbent assay. Our results show that treatment with IVIG in murine significantly reduced the clinical arthritis score (P < 0·001). Moreover, mode of action showed that IVIG significantly reduced circulating levels of inflammatory cytokines [interferon (IFN)-γ, interleukin (IL)-1ß, IL-17, IL-6, tumor necrosis factor (TNF)-α, P < 0·001], inhibiting anti-collagen antibodies (P < 0·001) in the plasma of collagen-induced arthritis mice. Importantly, histopathological examination revealed that IVIG treatment prevented the migration of inflammatory immune cells into the cartilage and synovium, reduced the extent of joint damage and preserved joint architecture. Our results proved for the first time the valuable anti-inflammatory treatment of IVIG in experimental RA. We propose IVIG therapy for a subgroup of patients with rheumatologically related diseases.


Subject(s)
Arthritis, Experimental/prevention & control , Arthritis, Rheumatoid/prevention & control , Cartilage/drug effects , Disease Models, Animal , Immunoglobulins, Intravenous/pharmacology , Synovial Membrane/drug effects , Animals , Arthritis, Experimental/immunology , Arthritis, Experimental/metabolism , Arthritis, Rheumatoid/immunology , Arthritis, Rheumatoid/metabolism , Cartilage/immunology , Cartilage/metabolism , Cytokines/blood , Humans , Immunoglobulins, Intravenous/administration & dosage , Immunoglobulins, Intravenous/immunology , Inflammation/immunology , Inflammation/metabolism , Inflammation/prevention & control , Inflammation Mediators/blood , Male , Mice, Inbred DBA , Neutrophil Infiltration/drug effects , Neutrophil Infiltration/immunology , Synovial Membrane/immunology , Synovial Membrane/metabolism
2.
J Laryngol Otol ; 133(8): 696-699, 2019 Aug.
Article in English | MEDLINE | ID: mdl-31290382

ABSTRACT

OBJECTIVE: To explore the use of fluorescence lifetime imaging microscopy in thyroid tissues, and to investigate how different thyroid lesions affect fluorescence lifetime. METHOD: Fluorescence lifetime measurements were taken of fresh frozen thyroid surgical specimens stained with fluorescein isothiocyanate tagged anti-thyroglobulin monoclonal antibodies. RESULTS: The mean fluorescence lifetime measurements in 12 patients - 3 with multinodular goitre, 4 with follicular adenoma, 4 with papillary thyroid carcinoma and 1 with follicular carcinoma - were 3.16 ns (range, 2.66-3.52 ns), 3.75 ns (range, 2.99-4.57 ns), 2.97 ns (range, 2.57-3.21 ns) and 3.61 ns, respectively. The fluorescence lifetime of follicular adenoma patients was higher than that of papillary thyroid carcinoma patients by 26 per cent (p = 0.058). The fluorescence lifetime in the follicular carcinoma patient was similar to the follicular adenoma group, but higher than in the papillary thyroid carcinoma group by 22 per cent (p = 0.01). CONCLUSION: Fluorescence lifetime measurements varied in different thyroid pathologies, possibly because of tissue-scale structural influences.


Subject(s)
Adenoma/diagnostic imaging , Goiter, Nodular/diagnostic imaging , Thyroid Gland/diagnostic imaging , Thyroid Neoplasms/diagnostic imaging , Adenoma/metabolism , Female , Fluorescein-5-isothiocyanate/pharmacology , Fluorescent Antibody Technique, Direct , Goiter, Nodular/metabolism , Humans , Male , Microscopy, Fluorescence , Thyroglobulin/antagonists & inhibitors , Thyroid Gland/pathology
3.
Clin Exp Immunol ; 193(2): 160-166, 2018 08.
Article in English | MEDLINE | ID: mdl-29698559

ABSTRACT

The role of helminth treatment in autoimmune diseases is growing constantly. Systemic lupus erythematosus (SLE) is a multi-system autoimmune disease with challenging treatment options. Tuftsin-phosphorylcholine (TPC) is a novel helminth-based compound that modulates the host immune network. This study was conducted to evaluate the potential value of TPC in ameliorating lupus nephritis in a murine model and specifically to compare the efficacy of TPC to the existing first-line therapy for SLE: corticosteroids (methylprednisolone). Lupus-prone NZBxW/F1 mice were treated with TPC (5 µg/mouse), methylprednisolone (MP; 5 mg/body weight) or phosphate-buffered saline (PBS) (control) three times per week once glomerulonephritis, defined as proteinuria of grade > 100 mg/dl, was established. Levels of anti-dsDNA autoantibodies were evaluated by enzyme-linked immunosorbent assay (ELISA), splenic cytokines were measured in vitro and the kidney microscopy was analysed following staining. TPC and MP treatments improved lupus nephritis significantly and prolonged survival in NZBxW/F1 mice. TPC-treated mice showed a significantly decreased level of proteinuria (P < 0·001) and anti-dsDNA antibodies (P < 0·001) compared to PBS-treated mice. Moreover, TPC and MP inhibited the production of the proinflammatory cytokines interferon IFN-γ, interleukin IL-1ß and IL-6 (P < 0·001) and enhanced expression of the anti-inflammatory cytokine IL-10 (P < 0·001). Finally, microscopy analysis of the kidneys demonstrated that TPC-treated mice maintained normal structure equally to MP-treated mice. These data indicate that the small molecule named TPC hinders lupus development in genetically lupus-prone mice equally to methylprednisolone in most of the cases. Hence, TCP may be employed as a therapeutic potential for lupus nephritis.


