RESUMO
Objective: To evaluate the long-term efficacy of percutaneous microwave ablation (MWA) therapy for hepatocellular carcinoma. Methods: 2054 cases with Barcelona Clinic Liver Cancer (BCLC) stage 0~B at the Fifth Medical Center of the Chinese People's Liberation Army General Hospital from January 2006 to September 2020 were retrospectively collected. All patients were followed up for at least 2 years. The primary endpoint of overall survival and secondary endpoints (tumor-related survival, disease-free survival, and postoperative complications) of patients treated with ultrasound-guided percutaneous MWA were analyzed. Kaplan-Meier method was used for stratified survival rate analysis. Fine-and-Gray competing risk model was used to analyze overall survival. Results: A total of 5 503 HCC nodules [mean tumor diameter (2.6±1.6) cm] underwent 3 908 MWAs between January 2006 and September 2020, with a median follow-up time of 45.6 (24.0 -79.2) months.The technical effectiveness rate of 5 375 tumor nodules was 97.5%. The overall survival rates at 5, 10, and 15-years were 61.6%, 38.8%, and 27.0%, respectively. The tumor-specific survival rates were 67.1%, 47.2%, and 37.7%, respectively. The free tumor survival rates were 25.8%, 15.7%, and 9.9%, respectively. The incidence rate of severe complications was 2.8% (108/3 908). Further analysis showed that the technical effectiveness and survival rate over the passing three time periods from January 2006-2010, 2011-2015, and 2016-September 2020 were significantly increased, with Pâ <â 0.001, especially for liver cancer 3.1~5.0 cm (Pâ <â 0.001). Conclusion: Microwave ablation therapy is a safe and effective method for BCLC stage 0-B, with significantly enhanced technical efficacy and survival rate over time.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Micro-Ondas , Humanos , Neoplasias Hepáticas/cirurgia , Neoplasias Hepáticas/terapia , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/cirurgia , Micro-Ondas/uso terapêutico , Estudos Retrospectivos , Taxa de Sobrevida , Resultado do Tratamento , Intervalo Livre de Doença , Ablação por Cateter/métodos , Feminino , Complicações Pós-Operatórias/epidemiologia , Masculino , Pessoa de Meia-IdadeRESUMO
OBJECTIVE: Intervertebral disc degeneration (IVDD) is the main cause of spine diseases, and apoptosis of nucleus pulposus (NP) cells is an important risk factor for the degeneration of intervertebral discs. Endoplasmic reticulum (ER) stress is involved in multiple apoptosis processes. This study investigated whether the specific COX-2 inhibitor parecoxib can inhibit NP cell apoptosis induced by ER stress. PATIENTS AND METHODS: Human NP cells were isolated from the disc tissue collected from IVDD patients. We used IL-1ß to establish an NP cell degenerated model. Degenerated levels were detected by the analysis of cell viability, collagen II, collagen X, aggrecan, TNF-α, IL-6, and MMP-13 expression. ER stress status was examined by GRP78 and CHOP expression. Apoptosis level was mainly indicated by the positive apoptotic cells and caspase-12 expression. CHOP-plasmid transfection was performed to overexpress the CHOP protein level. RESULTS: IL-1ß could induce the decrease of viability, collagen â ¡, aggrecan, but an increase of collagen X, TNF-α, IL-6, and MMP-13 in NP cells, as well as the upregulation of GRP78/PERK/caspase-12 and apoptosis level, which could be inhibited by parecoxib. Parecoxib could also suppress CHOP caused by COX-2 upregulation and apoptosis in NP cells. CONCLUSIONS: Parecoxib is a safe and efficient COX-2 inhibitor to NP cells, which could prevent NP cells apoptosis by suppressing ER stress.
