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1.
Mol Hum Reprod ; 23(3): 187-198, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28158750

RESUMO

Study Question: Can endometrial mesenchymal stromal cells (E-MSCs) differentiate into endothelial cells in an in vitro co-culture system with human umbilical vein endothelial cells (HUVECs)? Summary Answer: E-MSCs can acquire endothelial markers and function in a direct co-culture system with HUVECs. What is Known Already: E-MSCs have been identified in the human endometrium as well as in endometriotic lesions. E-MSCs appear to be involved in formation of the endometrial stromal vascular tissue and the support of tissue growth and vascularization. The use of anti-angiogenic drugs appears as a possible therapeutic strategy against endometriosis. Study Design, Size, Duration: This is an in vitro study comprising patients receiving surgical treatment of ovarian endometriosis (n = 9). Participants/Materials, Setting, Methods: E-MSCs were isolated from eutopic and ectopic endometrial tissue and were characterized for the expression of mesenchymal and endothelial markers by FACS analysis and Real-Time PCR. CD31 acquisition was evaluated by FACS analysis and immunofluorescence after a 48 h-direct co-culture with green fluorescent protein +-HUVECs. A tube-forming assay was set up in order to analyze the functional potential of their interaction. Finally, co-cultures were treated with the anti-angiogenic agent Cabergoline. Main Results and the Role of Chance: A subpopulation of E-MSCs acquired CD31 expression and integrated into tube-like structures when directly in contact with HUVECs, as observed by both FACS analysis and immunofluorescence. The isolation of CD31+ E-MSCs revealed significant increases in CD31, vascular endothelial growth factor receptor 2, TEK receptor tyrosine kinase and vascular endothelial-Cadherin mRNA expression levels with respect to basal and to CD31neg cells (P < 0.05). On the other hand, the expression of mesenchymal genes such as c-Myc, Vimentin, neuronal-Cadherin and sushi domain containing 2 remained unchanged. Cabergoline treatment induced a significant reduction of the E-MSC angiogenic potential (P < 0.05 versus control). Large Scale Data: Not applicable. Limitations, Reasons for Caution: Further studies are necessary to investigate the cellular and molecular mechanisms underlying the endothelial cell differentiation. Wider Implications of the Findings: E-MSCs may undergo endothelial differentiation, and be potentially involved in the development of endometriotic implants. Cell culture systems that more closely mimic the cellular complexity typical of endometriotic tissues in vivo are required to develop novel strategies for treatment. Study Funding/Competing Interest(s): This study was supported by the 'Research Fund ex-60%', University of Turin, Turin, Italy. All authors declare that their participation in the study did not involve actual or potential conflicts of interests.


Assuntos
Endometriose/patologia , Células-Tronco Mesenquimais/patologia , Modelos Biológicos , Neovascularização Patológica/patologia , Inibidores da Angiogênese/farmacologia , Biomarcadores/metabolismo , Cabergolina , Caderinas/genética , Caderinas/metabolismo , Comunicação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Endometriose/genética , Endometriose/metabolismo , Endometriose/cirurgia , Endométrio/metabolismo , Endométrio/patologia , Endométrio/cirurgia , Ergolinas/farmacologia , Feminino , Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Sindecana-2/genética , Sindecana-2/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Vimentina/genética , Vimentina/metabolismo
2.
Br J Cancer ; 108(12): 2525-36, 2013 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-23801032

RESUMO

BACKGROUND: Bone metastases represent a common and severe complication in breast cancer, and the involvement of cancer stem cells (CSCs) in the promotion of bone metastasis is currently under discussion. Here, we used a human-in-mice model to study bone metastasis formation due to primary breast CSCs-like colonisation. METHODS: Primary CD44⁺CD24⁻ breast CSCs-like were transduced by a luciferase-lentiviral vector and injected through subcutaneous and intracardiac (IC) routes in non-obese/severe-combined immunodeficient (NOD/SCID) mice carrying subcutaneous human bone implants. The CSCs-like localisation was monitored by in vivo luciferase imaging. Bone metastatic CSCs-like were analysed through immunohistochemistry and flow cytometry, and gene expression analyses were performed by microarray techniques. RESULTS: Breast CSCs-like colonised the human-implanted bone, resulting in bone remodelling. Bone metastatic lesions were histologically apparent by tumour cell expression of epithelial markers and vimentin. The bone-isolated CSCs-like were CD44⁻CD24⁺ and showed tumorigenic abilities after injection in secondary mice. CD44⁻CD24⁺ CSCs-like displayed a distinct bone tropism signature that was enriched in genes that discriminate bone metastases of breast cancer from metastases at other organs. CONCLUSION: Breast CSCs-like promote bone metastasis and display a CSCs-like bone tropism signature. This signature has clinical prognostic relevance, because it efficiently discriminates osteotropic breast cancers from tumour metastases at other sites.