Subject(s)
Anti-Inflammatory Agents/therapeutic use , Helminths/immunology , Kidney/pathology , Lupus Erythematosus, Systemic/drug therapy , Lupus Nephritis/drug therapy , Phosphorylcholine/analogs & derivatives , Phosphorylcholine/therapeutic use , Tuftsin/therapeutic use , Animals , Antibodies, Antinuclear/blood , Cytokines/metabolism , Disease Models, Animal , Drug Combinations , Female , Humans , Inflammation Mediators/metabolism , Kidney/drug effects , Methylprednisolone/therapeutic use , Mice , Mice, Inbred NZB , Phosphorylcholine/chemistry , Tuftsin/chemistry
4.
Oncogene ; 37(2): 218-230, 2018 01 11.
Article in English | MEDLINE | ID: mdl-28925390

ABSTRACT

Skin carcinogenesis is known to be a multi-step process with several stages along its malignant evolution. We hypothesized that transformation of normal epidermis to cutaneous squamous cell carcinoma (cSCC) is causally linked to alterations in microRNAs (miRNA) expression. For this end we decided to evaluate their alterations in the pathologic states ending in cSCC. Total RNA was extracted from formalin fixed paraffin embedded biopsies of five stages along the malignant evolution of keratinocytes towards cSCC: Normal epidermis, solar elastosis, actinic keratosis KIN1-2, advanced actinic keratosis KIN3 and well-differentiated cSCC. Next-generation small RNA sequencing was performed. We found that 18 miRNAs are overexpressed and 28 miRNAs are underexpressed in cSCC compared to normal epidermis. miR-424, miR-320, miR-222 and miR-15a showed the highest fold change among the overexpressed miRNAs. And miR-100, miR-101 and miR-497 showed the highest fold change among the underexpressed miRNAs. Heat map of hierarchical clustering analysis of significantly changed miRNAs and principle component analysis disclosed that the most prominent change in miRNAs expression occurred in the switch from 'early' stages; normal epidermis, solar elastosis and early actinic keratosis to the 'late' stages of epidermal carcinogenesis; late actinic keratosis and cSCC. We found several miRNAs with 'stage specific' alterations while others display a clear 'gradual', either progressive increase or decrease in expression along the malignant evolution of keratinocytes. The observed alterations focused in miRNAs involved in the regulation of AKT/mTOR or in those involved in epithelial to mesenchymal transition. We chose to concentrate on the evaluation of the molecular role of miR-497. We found that it induces reversion of epithelial to mesenchymal transition. We proved that SERPINE-1 is its biochemical target. The present study allows us to further study the pathways that are regulated by miRNAs along the malignant evolution of keratinocytes towards cSCC.


Subject(s)
Carcinoma, Squamous Cell/genetics , Cell Transformation, Neoplastic/genetics , MicroRNAs/genetics , Plasminogen Activator Inhibitor 1/genetics , Skin Neoplasms/genetics , Aged , Aged, 80 and over , Biopsy , Carcinoma, Squamous Cell/pathology , Cell Line, Tumor , Disease Progression , Epidermal Cells , Epidermis/pathology , Epithelial-Mesenchymal Transition/genetics , Female , Gene Expression Regulation, Neoplastic , High-Throughput Nucleotide Sequencing , Humans , Keratinocytes/pathology , Male , Middle Aged , Plasminogen Activator Inhibitor 1/metabolism , Primary Cell Culture , Sequence Analysis, RNA , Skin Neoplasms/pathology
5.
Scand J Rheumatol ; 46(3): 222-225, 2017 May.
Article in English | MEDLINE | ID: mdl-27440169

ABSTRACT

OBJECTIVES: Temporal artery biopsy (TAB) is performed in patients suspected of giant cell arteritis (GCA). Inadequate TAB specimen length is considered a possible explanation for a negative biopsy in patients with GCA. We investigated the association between specimen length and diagnostic yield of TAB. METHOD: We conducted a retrospective analysis of 240 patients who underwent TAB in a single hospital between 2000 and 2015. Patients were diagnosed with GCA based on positive TAB or, when TAB was negative, on clinical grounds that fulfilled the American College of Rheumatology (ACR) 1990 criteria. Baseline clinical and laboratory features and TAB length were obtained from medical records. Among patients diagnosed with GCA, the rate of TAB positivity was calculated according to biopsy length (< 5, 5-9, 10-14, and ≥ 20 mm). RESULTS: Out of 240 patients, 88 were diagnosed with GCA: 62 had a positive TAB and 26 were diagnosed based on clinical grounds despite a negative TAB. Among those who were diagnosed with GCA, the length of the TAB specimen was similar in those with a positive and a negative TAB (1.13 ± 1.68 mm vs. 1.15 ± 0.61 mm, respectively, p = 0.928). The TAB positivity rate was similar among all ranges of biopsy length [< 5 mm: 7/10 (70%); 5-9 mm: 22/31 (71%); 10-14 mm: 11/16 (69%); 15-19 mm: 11/16 (69%); ≥ 20 mm: 11/15 (73%, p = ns] and was similar to the overall biopsy positivity rate. CONCLUSIONS: Specimen length is not associated with diagnostic yield of TAB.


