RESUMO
Development of a new methodology to induce immunological chimerism after allogeneic hematopoietic cell (HC) transplantation in a rhesus macaque model is described. The chimeric state was achieved using a non-myeloablative, helical tomotherapy-based total lymphoid irradiation (TomoTLI) conditioning regimen followed by donor HC infusions between 1-haplotype matched donor/recipient pairs. The technique was tested as a feasibility study in an experimental group of seven rhesus macaques that received the novel TomoTLI tolerance protocol and HC allo-transplants. Two tomotherapy protocols were compared: TomoTLI (n = 5) and TomoTLI/total-body irradiation (TBI) (n = 2). Five of seven animals developed mixed chimerism. Three of five animals given the TomoTLI protocol generated transient mixed chimerism with no graft-versus-host disease (GVHD) with survival of 33, 152 and >180 days. However, the inclusion of belatacept in addition to a single fraction of TBI resulted in total chimerism and fatal GVHD in both animals, indicating an unacceptable conditioning regimen.
Assuntos
Quimerismo , Transplante de Células-Tronco Hematopoéticas , Tecido Linfoide/efeitos da radiação , Modelos Biológicos , Radioterapia de Intensidade Modulada/métodos , Animais , Doença Enxerto-Hospedeiro , Macaca mulatta , Modelos Animais , Transplante HomólogoRESUMO
The aim of this work was to study the effect of proton pencil beam scanning in the Bragg peak in the dose range of 0.1-1.5 Gy on the induction of cytogenetic damage in the bone marrow, reactive oxygen species (ROS) production in whole blood, and the state of lymphoid organs after total body irradiation of mice. Irradiation was carried out in the Prometeus proton synchrotron (Protvino) in the Bragg peak with proton energy at the output of 90-116 MeV. It was found that, under irradiation of mice in the range of low and medium doses of proton pencil beam scanning in the Bragg peak, the relative biological effectiveness (RBE) according to the criterion of cytogenetic changes was 1.15. In addition, it was found that the pathophysiological effect on the lymphoid organs and the production of ROS by blood cells were different as compared with the effect of X-rays.
Assuntos
Medula Óssea/efeitos da radiação , Tecido Linfoide/efeitos da radiação , Terapia com Prótons/métodos , Espécies Reativas de Oxigênio/metabolismo , Irradiação Corporal Total/métodos , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Tecido Linfoide/metabolismo , Tecido Linfoide/patologia , Masculino , Camundongos , Terapia com Prótons/efeitos adversos , Doses de Radiação , Eficiência Biológica Relativa , Irradiação Corporal Total/efeitos adversosRESUMO
Radiation induced lymphopenia is frequent and can be severe and durable. Although lymphocytes have long been known as highly radiosensitive cells, it is poorly characterized. Radiation-induced lymphopenia seems to affect lymphocyte subpopulations differently and seems to be influenced by radiation modalities. The depth and duration of lymphopenia depend on the location of the irradiation and the volumes of treatment. Importantly, radiation-induced lymphopenia has been associated with poorer prognosis in several tumor types. The knowledge about radiation-induced lymphopenia might lead to a rethinking of the modalities of radiotherapy and new approaches to restore lymphocytes counts.
Assuntos
Linfopenia/etiologia , Linfopenia/terapia , Linfócitos T/efeitos da radiação , Humanos , Subpopulações de Linfócitos/efeitos da radiação , Tecido Linfoide/efeitos da radiação , Neoplasias/sangue , Neoplasias/imunologia , Prognóstico , Tolerância a Radiação , Radioterapia/efeitos adversos , Radioterapia/métodos , Linfócitos T/fisiologiaRESUMO
High levels of uranium (U) exist in soil, water, and air in the Southwestern United States due, in part, to waste generated from more than 160,000 abandoned hard rock mines located in this region. As a result, many people living in this region are chronically exposed to U at levels that have been linked to detrimental health outcomes. In an effort to establish a relevant in vivo mouse model for future U immunotoxicity studies, we evaluated the tissue distribution of U in immune organs; blood, bone marrow, spleen, and thymus, as well as femur bones, kidneys, and liver, following a 60-d drinking water exposure to uranyl acetate (UA) in male and female C57BL/6J mice. Following the 60-d exposure, there was low overall tissue retention of U (<0.01%) at both the 5 and the 50 ppm (mg/L) oral concentrations. In both male and female mice, there was limited U accumulation in immune organs. U only accumulated at low concentrations in the blood and bone marrow of male mice (0.6 and 16.8 ng/g, respectively). Consistent with previous reports, the predominant sites of U accumulation were the femur bones (350.1 and 399.0 ng/g, respectively) and kidneys (134.0 and 361.3 ng/g, respectively) of male and female mice. Findings from this study provide critical insights into the distribution and retention of U in lymphoid tissues following chronic drinking water exposure to U. This information will serve as a foundation for immunotoxicological assessments of U, alone and in combination with other metals.