Assuntos
Neoplasias Ósseas/secundário , Osso e Ossos/metabolismo , Neoplasias da Mama/patologia , Carcinoma/patologia , Células-Tronco Neoplásicas/patologia , Transcriptoma , Adulto , Animais , Neoplasias Ósseas/genética , Osso e Ossos/patologia , Neoplasias da Mama/genética , Carcinoma/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Genes de Troca/genética , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Células-Tronco Neoplásicas/metabolismo , Especificidade de Órgãos/genética , Fenótipo , Transcriptoma/fisiologia
3.
J Dairy Sci ; 96(3): 1745-54, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23332842

RESUMO

Products of different origin, time of collection, and activities fall under the general term of colostrum and, therefore, great variability in composition as well as in the concentration of its components has been reported in the literature. In the present study, we describe the standardization of a bovine colostrum derivative and the characterization of its bioactive components. Evaluation of the most representative agents (lactoferrin, transferrin, IL-2, IFN-γ, tumor necrosis factor, IgG, and IgA) showed that a marked decrease in active components occurs after the first few hours. Bovine colostrum was, therefore, collected up to the fifth hour after delivery from Holstein cows, in the presence of preservatives, and immediately frozen. A protocol of centrifugation, filtration, and lyophilization was then applied to pools of colostrum from at least 30 cows to obtain a stable, sterile, standardized product. Preservatives were removed by dialysis. Evaluation of the active biological components of colostrum showed that the final product of colostrums contained significant and reproducible amounts of bioactive factors, including cytokines, immunomodulating factors, growth factors, and immunoglobulins. The final product appeared, therefore, as a sterile, pyrogen-free, standardized derivative of bovine colostrum with a high concentration of bioactive components.


Assuntos
Colostro/química , Animais , Carga Bacteriana/veterinária , Bovinos , Colostro/microbiologia , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Imunoglobulina A/análise , Imunoglobulina G/análise , Interferon gama/análise , Interleucina-2/análise , Lactoferrina/análise , Fatores de Tempo , Transferrina/análise , Fator de Necrose Tumoral alfa/análise
4.
J Cell Mol Med ; 14(6B): 1605-18, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19650833

RESUMO

Several studies indicate that adult stem cells may improve the recovery from acute tissue injury. It has been suggested that they may contribute to tissue regeneration by the release of paracrine factors promoting proliferation of tissue resident cells. However, the factors involved remain unknown. In the present study we found that microvesicles (MVs) derived from human liver stem cells (HLSC) induced in vitro proliferation and apoptosis resistance of human and rat hepatocytes. These effects required internalization of MVs in the hepatocytes by an alpha(4)-integrin-dependent mechanism. However, MVs pre-treated with RNase, even if internalized, were unable to induce hepatocyte proliferation and apoptosis resistance, suggesting an RNA-dependent effect. Microarray analysis and quantitative RT-PCR demonstrated that MVs were shuttling a specific subset of cellular mRNA, such as mRNA associated in the control of transcription, translation, proliferation and apoptosis. When administered in vivo, MVs accelerated the morphological and functional recovery of liver in a model of 70% hepatectomy in rats. This effect was associated with increase in hepatocyte proliferation and was abolished by RNase pre-treatment of MVs. Using human AGO2, as a reporter gene present in MVs, we found the expression of human AGO2 mRNA and protein in the liver of hepatectomized rats treated with MVs. These data suggested a translation of the MV shuttled mRNA into hepatocytes of treated rats. In conclusion, these results suggest that MVs derived from HLSC may activate a proliferative program in remnant hepatocytes after hepatectomy by a horizontal transfer of specific mRNA subsets.


Assuntos
Vesículas Citoplasmáticas/metabolismo , Hepatectomia , Regeneração Hepática/fisiologia , Fígado/citologia , Células-Tronco/metabolismo , Adulto , Animais , Apoptose , Proliferação de Células , Células Cultivadas , Regulação da Expressão Gênica , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Células-Tronco/citologia
5.
Am J Transplant ; 10(10): 2270-8, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20840477

