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Clinicians are increasingly using regenerative medicines to repair, replace, regenerate or rejuvenate lost, damaged or diseased genes, cells, tissues or organs. In South Africa, access to these novel gene therapies and cell and tissue-based products is limited. The human leukocyte antigen (HLA) diversity and a paucity of suitable HLA-identical unrelated donors, results in limited access to haematopoietic stem and progenitor cell transplantation (HSPCT). Cell-based products could increase this access. Genetic diversity can also manifest in local or region-specific rare congenital disorders, and in vivo gene therapies hold the promise of developing treatments and cures for these debilitating disorders. South Africa has a disproportionate mortality rate due to non-natural causes, with many surviving with permanent injuries and disabilities. Tissue-engineered cell-based products have the potential to restore many of those affected and improve quality of life and productivity. These factors create an urgency for South Africa to develop regenerative medicines to address the country's unique needs and to provide access to these new and innovative treatment modalities. Achieving this objective requires a well-coordinated effort by multiple stakeholders and role players. A critical component of a regenerative medicine ecosystem is establishing an enabling regulatory framework for these new classes of medicines. Here we provide a brief profile of South Africa, including its genetic diversity, economy, the impact of the burden of disease, health policy and the healthcare system. We address the regulation of medicines, how the existing framework can accommodate regenerative medicines, and the steps needed to establish a future regulatory framework.
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Qualidade de Vida , Medicina Regenerativa , Ecossistema , Terapia Genética , Humanos , África do SulRESUMO
The introduction of antiretroviral therapy (ART) and highly active antiretroviral therapy (HAART) has transformed human immunodeficiency virus (HIV)-1 into a chronic, well-managed disease. However, these therapies do not eliminate all infected cells from the body despite suppressing viral load. Viral rebound is largely due to the presence of cellular reservoirs which support long-term persistence of HIV-1. A thorough understanding of the HIV-1 reservoir will facilitate the development of new strategies leading to its detection, reduction, and elimination, ultimately leading to curative therapies for HIV-1. Although immune cells derived from lymphoid and myeloid progenitors have been thoroughly studied as HIV-1 reservoirs, few studies have examined whether mesenchymal stromal/stem cells (MSCs) can assume this function. In this review, we evaluate published studies which have assessed whether MSCs contribute to the HIV-1 reservoir. MSCs have been found to express the receptors and co-receptors required for HIV-1 entry, albeit at levels of expression and receptor localisation that vary considerably between studies. Exposure to HIV-1 and HIV-1 proteins alters MSC properties in vitro, including their proliferation capacity and differentiation potential. However, in vitro and in vivo experiments investigating whether MSCs can become infected with and harbour latent integrated proviral DNA are lacking. In conclusion, MSCs appear to have the potential to contribute to the HIV-1 reservoir. However, further studies are needed using techniques such as those used to prove that cluster of differentiation (CD)4+ T cells constitute an HIV-1 reservoir before a reservoir function can definitively be ascribed to MSCs. MSCs may contribute to HIV-1 persistence in vivo in the vasculature, adipose tissue, and bone marrow by being a reservoir for latent HIV-1. To harbour latent HIV-1, MSCs must express HIV-1 entry markers, and show evidence of productive or latent HIV-1 infection. The effect of HIV-1 or HIV-1 proteins on MSC properties may also be indicative of HIV-1 infection.
Assuntos
Infecções por HIV , HIV-1 , Células-Tronco Mesenquimais , Síndrome de Imunodeficiência Adquirida dos Símios , Vírus da Imunodeficiência Símia , Animais , Antirretrovirais/farmacologia , Antirretrovirais/uso terapêutico , Linfócitos T CD4-Positivos , Infecções por HIV/terapia , Humanos , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios/tratamento farmacológico , Vírus da Imunodeficiência Símia/genética , Latência ViralRESUMO
The discovery of human leucocyte antigen (HLA), serological matching and HLA-typing techniques, combined with the development of immunosuppressive medicines and improvements in infection control, have opened the way to cell, tissue and vascularised organ transplantation. Since the early 1960s, more than a million haematopoietic progenitor cell (HPC) transplantations have been performed worldwide to restore haematopoiesis and support immune system recovery after bone marrow ablation. HPC transplantation uses minimally manipulated autologous or allogeneic cells to restore the homologous functions of bone marrow. Research in biological sciences supported by new technologies is increasingly translated into therapeutic products intended to augment, repair, replace or regenerate genes, cells, tissues, organs and metabolic processes in the body. These products are referred to as regenerative medicine therapies or advanced therapy medicinal products, and include gene therapies, cell-based therapies and engineered tissue products.
