RESUMO
Polymorphisms of HLA genes, which play a crucial role in presenting peptides with diverse sequences in their peptide-binding pockets, are also thought to affect HLA gene expression, as many studies have reported associations between HLA gene polymorphisms and their expression levels. In this study, we devised an ectopic expression assay for the HLA class I genes in the context of the entire gene, and used the assay to show that the HLA-C*03:03:01 and C*04:01:01 polymorphic differences observed in association studies indeed cause different levels of RNA expression. Subsequently, we investigated the C*03:23N null allele, which was previously noted for its reduced expression, attributed to an alternate exon 3 3' splice site generated by G/A polymorphism at position 781 within the exon 3. We conducted a thorough analysis of the splicing patterns of C*03:23N, and revealed multiple aberrant splicing, including the exon 3 alternative splicing, which overshadowed its canonical counterpart. After confirming a significant reduction in RNA levels caused by the G781A alteration in our ectopic assay, we probed the function of the G-rich sequence preceding the canonical exon 3 3' splice site. Substituting the G-rich sequence with a typical pyrimidine-rich 3' splice site sequence on C*03:23N resulted in a marked elevation in RNA levels, likely due to the enhanced preference for the canonical exon 3 3' splice site over the alternate site. However, the same substitution led to a reduction in RNA levels for C*03:03:01. These findings suggested the dual roles of the G-rich sequence in RNA expression, and furthermore, underscore the importance of studying polymorphism effects within the framework of the entire gene, extending beyond conventional mini-gene reporter assays.
Assuntos
Antígenos HLA-C , Nucleotídeos , Antígenos HLA-C/genética , Sítios de Splice de RNA/genética , Splicing de RNA , Processamento AlternativoRESUMO
The mammalian brain contains multiple types of neuronal cells with complex assemblies and distinct structural and functional properties encoded by divergent gene programs. There is increasing evidence that alternative splicing (AS) plays fundamental roles in transcriptomic diversity and specifying synaptic properties of each neuronal cell type. However, the mechanisms underlying AS regulation and whether it controls synapse formation across GABAergic interneurons have not been fully elucidated. Here we show the differential expression levels of Sam68-like molecule 2 (SLM2), a major splicing regulator of neurexin (NRX), in GABAergic neuronal subtypes and its contribution to GABAergic synapse specification. Cortical SLM2 is strongly expressed not only in excitatory neurons but also in a subpopulation of GABAergic interneurons, especially in VIP-positive neurons that are originated from late-born caudal ganglionic eminence (GE)- derived cells. Using artificial synapse formation assay, we found that GE containing cortices form a strong synapse with LRRTM2, a trans-synaptic receptor of the alternatively spliced segment 4 (AS4)(-) of NRX. SLM2 knock-down reduced the NRX AS4(-) isoform expression and hence weaken LRRTM2-induced synapse formation. The addition of NRX AS4(-) was sufficient to rescue the synaptic formation by LRRTM2 in SLM2 knock-down neurons. Thus, our findings suggest a novel function of SLM2 in modifying network formation of a specific population of GABAergic interneurons and contribute to a better understanding of the roles AS plays in regulating synapse specificity and neuronal molecular diversity.
Assuntos
Processamento Alternativo , Neurônios GABAérgicos , Animais , Interneurônios , Mamíferos , Neurogênese , Sinapses/fisiologiaRESUMO
There has been an increase in the usage of heat-not-burn (HNB) cigarette products. However, their effects on alveolar epithelial cells (AECs) remain unknown. AECs are the target cells of conventional cigarette smoking-related respiratory diseases such as chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis and lung cancer whose pathogenesis involves oxidative stress. In this study, primary rat AECs were isolated, cultured and stimulated by HNB cigarette smoke extract (CSE). Our data indicate that rat AECs exposed to HNB CSE induced oxidative stress response genes (e.g. Hmox-1, Gsta1, Gsta3 and Nqo1). We also compared the oxidative stress response between two different types of AECs, alveolar type I-like (ATI-like) cells and type II (ATII) cells, and between two different types of cigarette, HNB cigarettes and conventional cigarettes. The expressions of Gsta1, Gsta3 and Nqo1 were higher in ATII cells than ATI-like cells in response to HNB and conventional cigarettes, but there was no significant difference in their expression levels between HNB cigarette and conventional cigarette. Taken together, our results suggest that HNB cigarettes have the similar potential as conventional cigarette products to induce oxidative stress response in AECs.
Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Fumar Cigarros/efeitos adversos , Estresse Oxidativo/efeitos dos fármacos , Alvéolos Pulmonares/efeitos dos fármacos , Células Epiteliais Alveolares/patologia , Animais , Modelos Animais de Doenças , Sistemas Eletrônicos de Liberação de Nicotina , Temperatura Alta/efeitos adversos , Humanos , Oxirredução/efeitos dos fármacos , Cultura Primária de Células , Alvéolos Pulmonares/patologia , Doença Pulmonar Obstrutiva Crônica/induzido quimicamente , Ratos , Fumaça/efeitos adversos , Nicotiana/efeitos adversosRESUMO
Neonicotinoids such as acetamiprid (ACE) belong to a new and widely used single class of pesticides. Neonicotinoids mimic the chemical structure of nicotine and share agonist activity with the nicotine acetylcholine receptor (nAchR). Neonicotinoids are widely considered to be safe in humans; however, they have recently been implicated in a number of human health disorders. A wide range of musculoskeletal and neuromuscular disorders associated with high doses of neonicotinoids administered to animals have also been reported. Consequently, we used a mouse model to investigate the response of the central nervous system to ACE treatment. Our results show that exposure to ACE-containing water for three or seven days (decuple and centuple of no observable adverse effect level (NOAEL)/day) caused a decrease in body weight in 10-week old A/JJmsSlc (A/J) mice. However, the treatments did not affect brain histology or expression of CD34. ACE concentrations were significantly higher in the midbrain of ACE-treated mice than that of the normal and vehicle groups. Expression levels of α7, α4, and ß2 nAChRs were found to be low in the olfactory bulb and midbrain of normal mice. Furthermore, in the experimental group (centuple ACE-containing water for seven days), ß2 nAChR expression decreased in many brain regions. Information regarding the amount of accumulated ACE and expression levels of the acetylcholine receptor in each region of the brain is important for understanding any clinical symptoms that may be associated with ACE exposure.
Assuntos
Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Piridinas/farmacologia , Piridinas/farmacocinética , Animais , Antígenos CD34/metabolismo , Peso Corporal/efeitos dos fármacos , Perfilação da Expressão Gênica , Imuno-Histoquímica , Inseticidas/metabolismo , Inseticidas/farmacocinética , Inseticidas/farmacologia , Masculino , Camundongos , Neonicotinoides , Piridinas/metabolismo , Receptores Nicotínicos/metabolismoRESUMO
Mitochondrial oxidative stress is considered as a key accelerator of fibrosis in various organs including the liver. However, the production of oxidative stress and progression of liver fibrosis may merely represent the independent consequences of hepatocellular injury caused by the primary disease. Because of a lack of appropriate experimental models to evaluate the sole effects of oxidative stress, it is virtually unknown whether this stress is causatively linked to the progression of liver fibrosis. Here, we examined the direct effects of mitochondrial reactive oxygen species (ROS) on the progression of high fat/calorie diet-induced steatohepatitis using Tet-mev-1 mice, in which a mutated succinate dehydrogenase transgene impairs the mitochondrial electron transport and generates an excess amount of ROS in response to doxycycline administration. Wild type and Tet-mev-1 mice that had been continuously given doxycycline-containing water were subsequently fed either normal chow or a cholesterol-free high-fat/high-sucrose diet for 4 months at approximately 1 or 2 years of age. Histopathological examinations indicated that neither the mitochondrial ROS induced in Tet-mev-1 mice nor the feeding of wild type animals with high-fat/high-sucrose diet alone caused significant liver fibrosis. Only when the Tet-mev-1 mice were fed a high-fat/high-sucrose diet, it induced lipid peroxidation in hepatocytes and enhanced hepatic CC chemokine expression. These events were accompanied by increased infiltration of CCR5-positive cells and activation of myofibroblasts, resulting in extensive liver fibrosis. Interestingly, this combinatorial effect of mitochondrial ROS and excess fat/calorie intake on liver fibrosis was observed only in 2-year-old Tet-mev-1 mice, not in the 1-year-old animals. Collectively, these results indicate that mitochondrial ROS in combination with excess fat/calorie intake accelerates liver fibrosis by enhancing CC chemokine production in aged animals. We have provided a good experimental model to explore how high fat/calorie intake increases the susceptibility to nonalcoholic steatohepatitis in aged individuals who have impaired mitochondrial adaptation.