RESUMO

The development of proteinuria has been observed in kidney-transplanted patients on m-TOR inhibitor (m-TORi) treatment. Recent studies suggest that m-TORi(s) may alter the behavior and integrity of glomerular podocytes. We analyzed renal biopsies from kidney-transplanted patients and evaluated the expression of nephrin, a critical component of the glomerular slit-diaphragm. In a group of patients on 'de novo' m-TORi-treatment, the expression of nephrin within glomeruli was significantly reduced in all cases compared to pretransplant donor biopsies. Biopsies from control transplant patients not treated with m-TORi(s) failed to present a loss of nephrin. In a group of patients subsequently converted to m-TORi-treatment, a protocol biopsy performed before introduction of m-TORi was also available. The expression of nephrin in the pre-m-TORi biopsies was similar to that observed in the pretransplant donor biopsies but was significantly reduced after introduction of m-TORi(s). Proteinuria increased after the m-TORi inititiation in this group. However, in some cases proteinuria remained normal despite reduction of nephrin. In vitro, sirolimus downregulated nephrin expression by human podocytes. Our results suggest that m-TORi(s) may affect nephrin expression in kidney-transplanted patients, consistently with the observation in vitro on cultured podocytes.


Assuntos
Glomérulos Renais/metabolismo , Transplante de Rim/efeitos adversos , Proteínas de Membrana/biossíntese , Sirolimo/efeitos adversos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Adulto , Idoso , Células Cultivadas , Humanos , Pessoa de Meia-Idade , Podócitos/metabolismo , Proteinúria/induzido quimicamente , Estudos Retrospectivos
6.
G Ital Nefrol ; 25(2): 161-8, 2008.
Artigo em Italiano | MEDLINE | ID: mdl-18350496

RESUMO

In the adult kidney, different populations of progenitor cells (or stem cells) have been identified. These cells may represent a remnant of embryonic stem cells in the adult tissue, or populations of bone-marrow-derived stem cells homed within the kidney and modified by the local microenvironment. This modification may be the expression of a partial commitment or of different degrees of maturation. Resident stem cells may account for the growth of the organ during development, for the physiological cell turnover, and for the repair of kidney damage. In addition, stem cells derived from the bone marrow and migrated through the circulation to the site of the damage may contribute to tissue repair. Preliminary studies suggest that this regenerative potential of stem cells could be exploited for therapeutic purposes by administration of ex vivo expanded stem cell populations or by development of strategies aimed to expand and differentiate local stem cells.


Assuntos
Nefropatias/cirurgia , Rim/citologia , Transplante de Células-Tronco , Células-Tronco , Animais , Humanos
7.
G Ital Nefrol ; 25(3): 297-305, 2008.
Artigo em Italiano | MEDLINE | ID: mdl-18473301

RESUMO

Renal cell carcinoma is characterized by intense angiogenesis associated with the inactivation of the von Hippel-Lindau oncosuppressor gene with consequent hyperexpression of proangiogenic factors. Functional and molecular characterization of renal tumor endothelial cells has demonstrated an increase in angiogenesis and cell survival. The proangiogenic phenotype was due to hyperactivation of the PI3K/Akt/mTor pathway, which downregulates the synthesis of the antiangiogenic factor thrombospondin-1. Moreover, renal tumor endothelial cells presented an immature and embryonic phenotype with expression of the embryonic kidney-specific gene PAX-2. It is conceivable that the endothelium present in renal carcinoma is heterogeneous, with a possible origin from adjacent vessels, resident or circulating stem cells, or from the tumor cells themselves. The relevance of the angiogenic process in renal carcinoma is underlined by the therapeutic effect of antiangiogenic drugs. Different drugs against VEGF, such as the anti-VEGF monoclonal antibody bevacizumab, and small molecule tyrosine-kinase inhibitors, such as sunitinib and sorafenib, showed a clinical effect in patients with metastatic carcinoma. However, antiangiogenic therapy, although beneficial, is not sufficient per se. These studies suggest a role for the angiogenic program in the growth and dissemination of renal carcinoma and indicate the need for new therapeutic strategies.


Assuntos
Carcinoma de Células Renais/irrigação sanguínea , Carcinoma de Células Renais/metabolismo , Fatores de Crescimento Endotelial/metabolismo , Neoplasias Renais/irrigação sanguínea , Neoplasias Renais/metabolismo , Neovascularização Patológica/metabolismo , Inibidores da Angiogênese/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados , Benzenossulfonatos/uso terapêutico , Bevacizumab , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Quimioterapia Combinada , Fatores de Crescimento Endotelial/biossíntese , Fatores de Crescimento Endotelial/genética , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Humanos , Indóis/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/genética , Niacinamida/análogos & derivados , Fator de Transcrição PAX2/metabolismo , Fenótipo , Compostos de Fenilureia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Piridinas/uso terapêutico , Pirróis/uso terapêutico , Sorafenibe , Sunitinibe , Trombospondina 1/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo
8.
G Ital Nefrol ; 25 Suppl 44: S3-S10, 2008.
Artigo em Italiano | MEDLINE | ID: mdl-19048579