Assuntos
Exposição Ambiental , Tecido Linfoide/efeitos da radiação , Compostos Organometálicos/administração & dosagem , Urânio/toxicidade , Animais , Sangue/efeitos da radiação , Medula Óssea/efeitos da radiação , Camundongos , Radiação , Sudoeste dos Estados Unidos , Baço/efeitos da radiação , Timo/efeitos da radiaçãoRESUMO
Swelling is the most common symptom of extremities lymphedema. Clinical evaluation and laboratory analysis were conducted after far infrared radiation (FIR) treatment on the main four components of lymphedema: fluid, fat, protein, and hyaluronan. Far infrared radiation is a kind of hyperthermia therapy with several and additional benefits as well as promoting microcirculation flow and improving collateral lymph circumfluence. Although FIR therapy has been applied for several years on thousands of lymphedema patients, there are still few studies that have reported the biological effects of FIR on lymphatic tissue. In this research, we investigate the effects of far infrared rays on the major components of lymphatic tissue. Then, we explore the effectiveness and safety of FIR as a promising treatment modality of lymphedema. A total of 32 patients affected by lymphedema in stage II and III were treated between January 2015 and January 2016 at our department. After therapy, a significant decrease of limb circumference measurements was noted and improving of quality of life was registered. Laboratory examination showed the treatment can also decrease the deposition of fluid, fat, hyaluronan, and protein, improving the swelling condition. We believe FIR treatment could be considered as both an alternative monotherapy and a useful adjunctive to the conservative or surgical lymphedema procedures. Furthermore, the real and significant biological effects of FIR represent possible future applications in wide range of the medical field.
Assuntos
Raios Infravermelhos/uso terapêutico , Terapia com Luz de Baixa Intensidade/métodos , Linfedema/radioterapia , Tecido Linfoide/efeitos da radiação , Humanos , Microcirculação , Qualidade de VidaRESUMO
ATM phosphorylation of Mdm2-S394 is required for robust p53 stabilization and activation in DNA-damaged cells. We have now utilized Mdm2(S394A) knockin mice to determine that phosphorylation of Mdm2-S394 regulates p53 activity and the DNA damage response in lymphatic tissues in vivo by modulating Mdm2 stability. Mdm2-S394 phosphorylation delays lymphomagenesis in Eµ-myc transgenic mice, and preventing Mdm2-S394 phosphorylation obviates the need for p53 mutation in Myc-driven tumorigenesis. However, irradiated Mdm2(S394A) mice also have increased hematopoietic stem and progenitor cell functions, and we observed decreased lymphomagenesis in sub-lethally irradiated Mdm2(S394A) mice. These findings document contrasting effects of ATM-Mdm2 signaling on p53 tumor suppression and reveal that destabilizing Mdm2 by promoting its phosphorylation by ATM would be effective in treating oncogene-induced malignancies, while inhibiting Mdm2-S394 phosphorylation during radiation exposure or chemotherapy would ameliorate bone marrow failure and prevent the development of secondary hematological malignancies.
Assuntos
Carcinogênese/metabolismo , Carcinogênese/efeitos da radiação , Oncogenes , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Radiação Ionizante , Animais , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Medula Óssea/patologia , Medula Óssea/efeitos da radiação , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/efeitos da radiação , Tecido Linfoide/metabolismo , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Camundongos Transgênicos , Fosforilação/efeitos da radiação , Fosfosserina/metabolismo , Estabilidade Proteica/efeitos da radiação , Proteínas Proto-Oncogênicas c-myc/metabolismo , Tolerância a Radiação/efeitos da radiação , Transdução de Sinais/efeitos da radiação , Proteína Supressora de Tumor p53/metabolismoRESUMO
Ionizing irradiation is used routinely to induce myeloablation and immunosuppression. However, it has not been possible to evaluate the extent of ablation without invasive biopsy. For lymphoid recovery, peripheral blood (PB) lymphocytes (PBLs) have been used for analysis, but they represent <2% of cells in lymphoid tissues (LTs). Using a combination of single-photon emission computed tomography imaging and a radiotracer ((99m)Tc-labeled rhesus immunoglobulin G1 anti-CD4R1 (Fab')2), we sequentially imaged CD4(+) cell recovery in rhesus macaques following total body irradiation (TBI) and reinfusion of vector-transduced, autologous CD34(+) cells. Our results present for the first time a sequential, real-time, noninvasive method to evaluate CD4(+) cell recovery. Importantly, despite myeloablation of circulating leukocytes following TBI, total depletion of CD4(+) lymphocytes in LTs such as the spleen is not achieved. The impact of TBI on LTs and PBLs is discordant, in which as few as 32.4% of CD4(+) cells were depleted from the spleen. In addition, despite full lymphocyte recovery in the spleen and PB, lymph nodes have suboptimal recovery. This highlights concerns about residual disease, endogenous contributions to recovery, and residual LT damage following ionizing irradiation. Such methodologies also have direct application to immunosuppressive therapy and other immunosuppressive disorders, such as those associated with viral monitoring.