Assuntos
Quimiocinas/biossíntese , Hepatopatia Gordurosa não Alcoólica/metabolismo , Estresse Oxidativo , Animais , Células Cultivadas , Dieta Hiperlipídica/efeitos adversos , Progressão da Doença , Ingestão de Energia , Ontologia Genética , Hepatócitos/metabolismo , Peroxidação de Lipídeos , Fígado/imunologia , Fígado/metabolismo , Macrófagos/metabolismo , Masculino , Potencial da Membrana Mitocondrial , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Anotação de Sequência Molecular , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/imunologia , Hepatopatia Gordurosa não Alcoólica/patologia , Espécies Reativas de Oxigênio/metabolismo , Receptores CCR5/metabolismoRESUMO
INTRODUCTION: One of the causes for poor vasculogenesis of diabetes mellitus (DM) is known to rise from the dysfunction of bone marrow-derived endothelial progenitor cells (BM EPCs). However, the origin of its cause is less understood. We aimed to investigate the effect of oxidative stress in early stage of diabetic BM-EPC and whether its vasculogenic dysfunction is caused by oxidative stress. METHODS: Bone marrow c-Kit+Sca-1+Lin- (BM-KSL) cells were sorted from control and streptozotocin-induced diabetic C57BL6J mice by flow cytometry. BM-KSLs were then assessed for vasculogenic potential (colony forming assay; EPC-CFA), accumulation of intracellular ROS (CM-H2DCFDA), carbonylated protein (ELISA), anti-oxidative enzymes expression (RT-qPCR) and catalase activity (Amplex Red). RESULTS: Compared to control, DM BM-KSL had significantly lower EPC-CFUs in both definitive EPC-CFU and total EPC-CFU (p < 0.05). Interestingly, the oxidative stress level of DM BM-KSL was comparable and was not significantly different to control followed by increased in anti-oxidative enzymes expression and catalase activity. CONCLUSIONS: Primitive BM-EPCs showed vasculogenic dysfunction in early diabetes. However the oxidative stress is not denoted as the major initiating factor of its cause. Our results suggest that primitive BM-KSL cell has the ability to compensate oxidative stress levels in early diabetes by increasing the expression of anti-oxidative enzymes.
RESUMO
Sequences of long-interspersed elements (LINE-1, L1) make up â¼17% of the human genome. De novo insertions of retrotransposition-active L1s can result in genetic diseases. It has been recently shown that the homozygous inactivation of two adjacent genes SLCO1B1 and SLCO1B3 encoding organic anion transporting polypeptides OATP1B1 and OATP1B3 causes a benign recessive disease presenting with conjugated hyperbilirubinemia, Rotor syndrome. Here, we examined SLCO1B1 and SLCO1B3 genes in six Japanese diagnosed with Rotor syndrome on the basis of laboratory data and laparoscopy. All six Japanese patients were homozygous for the c.1738C>T nonsense mutation in SLCO1B1 and homozygous for the insertion of a â¼6.1-kbp L1 retrotransposon in intron 5 of SLCO1B3, which altogether make up a Japanese-specific haplotype. RNA analysis revealed that the L1 insertion induced deleterious splicing resulting in SLCO1B3 transcripts lacking exon 5 or exons 5-7 and containing premature stop codons. The expression of OATP1B1 and OATP1B3 proteins was not detected in liver tissues. This is the first documented case of a population-specific polymorphic intronic L1 transposon insertion contributing to molecular etiology of recessive genetic disease. Since L1 activity in human genomes is currently seen as a major source of individual genetic variation, further investigations are warranted to determine whether this phenomenon results in other autosomal-recessive diseases.