Subject(s)
Giant Cell Arteritis/pathology , Temporal Arteries/pathology , Aged , Aged, 80 and over , Biopsy/methods , Female , Giant Cell Arteritis/diagnosis , Humans , Male , Middle Aged , Organ Size , Retrospective Studies
6.
Clin Exp Immunol ; 184(1): 19-28, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26618631

ABSTRACT

Treatment with helminthes and helminthes ova improved the clinical symptoms of several autoimmune diseases in patients and in animal models. Phosphorylcholine (PC) proved to be the immunomodulatory molecule. We aimed to decipher the tolerogenic potential of tuftsin-PC (TPC), a novel helminth-based compound in collagen-induced arthritis (CIA) a mouse model of rheumatoid arthritis (RA). CIA DBA/1 mice were treated with TPC subcutaneously (5 µg/0.1 ml) or orally (250 µg/0.1 ml), starting prior to disease induction. The control groups were treated with PBS. Collagen antibodies were tested by enzyme-linked immunosorbent assay (ELISA), cytokine protein levels by ELISA kits and regulatory T (Treg ) and regulatory B (Breg ) cell phenotypes by fluorescence-activated cell sorter (FACS). TPC-treated mice had a significantly lower arthritis score of 1.5 in comparison with control mice 11.8 (P < 0.0001) in both subcutaneous and orally treated groups at day 31. Moreover, histology analysis demonstrated highly inflamed joints in control mice, whereas TPC-treated mice maintained normal joint structure. Furthermore, TPC decreased the titres of circulating collagen II antibodies in mice sera (P < 0.0001), enhanced expression of IL-10 (P < 0.0001) and inhibited production of tumour necrosis factor (TNF)-α, interleukin (IL)-17 and IL-1ß (P < 0.0001). TPC significantly expanded the CD4(+) CD25(+) forkhead box protein 3 (FoxP3(+) ) Treg cells and CD19(+) IL-10(+) CD5(high) CD1d(high) T cell immunoglobulin mucin-1 (TIM-1(+) ) Breg cell phenotypes (P < 0.0001) in treated mice. Our data indicate that treatment with TPC attenuates CIA in mice demonstrated by low arthritic score and normal joints histology. TPC treatment reduced proinflammatory cytokines and increased anti-inflammatory cytokine expression, as well as expansion of Treg and Breg cells. Our results may lead to a new approach for a natural therapy for early rheumatoid arthritis onset.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Antibodies/blood , Arthritis, Experimental/drug therapy , Phosphorylcholine/pharmacology , Tuftsin/pharmacology , Administration, Oral , Animals , Arthritis, Experimental/immunology , Arthritis, Experimental/pathology , B-Lymphocytes, Regulatory/drug effects , B-Lymphocytes, Regulatory/immunology , B-Lymphocytes, Regulatory/pathology , CD4-Positive T-Lymphocytes/drug effects , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/pathology , Collagen Type II/blood , Collagen Type II/immunology , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/immunology , Gene Expression , Hepatitis A Virus Cellular Receptor 1 , Immunophenotyping , Injections, Subcutaneous , Interleukin-10/genetics , Interleukin-10/immunology , Interleukin-17/genetics , Interleukin-17/immunology , Interleukin-1beta/genetics , Interleukin-1beta/immunology , Joints/drug effects , Joints/immunology , Joints/pathology , Male , Membrane Proteins/genetics , Membrane Proteins/immunology , Mice , Mice, Inbred DBA , T-Lymphocytes, Regulatory/drug effects , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/pathology , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunology
7.
Oncogene ; 35(4): 438-47, 2016 Jan 28.
Article in English | MEDLINE | ID: mdl-25915843

ABSTRACT

Growth factors of the epidermal growth factor (EGF)/neuregulin family are involved in tumor progression and, accordingly, antibodies that intercept a cognate receptor, epidermal growth factor receptor (EGFR)/ERBB1, or a co-receptor, HER2, have been approved for cancer therapy. Although they might improve safety and delay onset of chemoresistance, no anti-ligand antibodies have been clinically approved. To identify suitable ligands, we surveyed fluids from ovarian and lung cancer patients and found that amphiregulin (AREG) is the most abundant and generalized ligand secreted by advanced tumors. AREG is a low affinity EGFR ligand, which is upregulated following treatment with chemotherapeutic drugs. Because AREG depletion retarded growth of xenografted ovarian tumors in mice, we generated a neutralizing monoclonal anti-AREG antibody. The antibody inhibited growth of ovarian cancer xenografts and strongly enhanced chemotherapy efficacy. Taken together, these results raise the possibility that AREG and other low- or high-affinity binders of EGFR might serve as potential targets for cancer therapy.


Subject(s)
Antibodies, Monoclonal/pharmacology , EGF Family of Proteins/genetics , EGF Family of Proteins/metabolism , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Amphiregulin , Animals , Antibodies, Monoclonal/immunology , Antineoplastic Agents/pharmacology , Culture Media, Conditioned/analysis , EGF Family of Proteins/immunology , ErbB Receptors/metabolism , Female , Gene Expression Regulation, Neoplastic/drug effects , Humans , Mice, Nude , Molecular Targeted Therapy/methods , Ovarian Neoplasms/genetics , Transforming Growth Factor alpha/metabolism , Transforming Growth Factor alpha/pharmacology , Tumor Cells, Cultured , Ubiquitination , Xenograft Model Antitumor Assays
8.
Clin Exp Immunol ; 182(3): 241-50, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26132809