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Tecido Linfoide/fisiologia , Tomografia Computadorizada de Emissão de Fóton Único , Condicionamento Pré-Transplante , Animais , Medula Óssea/efeitos da radiação , Antígenos CD4/genética , Contagem de Linfócito CD4 , Sistemas Computacionais , Genes Reporter , Genes Sintéticos , Vetores Genéticos , Proteínas de Fluorescência Verde/análise , Proteínas de Fluorescência Verde/genética , Imunoglobulina G/genética , Lentivirus/genética , Linfonodos/imunologia , Linfonodos/efeitos da radiação , Tecido Linfoide/diagnóstico por imagem , Tecido Linfoide/efeitos da radiação , Macaca mulatta , Imagem Multimodal , Especificidade de Órgãos , Quimera por Radiação , Baço/imunologia , Baço/efeitos da radiação , Tomografia Computadorizada por Raios X , Transdução Genética , Transplante Autólogo , Irradiação Corporal TotalRESUMO
The acute period after total body irradiation (TBI) is associated with an increased risk of infection, principally resulting from the loss of hematopoietic stem cells, as well as disruption of mucosal epithelial barriers. Although there is a return to baseline infection control coinciding with the apparent progressive recovery of hematopoietic cell populations, late susceptibility to infection in radiation-sensitive organs such as lung and kidney is known to occur. Indeed, pulmonary infections are particularly prevalent in hematopoietic cell transplant (HCT) survivors, in both adult and pediatric patient populations. Preclinical studies investigating late outcomes from localized thoracic irradiation have indicated that the mechanisms underlying pulmonary delayed effects are multifactorial, including exacerbated and persistent production of pro-inflammatory molecules and abnormal cross-talk among parenchymal and infiltrating immune and inflammatory cell populations. However, in the context of low-dose TBI, it is not clear whether the observed exacerbated response to infection remains contingent on these same mechanisms. It is possible instead, that after systemic radiation-induced injury, the susceptibility to infection may be independently related to defects in alternative organs that are revealed only through the challenge itself; indeed, we have hypothesized that this defect may be due to radiation-induced chronic effects in the structure and function of secondary lymphoid organs (SLO). In this study, we investigated the molecular and cellular alterations in SLO (i.e., spleen, mediastinal, inguinal and mesenteric lymph nodes) after TBI, and the time points when there appears to be immune competence. Furthermore, due to the high incidence of pulmonary infections in the late post-transplantation period of bone marrow transplant survivors, particularly in children, we focused on outcomes in mice irradiated as neonates, which served as a model for a pediatric population, and used the induction of adaptive immunity against influenza virus as a functional end point. We demonstrated that, in adult animals irradiated as neonates, high endothelial venule (HEV) expansion, generation of follicular helper T cells (TFH) and formation of splenic germinal centers (GC) were rapidly and, more importantly, persistently impaired in SLO, suggesting that the early-life exposure to sublethal radiation had long-lasting effects on the induction of humoral immunity. Although the neonatal TBI did not affect the overall outcome from influenza infection in the adults at the earlier time points assessed, we believe that they nonetheless contribute significantly to the increased mortality observed at subsequent late time points. Furthermore, we speculate that the detrimental and persistent impact on the induction of CD4 T- and B-cell responses in the mediastinal lymph nodes will decrease the animals' ability to respond to other aerial pathogens. Since many of these pathogens are normally cleared by antibodies, our findings provide an explanation for the susceptibility of survivors of childhood HCT to life-threatening respiratory tract infections. These findings have implications regarding the need for increased monitoring in pediatric hematopoietic cell transplant patients, since they indicate that there are ongoing and cumulative defects in SLO, which, importantly, develop during the immediate and early postirradiation period when patients may appear immunologically competent. The identification of changes in immune-related signals may offer bioindicators of progressive dysfunction, and of potential mechanisms that could be targeted so as to reduce the risk of infection from extracellular pathogens. Furthermore, these results support the potential susceptibility of the pediatric population to infection after sublethal irradiation in the context of a nuclear or radiological event.