Assuntos
Doenças Genéticas Inatas/genética , Hiperbilirrubinemia Hereditária/genética , Íntrons , Elementos Nucleotídeos Longos e Dispersos , Adulto , Feminino , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado , Masculino , Pessoa de Meia-Idade , Transportadores de Ânions Orgânicos/genética , Transportadores de Ânions Orgânicos Sódio-Independentes/genética , Fenótipo , Retroelementos , Membro 1B3 da Família de Transportadores de Ânion Orgânico Carreador de SolutoRESUMO
Recent advances in cancer biology have revealed that many malignancies possess a hierarchal system, and leukemic stem cells (LSC) or leukemia-initiating cells (LIC) appear to be obligatory for disease progression. Acute promyelocytic leukemia (APL), a subtype of acute myeloid leukemia characterized by the formation of a PML-RARα fusion protein, leads to the accumulation of abnormal promyelocytes. In order to understand the precise mechanisms involved in human APL leukemogenesis, we established a humanized in vivo APL model involving retroviral transduction of PML-RARA into CD34(+) hematopoietic cells from human cord blood and transplantation of these cells into immunodeficient mice. The leukemia well recapitulated human APL, consisting of leukemic cells with abundant azurophilic abnormal granules in the cytoplasm, which expressed CD13, CD33 and CD117, but not HLA-DR and CD34, were clustered in the same category as human APL samples in the gene expression analysis, and demonstrated sensitivity to ATRA. As seen in human APL, the induced APL cells showed a low transplantation efficiency in the secondary recipients, which was also exhibited in the transplantations that were carried out using the sorted CD34- fraction. In order to analyze the mechanisms underlying APL initiation and development, fractionated human cord blood was transduced with PML-RARA. Common myeloid progenitors (CMP) from CD34(+)/CD38(+) cells developed APL. These findings demonstrate that CMP are a target fraction for PML-RARA in APL, whereas the resultant CD34(-) APL cells may share the ability to maintain the tumor.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Leucemia Promielocítica Aguda/patologia , Proteínas de Fusão Oncogênica/metabolismo , Animais , Antígenos CD34/metabolismo , Diferenciação Celular , Modelos Animais de Doenças , Sangue Fetal/citologia , Células-Tronco Hematopoéticas/citologia , Humanos , Leucemia Promielocítica Aguda/mortalidade , Antígenos Comuns de Leucócito/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Proteínas de Fusão Oncogênica/genética , Taxa de Sobrevida , Transplante HeterólogoRESUMO
Activation-induced cytidine deaminase (AID) is essential for class switch recombination and somatic hypermutation. Its deregulated expression acts as a genomic mutator that can contribute to the development of various malignancies. During treatment with imatinib mesylate (IM), patients with chronic myeloid leukemia often develop hypogammaglobulinemia, the mechanism of which has not yet been clarified. Here, we provide evidence that class switch recombination on B-cell activation is apparently inhibited by IM through down-regulation of AID. Furthermore, expression of E2A, a key transcription factor for AID induction, was markedly suppressed by IM. These results elucidate not only the underlying mechanism of IM-induced hypogammaglobulinemia but also its potential efficacy as an AID suppressor.
Assuntos
Citidina Desaminase/antagonistas & inibidores , Switching de Imunoglobulina/efeitos dos fármacos , Piperazinas/farmacologia , Pirimidinas/farmacologia , Animais , Benzamidas , Citidina Desaminase/metabolismo , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/imunologia , Avaliação Pré-Clínica de Medicamentos , Mesilato de Imatinib , Imunossupressores/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Piperazinas/uso terapêutico , Pirimidinas/uso terapêutico , Recombinação Genética/efeitos dos fármacos , Recombinação Genética/imunologia , Ovinos , Hipermutação Somática de Imunoglobulina/efeitos dos fármacos , Resultado do TratamentoRESUMO
OBJECTIVE: Hematopoietic stem cells (HSCs) consist of heterogenous subpopulations, one of which is CD34(-) HSCs. Recent development of successful engraftment by intra-bone marrow transplantation revealed severe combined immunodeficiency (scid) mouse-repopulating cell (SRC) activity in human CD34(-) cord blood (CB) cells. On the other hand, CD34(-) cells from bone marrow (BM) cells remain relatively undefined. Here, we investigated pre-SRC populations in human BM CD34(-) cells and the effect of the niche-related factor, angiopoietin-1, on them. METHODS: Two populations in BM CD34(-) cells (namely M cells and S cells) were purified by flow cytometry. Then, they were cocultured with six growth factors on the hematopoietic-supportive mouse BM stromal cell line, HESS-5 or AHESS-5 that were engineered to produce human angiopoietin-1, because we detected Tie2 expression on M cells and S cells. Cultured cells were assessed for their in vitro and in vivo hematopietic activities. RESULTS: After 7 days in coculture, AHESS-5 was stronger more effective than HESS-5 in converting M and S cells to CD34(+) cells (M cells: 67.4% vs 17.5%, n =6, p < 0.001) (S cells: 42.3% vs 2.3%, n = 6, p < 0.001). Furthermore, both M and S cells were able to engraft in immunodeficient mice after they were cocultured on AHESS-5. CONCLUSIONS: Results suggest that angiopoietin-1 supports SRC activities in human CD34(-) BM cells, as murine studies demonstrated. Furthermore, identification of previously undetected subpopulations of BM CD34(-) HSCs unveils heterogenous components in the stem cell pool.