ABSTRACT

Administration of intravenous immunoglobulin (IVIg) is a recognized safe and efficient immunomodulation therapy for many autoimmune diseases. Anti-idiotypic antibody binding to pathogenic autoantibodies was proposed as one of the mechanisms attributed to the protective activity of IVIg in autoimmunity. The aim of this study was to fractionate the anti-anti-citrullinated protein anti-idiotypic-antibodies (anti-ACPA) from an IVIg preparation and to test it as a treatment for collagen-induced arthritis in mice. IVIg was loaded onto an ACPA column. The eluted fraction was defined as ACPA-specific-IVIg (ACPA-sIVIg). Collagen-induced-arthritis (CIA) was induced in mice. Mice were treated weekly with ACPA-sIVIg, low-dose-IVIg, high-dose-IVIg and phosphate-buffered saline (PBS). Sera-ACPA titres, anti-collagen anitbodies and cytokine levels were analysed by enzyme-linked immunosorbent assay (ELISA); antibody-forming-cell activity by enzyme-linked imunospot (ELISPOT) assay; and expansion of regulatory T cell (Treg ) population by fluorescence activated cell sorter (FACS). ACPA-sIVIg inhibited ACPA binding to citrullinated-peptides (CCP) in vitro 100 times more efficiently than the IVIg compound. ACPA-sIVIg was significantly more effective than the IVIg-preparation in attenuating the development of collagen-induced arthritis. Splenocytes from CIA mice treated with ACPA-sIVIg reduced the ACPA and anti-collagen-antibody titres, including the number of anti-collagen and ACPA antibody-forming cells. In parallel, splenocytes from ACPA-sIVIg treated mice secreted higher levels of anti-inflammatory cytokines and lower proinflammatory cytokines. The ACPA-sIVIg inhibitory potential was accompanied with expansion of the Treg population. Low-dose IVIg did not affect the humoral and cellular response in the CIA mice in comparison to the PBS-treated mice. Based on our results, IVIg may be considered as a safe compound for treating patients with rheumatoid arthritis by neutralizing pathogenic autoantibodies, reducing proinflammatory cytokines and expanding the Treg population.


Subject(s)
Arthritis, Experimental/drug therapy , Arthritis, Experimental/immunology , Immunoglobulins, Intravenous/pharmacology , T-Lymphocytes, Regulatory/immunology , Animals , Antibodies, Anti-Idiotypic/blood , Antibodies, Anti-Idiotypic/immunology , Arthritis, Experimental/blood , Arthritis, Experimental/pathology , Immunoglobulins, Intravenous/immunology , Mice , Spleen/immunology , Spleen/metabolism , Spleen/pathology , T-Lymphocytes, Regulatory/metabolism
9.
Clin Exp Rheumatol ; 33(2 Suppl 89): S-84-9, 2015.
Article in English | MEDLINE | ID: mdl-26016755

ABSTRACT

OBJECTIVES: Temporal artery biopsy (TAB) is performed in cases of suspected giant cell arteritis (GCA), and is the gold-standard for diagnosis of the disease. Current American College of Rheumatology (ACR) classification criteria may aid in the diagnosis of GCA. We aimed to assess whether TAB is essential in all cases of suspected GCA, or whether ACR criteria can replace the need for this procedure in some cases. METHODS: Retrospective analysis of 216 patients who underwent TAB in a single hospital between 2000 and 2013. Pre-TAB and post-TAB ACR criteria were calculated. Sensitivity and specificity of ACR criteria for the diagnosis of GCA were assessed. RESULTS: Overall, 55 patients had histological evidence of GCA.Out of 161 patients with negative TAB findings, 34 were diagnosed with GCA, and 127 were not diagnosed with GCA. Sensitivity of TAB for the diagnosis of GCA was 61.7%. Sensitivity and specificity of ACR criteria for diagnosis of GCA before performing TAB were 68.5% and 58%, respectively. Sensitivity and specificity of ACR criteria after performing TAB biopsy were 89.8% and 64.5%, respectively. CONCLUSIONS: Temporal artery biopsy should be performed in the majority of patients with suspected GCA, and may be obviated only in patients with a pre-TAB ACR score of ≤ 1. In all other cases, when GCA is suspected, ACR criteria should not be a substitute to TAB, as they are not highly specific.


Subject(s)
Giant Cell Arteritis/pathology , Temporal Arteries/pathology , Age Factors , Aged , Aged, 80 and over , Biopsy , Blood Sedimentation , Cohort Studies , Female , Giant Cell Arteritis/complications , Giant Cell Arteritis/diagnosis , Headache/etiology , Humans , Male , Middle Aged , Retrospective Studies , Sensitivity and Specificity
10.
Br J Cancer ; 110(9): 2269-76, 2014 Apr 29.
Article in English | MEDLINE | ID: mdl-24667644

ABSTRACT

BACKGROUND: Challenges in developing drugs for pancreatic ductal adenocarcinoma (PDAC) include obtaining metastatic cancer tissue for research and validating biomarkers predicative for personalised therapeutic decisions. We have recently developed a novel therapeutic model for PDAC to address these challenges based on the isolation of viable PDAC cells derived from ascites fluid. METHODS: Ascites fluid was obtained from PDAC patients undergoing palliative paracentesis. Ascites-derived PDAC primary cells were isolated, cultured and characterised in ovo and in vitro. RESULTS: We successfully established ascites-derived primary cell cultures within 2-7 days from 92% (93 out of 101) of the ascites fluid samples obtained (from 36 different patients). Homogeneous epithelial PDAC-enriched cell cultures were identified and characterised. We observed a wide range in doubling times and migration properties among the different patient-derived cell cultures. The diverse nature of each individual patient's cell cultures was further demonstrated by differences in therapeutic susceptibility and resistance. The tumorigenicity and invasiveness of the cells were demonstrated in vivo using chicken chorioallantoic membrane grafts. CONCLUSIONS: We have developed a unique ascites-derived PDAC primary cell culture model. This model has the potential to study signalling pathways in PDAC progression and to evaluate targeted therapies for the individual patient expeditiously, thereby supporting personalised treatment decisions.