Assuntos
Apoptose/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Isoxazóis/farmacologia , Núcleo Pulposo/efeitos dos fármacos , Adulto , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Chaperona BiP do Retículo Endoplasmático , Feminino , Humanos , Interleucina-1beta/antagonistas & inibidores , Interleucina-1beta/metabolismo , Masculino , Núcleo Pulposo/metabolismo , Núcleo Pulposo/patologiaRESUMO
Oxidative stress (OS) induces osteoblast apoptosis, which plays a crucial role in the initiation and progression of osteoporosis. Although OS is closely associated with mitochondrial dysfunction, detailed mitochondrial mechanisms underlying OS-induced osteoblast apoptosis have not been thoroughly elucidated to date. In the present study, we found that mitochondrial abnormalities largely contributed to OS-induced osteoblast apoptosis, as evidenced by enhanced production of mitochondrial reactive oxygen species; considerable reduction in mitochondrial respiratory chain complex activity, mitochondrial membrane potential, and adenosine triphosphate production; abnormality in mitochondrial morphology; and alteration of mitochondrial dynamics. These mitochondrial abnormalities were primarily mediated by an imbalance in mitochondrial fusion and fission through a protein kinase B- (AKT-) glycogen synthase kinase 3ß- (GSK3ß-) optic atrophy 1- (OPA1-) dependent mechanism. Hydroxytyrosol (3,4-dihydroxyphenylethanol (HT)), an important compound in virgin olive oil, significantly prevented OS-induced osteoblast apoptosis. Specifically, HT inhibited OS-induced mitochondrial dysfunction by decreasing OPA1 cleavage and by increasing AKT and GSK3ß phosphorylation. Together, our results indicate that the AKT-GSK3ß signaling pathway regulates mitochondrial dysfunction-associated OPA1 cleavage, which may contribute to OS-induced osteoblast apoptosis. Moreover, our results suggest that HT could be an effective nutrient for preventing osteoporosis development.
Assuntos
GTP Fosfo-Hidrolases/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Mitocôndrias/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoporose/metabolismo , Álcool Feniletílico/análogos & derivados , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/patologia , Osteoblastos/patologia , Osteoporose/patologia , Estresse Oxidativo/fisiologia , Álcool Feniletílico/farmacologia , Transdução de Sinais , TransfecçãoRESUMO
Advanced glycation end products (AGEs) can stimulate osteoblast apoptosis and have a critical role in the pathophysiology of diabetic osteoporosis. Mitochondrial abnormalities are closely related to osteoblast dysfunction. However, it remains unclear whether mitochondrial abnormalities are involved in AGE-induced osteoblastic cell apoptosis. Silibinin, a major flavonolignan compound of silimarin, has strong antioxidant and mitochondria-protective properties. In the present study, we explored the possible mitochondrial mechanisms underlying AGE-induced apoptosis of osteoblastic cells and the effect of silibinin on osteoblastic cell apoptosis. We demonstrated that mitochondrial abnormalities largely contributed to AGE-induced apoptosis of osteoblastic cells, as evidenced by enhanced mitochondrial oxidative stress, conspicuous reduction in mitochondrial membrane potential and adenosine triphosphate production, abnormal mitochondrial morphology, and altered mitochondrial dynamics. These AGE-induced mitochondrial abnormalities were mainly mediated by the receptor of AGEs (RAGE). In addition, we found that silibinin directly downregulated the expression of RAGE and modulated RAGE-mediated mitochondrial pathways, thereby preventing AGE-induced apoptosis of osteoblastic cells. This study not only provides a new insight into the mitochondrial mechanisms underlying AGE-induced osteoblastic cell apoptosis, but also lays a foundation for the clinical use of silibinin for the prevention or treatment of diabetic osteoporosis.