RESUMO

Much study has been dedicated to the understanding of the mechanisms leading to the progression of renal injury and to the development of strategies to limit this progression or possibly induce tissue regeneration. Among several identified mechanisms, the role of angiotensin II is widely recognized. Moreover, the progression of glomerular damage is characterized by capillary loss, reduction of the proliferative response, and production of antiangiogenic factors. Several lines of evidence support the potential effect of therapeutic startegies aimed at interfering with angiotensin II or stimulating angiogenesis in order to reduce the progression of renal injury. Recent work has underlined the potential of strategies involving the use of stem cells. Different populations of stem cells have been identified in the adult kidney. During renal injury, stem cells derived from the bone marrow that migrate through the circulation to the kidney may contribute to tissue repair. The regenerative potential of stem cells could be exploited by administration of ex vivo expanded stem cell populations or by the development of techniques to expand and differentiate local stem cells.


Assuntos
Transplante de Células-Tronco Hematopoéticas , Nefropatias/fisiopatologia , Nefropatias/cirurgia , Rim/lesões , Diferenciação Celular , Proliferação de Células , Progressão da Doença , Células Epiteliais/metabolismo , Humanos , Itália , Nefropatias/patologia , Glomérulos Renais/patologia , Receptor Tipo 2 de Angiotensina/deficiência , Regeneração , Resultado do Tratamento
9.
Sci Rep ; 7(1): 16875, 2017 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-29203902

RESUMO

Injection of amniotic fluid stem cells (AFSC) delays the course of progression of renal fibrosis in animals with Alport Syndrome, enhancing kidney function and improving survival. The mechanisms responsible for these protective outcomes are still largely unknown. Here, we showed that vascular endothelial growth factor (VEGF) signaling within the glomeruli of Alport mice is strongly elevated early on in the disease, causing glomerular endothelial cell damage. Intraventricular injected AFSC that homed within the glomeruli showed strong modulation of the VEGF activity, particularly in glomerular endothelial cells. To investigate this phenomenon we hypothesized that extracellular vesicles (EVs) produced by the AFSC could be responsible for the observed renoprotection. AFSC derived EVs presented exosomal and stem cell markers on their surface membrane, including VEGFR1 and VEGFR2. EVs were able to modulate VEGF in glomerular endothelial cells by effectively trapping the excess VEGF through VEGFR1-binding preventing cellular damage. In contrast, VEGFR1/sVEGFR1 knockout EVs failed to show similar protection, thus indicating that VEGF trapping is a potentially viable mechanism for AFSC-EV mediated renoprotection. Taken together, our findings establish that EVs secreted by AFSC could target a specific signaling pathway within the glomerulus, thus representing a new potential glomerulus-specific targeted intervention.


Assuntos
Células Endoteliais/metabolismo , Vesículas Extracelulares , Células-Tronco/metabolismo , Líquido Amniótico/citologia , Animais , Células Cultivadas , Técnicas de Cocultura , Creatinina/sangue , Modelos Animais de Doenças , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Vesículas Extracelulares/metabolismo , Glomérulos Renais/citologia , Camundongos , Nefrite Hereditária/metabolismo , Nefrite Hereditária/patologia , Proteinúria/patologia , Transdução de Sinais , Células-Tronco/citologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
10.
Int J Immunopathol Pharmacol ; 19(1): 81-96, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16569346

RESUMO

Kaposi's sarcoma (KS) is a malignancy associated to conditions of immune system impairment such as HIV-1 infection and post-transplantation therapy. Here we report that HIV-1-Tat protein, at concentrations well below those detected in AIDS patients, up-regulates the expression of both CD40 and CD154 on KS cells. This occurred also in the presence of vincristine, that at doses shown to induce apoptosis decreased the expression of both CD40 and CD154 on KS cells. The treatment with a soluble CD40-muIg fusion protein (CD40 fp) that prevents the binding of CD154 with cell surface CD40, as well as the transfection with a vector for soluble CD40 (KS sCD40), decreased the anti-apoptotic effect of Tat. Moreover, Tat-induced motility of KS cells was inhibited by soluble CD40 fp. Tat also enhanced the expression of intracellular proteins known to transduce signals triggered by CD40 engagement, in particular TRAF-3. Tat as well as soluble CD154 (sCD154) prevented vincristine-induced reduction of TRAF-3 in KS cells transfected with a vector for neomycin resistance (KS psv-neo), but not in KS sCD40. Immunoprecipitation studies showed that Tat induced CD40 / TRAF-3 association and that this binding was abrogated upon the incubation with the soluble CD40 fp. These data suggest that Tat activates the CD40-CD154 pathway by enhancing the membrane expression of CD40 and in particular of CD154, and by activating the TRAF-3-dependent signaling pathway of CD40. These findings indicate that the CD40-CD154 pathway mediates the anti-apoptotic and migratory effects of HIV-1- Tat, suggesting the potential therapeutic benefits of blocking CD40 activation in HIV-1-associated KS.