Assuntos
Animais Recém-Nascidos , Tecido Linfoide/efeitos da radiação , Irradiação Corporal Total , Animais , Quimiocinas/metabolismo , Citocinas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Tecido Linfoide/anatomia & histologia , Tecido Linfoide/metabolismo , Tecido Linfoide/fisiologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Although SIGN-R1-mediated complement activation pathway has been shown to enhance the systemic clearance of apoptotic cells, the role of SIGN-R1 in the clearance of radiation-induced apoptotic cells has not been characterized and was investigated in this study. Our data indicated that whole-body γ-irradiation of mice increased caspase-3(+) apoptotic lymphocyte numbers in secondary lymphoid organs. Following γ-irradiation, SIGN-R1 and complements (C4 and C3) were simultaneously increased only in the mice spleen tissue among the assessed tissues. In particular, C3 was exclusively activated in the spleen. The delayed clearance of apoptotic cells was markedly prevalent in the spleen and liver of SIGN-R1 KO mice, followed by a significant increase of CD11b(+) cells. These results indicate that SIGN-R1 and complement factors play an important role in the systemic clearance of radiation-induced apoptotic innate immune cells to maintain tissue homeostasis after γ-irradiation.
Assuntos
Apoptose/fisiologia , Moléculas de Adesão Celular/fisiologia , Proteínas do Sistema Complemento/fisiologia , Lectinas Tipo C/fisiologia , Receptores de Superfície Celular/fisiologia , Irradiação Corporal Total , Animais , Apoptose/efeitos da radiação , Raios gama , Humanos , Linfócitos/citologia , Linfócitos/efeitos da radiação , Tecido Linfoide/citologia , Tecido Linfoide/efeitos da radiação , Macrófagos/citologia , Macrófagos/efeitos da radiação , Camundongos Endogâmicos C57BLRESUMO
PURPOSE: The threat of radiation exposure requires a mechanistic understanding of radiation-induced immune injury and recovery. The study objective was to evaluate responses to ionizing radiation in ovariectomized (surgically post-menopausal) female cynomolgus macaques. MATERIALS AND METHODS: Animals received a single total-body irradiation (TBI) exposure at doses of 0, 2 or 5 Gy with scheduled necropsies at 5 days, 8 weeks and 24 weeks post-exposure. Blood and lymphoid tissues were evaluated for morphologic, cellular, and molecular responses. RESULTS: Irradiated animals developed symptoms of acute hematopoietic syndrome, and reductions in thymus weight, thymopoiesis, and bone marrow cellularity. Acute, transient increases in plasma monocyte chemoattractant protein 1 (MCP-1) were observed in 5 Gy animals along with dose-dependent alterations in messenger ribonucleic acid (mRNA) signatures in thymus, spleen, and lymph node. Expression of T cell markers was lower in thymus and spleen, while expression of macrophage marker CD68 (cluster of differentiation 68) was relatively elevated in lymphoid tissues from irradiated animals. CONCLUSIONS: Ovariectomized female macaques exposed to moderate doses of radiation experienced increased morbidity, including acute, dose-dependent alterations in systemic and tissue-specific biomarkers, and increased macrophage/T cell ratios. The effects on mortality exceeded expectations based on previous studies in males, warranting further investigation.
Assuntos
Lesões Experimentais por Radiação/genética , Lesões Experimentais por Radiação/imunologia , Irradiação Corporal Total/efeitos adversos , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Quimiocina CCL2/sangue , Relação Dose-Resposta à Radiação , Feminino , Perfilação da Expressão Gênica , Tecido Linfoide/imunologia , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Macaca fascicularis , Macrófagos/imunologia , Macrófagos/efeitos da radiação , Ovariectomia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Lesões Experimentais por Radiação/patologia , Linfócitos T/imunologia , Linfócitos T/efeitos da radiaçãoRESUMO
Total lymphoid irradiation (TLI) with antithymocyte globulin (ATG) is a unique regimen that prepares recipients for allogeneic hematopoietic cell transplantation by targeting lymph nodes, while sparing large areas of the bone marrow. TLI is reported to increase the frequency of CD4(+)CD25(+)FoxP3(+) T-regulatory cells (Treg) relative to conventional T cells. In this study, barriers to hematopoietic stem cell (HSC) engraftment following this nonmyeloablative conditioning were evaluated. TLI/ATG resulted in profound lymphoablation but endogenous host HSC remained. Initial donor HSC engraftment occurred only in radiation exposed marrow sites, but gradually distributed to bone marrow outside the radiation field. Sustained donor engraftment required host lymphoid cells insofar as lymphocyte deficient Rag2γc(-/-) recipients had unstable engraftment compared with wild-type. TLI/ATG treated wild-type recipients had increased proportions of Treg that were associated with increased HSC frequency and proliferation. In contrast, Rag2γc(-/-) recipients who lacked Treg did not. Adoptive transfer of Treg into Rag2γc(-/-) recipients resulted in increased cell cycling of endogenous HSC. Thus, we hypothesize that Treg influence donor engraftment post-TLI/ATG by increasing HSC cell cycling, thereby promoting the exit of host HSC from the marrow niche. Our study highlights the unique dynamics of donor hematopoiesis following TLI/ATG, and the effect of Treg on HSC activity.