Assuntos
Medicina Regenerativa/tendências , Terapia Baseada em Transplante de Células e Tecidos/tendências , Terapia Genética/tendências , Transplante de Células-Tronco Hematopoéticas , Teste de Histocompatibilidade , Humanos , África do Sul , Engenharia Tecidual/tendênciasRESUMO
The growing need for haematopoietic stem cell transplantation (HSCT) is reflected in the increasing number of transplants performed globally each year. HSCT provides life-changing and potentially curative therapy for a range of pathologies including haematological malignancies; other indications include certain congenital and acquired disorders of the haematopoietic system, autoimmune conditions and hereditary diseases. The primary goals of HSCT are either to replace haematopoietic stem and progenitor cells (HSPC) following myeloablative chemotherapy or to cure the original pathology with allogeneic HSPCs. Success depends on optimal outcomes at various stages of the procedure including mobilisation of marrow stem/progenitor cells for harvesting from the patient or donor, long-term and sustainable engraftment of these cells in the recipient, and prevention of graft-versus-host disease in the case of allogeneic HSCT. Challenges in South Africa include high cost, limited infrastructure and lack of appropriately trained staff, as well as limitations in securing suitable haematopoietic stem cell donors. This review aims to provide an overview of HSCT and some of the challenges that are faced in the South African context.
Assuntos
Doenças Hematológicas/terapia , Neoplasias Hematológicas/terapia , Transplante de Células-Tronco Hematopoéticas/métodos , Doenças Autoimunes/terapia , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , África do SulRESUMO
Cellular therapy has become a billion-dollar industry and is set to become one of the therapeutic pillars of healthcare in the 21st century. Adult stem cells, which include haematopoietic stem and progenitor cells (HSPCs) and mesenchymal stromal/stem cells (MSCs), is one of the major cell types currently under investigation for use in cell therapy. This review focuses on HSPCs and MSCs and discusses their heterogeneous nature and the problems faced in expanding these cells to therapeutic numbers for use in clinical applications.
Assuntos
Transplante de Células-Tronco Hematopoéticas/métodos , Células-Tronco Hematopoéticas/citologia , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/citologia , Adulto , Animais , HumanosRESUMO
Human immunodeficiency virus (HIV) infection not only leads to a compromised immune system, but also disrupts normal haematopoiesis, resulting in the frequent manifestation of cytopenias (anaemia, thrombocytopenia and neutropenia). Although there is a definite association between the severity of cytopenia and HIV disease stage, this relationship is not always linear. For example, cytopenias such as thrombocytopenia may occur during early stages of infection. The aetiology of these haematological abnormalities is complex and multifactorial, including drug-induced impaired haematopoiesis, bone marrow suppression due to infiltration of infectious agents or malignant cells, HIV-induced impaired haematopoiesis, and several other factors. In this review, we describe the frequencies of anaemia, thrombocytopenia and neutropenia reported for HIV-infected, treatment-naïve cohorts studied in eastern and southern sub-Saharan African countries. We present a rational approach for the use of diagnostic tests during the workup of HIV-infected patients presenting with cytopenia, and discuss how HIV impacts on haematopoietic stem/progenitor cells (HSPCs) resulting in impaired haematopoiesis. Finally, we describe the direct and indirect effects of HIV on HSPCs which result in defective haematopoiesis leading to cytopenias.