Assuntos
Apoptose/efeitos dos fármacos , Ligante de CD40/biossíntese , Movimento Celular/efeitos dos fármacos , Produtos do Gene tat/farmacologia , Sarcoma de Kaposi/metabolismo , Western Blotting , Antígenos CD40/biossíntese , Antígenos CD40/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/ultraestrutura , Sobrevivência Celular/efeitos dos fármacos , DNA de Neoplasias/biossíntese , DNA de Neoplasias/isolamento & purificação , HIV-1/metabolismo , Humanos , Imunoprecipitação , Marcação In Situ das Extremidades Cortadas , Indicadores e Reagentes , Análise de Sequência com Séries de Oligonucleotídeos , Transfecção , Produtos do Gene tat do Vírus da Imunodeficiência Humana
11.
Sci Rep ; 6: 32602, 2016 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-27586846

RESUMO

Purinergic signaling is involved in inflammation and cancer. Extracellular ATP accumulates in tumor interstitium, reaching hundreds micromolar concentrations, but its functional role on tumor vasculature and endothelium is unknown. Here we show that high ATP doses (>20 µM) strongly inhibit migration of endothelial cells from human breast carcinoma (BTEC), but not of normal human microvascular EC. Lower doses (1-10 µM result ineffective. The anti-migratory activity is associated with cytoskeleton remodeling and is significantly prevented by hypoxia. Pharmacological and molecular evidences suggest a major role for P2X7R and P2Y11R in ATP-mediated inhibition of TEC migration: selective activation of these purinergic receptors by BzATP mimics the anti-migratory effect of ATP, which is in turn impaired by their pharmacological or molecular silencing. Downstream pathway includes calcium-dependent Adenilyl Cyclase 10 (AC10) recruitment, cAMP release and EPAC-1 activation. Notably, high ATP enhances TEC-mediated attraction of human pericytes, leading to a decrease of endothelial permeability, a hallmark of vessel normalization. Finally, we provide the first evidence of in vivo P2X7R expression in blood vessels of murine and human breast carcinoma. In conclusion, we have identified a purinergic pathway selectively acting as an antiangiogenic and normalizing signal for human tumor-derived vascular endothelium.


Assuntos
Neoplasias da Mama/patologia , Movimento Celular , AMP Cíclico/metabolismo , Células Endoteliais/patologia , Receptores Purinérgicos P2X7/metabolismo , Receptores Purinérgicos P2/metabolismo , Transdução de Sinais , Trifosfato de Adenosina/farmacologia , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Feminino , Humanos , Camundongos Endogâmicos BALB C , Modelos Biológicos , Transdução de Sinais/efeitos dos fármacos
12.
Int J Immunopathol Pharmacol ; 18(2): 327-37, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-15888255

RESUMO

Deposition of atherogenic lipoproteins is associated with various glomerular diseases. In particular, oxidized LDL (oxLDL) may affect mesangial cells and favour the development of glomerulosclerosis. The aim of the present study was to investigate on cultured human mesangial cells (HMC) whether oxLDL induces apoptosis by a mechanism dependent on the inhibition of Akt survival pathway, and whether the engagement of mesangial CD40 by its ligand CD154 inhibits the apoptotic effect of oxLDL. Tunel assays demonstrated that incubation of HMC for 24 h with oxLDL, but not with unmodified LDL, induced a dose-dependent increase in apoptosis of HMC associated with a decrease in Akt phosphorylation. Enzymatic kinase assay showed that also the Akt activity was reduced in a dose-dependent manner by treatment with oxLDL. Stimulation of mesangial CD40 with sCD154 rescued HMC from oxLDL-dependent apoptosis, while two unrelated pharmacological inhibitors of PI3K LY294002 and wortmannin abrogated this anti-apoptotic effect, suggesting an involvement of the PI3K/Akt pathway. Moreover CD40 stimulation maintained an elevated phosphorylation of Akt and preserved its enzymatic activity in the presence of oxLDL. Indeed, CD154 induced a rapid enhancement in Akt enzymatic activity, that was temporarily correlated with the association of CD40 with TRAF3, TRAF6, c-Cbl and the p85 subunit of PI3K. In conclusion, these results suggest that CD40 stimulation protects HMC from toxic effects of oxLDL by promoting PI3K/Akt-dependent cell survival.