Subject(s)
Ascites/pathology , Ascitic Fluid/pathology , Carcinoma, Pancreatic Ductal/metabolism , Pancreatic Neoplasms/pathology , Precision Medicine , Primary Cell Culture/methods , Xenograft Model Antitumor Assays , Animals , Cell Separation , Chick Embryo , Chorioallantoic Membrane , Epithelial-Mesenchymal Transition , Humans , Molecular Targeted Therapy , Pancreatic Neoplasms
11.
Cell Death Differ ; 18(2): 271-81, 2011 Feb.
Article in English | MEDLINE | ID: mdl-20689556

ABSTRACT

A mutation within one allele of the p53 tumor suppressor gene can inactivate the remaining wild-type allele in a dominant-negative manner and in some cases can exert an additional oncogenic activity, known as mutant p53 'gain of function' (GOF). To study the role of p53 mutations in prostate cancer and to discriminate between the dominant-negative effect and the GOF activity of mutant p53, we measured, using microarrays, the expression profiles of three immortalized prostate epithelial cultures expressing wild-type, inactivated p53 or mutated p53. Analysis of these gene expression profiles showed that both inactivated p53 and p53(R175H) mutant expression resulted in the upregulation of cell cycle progression genes. A second group, which was upregulated exclusively by mutant p53(R175H), was predominantly enriched in developmental genes. This group of genes included the Twist1, a regulator of metastasis and epithelial-mesenchymal transition (EMT). Twist1 levels were also elevated in metastatic prostate cancer-derived cell line DU145, in immortalized lung fibroblasts and in a subset of lung cancer samples, all in a mutant p53-dependent manner. p53(R175H) mutant bearing immortalized epithelial cells showed typical features of EMT, such as higher expression of mesenchymal markers, lower expression of epithelial markers and enhanced invasive properties in vitro. The mechanism by which p53(R175H) mutant induces Twist1 expression involves alleviation of the epigenetic repression. Our data suggest that Twist1 expression might be upregulated following p53 mutation in cancer cells.


Subject(s)
Epithelial-Mesenchymal Transition , Nuclear Proteins/metabolism , Prostatic Neoplasms/metabolism , Tumor Suppressor Protein p53/metabolism , Twist-Related Protein 1/metabolism , Amino Acid Substitution , Cell Line, Transformed , Cell Line, Tumor , Epigenesis, Genetic , Histones/metabolism , Humans , Male , Mutation , Nuclear Proteins/genetics , Polycomb Repressive Complex 1 , Promoter Regions, Genetic , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , Proto-Oncogene Proteins/metabolism , Repressor Proteins/metabolism , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/physiology , Twist-Related Protein 1/genetics , Up-Regulation
12.
Clin Exp Immunol ; 162(3): 543-9, 2010 Dec.
Article in English | MEDLINE | ID: mdl-20964642

ABSTRACT

Pemphigus vulgaris is a rare life-threatening autoimmune bullous disease caused by immunoglobulin G (IgG) autoantibodies directed against desmogleins 1 and 3. Previously, we showed that intravenous immunoglobulin (IVIG) ameliorates anti-desmoglein-induced experimental pemphigus vulgaris in newborn naive mice. The aim of this study was to examine the efficacy of anti-anti-desmoglein-specific IVIG in a similar model. Pemphigus-vulgaris-specific IVIG (PV-sIVIG) was affinity-purified from IVIG on a column of single-chain variable fragment (scFv) anti-desmogleins 1 and 3. The anti-idiotypic activity of PV-sIVIG was confirmed by enzyme-linked immunosorbent assay, inhibition assay. After induction of pemphigus by injection of anti-desmogleins 1 and 3 scFv to newborn mice, the animals were treated with PV-sIVIG, IVIG (low or high dose) or IgG from a healthy donor (n = 10 each). The skin was examined 24-48 h later, and samples of affected areas were analysed by histology and immunofluorescence. In vitro study showed that PV-sIVIG significantly inhibited anti-desmogleins 1 and 3 scFv binding to recombinant desmoglein-3 in a dose-dependent manner. Specificity was confirmed by inhibition assay. In vivo analysis revealed cutaneous lesions of pemphigus vulgaris in mice injected with normal IgG (nine of 10 mice) or low-dose IVIG (nine of 10 mice), but not in mice treated with PV-sIVIG (none of 10) or high-dose IVIG (none of 10). On immunopathological study, PV-sIVIG and regular IVIG prevented the formation of acantholysis and deposition of IgG in intercellular spaces. In conclusion, the PV-sIVIG preparation is more effective than native IVIG in inhibiting anti-desmoglein-induced pemphigus vulgaris in mice and might serve as a future therapy in patients with the clinical disease.