Assuntos
Infecções por HIV/complicações , Hematopoese , Anemia/diagnóstico , Anemia/etiologia , Células-Tronco Hematopoéticas/citologia , Humanos , Neutropenia/diagnóstico , Neutropenia/etiologia , Células-Tronco/citologia , Trombocitopenia/diagnóstico , Trombocitopenia/etiologiaRESUMO
Autologous and allogeneic haematopoietic stem cell (HSC) transplantation has been performed in patients with various malignant and non-malignant haematological disorders for more than 50 years. Ex vivo gene modification of HSCs for autologous transplantation opens up new therapeutic avenues for genetic and infectious diseases. Major advances have been made over the last three decades with respect to gene modification of HSCs and transplantation strategies, ultimately culminating in the approval of two such therapies in Europe (Strimvelis for a rare primary immune deficiency, and LentiGlobin for beta-thalassaemia). Newer gene-modifying technologies and treatment regimens have also recently come to the fore, which hold great promise for the development of safer and more effective treatments. We provide an overview of the current state of gene-modified HSC therapies, highlighting success stories, limitations and important considerations for achieving successful translation of these therapies to the clinic.
Assuntos
Terapia Genética/métodos , Doenças Hematológicas/terapia , Transplante de Células-Tronco Hematopoéticas/métodos , Células-Tronco Hematopoéticas/citologia , HumanosRESUMO
Neurological disease encompasses a diverse group of disorders of the central and peripheral nervous systems, which collectively are the leading cause of disease burden globally. The scope of treatment options for neurological disease is limited, and drug approval rates for improved treatments remain poor when compared with other therapeutic areas. Stem cell therapy provides hope for many patients, but should be tempered with the realisation that the scientific and medical communities are still to fully unravel the complexities of stem cell biology, and to provide satisfactory data that support the rational, evidence-based application of these cells from a therapeutic perspective. We provide an overview of the application of stem cells in neurological disease, starting with basic principles, and extending these to describe the clinical trial landscape and progress made over the last decade. Many forms of stem cell therapy exist, including the use of neural, haematopoietic and mesenchymal stem cells. Cell therapies derived from differentiated embryonic stem cells and induced pluripotent stem cells are also starting to feature prominently. Over 200 clinical studies applying various stem cell approaches to treat neurological disease have been registered to date (Clinicaltrials.gov), the majority of which are for multiple sclerosis, stroke and spinal cord injuries. In total, we identified 17 neurological indications in clinical stage development. Few studies have progressed into large, pivotal investigations with randomised clinical trial designs. Results from such studies will be essential for approval and application as mainstream treatments in the future.
Assuntos
Terapia Baseada em Transplante de Células e Tecidos/métodos , Doenças do Sistema Nervoso/terapia , Transplante de Células-Tronco/métodos , Humanos , Doenças do Sistema Nervoso/fisiopatologia , Ensaios Clínicos Controlados Aleatórios como Assunto , Células-Tronco/citologiaRESUMO
The probability of a Black African finding a matched unrelated donor for a hematopoietic stem cell transplant is minimal due to the high degree of genetic diversity amongst individuals of African origin. This problem could be resolved in part by the establishment of a public cord blood (CB) stem cell bank. The high prevalence of human immunodeficiency virus (HIV) amongst women attending antenatal clinics in sub-Saharan Africa together with the risk of mother-to-child transmission increases the risk of transplant transmissible infection. In addition to screening the mother in a period inclusive of 7 days prior to the following delivery, we propose that all CB units considered for storage undergo rigorous and reliable screening for HIV. The Ultrio-plus® assay is a highly specific and sensitive test for detecting HIV, hepatitis-B and hepatitis-C viruses in peripheral blood. We validated the Ultrio-plus® assay for analytical sensitivity in detecting HIV in CB at the level of detection of the assay. Until more comprehensive and sensitive methods are developed, the sensitivity and reliability of the Ultrio-plus® assay suggest that it could be used for the routine screening of CB units in conjunction with currently recommended maternal screening to reduce the risk of transplant transmissible infection.
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Cystic fibrosis (CF), one of the most commonly observed and diagnosed fatal monogenic disorders globally, was initially thought to affect individuals of Caucasian/European descent almost exclusively. It is increasingly appreciated, however, that non-Caucasian populations are also affected by this condition. Although this has been known in South Africa (SA) for over two decades, a large disparity still exists in data pertaining to the different population groups in the country. This article seeks to highlight existing published data on CF in SA populations and reflects on the means through which these have been generated over the years. Additionally, the article briefly discusses the consequences of incomplete data and how this could potentially be addressed in the future through innovative and collaborative approaches.