Assuntos
Apoptose/efeitos dos fármacos , Antígenos CD40/metabolismo , Mesângio Glomerular/citologia , Lipoproteínas LDL/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Ligante de CD40/farmacologia , Células Cultivadas , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Mesângio Glomerular/efeitos dos fármacos , Mesângio Glomerular/enzimologia , Humanos , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação , Proteínas Proto-Oncogênicas c-akt , Proteínas Recombinantes/farmacologia
13.
Int J Oncol ; 8(3): 543-8, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21544394

RESUMO

The monoclonal antibody (P-3 mAb) and the rabbit immune sera (RIS) recognizing a 14 kDa perchloric acid soluble protein extracted from goat liver (UK 114), locate this antigen on the cell membrane of several human cancer cell lines. UK 114-positive cells (e.g. HT 29 and KATO VI cells) undergo antibody-dependent cytolysis bl vitro. In nu/nu mice bearing xenografted HT 29 cells, tumor growth was markedly impaired by peri-tumoral injection of anti-UK 114 antibodies. These experiments suggest that human tumors expressing UK 114 over the cell membrane may undergo antibody-mediated cytolysis and growth control.

14.
Int J Oncol ; 10(4): 779-85, 1997 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21533445

RESUMO

UK114 is a 14 kDa protein identified in the perchloric acid soluble extract of goat liver. Anti-UK114 antibodies identify this protein as expressed by the membrane of human cancer cells. In the present study, anti-UK114 antibodies were detected in the sera of cancer patients by ELISA and by immunofluorescence tests using human cancer cell lines as a target. UK114 protein is antigenic in cancer patients. Human anti-UK114 antibodies have cytolytic effects in vitro and their presence correlates with inhibition of growth of human carcinoma cells xenografted in nu/nu mice. This finding may open new prospects for immunotherapy.

15.
Virchows Arch ; 431(5): 323-8, 1997 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9463573

RESUMO

A perchloric acid-soluble protein extracted from goat liver and designated as UK 114 is known to be expressed over the cell membrane of (some) human cancer cell lines. This protein is antigenic, and specific antibodies elicit complement-dependent cytolysis of neoplastic target cells. In this study we demonstrate that administration of UK 114, either pure or as a crude extract (designated UK 101), inhibits the growth of mammary carcinomas induced in female Sprague-Dawley rats by dimethylbenzanthracene (DMBA). The mechanism of the tumour inhibitory activity of UK 114 is probably related to induction of immunosurveillance.


Assuntos
Adenocarcinoma/tratamento farmacológico , Fígado/química , Neoplasias Mamárias Animais/tratamento farmacológico , Proteínas/farmacologia , 9,10-Dimetil-1,2-benzantraceno , Adenocarcinoma/induzido quimicamente , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Antígenos de Neoplasias/metabolismo , Proteínas do Sistema Complemento/imunologia , Citotoxicidade Imunológica/imunologia , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Cabras , Neoplasias Mamárias Animais/induzido quimicamente , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Proteínas/imunologia , Proteínas/isolamento & purificação , Ratos , Ratos Sprague-Dawley
16.
Sci Total Environ ; 270(1-3): 179-90, 2001 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-11327392

RESUMO

BACKGROUND: A possible relationship between Silica (Si) exposure and antineutrophil cytoplasm antibodies (ANCA)-associated vasculitis has been reported. Furthermore, tuberculosis (TBC) has been frequently described in patients with silicosis, and TBC infection shares with ANCA-associated vasculitis the formation of granulomas. Therefore, an intriguing network including Silica, Vasculitis, TBC and ANCA might be hypothesized. The aim of this work was to further investigate these correlations using both epidemiological and pathogenic approaches. METHODS: Study I--epidemiological study. A case-control study to compare the occupational histories of 31 cases of biopsy proven vasculitis (18 pauci-immune crescentic glomerulonephritis, 9 microscopic polyangitis, 4 Wegener's granulomatosis) with those of 58 age, sex and residence-matched controls (affected by other kidney diseases), was performed. Occupational Health physicians designed an appropriate questionnaire in order to evaluate a wide spread of exposures and calculate their entity by the product of Intensity x Frequency x Duration. Study II--tuberculosis association. A case-control study to evaluate the frequency of a previous history of tuberculosis (TBC) in 45 patients with vasculitis and 45 controls were performed. Study III--ANCA positivity. A case-control study to evaluate the presence of ANCA was performed by testing blood samples of 64 people with previous professional exposure and 65 sex/age matched patients hospitalized in a General Medicine Unit. Furthermore, the same evaluation was made in a pilot study in 16 patients with ongoing or previous TBC. Study IV--experimental study. The oxygen free radicals (OFR) and IL-12 production (both involved in the pathogenesis of vasculitis) from human phagocytic cells stimulated with an amorphous (diatomaceous earth) and a crystalline (quartz) form of Si at the doses of 10 and 100 microg ml(-1) was evaluated. RESULTS: Study I--a positive history of exposure to Si resulted in significantly more present in cases (14/31 = 45%) than in controls (14/58 = 24%, P = 0.04, OR = 2.4) and no other significant exposure association was found (including asbestos, mineral oil, formaldehyde, diesel and welding fumes, grain and wood dust, leather, solvents, fungicides, bitumen, lead and paint). Study II--past TBC infection was significantly more present in patients with vasculitis (12/45 = 26%) than in controls (4/45 = 8%, P < 0.05). Study III--ANCA was present in 2/64 exposed people (vs. 0/65 controls, P = NS) and 0/16 patients with TBC. Study IV--both amorphous and crystalline Si forms represented a stimulus for OFR and IL-12 production, but quartz resulted as a greater inductor. CONCLUSIONS: We conclude that Si exposure might be a risk factor for ANCA-associated vasculitis, possibly enhancing endothelial damage by phagocyte generation of oxygen free radicals and Th1 differentiation by an excessive IL-12 phagocyte production. Frequency of TBC was significantly higher in vasculitis patients. ANCA was not frequent in the preliminary examination of people with previous professional exposure or patients with TBC, but the number of samples evaluated is too small to allow conclusions.