Subject(s)
Antibodies, Anti-Idiotypic/administration & dosage , Immunoglobulins, Intravenous/administration & dosage , Pemphigus/drug therapy , Single-Chain Antibodies/metabolism , Skin/drug effects , Acantholysis/prevention & control , Animals , Animals, Newborn , Antibodies, Anti-Idiotypic/adverse effects , Antibodies, Anti-Idiotypic/immunology , Antibodies, Anti-Idiotypic/isolation & purification , Autoantibodies/administration & dosage , Autoantibodies/immunology , Autoantibodies/metabolism , Desmoglein 1/immunology , Desmoglein 3/immunology , Disease Models, Animal , Epitopes , Humans , Immunoglobulins, Intravenous/adverse effects , Immunoglobulins, Intravenous/immunology , Immunoglobulins, Intravenous/isolation & purification , Mice , Mice, Inbred C57BL , Pemphigus/immunology , Pemphigus/physiopathology , Protein Engineering , Single-Chain Antibodies/administration & dosage , Single-Chain Antibodies/genetics , Skin/immunology , Skin/metabolism , Skin/pathology
13.
Clin Exp Immunol ; 161(2): 276-83, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20456418

ABSTRACT

Eotaxin-2 is a potent chemoattractant for eosinophils, basophils and T helper type 2 (Th2) lymphocytes. The eotaxin-2/CCL24 receptor CCR3 is expressed in human brain, skin, endothelium and macrophages. The aim of the current study was to evaluate the protective effect of a monoclonal anti-eotaxin-2 antibody on the development of adjuvant-induced arthritis in rats (AIA). Adjuvant arthritis was induced in Lewis rats by intradermal injection of incomplete Freund's adjuvant +Mycobacterium tuberculosis. Rats were treated by intraperitoneal (i.p.) injection with three monoclonal antibodies against eotaxin-2 (G7, G8, D8) three times per week. Controls were treated with total mouse immunoglobulin G (IgG), methotrexate (MTX) or phosphate-buffered saline (PBS). Arthritis severity was evaluated by measuring ankle swelling, arthritic score, whole animal mobility and body weight. Sample joints were obtained for pathological evaluation and postmortem X-ray of ankle joints was performed to document erosions. Significant inhibition of arthritis was observed in rats treated with anti-eotaxin-2 antibodies compared to those treated with immunoglobulin or PBS. Inhibition was manifest in ankle diameter, arthritic score and mobility score. The antibody marked D8 showed the greatest efficacy. The effect was observed both in animals treated before the appearance of arthritis and in those where treatment was begun after development of joint inflammation. Combined treatment with D8 and MTX caused additional protection. Significant reduction of inflammation in D8-treated animals was also demonstrated in pathological and X-ray examinations. Inhibition of eotaxin-2 by monoclonal antibodies has a significant protective effect in adjuvant arthritis. These results may introduce a novel therapeutic target in rheumatoid arthritis and additional inflammatory joint disorders.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Arthritis, Experimental/drug therapy , Arthritis, Experimental/prevention & control , Chemokine CCL24/antagonists & inhibitors , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/pharmacology , Arthritis, Experimental/diagnosis , Arthritis, Experimental/pathology , Arthrography , Body Weight/drug effects , Chemokine CCL24/immunology , Dose-Response Relationship, Drug , Drug Therapy, Combination/methods , Freund's Adjuvant/administration & dosage , Freund's Adjuvant/immunology , Joints/drug effects , Joints/pathology , Locomotion/drug effects , Male , Methotrexate/administration & dosage , Methotrexate/pharmacology , Methotrexate/therapeutic use , Rats , Rats, Inbred Lew , Tarsus, Animal/drug effects , Tarsus, Animal/pathology
15.
Gut ; 58(3): 396-403, 2009 Mar.
Article in English | MEDLINE | ID: mdl-18832521

ABSTRACT

OBJECTIVE: The onset of the effect of thiopurines is delayed for several months. The aim of this study was to investigate immune mechanisms for this delay. METHODS: The effects of thiopurines on human peripheral blood T cells and on lamina propria lymphocytes were investigated for apoptosis induction by Annexin V/propidium iodide (PI) and for cytokine secretion by intracellular staining and ELISA assays. To investigate the mechanism of the effect of thiopurines in vivo, Balb/C mice were co-immunised with HEL/OVA (hen egg lysozyme/ovalbumin) antigens, and then repeatedly challenged by HEL only, while being treated by mercaptopurine or vehicle alone for either 4 or 20 weeks. The memory response of CD4+ splenocytes towards HEL/OVA was then determined by CFSE (carboxyfluorescein succinimidyl ester) dilution. RESULTS: Thiopurines arrested the proliferation of stimulated T cells but did not enhance the apoptosis of either resting T cells or activated T cells until day 5 poststimulation. Despite the proliferation arrest, stimulated T cells successfully differentiated into effector cells, as evidenced by their capacity for proinflammatory cytokine secretion, potent adhesion and cytotoxicity. Prolonged mercaptopurine treatment of mice for 20 weeks selectively reduced the CD4+ memory response to a repeatedly encountered HEL antigen, but did not affect the T cell memory pool to the previously presented OVA antigen. A shorter, 4 weeks, treatment with mercaptopurine did not inhibit the memory response to either antigen. CONCLUSIONS: T cells arrested from cycling by thiopurines can still differentiate into potent effector cells capable of propagating the inflammatory process. Thiopurine treatment results in depletion of antigen-specific memory T cells, but this effect is dependent upon repeated encounters with the antigen over a prolonged time course.