Assuntos
Angiopoietina-1/fisiologia , Antígenos CD34/imunologia , Células da Medula Óssea/citologia , Hematopoese , Western Blotting , Células da Medula Óssea/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Imuno-HistoquímicaRESUMO
To characterize human hematopoietic stem cells (HSCs), xenotransplantation techniques such as the severe combined immunodeficiency (SCID) mouse repopulating cell (SRC) assay have proven the most reliable methods thus far. While SRC quantification by limiting dilution analysis (LDA) is the gold standard for measuring in vitro expansion of human HSCs, LDA is a statistical method and does not directly establish that a single HSC has self-renewed in vitro. This would require a direct clonal method and has not been done. By using lentiviral gene marking and direct intra-bone marrow injection of cultured CD34+ CB cells, we demonstrate here the first direct evidence for self-renewal of individual SRC clones in vitro. Of 74 clones analyzed, 20 clones (27%) divided and repopulated in more than 2 mice after serum-free and stroma-dependent culture. Some of the clones were secondary transplantable. This indicates symmetric self-renewal divisions in vitro. On the other hand, 54 clones (73%) present in only 1 mouse may result from asymmetric divisions in vitro. Our data demonstrate that current ex vivo expansion conditions result in reliable stem cell expansion and the clonal tracking we have employed is the only reliable method that can be used in the development of clinically appropriate expansion methods.
Assuntos
Proliferação de Células , Células-Tronco Hematopoéticas/metabolismo , Animais , Antígenos CD34/metabolismo , Técnicas de Cultura de Células/métodos , Células Cultivadas , Transplante de Células-Tronco Hematopoéticas/métodos , Células-Tronco Hematopoéticas/citologia , Humanos , Lentivirus , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transdução Genética , Transplante HeterólogoRESUMO
Hematopoiesis is maintained by specific interactions between both hematopoietic and nonhematopoietic cells. Whereas hematopoietic stem cells (HSCs) have been extensively studied both in vitro and in vivo, little is known about the in vivo characteristics of stem cells of the nonhematopoietic component, known as mesenchymal stem cells (MSCs). Here we have visualized and characterized human MSCs in vivo following intramedullary transplantation of enhanced green fluorescent protein-marked human MSCs (eGFP-MSCs) into the bone marrow (BM) of nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice. Between 4 to 10 weeks after transplantation, eGFP-MSCs that engrafted in murine BM integrated into the hematopoietic microenvironment (HME) of the host mouse. They differentiated into pericytes, myofibroblasts, BM stromal cells, osteocytes in bone, bone-lining osteoblasts, and endothelial cells, which constituted the functional components of the BM HME. The presence of human MSCs in murine BM resulted in an increase in functionally and phenotypically primitive human hematopoietic cells. Human MSC-derived cells that reconstituted the HME appeared to contribute to the maintenance of human hematopoiesis by actively interacting with primitive human hematopoietic cells.
Assuntos
Diferenciação Celular/fisiologia , Hematopoese/fisiologia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/fisiologia , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , Humanos , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCIDRESUMO
While mononuclear cells isolated from murine skeletal muscle were shown to be capable of hematopoietic activity, similar hematopoietic cells (HC) recently were reported to exist in primate muscle. We investigated muscle-derived HC from young and adult human subjects. Although hematopoietic stem cells were rare in muscle, their frequency nonetheless was approximately four times greater than in peripheral blood. These cells in muscle appeared to originate from CD45(+) bone marrow cells. Our results suggested an additional function of human skeletal muscle as a reservoir of HC.