Assuntos
Fibrose Cística/diagnóstico , Humanos , África do SulRESUMO
The fields of cell and gene therapy are moving rapidly towards providing innovative cures for incurable diseases. A current and highly topical example is immunotherapies involving T-cells that express chimeric antigen receptors (CAR T-cells), which have shown promise in the treatment of leukaemia and lymphoma. These new medicines are indicative of the changes we can anticipate in the practice of medicine in the near future. Despite their promise, they pose challenges for introduction into the healthcare sector in South Africa (SA), including: (i) that they are technologically demanding and their manufacture is resource intensive; (ii) that the regulatory system is underdeveloped and likely to be challenged by ethical, legal and social requirements that accompany these new therapies; and (iii) that costs are likely to be prohibitive, at least initially, and before economies of scale take effect. Investment should be made into finding novel and innovative ways to introduce these therapies into SA sooner rather than later to ensure that SA patients are not excluded from these exciting new opportunities.
Assuntos
Terapia Baseada em Transplante de Células e Tecidos/economia , Difusão de Inovações , Terapia Genética/economia , Custos de Cuidados de Saúde , Acessibilidade aos Serviços de Saúde , Leucemia/terapia , Linfoma/terapia , Classe Social , Antígenos CD19/economia , Antígenos CD19/uso terapêutico , Produtos Biológicos , Terapia Baseada em Transplante de Células e Tecidos/ética , Terapia Genética/ética , Terapia Genética/legislação & jurisprudência , Humanos , Imunoterapia Adotiva/economia , Imunoterapia Adotiva/ética , Imunoterapia Adotiva/legislação & jurisprudência , Receptores de Antígenos de Linfócitos T/uso terapêutico , África do SulRESUMO
Acute myeloid leukemia (AML) is characterized by proliferation of the myeloid lineage and accumulation of immature hematopoietic cells in the bone marrow and is typified by marked heterogeneity both in response to treatment and survival. AMLprofiler is a qualitative in vitro diagnostic microarray incorporating seven molecular biomarkers used to diagnose and predict posttherapy survival rates. In this study, we compared AMLprofiler to routine AML diagnostic methodologies employed in South Africa, focusing on consistency of the results, cost, and time to result. RNA was isolated from bone marrow and peripheral blood samples from patients with de novo AML and was processed using Affymetrix Gene Profiling Reagent kits. The results from AMLprofiler and standard methodologies were highly comparable. In addition, many samples were determined to be positive for biomarkers not routinely investigated in South Africa, namely, CEBPA double mutants, NPM1 variants, and altered expression levels of BAALC and EVI1. 38% of samples presented with no positive biomarker; AMLprofiler nonetheless enabled 26% of AML patients to be classified into either favorable or poor prognostic categories. This study highlights the comprehensive nature of the microarray. Decreased time to result and refinement of risk stratification are notable benefits.
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The lymphatic system is the primary pathway of metastasis for most human cancers. Recent research efforts in studying lymphangiogenesis have suggested the existence of a relationship between lymphatic vessel density and patient survival. However, current methodology of lymphangiogenesis quantification is still characterised by high intra- and interobserver variability. For the amount of lymphatic vessels in a tumour to be a clinically useful parameter, a reliable quantification technique needs to be developed. With this consensus report, we therefore would like to initiate discussion on the standardisation of the immunohistochemical method for lymphangiogenesis assessment.