Assuntos
Sobrevivência de Enxerto/imunologia , Hematopoese/imunologia , Linfócitos T Reguladores/imunologia , Condicionamento Pré-Transplante/métodos , Animais , Medula Óssea/imunologia , Medula Óssea/metabolismo , Medula Óssea/efeitos da radiação , Sobrevivência de Enxerto/genética , Hematopoese/genética , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Tecido Linfoide/imunologia , Tecido Linfoide/efeitos da radiação , Camundongos , Camundongos Knockout , Linfócitos T Reguladores/metabolismo , Doadores de Tecidos , Quimeras de Transplante , Transplante HomólogoRESUMO
Lymphoid organs exhibit complex structures tightly related to their function. Surprisingly, although the thymic medulla constitutes a specialized microenvironment dedicated to the induction of T cell tolerance, its three-dimensional topology remains largely elusive because it has been studied mainly in two dimensions using thymic sections. To overcome this limitation, we have developed an automated method for full organ reconstruction in three dimensions, allowing visualization of intact mouse lymphoid organs from a collection of immunolabeled slices. We validated full organ reconstruction in three dimensions by reconstructing the well-characterized structure of skin-draining lymph nodes, before revisiting the complex and poorly described corticomedullary organization of the thymus. Wild-type thymi contain ~200 small medullae that are connected to or separated from a major medullary compartment. In contrast, thymi of immunodeficient Rag2(-/-) mice exhibit only ~20 small, unconnected medullary islets. Upon total body irradiation, medullary complexity was partially reduced and then recovered upon bone marrow transplantation. This intricate topology presents fractal properties, resulting in a considerable corticomedullary area. This feature ensures short distances between cortex and medulla, hence efficient thymocyte migration, as assessed by mathematical models. Remarkably, this junction is enriched, particularly in neonates, in medullary thymic epithelial cells expressing the autoimmune regulator. The emergence of a major medullary compartment is induced by CD4(+) thymocytes via CD80/86 and lymphotoxin-α signals. This comprehensive three-dimensional view of the medulla emphasizes a complex topology favoring efficient interactions between developing T cells and autoimmune regulator-positive medullary thymic epithelial cells, a key process for central tolerance induction.
Assuntos
Imageamento Tridimensional , Timo/citologia , Timo/patologia , Animais , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Transplante de Medula Óssea , Antígenos CD4/metabolismo , Células Epiteliais/metabolismo , Imageamento Tridimensional/métodos , Tecido Linfoide/citologia , Tecido Linfoide/efeitos dos fármacos , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Camundongos , Camundongos Knockout , Timócitos/metabolismo , Timo/efeitos dos fármacos , Timo/efeitos da radiaçãoRESUMO
BACKGROUND: Allogeneic bone marrow transplantation (allo-BMT) is a potentially curative therapy for a variety of hematologic diseases, but benefits, including graft-versus-tumor (GVT) activity are limited by graft-versus-host-disease (GVHD). Carcinoembryonic antigen related cell adhesion molecule 1 (Ceacam1) is a transmembrane glycoprotein found on epithelium, T cells, and many tumors. It regulates a variety of physiologic and pathological processes such as tumor biology, leukocyte activation, and energy homeostasis. Previous studies suggest that Ceacam1 negatively regulates inflammation in inflammatory bowel disease models. METHODS: We studied Ceacam1 as a regulator of GVHD and GVT after allogeneic bone marrow transplantation (allo-BMT) in mouse models. In vivo, Ceacam1(-/-) T cells caused increased GVHD mortality and GVHD of the colon, and greater numbers of donor T cells were positive for activation markers (CD25(hi), CD62L(lo)). Additionally, Ceacam1(-/-) CD8 T cells had greater expression of the gut-trafficking integrin α(4)ß(7), though both CD4 and CD8 T cells were found increased numbers in the gut post-transplant. Ceacam1(-/-) recipients also experienced increased GVHD mortality and GVHD of the colon, and alloreactive T cells displayed increased activation. Additionally, Ceacam1(-/-) mice had increased mortality and decreased numbers of regenerating small intestinal crypts upon radiation exposure. Conversely, Ceacam1-overexpressing T cells caused attenuated target-organ and systemic GVHD, which correlated with decreased donor T cell numbers in target tissues, and mortality. Finally, graft-versus-tumor survival in a Ceacam1(+) lymphoma model was improved in animals receiving Ceacam1(-/-) vs. control T cells. CONCLUSIONS: We conclude that Ceacam1 regulates T cell activation, GVHD target organ damage, and numbers of donor T cells in lymphoid organs and GVHD target tissues. In recipients of allo-BMT, Ceacam1 may also regulate tissue radiosensitivity. Because of its expression on both the donor graft and host tissues, this suggests that targeting Ceacam1 may represent a potent strategy for the regulation of GVHD and GVT after allogeneic transplantation.