Subject(s)
Apoptosis/drug effects , Azathioprine/therapeutic use , Caspases, Effector/drug effects , Immunosuppressive Agents/therapeutic use , Inflammatory Bowel Diseases/drug therapy , T-Lymphocytes/drug effects , Animals , Apoptosis/immunology , Caspases, Effector/immunology , Cell Proliferation/drug effects , Drug Design , Female , Humans , Immunologic Memory , Inflammatory Bowel Diseases/immunology , Male , Mice , Mice, Inbred BALB C , Models, Biological , T-Lymphocytes/immunology
16.
Pathobiology ; 75(6): 346-55, 2008.
Article in English | MEDLINE | ID: mdl-19096230

ABSTRACT

BACKGROUND: One strategy to increase tissue specificity of gene therapy is to use promoters or enhancers. OBJECTIVES: (1) To enhance the selectivity of a murine preproendothelin-1 (PPE-1) promoter in tumor angiogenesis by using a positive endothelial transcription-binding element. (2) To test the specificity and efficiency of the modified PPE-1 promoter [PPE-1(3X)] in vitro and in vivo by using reporter genes, and the therapeutic gene herpes simplex virus-thymidine kinase (HSV-TK) in a mouse model of Lewis lung carcinoma (LLC). RESULTS: The modified PPE-1 promoter specifically induced expression in the tumor angiogenic vascular bed with a 35-fold higher expression compared to the normal vasculare bed of the lung. Thus, when the HSV-TK gene controlled by the modified PPE-1 promoter was used systemically, it induced tumor-specific necrosis, apoptosis and mononuclear infiltrates, leading to massive destruction of the neovasculature of the pulmonary metastasis, which suppressed metastasis development. CONCLUSIONS: These results show that an adenoviral vector armed with HSV-TK controlled by the endothelial-selective murine PPE-1(3X) promoter is efficient and safe to target tumor neovasculature.


Subject(s)
Carcinoma, Lewis Lung/therapy , Endothelin-1/genetics , Genetic Therapy/methods , Neovascularization, Pathologic/therapy , Promoter Regions, Genetic , Simplexvirus/enzymology , Thymidine Kinase/genetics , Adenoviridae/genetics , Animals , Carcinoma, Lewis Lung/blood supply , Endothelium, Vascular/metabolism , Genes, Viral/genetics , Genetic Vectors , Lung/blood supply , Male , Mice , Mice, Inbred C57BL , Simplexvirus/genetics , Thymidine Kinase/metabolism
17.
Cancer Gene Ther ; 15(8): 535-42, 2008 Aug.
Article in English | MEDLINE | ID: mdl-18421310

ABSTRACT

Ad-PPE-Fas-c is an adenovector that expresses Fas-c under the control of the modified pre-proendothelin-1 (PPE-1) promoter. Fas-c is a chimeric death receptor containing the extracellular portion of tumour necrosis factor 1 receptor (TNFR1) and the transmembrane and intracellular portion of Fas. We recently demonstrated that Ad-PPE-Fas-c induced Fas-receptor-mediated endothelial cell apoptosis. Previously, doxorubicin was shown to enhance Fas-receptor clustering and the induction of its cascade. Therefore, the goal of this work was to test whether doxorubicin augments the capacity of Ad-PPE-Fas-c to induce endothelial cell apoptosis and to elucidate whether either the death-receptor-mediated apoptotic cascade or the mitochondria-associated apoptotic cascade is involved in the combined treatment effect. We found that a combined treatment of Ad-PPE-Fas-c and doxorubicin synergistically induced a reduction in endothelial cell viability and apoptosis. z-IETD-FMK, a caspase-8 inhibitor, and z-LEHD-FMK, a caspase-9 inhibitor, significantly decreased apoptosis induced by the combined treatment. Systemically administered combined therapy significantly reduced the lung metastases burden (70%) in mice as compared to each treatment alone. Thus, a combined treatment of Ad-PPE-Fas-c gene therapy and chemotherapy may be effective in the treatment of metastatic diseases and both the Fas cascade and the mitochondria-associated cascade are essential for this effect.


Subject(s)
Apoptosis/drug effects , Caspase 8/metabolism , Caspase 9/metabolism , Doxorubicin/administration & dosage , Endothelium, Vascular/drug effects , Genetic Therapy , Neoplasm Metastasis/prevention & control , Neovascularization, Pathologic/prevention & control , Adenoviridae/genetics , Animals , Cattle , Cells, Cultured , Endothelium, Vascular/cytology , Genetic Vectors , Mice , Mice, Inbred C57BL
18.
Scand J Immunol ; 60(5): 529-34, 2004 Nov.
Article in English | MEDLINE | ID: mdl-15541047

ABSTRACT

TCRDV1-positive lymphocytes, which infiltrate colon carcinomas, were recently shown to be cytolytic for tumour cells. However, the immune compartment from which these cells originate is unknown. The aim of the present studies was to determine the origin of TCRDV1-positive cells in colonic neoplasia. Biopsies were obtained from normal colon, polyps or carcinomas, concurrently with a sample from the peripheral blood. RNA was extracted and a TCRDV1-specific reverse transcriptase-polymerase chain reaction (RT-PCR) was performed. Amplification products were analysed by a CDR3 display and sequence analysis. In five out of six patients, the TCRDV1 CDR3 display of the whole cell population within the neoplastic tissue was distinct from that in the normal mucosa and the peripheral blood. The nucleotide sequences of CDR3 domains from the three compartments were distinct as well. In one patient, a pattern similar to the CDR3 display was detected in neoplastic and normal intestinal tissues. However, using junction-specific RT-PCR of CDR3 sequences derived from the neoplastic cells, such sequences could be detected in all three compartments. These findings suggest that in contrast to the current paradigm, a unique TCRDV1 cell population circulates in the peripheral blood and normal intestinal tissue and infiltrates colon neoplasia rather than being restricted to a single compartment as previously thought.