Assuntos
Anticorpos Anticitoplasma de Neutrófilos/sangue , Doenças Autoimunes/epidemiologia , Dióxido de Silício/toxicidade , Tuberculose/complicações , Vasculite/epidemiologia , Vasculite/imunologia , Adulto , Idoso , Anticorpos Anticitoplasma de Neutrófilos/efeitos dos fármacos , Doenças Autoimunes/patologia , Estudos de Casos e Controles , Feminino , Radicais Livres/metabolismo , Humanos , Interleucina-12/metabolismo , Masculino , Pessoa de Meia-Idade , Exposição Ocupacional/efeitos adversos , Projetos Piloto , Vasculite/patologia
17.
Minerva Urol Nefrol ; 48(1): 1-6, 1996 Mar.
Artigo em Italiano | MEDLINE | ID: mdl-8848762

RESUMO

Recent experimental studies allowed the identification of several mechanisms of immune deposit formation, which are able to reproduce the morphological and clinical pattern of human glomerulonephritis. Moreover, it was shown that most of the lesions considered, in the past, as due to circulating immune complexes (IC), are instead caused by the "in situ" formation of IC. As a result of these studies, the following schematic classification was proposed: 1) immune deposits formed by glomerular localization of IC primarily formed in the circulation; 2) immune deposits formed "in situ" by reaction of circulating antibodies with fixed structural antigens; 3) immune deposits formed "in situ" by antibodies reactive with movable structural antigens; 4) immune deposits formed "in situ" by antibodies reactive with sequestered antigens leaking out of tissues; 5) IC formed "in situ" by antibodies reactive with exogenous or non-glomerular endogenous antigens planted in the glomeruli; 6) ANCA-associated glomerular disease.


Assuntos
Glomerulonefrite/imunologia , Anticorpos Anticitoplasma de Neutrófilos , Complexo Antígeno-Anticorpo/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Reações Antígeno-Anticorpo , Autoanticorpos/imunologia , Biomarcadores , Humanos
18.
Reprod Sci ; 21(3): 296-304, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24037753

RESUMO

The cyclical arrival of endometrial cells into the abdominal cavity through retrograde flux at menstruation represents the etiopathogenetic basis of endometriosis. The endometrium has peculiar regenerative properties linked to the presence of adult stem cells similar to mesenchymal stem cells (MSCs). Once in the abdominal cavity, these MSCs could proliferate, invade, and differentiate into endometrial cells, finally generating ectopic implants. As only differentiated endometrial cells, and not endometrial MSCs, possess steroid hormone receptors, MSCs could be responsible for the high rate of persistence/recurrence of the disease after hypoestrogenism-inducing therapies. Even angiogenesis promoted by MSCs could play an important role, as survival and proliferation of endometriotic tissue depend on the formation of new blood vessels. Inhibition of angiogenesis represents, in fact, a new, promising therapeutic approach for the disease. Further, medications directly targeting endometriosis MSCs could be effective, alone or in association with hormonal treatments, in increasing the success of medical treatment.