Assuntos
Técnicas Imunoenzimáticas/métodos , Linfangiogênese , Vasos Linfáticos/patologia , Neoplasias/patologia , Biomarcadores Tumorais/metabolismo , Células Endoteliais/patologia , Humanos , Metástase Linfática , Vasos Linfáticos/metabolismo , Glicoproteínas de Membrana/metabolismo , Neoplasias/irrigação sanguínea , Fator A de Crescimento do Endotélio Vascular/metabolismoAssuntos
Vasos Linfáticos/metabolismo , Vasos Linfáticos/patologia , Sarcoma/metabolismo , Sarcoma/patologia , Fator C de Crescimento do Endotélio Vascular/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Humanos , Imuno-Histoquímica , Metástase Linfática , Sarcoma/secundárioRESUMO
Primary human brain tumours account for approximately 2% of all cancers. High levels of expression of vascular endothelial growth factor-A (VEGF-A), a potent angiogenic factor, are linked to poor prognosis. In contrast, the potential role in human brain tumour biology of newer VEGF family members, VEGF-C and VEGF-D, both of which are lymphangiogenic factors, is poorly understood. In the present study, the expression of all VEGFs (VEGF-A, -B, -C, and -D) and their receptors (VEGFR-1, -2, and -3) has been assessed in 39 primary human brain tumours. The well-established findings were confirmed with VEGF-A. Surprisingly, however, VEGF-C and VEGF-D, as well as VEGFR-3, were expressed in some tumour types such as haemangioblastomas and glioblastomas, despite their lack of lymphatic vessels. VEGF-C and VEGFR-3 transcripts were localized to the tumour palisade around necrotic areas in glioblastomas and were evenly distributed throughout haemangioblastomas. VEGF-C protein was localized by immunohistochemistry to the palisade layer in glioblastomas. More than 50% of VEGF-C-positive cells also expressed the intermediate-stage inflammatory macrophage marker CD163; however, a significant proportion of VEGF-C-positive cells were CD163-negative. These data demonstrate the presence of molecules, primarily described as regulators of lymphangiogenesis, in normal human brain and brain tumours that are devoid of lymphatics. Their localization in macrophages points to a role in tumour-associated inflammation.
Assuntos
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Hemangioblastoma/metabolismo , Fator C de Crescimento do Endotélio Vascular/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Expressão Gênica , Glicoproteínas/metabolismo , Humanos , Hibridização In Situ/métodos , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Reação em Cadeia da Polimerase/métodos , RNA Mensageiro/genética , RNA Neoplásico/genética , Estudos Retrospectivos , Fator A de Crescimento do Endotélio Vascular/biossíntese , Fator A de Crescimento do Endotélio Vascular/genética , Fator B de Crescimento do Endotélio Vascular/biossíntese , Fator B de Crescimento do Endotélio Vascular/genética , Fator D de Crescimento do Endotélio Vascular/biossíntese , Fator D de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Proteínas de Transporte VesicularRESUMO
BACKGROUND: Oesophageal adenocarcinoma is an aggressive neoplasm with poor prognosis as a result of early lymph node metastasis. AIMS: To measure lymphatic vessel density (LVD) in the neoplastic progression from Barrett's metaplasia to adenocarcinoma and determine whether LVD can predict the risk of cancer. In addition, to correlate LVD with lymph node metastasis and assess whether LVD could be used as a prognostic indicator for outcome or survival. METHODS: LVD and microvascular density (MVD) were assessed after immunohistochemical staining of vessels in Barrett's metaplasia, dysplasia, and adenocarcinoma tissues and were correlated with clinicopathological features. RESULTS: LVD was significantly reduced in adenocarcinoma, being half that seen in normal stomach/oesophagus or metaplasia/dysplasia. LVD did not correlate with tumour grade, stage, or clinical outcome; however, patients who had either lymph node metastasis or invasion of tumour cells into peritumorous lymphatic vessels had a significantly worse overall survival. MVD was also assessed as a prognostic marker; its increase appeared to be linked more with the development of Barrett's metaplasia than adenocarcinoma. CONCLUSIONS: The reduction in lymphatic vessel numbers was not useful for determining disease outcome in the patient group studied. It is the entry of tumour cells into pre-existing peritumorous lymphatic vessels that confers a significantly worse overall survival.