Assuntos
Transplante de Medula Óssea/imunologia , Antígeno Carcinoembrionário/metabolismo , Doença Enxerto-Hospedeiro/imunologia , Efeito Enxerto vs Tumor/imunologia , Animais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos da radiação , Polaridade Celular/efeitos da radiação , Proliferação de Células/efeitos da radiação , Citotoxicidade Imunológica/efeitos da radiação , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Doença Enxerto-Hospedeiro/complicações , Doença Enxerto-Hospedeiro/mortalidade , Efeito Enxerto vs Tumor/efeitos da radiação , Humanos , Integrinas/metabolismo , Intestino Delgado/patologia , Intestino Delgado/efeitos da radiação , Ativação Linfocitária/imunologia , Ativação Linfocitária/efeitos da radiação , Contagem de Linfócitos , Tecido Linfoide/citologia , Tecido Linfoide/efeitos da radiação , Camundongos , Especificidade de Órgãos/imunologia , Especificidade de Órgãos/efeitos da radiação , Lesões Experimentais por Radiação/complicações , Lesões Experimentais por Radiação/metabolismo , Lesões Experimentais por Radiação/patologia , Radiação Ionizante , Transplante HomólogoRESUMO
OBJECTIVE: Hematopoietic regeneration is regulated by cell survival proteins, such as the Bcl-2 family. Bid, a BH3-only protein of the Bcl-2 family, has multiple cellular functions and is involved in a variety of physiological or pathological conditions. We attempted to define its role in hematopoietic cell repopulation under the stress condition of bone marrow transplantation. MATERIALS AND METHODS: We performed conventional or competitive bone marrow transplantation with donor hematopoietic cells from Bid(-/-) or Bid(+/+) mice. Flow cytometry was used for quantification of hematopoietic stem cells, hematopoietic progenitor cells, and differentiated cells in different lineages (T, B, and myeloid cells). Single cell culture and homing assays were performed to further evaluate hematopoietic stem cell functions. Hematopoietic progenitor cells were also measured by the colony-forming cell culture. RESULTS: Contrary to the widely recognized role of Bid as a pro-apoptotic protein, the absence of Bid significantly reduced the reconstitution of donor hematopoietic cells in γ-irradiated recipients. Interestingly, however, numbers of hematopoietic stem cells and hematopoietic progenitor cells and their functions were not overtly altered. Instead, the regeneration of donor T and B cells was significantly impaired in the absence of Bid. Further analysis indicated an accumulation of the triple-negative T-cell population in the thymus, and pro-B cells in the bone marrow. CONCLUSIONS: Our current study demonstrates a positive impact of Bid on hematopoietic regeneration mainly due to its unique effects on donor lymphopoiesis in the transplant recipients.
Assuntos
Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/fisiologia , Raios gama , Tecido Linfoide/efeitos da radiação , Regeneração , Animais , Transplante de Medula Óssea , Citometria de Fluxo , Tecido Linfoide/patologia , Camundongos , Camundongos TransgênicosRESUMO
AIMS: Radiation therapy (RT) is used in the treatment of approximately half of all cancer patients. Although there have been great improvements in tumor localization and the technical accuracy of RT delivery, some RT patients still have idiosyncratic hypersensitivity to ionizing radiation (IR) in their normal tissues. Although much effort has been expended in the search for assays that could detect radiosensitive individuals prior to treatment and facilitate tailored therapy; a suitable and clinically practical predictive assay has yet to be realized. Since DNA double-strand breaks (DSB) are a major lesion caused by IR, we hypothesized that radiation hypersensitive individuals might be deficient in the repair of such lesions. METHODS: To test this hypothesis we quantitatively and functionally characterized DSB repair of the two major non-homologous end-joining (NHEJ) sub-pathways in a pilot study using a plasmid repair reconstitution assay in lymphoblastoid and fibroblast cell lines from radiosensitive cancer patients and controls. Experiments using well-characterized mammalian DSB repair mutants demonstrated the ability of the assay to distinguish NHEJ sub-pathways. The proportion of direct end-joining repair compared with that of microhomology-directed repair was used as a functional end-point of DSB repair competence in the different cell lines. RESULTS: We found that the overall level of NHEJ sub-pathway repair competency was similar in cell lines from radiosensitive patients and controls. CONCLUSION: These data suggest that this assay in these cell lineages has limited usefulness as a predictive screen for the endogenous DNA DSB repair competency of radiosensitive cancer patients' cells but can usefully characterize major cellular DSB repair phenotypes.