Assuntos
Células-Tronco Adultas/patologia , Inibidores da Angiogênese/uso terapêutico , Endometriose/patologia , Endométrio/patologia , Neovascularização Patológica/patologia , Adulto , Células-Tronco Adultas/efeitos dos fármacos , Inibidores da Angiogênese/farmacologia , Animais , Endometriose/tratamento farmacológico , Endométrio/efeitos dos fármacos , Feminino , Humanos , Neovascularização Patológica/tratamento farmacológico , Niacinamida/análogos & derivados , Niacinamida/farmacologia , Niacinamida/uso terapêutico , Compostos de Fenilureia/farmacologia , Compostos de Fenilureia/uso terapêutico , Sorafenibe
19.
Oncogene ; 31(2): 200-12, 2012 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-21685934

RESUMO

Changes in intracellular calcium [Ca(2+)](i) levels control critical cytosolic and nuclear events that are involved in the initiation and progression of tumor angiogenesis in endothelial cells (ECs). Therefore, the mechanism(s) involved in agonist-induced Ca(2+)(i) signaling is a potentially important molecular target for controlling angiogenesis and tumor growth. Several studies have shown that blood vessels in tumors differ from normal vessels in their morphology, blood flow and permeability. We had previously reported a key role for arachidonic acid (AA)-mediated Ca(2+) entry in the initial stages of tumor angiogenesis in vitro. In this study we assessed the mechanism involved in AA-induced EC migration. We report that TRPV4, an AA-activated channel, is differentially expressed in EC derived from human breast carcinomas (BTEC) as compared with 'normal' EC (HMVEC). BTEC display a significant increase in TRPV4 expression, which was correlated with greater Ca(2+) entry, induced by AA or 4αPDD (a selective TRPV4 agonist) in the tumor-derived ECs. Wound-healing assays revealed a key role of TRPV4 in regulating cell migration of BTEC but not HMVEC. Knockdown of TRPV4 expression completely abolished AA-induced BTEC migration, suggesting that TRPV4 mediates the pro-angiogenic effects promoted by AA. Furthermore, pre-incubation of BTEC with AA induced actin remodeling and a subsequent increase in the surface expression of TRPV4. This was consistent with the increased plasma membrane localization of TRPV4 and higher AA-stimulated Ca(2+) entry in the migrating cells. Together, the data presented herein demonstrate that: (1) TRPV4 is differentially expressed in tumor-derived versus 'normal' EC; (2) TRPV4 has a critical role in the migration of tumor-derived but not 'normal' EC migration; and (3) AA induces actin remodeling in BTEC, resulting in a corresponding increase of TRPV4 expression in the plasma membrane. We suggest that the latter is critical for migration of EC and thus in promoting angiogenesis and tumor growth.


Assuntos
Actinas/metabolismo , Ácido Araquidônico/metabolismo , Movimento Celular/fisiologia , Endotélio Vascular/patologia , Neoplasias/irrigação sanguínea , Canais de Cátion TRPV/fisiologia , Humanos
20.
Leukemia ; 26(5): 1081-90, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22024722

RESUMO

Angiogenesis is considered a hallmark of multiple myeloma (MM) progression. In the present study, we evaluated the morphological and functional features of endothelial cells (ECs) derived from bone marrow (BM) of patients affected by MM (MMECs). We found that MMECs compared with normal BM ECs (BMECs) showed increased expression of syndecan-1. Silencing of syndecan-1 expression by RNA interference technique decreased in vitro EC survival, proliferation and organization in capillary-like structures. In vivo, in severe combined immunodeficient mice, syndecan-1 silencing inhibited MMEC organization into patent vessels. When overexpressed in human umbilical vein ECs and BMECs, syndecan-1 induced in vitro and in vivo angiogenic effects. Flow-cytometric analysis of MMECs silenced for syndecan-1 expression indicated a decreased membrane expression of vascular endothelial growth factor (VEGF) receptor-2 (VEGFR-2). Immunoprecipitation and confocal analysis showed colocalization of VEGFR-2 with syndecan-1. Absence of nuclear translocation of VEGFR-2 in syndecan-1-knockdown cells together with the shift from perinuclear localization to recycling compartments suggest a role of syndecan-1 in modulation of VEGFR-2 localization. This correlated with an in vitro decreased VEGF-induced invasion and motility. These results suggest that syndecan-1 may contribute to the highly angiogenic phenotype of MMECs by promoting EC proliferation, survival and modulating VEGF-VEGFR-2 signalling.


Assuntos
Endotélio/patologia , Mieloma Múltiplo/patologia , Neovascularização Patológica , Sindecana-1/fisiologia , Animais , Células Cultivadas , Citometria de Fluxo , Inativação Gênica , Humanos , Imunoprecipitação , Camundongos , Mieloma Múltiplo/irrigação sanguínea , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Sindecana-1/genética , Células Tumorais Cultivadas , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
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