Subject(s)
Carcinoma/immunology , Colonic Neoplasms/immunology , Intestinal Mucosa/immunology , Lymphocytes/immunology , Receptors, Antigen, T-Cell/immunology , Complementarity Determining Regions/genetics , Complementarity Determining Regions/immunology , Humans , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Antigen, T-Cell/genetics
19.
J Clin Pathol ; 57(7): 712-6, 2004 Jul.
Article in English | MEDLINE | ID: mdl-15220363

ABSTRACT

BACKGROUND: Hirschsprung's disease (HD) is a congenital disorder characterised by the absence of ganglion cells in the large bowel, leading to functional obstruction and colonic dilatation proximal to the affected segment. A subclass of nerve cell bodies in both submucosa and myenteric ganglia of the human gastrointestinal tract were found to show immunopositivity for calretinin, a calcium binding protein, which plays an important role in the organisation and functioning of the central nervous system. AIM: To investigate calretinin immunostaining in ganglionic and aganglionic HD colon specimens, and compare it with staining for S100, neurone specific enolase, and c-kit. METHODS: Ten large bowel, full thickness specimens from patients with classic rectosigmoid HD were selected from the pathology repository. In total, 54 paraffin wax blocks-24 from the ganglionic zone, 17 from the aganglionic zone, and 13 from the transitional zone-were processed. RESULTS: Calretinin was not expressed in aganglionic segments of HD and associated nerve fibres, whereas in ganglionic HD segments and in normal colon both ganglion cells and nerve fibres were immunopositive. In addition, c-kit showed an altered distribution in the interstitial cells of Cajal. The transitional zone showed a broad spectrum of histomorphological and immunohistochemical patterns of both calretinin and c-kit expression. CONCLUSION: The absence of calretinin expression may serve as a diagnostic aid in identifying aganglionic segments in HD.


Subject(s)
Hirschsprung Disease/diagnosis , S100 Calcium Binding Protein G/metabolism , Adult , Biomarkers/analysis , Calbindin 2 , Child, Preschool , Colon/innervation , Colon/metabolism , Colon/pathology , Ganglia/metabolism , Hirschsprung Disease/metabolism , Hirschsprung Disease/pathology , Humans , Infant , Infant, Newborn , Nerve Fibers/metabolism , Nerve Fibers/pathology , Phosphopyruvate Hydratase/analysis , Proto-Oncogene Proteins c-kit/analysis , S100 Proteins/analysis
20.
Int J Gynecol Cancer ; 14(2): 251-8, 2004.
Article in English | MEDLINE | ID: mdl-15086724

ABSTRACT

OBJECTIVE: To analyze the somatic pattern of p53 expression and BRCA germline mutation status in Israeli patients with both ovarian (OvCa) and breast cancer (BrCa). METHODS: The study group comprised 43 Israeli patients with OvCa, all of whom had previous primary BrCa. p53 immunohistochemistry (IHC) on all available archival tissues and genotyping for the three predominant Jewish germline BRCA1-2 mutations were carried out. Samples from 64 patients with solitary OvCa and 61 with solitary BrCa were similarly analyzed as controls. RESULTS: p53 expression pattern and the immunopositivity rate were similar in the ovarian and breast tumors within the study group and in the two control groups: positive p53 staining was detected in 68% of ovarian tumors in the study group compared with 71.9% in the controls, and in 19.4% of the BrCa tissues versus 21.3% in the controls. Within the study group, advanced stage OvCa had a higher rate of p53 expression (84%) compared to early stage disease (38.5%) (P = 0.006). This difference was not apparent in the solitary OvCa control group. OvCa in BRCA1-2 mutation carriers from the study group were more likely to display positive p53 staining (79%), especially in tumors diagnosed before the age of 60 (90%) compared with the OvCa of noncarriers (60%), but this difference was statistically insignificant. The p53 expression rate in BrCa samples from the study group was not associated with BRCA1-2 mutation status. CONCLUSIONS: Positive p53 expression, detected by IHC, in OvCa patients with previous primary BrCa is significantly higher in advanced stage disease in BRCA1-2 mutation carriers. There is a higher positive p53 expression somatically in OvCa in BRCA1-2 carriers in whom OvCa was diagnosed before the age of 60 years, although this trend is not statistically significant. These observations suggest that somatic p53 inactivation may be an important event in ovarian tumorigenesis in this subset of patients.


Subject(s)
Breast Neoplasms/genetics , Genes, BRCA1 , Genes, p53/genetics , Neoplasms, Multiple Primary/genetics , Ovarian Neoplasms/genetics , Aged , Breast Neoplasms/pathology , Case-Control Studies , Cystadenocarcinoma, Serous/genetics , Cystadenocarcinoma, Serous/pathology , Female , Gene Expression Regulation, Neoplastic , Genotype , Humans , Immunohistochemistry , Israel , Jews/genetics , Middle Aged , Mutation , Neoplasms, Multiple Primary/pathology , Neoplastic Syndromes, Hereditary/genetics , Neoplastic Syndromes, Hereditary/pathology , Ovarian Neoplasms/pathology , White People/genetics
SELECTION OF CITATIONS
SEARCH DETAIL
...