Assuntos
Apoptose/efeitos dos fármacos , Produtos Finais de Glicação Avançada/toxicidade , Mitocôndrias/metabolismo , Osteoblastos/patologia , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Transdução de Sinais , Silibina/farmacologia , Animais , Benzamidas/farmacologia , Linhagem Celular , Forma Celular/efeitos dos fármacos , Ciclosporina/farmacologia , Camundongos , Compostos Organofosforados/farmacologia , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Ubiquinona/análogos & derivados , Ubiquinona/farmacologiaRESUMO
PurposeCD59 complement regulator and complement factor H (CFH) have important roles in complement activation pathways, which are known to affect the development of uveitis. The present study was performed to investigate whether an association exists between CD59 and CFH genetic polymorphisms and acute anterior uveitis (AAU).MethodsA total of 600 individuals (300 patients diagnosed with AAU and 300 healthy controls) were recruited for this case-control study. Five single-nucleotide polymorphisms (SNPs) in CD59 (rs831626, rs12272807, rs831625, rs11585, and rs12576440) and CFH-rs1065489 were genotyped using Sequenom MassARRAY technology. Allele and genotype frequencies were statistically compared between patients and controls using χ2 test. Analyses were stratified for gender, human leukocyte antigen (HLA)-B27, and ankylosing spondylitis (AS) status.ResultsNo significant association was found between any of the six polymorphisms and AAU. In HLA-B27-negative AAU patients, the frequencies of the G allele and GG homozygosity were lower in CD59-rs831626 when compared with controls (P=0.032). There were also significant decreases in the frequencies of T allele and TT homozygosity in CFH-rs1065489 in AAU patients with AS compared with controls (P=0.002). Furthermore, the frequencies of the T allele and TT homozygosity in CFH-rs1065489 were lower in the AAU male patients with AS compared with controls (P=0.015).ConclusionOur results revealed that SNPs CD59-rs831626 and CFH-rs1065489 were associated with the susceptibility of AAU. The influence on AAU could be gender specific and dependent on the HLA-B27 and AS status. No positive results were found in the overall group.
Assuntos
Antígenos CD59/genética , Polimorfismo de Nucleotídeo Único , Uveíte Anterior/genética , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Povo Asiático/genética , Estudos de Casos e Controles , Criança , China/epidemiologia , Fator H do Complemento/genética , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da PolimeraseRESUMO
Pyrroloquinoline quinone (PQQ) is linked to fundamental biological processes such as mitochondrial biogenesis and lipid metabolism. PQQ may also function as an essential micronutrient during animal development. Recent studies have shown the therapeutic potential of PQQ for several age-related diseases due to its antioxidant capacity. However, whether PQQ can promote longevity is unknown. Here, we investigate the effects of PQQ on oxidative stress resistance as well as lifespan modulation in Caenorhabditis elegans. We find that PQQ enhances resistance to oxidative stress and extends the lifespan of C. elegans at optimal doses. The underlying molecular mechanism involves the increased activities of the primary lifespan extension transcriptional factors DAF-16/FOXO, the conserved oxidative stress-responsive transcription factor SKN-1/Nrf2, and upregulation of daf-16, skn-1 downstream targets including sod-3, hsp16.2, gst-1 and gst-10. Our findings uncover a novel role of PQQ in longevity, supporting PQQ as a possible dietary supplement for overall health improvement.