Assuntos
Adenocarcinoma/patologia , Esôfago de Barrett/patologia , Neoplasias Esofágicas/patologia , Vasos Linfáticos/patologia , Lesões Pré-Cancerosas/patologia , Adenocarcinoma/secundário , Adenocarcinoma/cirurgia , Adulto , Idoso , Idoso de 80 Anos ou mais , Progressão da Doença , Neoplasias Esofágicas/cirurgia , Esôfago/patologia , Feminino , Seguimentos , Humanos , Metástase Linfática , Masculino , Metaplasia/patologia , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Análise de SobrevidaRESUMO
Increased understanding of the mechanisms of angiogenesis and lymphangiogenesis has provided a glimpse at some of the molecules involved in the pathophysiology of hemangiomas and vascular malformations. This review focuses on recent advances in our understanding of the mechanisms of angiogenesis/lymphangiogenesis and the differentiation of arterial, venous, and lymphatic vessels. We integrate this knowledge with new data obtained from genetic studies in humans, which have revealed a number of heretofore-unsuspected candidates involved in the development of familial vascular anomalies. We present a common infantile vascular tumor, hemangioma, and then focus on hereditary familial vascular and lymphatic malformations. We also summarize transgenic mouse models for some of these malformations. It seems reasonable to believe that novel therapeutic strategies will soon emerge for the treatment of hemangiomas and vascular malformations.
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Vasos Sanguíneos/anormalidades , Animais , Malformações Arteriovenosas/embriologia , Malformações Arteriovenosas/genética , Malformações Arteriovenosas/patologia , Vasos Sanguíneos/embriologia , Anormalidades Congênitas/embriologia , Anormalidades Congênitas/genética , Feminino , Substâncias de Crescimento/genética , Substâncias de Crescimento/fisiologia , Hemangioma/embriologia , Hemangioma/genética , Hemangioma/patologia , Humanos , Recém-Nascido , Linfangiogênese/genética , Sistema Linfático/anormalidades , Sistema Linfático/metabolismo , Linfedema/classificação , Linfedema/genética , Masculino , Camundongos , Camundongos Transgênicos , Neovascularização Fisiológica/genética , Receptores de Fatores de Crescimento/genética , Receptores de Fatores de Crescimento/fisiologia , Remissão EspontâneaRESUMO
Tufted angioma is a rare benign vascular lesion of unknown etiology which mainly affects children under 5 years. It is characterized by nodules or tufts of capillary-sized vessels in the dermis. Here we report the second familial occurrence of tufted angioma, with a mode of inheritance compatible with a monogenic autosomal dominant trait with reduced penetrance. A preliminary investigation was performed to exclude association between the predisposition and certain candidate genes including KDR (kinase insert domain receptor), TEK (TEK tyrosine kinase endothelial), ACVRL1 (activin receptor-like kinase 1), ENG (endoglin) and FLT4 (fms-like tyrosine kinase 4). KDR, ENG and FLT4 were all compatible with linkage, with haplotypes being shared between three affected individuals and the one obligate carrier available for testing. TEK and ACVRL1 could essentially be excluded. Finally, we provide definitive evidence for the existence of both blood and lymphatic vascular elements in the lesion.
Assuntos
Predisposição Genética para Doença , Hemangioma/genética , Receptores de Ativinas Tipo I/genética , Receptores de Activinas Tipo II , Antígenos CD , Pré-Escolar , Endoglina , Feminino , Genes Dominantes , Hemangioma/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Repetições de Microssatélites , Linhagem , Molécula-1 de Adesão Celular Endotelial a Plaquetas/biossíntese , Receptor TIE-2/genética , Receptores de Superfície Celular , Neoplasias Cutâneas/genética , Molécula 1 de Adesão de Célula Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genéticaRESUMO