Assuntos
Células-Tronco Hematopoéticas/fisiologia , Músculo Esquelético/citologia , Adolescente , Adulto , Idoso , Animais , Células da Medula Óssea/fisiologia , Linhagem da Célula , Criança , Feminino , Citometria de Fluxo , Imunofluorescência , Transplante de Células-Tronco Hematopoéticas , Humanos , Imunofenotipagem , Antígenos Comuns de Leucócito/metabolismo , Masculino , Camundongos , Pessoa de Meia-IdadeRESUMO
In multiunit cord blood transplantation, hematopoietic stem cells from each unrelated cord blood (UCB) unit competitively reconstitute the hematopoietic system in a recipient. To evaluate the fate of the progeny of each UCB unit and to determine the effects of graft-versus-graft reaction, we established a novel competitive repopulation assay using NOD/SCID/gammac(null) mice in which human T lymphocytes develop from CD34+ cells. CD34+ cells from each UCB unit were labeled with recombinant lentivirus vectors carrying genes encoding either enhanced green fluorescent protein (EGFP) or enhanced yellow fluorescent protein (EYFP). Hematopoietic chimerism composed of both EGFP+ and EYFP+ cells was stably maintained up to 6 months after transplantation with purified CD34+ cells; the ratio of EGFP+ to EYFP+ cells in peripheral blood and bone marrow posttransplantation was equivalent to the ratio of these cells at transplantation. However, when mononuclear cells from two UCB units were cotransplanted with CD34+ cells, engraftment was highly competitive, with cells from only one or the other of the two UCB units surviving. Further subfractionations of mononuclear cells indicate that the skewed chimerism that is often observed in clinical multiunit cord blood transplantation may be mediated by the cooperation of both CD4+ and CD8+ T cells. The assay established here will be a useful tool for analyzing hematopoietic reconstitution in clinical multiunit cord blood transplantation.
Assuntos
Sangue Fetal/citologia , Transplante de Células-Tronco Hematopoéticas/métodos , Lentivirus/genética , Animais , Antígenos CD34/análise , Proteínas de Bactérias/análise , Proteínas de Bactérias/genética , Diferenciação Celular , Ensaio de Unidades Formadoras de Colônias , Marcadores Genéticos , Vetores Genéticos/genética , Doença Enxerto-Hospedeiro/prevenção & controle , Proteínas de Fluorescência Verde/análise , Proteínas de Fluorescência Verde/genética , Células-Tronco Hematopoéticas/virologia , Subunidade gama Comum de Receptores de Interleucina , Leucócitos Mononucleares/fisiologia , Leucócitos Mononucleares/transplante , Proteínas Luminescentes/análise , Proteínas Luminescentes/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Receptores de Interleucina-7/genética , Quimeras de TransplanteRESUMO
Acute tumor lysis syndrome (TLS) is a condition that results from the rapid destruction of tumor cells accompanied with a massive release of cellular breakdown products. Acute renal failure resulting from TLS has been reported in cases of hematologic malignancies, spontaneous or treatment induced, the latter especially by chemotherapy. We present the case of a patient with diffuse large B-cell lymphoma who developed radiotherapy-induced TLS and subsequently acute renal failure. He presented with a large mediastinal tumor compressing the airway, thus causing dyspnea. After 6 Gy/3 fractions/3 days of palliative radiotherapy for the tumor, a decrease in urine volume was noted as well as rapid tumor shrinking. Because this patient died despite previous prophylaxic anti-uric acid treatment and hemodialysis, his case illustrates the need to anticipate the development of acute renal failure, even though there may be no remarkable serum uric acid elevation after the initiation of radiotherapy.
Assuntos
Linfoma Difuso de Grandes Células B/radioterapia , Neoplasias do Mediastino/radioterapia , Cuidados Paliativos/métodos , Radioterapia/efeitos adversos , Síndrome de Lise Tumoral/etiologia , Doença Aguda , Injúria Renal Aguda/etiologia , Idoso , Alopurinol/administração & dosagem , Antimetabólitos/administração & dosagem , Creatinina/sangue , Progressão da Doença , Evolução Fatal , Humanos , Masculino , Mediastino/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Ácido Úrico/urinaRESUMO
OBJECTIVE: Recent studies have shown that bone marrow (BM) contains cells capable of differentiating into myocytes in vivo. However, addition of demethylation drugs has been necessary to induce myocyte differentiation from BM cells in vitro, and precise mechanisms of BM cells' conversion to myocytes and the origin of those cells have not been established. We investigated the expression of myogenic markers during differentiation and maturation of myocytes from BM-derived multipotent adult progenitor cells (MAPC) under physiological culture condition. MATERIALS AND METHODS: Frozen BM samples from 21 healthy donors were used as a source of MAPC. To induce myocyte differentiation MAPC was cultured in the presence of 5% FCS, VEGF, bFGF, and IGF-1, and the expressions of myocyte markers were examined at various time points. We also investigated engraftment and differentiation of MAPC-derived myocytes in vivo. RESULTS: Frozen BM-derived MAPC, cultured under the physiological myogenic condition, demonstrated spatial expression patterns of several myocyte markers similar to that of authentic myocyte differentiation. When injected into murine muscles, MAPC treated with the myogenic condition engrafted and differentiated into myocyte marker-positive cells and myotubes in vivo. CONCLUSION: For the first time, we were able to induce myocyte formation from BM cells under the physiological condition in vitro and demonstrated that treating cells with this condition prior to intramuscular injection increased efficiency of engraftment and differentiation in vivo.