Assuntos
Envelhecimento/fisiologia , Caenorhabditis elegans/fisiologia , Longevidade/fisiologia , Estresse Oxidativo , Cofator PQQ/farmacologia , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ligação a DNA/genética , Fatores de Transcrição Forkhead/genética , RNA Mensageiro/genética , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/genética , Fatores de Transcrição/genéticaRESUMO
INTRODUCTION: The aim of this study was to determine the prevalence of ß-globin gene cluster deletions in individuals with increased Hb F levels in a Chinese population. METHODS: Subjects with HbF levels ≥ 10% were selected for further investigation. Gap-PCR was used to screen for three common ß-globin gene cluster deletions: Chinese ((A) γδß)(0)-thalassemia, Southeast Asian (SEA) deletion and Hb Lepore. Multiplex ligation-dependent probe amplification (MLPA) was used to analyze dosage changes of the ß-globin gene cluster for those not associated with one of the three common deletions. RESULTS: One hundred and thirty-one individuals had an increased Hb F level; among these, 51 (38.9%) were showed to have Chinese ((A) γδß)(0)-thalassemia (n = 37) or SEA deletion (n = 14). A single case of Hb Lepore-Boston-Washington was detected. MLPA only detected 2 deletions in three cases of the remaining 80 patients. Gap-PCR confirmed that they included a 1357 bp ß-globin gene deletion (NG_000007.3:g.69997_71353del1357) in one case and a HBG2-HBG1 fusion gene consisting of exons 1 and 2 of HBG2 ((G) γ-globin gene) and exon 3 of HBG1 ((A)γ-globin gene) (HBG2:c.315 + 573_HBG1: c.315 + 572del) in two cases. CONCLUSION: The Chinese ((A) γδß)(0)-thalassemia and SEA deletion are the most common large deletions of ß-globin gene cluster in Chinese. Gap-PCR for the detection of these two deletions should be used in thalassemia screening program in China where the incidence of ß-thalassemia is high.
Assuntos
Povo Asiático/genética , Hemoglobina Fetal/genética , Genética Populacional , Família Multigênica , Deleção de Sequência , Globinas beta/genética , China , Índices de Eritrócitos , Feminino , Humanos , Masculino , Tipagem Molecular/métodos , Talassemia beta/diagnóstico , Talassemia beta/epidemiologia , Talassemia beta/genéticaRESUMO
Poly(3,4-ethylenedioxythiophene) (PEDOT) doped with poly(styrene sulfonate) (PSS) has a variety of chemical and biomedical applications. The application of PEDOT/PSS polymers in drug delivery has attracted attention. However, whether conducting polymers of PEDOT/PSS could be used for dopamine delivery has not clear. In the present study, the PEDOT/PSS coatings incorporated with dopamine were fabricated on 0.5 mm diameter platinum electrodes, electrochemical properties, and dopamine delivery capacities of these electrodes were evaluated in vitro and in vivo through implanting these electrodes into brain striatum area. The findings demonstrated that the PEDOT/PSS/dopamine coatings on platinum electrodes could reduce electrodes impedances, increase charge storage capacities, and release significant levels of dopamine upon electrical stimulation of these electrodes. These results indicated that polymers of PEDOT/PSS/dopamine could be used for dopamine delivery, implicating potential application of PEDOT/PSS/dopamine-coated implantable electrodes in the treatment of some diseases associated with dopamine deficits, such as, electrodes for the treatment of Parkinson's disease during deep brain stimulation.
Assuntos
Materiais Revestidos Biocompatíveis/química , Dopaminérgicos/administração & dosagem , Dopamina/administração & dosagem , Sistemas de Liberação de Medicamentos/instrumentação , Eletrodos Implantados , Poliestirenos/química , Tiofenos/química , Animais , Encéfalo/cirurgia , Dopamina/química , Dopaminérgicos/química , Estimulação Elétrica , Humanos , Técnicas In Vitro , Masculino , Ratos Sprague-DawleyRESUMO
INTRODUCTION: Neovasculature imaging is a promising approach for tumor diagnosis. We constructed tumor neovasculature targeted paramagnetic nanoliposomes with RGD10, F56, and K237 peptides, which can bind to Integrin αvß3 and VEGFR-1, VEGFR-2, respectively, and compared their potential value as MRI contrast agents for detecting small tumors in animal models. MATERIALS AND METHODS: Peptide-Ahx-palmitic acid conjugate was synthesized using Fmoc solid-phase synthesis chemistry. Targeted paramagnetic nanoliposomes were prepared by the thin film dispersion-sonication method. The tumor signal enhancements of liposome particles were evaluated by MRI in a xenograft mice model. RESULTS: The apparent affinity constants of RGD10, K237, and F56 peptides binding to their cell receptors were 9.15 × 10(7), 6.01 × 10(7), and 3.85 × 10(7) mol/L, respectively. RGD10 and K237 targeted paramagnetic nanoliposomes have shown much greater tumor-specific MRI signal enhancement in xenograft of the nude mice compared to F56 targeted paramagnetic nanoliposome. Tumor signal enhancement rate (SER %) increased 2.21 ± 0.09 and 1.82 ± 0.05 fold, respectively, for RGD10 and K237 compared to non-targeted control in T1 weighted MR image. CONCLUSION: RGD10 and K237 targeted paramagnetic nanoliposomes can be developed as potential tumor-specific MRI contrast agents and are helpful for tumor detection.