Assuntos
Linhagem da Célula , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Reparo do DNA/efeitos da radiação , Fibroblastos/efeitos da radiação , Tecido Linfoide/efeitos da radiação , Neoplasias/patologia , Tolerância a Radiação/genética , Estudos de Casos e Controles , Células Cultivadas , Reparo do DNA/genética , Humanos , Tecido Linfoide/citologia , Neoplasias/genética , Neoplasias/radioterapia , Radiação IonizanteRESUMO
A previous in vitro study showed that sphingosine-1-phosphate (S1P), a ceramide antagonist, preserved endothelial cells in culture from radiation-induced apoptosis. We proposed to validate the role of S1P in tissue radioprotection by inhibiting acute gastrointestinal (GI) syndrome induced by endothelial cell apoptosis after high dose of radiation. Retro-orbital S1P was injected in mice exposed to 15 Gy, a dose-inducing GI syndrome within 10 days. Overall survival and apoptosis on intestines sections were studied. Intestinal cell type targeted by S1P and early molecular survival pathways were researched using irradiated in vitro cell models and in vivo mouse models. We showed that retro-orbital S1P injection before irradiation prevented GI syndrome by inhibiting endothelium collapse. We defined endothelium as a specific therapeutic target because only these cells and not intestinal epithelial cells, or B and T lymphocytes, were protected. Pharmacologic approaches using AKT inhibitor and pertussis toxin established that S1P affords endothelial cell protection in vitro and in vivo through a mechanism involving AKT and 7-pass transmembrane receptors coupled to Gi proteins. Our results provide strong pharmacologic and mechanistic proofs that S1P protects endothelial cells against acute radiation enteropathy.
Assuntos
Apoptose/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Animais , Apoptose/efeitos da radiação , Western Blotting , Medula Óssea/efeitos dos fármacos , Medula Óssea/patologia , Medula Óssea/efeitos da radiação , Linhagem Celular , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/patologia , Trato Gastrointestinal/efeitos da radiação , Humanos , Imuno-Histoquímica , Intestino Delgado/citologia , Intestino Delgado/efeitos da radiação , Tecido Linfoide/efeitos dos fármacos , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/efeitos dos fármacos , Fosforilação/efeitos da radiação , Lesões Experimentais por Radiação/tratamento farmacológico , Esfingosina/farmacologia , SíndromeRESUMO
Over-expression of manganese superoxide dismutase (MnSOD) protects tissues from radiation. M40403 is a stable non-peptidyl mimetic of MnSOD that crosses cell membranes and is effective in reducing experimental inflammation. Male BALB/c mice were injected intraperitoneally (i.p.) and subcutaneously (s.c.) with M40403, 30 min before 6.5, 7.5 and 8.5 Gy total body irradiation (TBI). Whereas all control injected mice died after receiving 8.5 Gy TBI by day 17, 30 day survival of mice pre-treated i.p. with 40, 30, 20 or 10 mg/kg was 100%, 90%, 81% and 25%, respectively. The Dose Reduction Factor 50/30 for animals treated with 30 mg M40403 s.c. 30 min prior to TBI was 1.41. Decreased apoptosis of the large and particularly the small bowel and marked recovery of both lymphoid and hematopoietic tissues occurred in the M40403 pre-treated animals. M40403 is effective in reducing TBI-induced tissue destruction and has potential as a new radioprotective agent.
Assuntos
Materiais Biomiméticos/farmacologia , Sistema Hematopoético/efeitos dos fármacos , Tecido Linfoide/efeitos dos fármacos , Compostos Organometálicos/farmacologia , Protetores contra Radiação/farmacologia , Superóxido Dismutase , Irradiação Corporal Total , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Relação Dose-Resposta à Radiação , Sistema Hematopoético/patologia , Sistema Hematopoético/efeitos da radiação , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Masculino , Manganês , Camundongos , Camundongos Endogâmicos BALB C , Compostos Organometálicos/química , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Protetores contra Radiação/química , Taxa de SobrevidaRESUMO
A comparative analysis of low linear energy transfer (LET) gamma-radiation-induced damage in the lymphatic tissue of a tropical seasonal breeder, Indian palm squirrel (Funambulus pennanti), during its reproductively active phase (RAP) and inactive phase (RIP) was performed with simultaneous investigation of the effects of long-term melatonin pre-treatment (100 microg/100 g body weight). A total of 120 squirrels (60 during RAP and 60 during RIP) were divided into 12 groups and sacrificed at 4, 24, 48, 72 and 168 h following 5 Gy gamma-radiation exposure; control groups were excluded from exposure. Total leukocyte count and absolute lymphocyte count (ALC) and melatonin only of peripheral blood, stimulation index, thiobarbituric-acid-reactive substances (TBARS) level, superoxide dismutase (SOD) activity, and the apoptotic index of spleen as analysed by terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labelling (TUNEL) noted at observed time-points were significantly reduced in melatonin pre-treated groups during RAP and RIP. Long-term melatonin pre-treatment mitigated radiation-induced alterations more prominently during RIP, as assessed by ALC, TBARS, SOD, TUNEL and caspase-3 activity, at some time-points. Our results demonstrate an inhibitory role of melatonin on caspase-3 activity in splenocytes during RAP and RIP following gamma-radiation-induced caspase-mediated apoptosis. Hence, we propose that melatonin might preserve the viability of immune cells of a seasonal breeder against background radiation, which is constantly present in the environment.