Assuntos
Células da Medula Óssea , Células-Tronco Multipotentes/citologia , Células Musculares/citologia , Células Musculares/transplante , Adolescente , Adulto , Animais , Antígenos CD/análise , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Linhagem da Célula , Transplante de Células , Criança , Sobrevivência de Enxerto , Humanos , Camundongos , Pessoa de Meia-Idade , Fibras Musculares Esqueléticas/citologia , Transplante HeterólogoRESUMO
Although malignant lymphoma of the breast is a rare disease, we recently experienced a high incidence of central nervous system (CNS) involvement. Thus, we retrospectively reviewed clinical records of 921 patients with breast tumors, treated from 1990 to 2002, to examine CNS involvement. Eight patients were found to have malignant lymphomas during this period. Of these, four patients ranging from 46 to 80 years of age had CNS involvement, one with Burkitt type and three with diffuse large B-cell lymphoma. The patients received surgery and/or chemotherapy and/or radiotherapy. The four other patients without CNS involvement are alive without tumor, whereas three of four patients with CNS involvement died of the disease. It should be noted that CNS involvement in breast lymphoma is not so rare. It is difficult, but important, to determine whether malignant lymphoma of the breast originates in the breast or is of systemic origin. Although brain metastasis could be treated effectively by radiotherapy and/or chemotherapy, the prognosis was poor. Further intensive treatment is required for breast malignant lymphoma with CNS involvement.
Assuntos
Neoplasias da Mama/terapia , Neoplasias do Sistema Nervoso Central/terapia , Linfoma não Hodgkin/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/patologia , Linfoma de Burkitt/patologia , Linfoma de Burkitt/terapia , Terapia Combinada , Feminino , Humanos , Linfoma de Células B/patologia , Linfoma de Células B/terapia , Linfoma Difuso de Grandes Células B/patologia , Linfoma Difuso de Grandes Células B/terapia , Linfoma não Hodgkin/patologia , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
To measure the ability of human hematopoietic stem cells (HSCs), the SCID-repopulating cell (SRC) assay has been widely used. Conventionally, human HSCs are transplanted into a nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse via a tail vein. However, those cells must go through various obstacles until they reach the mouse marrow environment, which could explain the generally low homing efficiency in this system. Thus, the capability of HSCs may not be studied accurately by this intravenous transplantation method. In our attempt to reveal actual SRC potential, ie, self-renewal and multilineage differentiation in recipient bone marrow, we introduced cells into mouse marrow directly (intrabone marrow [iBM]) to minimize the effect of factors that may interfere with the homing of HSCs and compared the results obtained by intravenous and iBM methods. When cord blood CD34(+)CD38(-) cells were transplanted in NOD/SCID mice by iBM, a 15-fold higher frequency of SRC, 1 in 44 CD34(+)CD38(-) cells, was achieved compared with 1 in 660 by the intravenous method. Furthermore, the iBM transplant showed high levels of engraftment in the secondary transplantation. Pretreatment of CD34(+) cells with antibodies that block either very late antigen 4 (VLA-4) or VLA-5 reduced engraftment partially, whereas blockage of both molecules resulted in complete inhibition of engraftment, which suggests that VLA-4 and VLA-5 are involved in different processes in engraftment or have complementary roles. Our results indicate that the iBM injection strategy is a more sensitive and direct way to measure the capability of human SRCs and is useful to investigate the interaction of HSCs and marrow environment in vivo.