To date, numerous genes have been identified which are involved in both tumour neovascularisation (angiogenesis) and tumour cell invasion, and most of them are also expressed to some extent under normal physiological conditions. However, little is known about how these genes co-express in these settings. This study was undertaken to quantitate mRNA levels in normal and malignant cervical tissues of nine selected genes (VEGF(121), VEGF(165), VEGF(189), VEGF-C, eIF-4E, b-FGF, TSP-2, MMP-2 and MMP-9) implicated in the above processes using real-time quantitative RT-PCR. In addition, the Spearman's rank correlation was used to determine their co-expression patterns. The transcript levels for the different VEGF-A splice variants (VEGF(121), VEGF(165), VEGF(189)) were at least 10-fold higher in the cancer cases, with the highest levels in the primary tumours demonstrating lympho-vascular space involvement. The lymphangiogenic factor VEGF-C and MMP-9 were upregulated 130- and 80-fold respectively in cervical cancers. The highest levels of VEGF-C mRNA were found in the lymph-node positive group. The transcript levels for b-FGF were similar in normal cervical tissue and early-stage cervical cancer, however, higher levels were found in the cervical cancers with advanced stage disease. Comparing gene transcript levels between recurrent and non-recurrent cervical cancer patients revealed significant differences (P=0.038) in transcript levels for the angiogenesis inhibitor TSP-2, with the highest levels in non-recurrent cases. Co-expression pattern analysis in normal cervical tissue revealed highly significant co-expressions (P<0.0001) between TSP-2 and most other genes analysed (VEGF(121), VEGF(165), VEGF-C, b-FGF and MMP-2). In cervical cancer, TSP-2 appears only to be highly co-expressed with MMP-2 (P<0.0001). In contrast to normal cervical tissue, we found a highly significant co-expression (P<0.0001) between MMP-9 and VEGF(189) in cervical cancer. The combined application of real-time quantitative RT-PCR and Spearman's rank correlation identifies gene transcripts which are simultaneously co-expressed. Our results revealed a significant co-expression between the angiogenesis inhibitor TSP-2 and most other genes analysed in normal cervical tissue. In cervical cancer, we found a strong upregulation of VEGF-C and MMP-9 mRNA, with a highly significant co-expression between MMP-9 and VEGF(189).
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Regulação Neoplásica da Expressão Gênica , Modelos Genéticos , Invasividade Neoplásica/genética , Proteínas de Neoplasias/genética , Neovascularização Patológica/genética , Neovascularização Fisiológica/genética , Neoplasias do Colo do Útero/genética , Colo do Útero/metabolismo , Fatores de Crescimento Endotelial/biossíntese , Fatores de Crescimento Endotelial/genética , Fator de Iniciação 4E em Eucariotos , Feminino , Fatores de Crescimento de Fibroblastos/biossíntese , Fatores de Crescimento de Fibroblastos/genética , Humanos , Linfocinas/biossíntese , Linfocinas/genética , Metaloproteinase 2 da Matriz/biossíntese , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/biossíntese , Metaloproteinase 9 da Matriz/genética , Proteínas de Neoplasias/biossíntese , Recidiva Local de Neoplasia , Fatores de Iniciação de Peptídeos/biossíntese , Fatores de Iniciação de Peptídeos/genética , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética , RNA Mensageiro/biossíntese , RNA Ribossômico 18S/biossíntese , RNA Ribossômico 18S/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcrição Gênica , Neoplasias do Colo do Útero/metabolismo , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
Exponential tumor growth is angiogenesis-dependent. Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are potent angiogenic inducers that act synergistically in vivo and in vitro. We assessed the effect of specific inhibitors of VEGF receptor (VEGFR)-2 tyrosine kinase activity in in vivo and in vitro models of VEGF- and bFGF-induced angiogenesis. In an implant mouse model of angiogenesis, VEGFR-2 inhibitors completely blocked angiogenesis induced by VEGF, and, surprisingly, also inhibited the effect of bFGF to various extents. In vitro, VEGF- and bFGF-induced bovine microvascular and aortic endothelial (BME and BAE) cell collagen gel invasion could be blocked by the VEGFR-2 inhibitors by 100 and approximately 90%, respectively. Similar results were obtained with VEGFR-1-IgG and VEGFR-3-IgG fusion proteins and with VEGFR-2 blocking antibodies. Both BME and BAE cells produce VEGF and VEGF-C, which is not modulated by bFGF. Thus, the unexpected inhibition of bFGF-induced angiogenesis by VEGFR-2 antagonists reveals a requirement for endogenous VEGF and VEGF-C in this process. These findings broaden the spectrum of mediators of angiogenesis that can be inhibited by VEGFR-2 antagonists and highlight the importance of these compounds as agents for inhibiting tumor growth sustained by both VEGF and bFGF.