Assuntos
Antioxidantes/farmacologia , Raios gama/efeitos adversos , Transferência Linear de Energia , Tecido Linfoide/efeitos da radiação , Melatonina/farmacologia , Melatonina/fisiologia , Lesões Experimentais por Radiação/prevenção & controle , Reprodução/fisiologia , Baço/efeitos dos fármacos , Animais , Antioxidantes/administração & dosagem , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Caspase 3/metabolismo , Ritmo Circadiano/imunologia , Fragmentação do DNA , Contagem de Leucócitos , Linfócitos/efeitos dos fármacos , Linfócitos/imunologia , Linfócitos/efeitos da radiação , Tecido Linfoide/efeitos dos fármacos , Masculino , Melatonina/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Eficiência Biológica Relativa , Reprodução/imunologia , Sciuridae/imunologia , Sciuridae/fisiologia , Comportamento Sexual Animal/efeitos dos fármacos , Baço/metabolismo , Baço/efeitos da radiação , Superóxido Dismutase/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Fatores de TempoRESUMO
In breast cancer radiotherapy, the internal mammary lymphatic chain is treated in the target volume in a group of patients with high-risk criteria. Because of the variability of the anatomic region and structures in the irradiation field, there are a number of different techniques in breast radiotherapy. While irradiating the target volume, we also consider minimizing the dose to critical structures such as heart, lung, and contralateral breast tissue. In this study, we evaluated the dose distribution of different radiotherapy techniques in patients with left-sided breast cancer who had breast-conserving surgery. A three-dimensional computerized planning system (3DCPS) was used for each patient to compare wide-field, oblique photon-electron, and perpendicular photon-electron techniques in terms of dose homogeneities in the target volume; the doses received by the contralateral breast, heart, and lung; and the coverage of the internal mammary chain. Data from 3DCPS were controlled by the Rando-phantom and thermoluminescence dosimetry. Critical structures were irradiated with acceptable dose percentages in addition to the internal mammary chain with both wide-field and photon-electron techniques. We detected more frequent hot spots in the oblique photon-electron technique than in the other techniques, and this situation necessitated changing the junctions. The wide-field technique was easy to perform and exposed less radiation dose to the heart than photon-electron techniques. In conclusion, we suggest the use of the wide-field technique in breast irradiation when the internal mammary area is in the target volume.
Assuntos
Artefatos , Neoplasias da Mama/radioterapia , Planejamento da Radioterapia Assistida por Computador/métodos , Mama/efeitos dos fármacos , Mama/patologia , Neoplasias da Mama/cirurgia , Relação Dose-Resposta à Radiação , Feminino , Coração/efeitos da radiação , Humanos , Pulmão/efeitos da radiação , Tecido Linfoide/efeitos da radiação , Lesões por Radiação/prevenção & controle , Dosagem Radioterapêutica , Planejamento da Radioterapia Assistida por Computador/normas , Dosimetria TermoluminescenteRESUMO
BACKGROUND: Chronic alcoholics have increased susceptibility to and severity of infection, which are likely to be a result of impaired immune defense mechanisms. The contribution of dendritic cells (DC) to these immune defense changes is not well understood. Alterations in DC numbers, dendropoiesis, and lifespan have not been specifically studied in vivo in chronic ethanol (EtOH) exposure models. As DC play an essential role in initiating immune responses, alterations in these DC characteristics would help explain changes observed in adaptive immune responses. METHODS: Mice received 20% EtOH (w/v) in the drinking water for up to 28 weeks, with mouse chow ad libitum. In EtOH-fed and water control mice, DC were enumerated by flow cytometry. The effect of EtOH on DC precursor numbers was determined by differentiation in vitro in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4, and the effect of an EtOH environment on untreated DC differentiation was measured following bone marrow transfer to irradiated hosts. DC turnover rate was also examined by bromodeoxyuridine incorporation and loss. RESULTS: The percentage and absolute numbers of DC were decreased in spleen and increased in thymus beginning as early as 4 weeks of EtOH feeding. In addition, the overall cellularity of spleen and thymus were altered by this regimen. However, chronic EtOH consumption did not adversely affect DC precursor numbers, differentiation abilities, or turnover rates. CONCLUSIONS: Decreased splenic DC numbers observed following chronic murine EtOH consumption are not because of altered DC precursor numbers or differentiation, nor increased DC turnover rate. Similarly, increased thymic DC numbers are not the result of alterations in DC precursor differentiation or turnover rate. Compartment size plays a role in determining splenic and thymic DC numbers following chronic EtOH feeding. EtOH-induced alterations in total DC numbers provide several mechanisms to partially explain why chronic alcoholics have increased susceptibility to infections.