Assuntos
Medula Óssea , Transplante de Células-Tronco de Sangue do Cordão Umbilical/métodos , Células-Tronco Hematopoéticas/citologia , Transplante Heterólogo/métodos , Animais , Linhagem da Célula , Movimento Celular , Feminino , Citometria de Fluxo , Genes Reporter , Sobrevivência de Enxerto , Proteínas de Fluorescência Verde , Humanos , Recém-Nascido , Injeções , Injeções Intravenosas , Integrina alfa4beta1/fisiologia , Integrina alfa5beta1/fisiologia , Proteínas Luminescentes/análise , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Especificidade de Órgãos , Receptores CXCR4/fisiologia , Proteínas Recombinantes de Fusão/análise , Cauda/irrigação sanguínea , Quimeras de TransplanteRESUMO
PURPOSE: To assess the effect of hepatic artery occlusion with or without hepatic venous outflow interruption on coagulation diameter during percutaneous microwave coagulation therapy (PMCT) for hepatocellular carcinoma (HCC) by a prospective and randomized trial. MATERIALS AND METHODS: Thirty-one patients with 36 HCCs (10-38 mm in diameter) were randomly separated into two treatment groups as follows: group 1 (14 tumors in 14 patients) was treated with PMCT in conjunction with both segmental hepatic artery embolization with gelatin sponge particles and temporary interruption of hepatic venous flow by means of a 6-F balloon catheter to reduce the portal venous flow; group 2 (22 tumors in 17 patients) was treated with PMCT with segmental hepatic artery embolization only. PMCT under ultrasound (US) guidance was performed with 2,450 MHz of microwave frequency at 40-60 W and a needle applicator 1.6 mm in diameter. The coagulated area was measured at the maximum diameter perpendicular to the needle tract on enhanced computed tomography (CT) performed immediately after PMCT. The local effect of the treatment was evaluated by follow-up enhanced CT (6-33 mo). RESULTS: Patients in group 1 had a significantly larger coagulation area (mean +/-SD = 42.9 mm +/- 8.3), with coagulation times of 5.3 min +/- 1.4, than patients in group 2 (32.6 mm +/- 8.0), with coagulation times of 4.2 min +/- 1.3 (P <.05). Follow-up enhanced CT showed no local enhancement of the tumor, indicating complete necrosis and no local recurrence, except for four tumors. There were no major complications after PMCT except liver abscess that developed after PMCT in one patient with pneumobilia. CONCLUSIONS: PMCT with combined hepatic arterial embolization and temporary hepatic venous flow interruption can coagulate significantly larger volumes of tumor than PMCT with only hepatic arterial embolization.
Assuntos
Carcinoma Hepatocelular/terapia , Neoplasias Hepáticas/terapia , Fígado/irrigação sanguínea , Micro-Ondas/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Hepatocelular/irrigação sanguínea , Terapia Combinada , Embolização Terapêutica , Feminino , Humanos , Neoplasias Hepáticas/irrigação sanguínea , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Radiografia Intervencionista , Estatísticas não Paramétricas , Resultado do TratamentoRESUMO
Our purpose in this study was to determine the influence of radiotherapy, especially brachytherapy, on the activity of natural killer cells (NK). We examined changes in NK activity before and after radiotherapy in 27 patients who underwent radiotherapy with or without brachytherapy, comprising of 16 cases of cervical cancer (three recurrences), 5 of prostate cancer, 4 of esophageal cancer and 2 of tongue cancer. Fourteen intracavitary procedures (for 13 cervical cancers and 1 esophageal cancer) and 10 interstitial brachytherapy (for 3 cases of recurrent cervical cancer, 5 of prostate cancer and 2 of tongue cancer) were performed with Ir-192 microSelectron HDR and Selectron Cs-137. External radiotherapy consisted of 10 MV X-ray administration for 13 cases of cervical cancer and 4 of esophageal cancer. The number of white blood cell was reduced by radiotherapy from 5065 +/- 2002 count/ml to 4281 +/- 1392 count/ml (p=0.02), that of lymphocytes from 1518 +/- 817 to 762 +/- 409 /ml (p<0.0001), and that of CD 16+ cells from 274 +/- 197 to 14 +/- 96 (p=0.03). No significant change was observed in the number of CD 56+ cells (274 +/- 166 to 211 +/- 153 /ml). Overall NK activity was reduced by radiotherapy from 37 +/- 19% to 30 +/- 19% (p=0.001). External radiotherapy with or without brachytherapy reduced NK activity from 33 +/- 18% to 23 +/- 16% (p=0.004). However interstitial brachytherapy produced little change in NK activity from 42 +/- 18 to 39 +/- 19%). Radiotherapy reduced the number of white blood cell, lymphocyte and CD 16+ cells. Although external radiotherapy suppressed NK activity, only brachytherapy showed little influence on NK activity alteration.