Assuntos
Diagnóstico por Imagem/métodos , Lipossomos , Imageamento por Ressonância Magnética/métodos , Nanopartículas Metálicas , Neoplasias Experimentais/patologia , Neovascularização Patológica , Animais , Sistemas de Liberação de Medicamentos/métodos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neovascularização Patológica/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Fascin, an actin-bundling protein, induces membrane protrusions and increases cell motility in various transformed cells. The overexpression of fascin in esophageal squamous cell carcinoma (ESCC) has been described only recently, but the roles and mechanism still remained unclear. Here, by using RNA interference (RNAi), we have stably silenced the expression of the fascin in EC109 cells, an ESCC cell line. Down-regulation of fascin resulted in a suppression of cell proliferation and as well as a decrease in cell invasiveness. Furthermore, we revealed that fascin might have functions in regulating tumor growth in vivo. The effect of fascin on cell invasiveness correlated with the activation of matrix metalloproteases such as MMP-2 and MMP-9. We examined that fascin down-expression also led to a decrease of c-erbB-2 and beta-catenin at the protein level. These results suggested that fascin might play crucial roles in regulating neoplasm progression of ESCC.
Assuntos
Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Proteínas de Transporte/fisiologia , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patologia , Proteínas dos Microfilamentos/fisiologia , Animais , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Proliferação de Células , Proteínas do Citoesqueleto/metabolismo , Regulação para Baixo , Células HeLa , Humanos , Camundongos , Camundongos Nus , Proteínas dos Microfilamentos/antagonistas & inibidores , Proteínas dos Microfilamentos/genética , Invasividade Neoplásica , Interferência de RNA , Receptor ErbB-2/metabolismo , Transativadores/metabolismo , beta CateninaRESUMO
To examine certain characteristics of multistep carcinogenesis, we studied telomerase activity and malignant phenotypes in the immortal, premalignant and malignant stages of esophageal epithelial cells induced by HPV. An immortalized human fetal esophageal epithelial cell line (SHEE) was induced by E6E7 genes of human papillomavirus (HPV) type 18. Cells in the 10th passage, (SHEE10), 31st passage (SHEE31), 61st passage (SHEE61) and SHEE61A which were selected and expanded from anchorage-independent growth colonies of SHEE61, were examined as follows: cell morphology by electron-microscopy; the cell cycle by flow cytometry, telomerase activity by TRAP assay, tumorigenic detection including anchorage-independent growth by soft agar culture and tumor formation by inoculating cells into SCID and nude mice, and detection of HPV18 E6E7 oncoprotein by Western blot. The morphology of the SHEE10 cells exhibited good differentiation, the SHEE60 and SHEE61A cells were relatively poorly differentiated, and the SHEE31 cells were differentiated in two distinct ways. The telomerase was activated in SHEE31, SHEE61 and SHEE61A, but not in SHEE10 cells. SHEE61 and SHEE61A cells were weakened in contact-inhibition and increased in anchorage-independent growth. Inoculated into SCID and nude mice, the cells of the earlier two passages could not develop tumors; the SHEE61 developed one tumor in four SCID mice, but not in nude mice, and the SHEE61A cells developed tumors in both strains of immunodeficient mice. HPV18 E6E7 DNA detection by Western blotting was positive in all cell passages. In the process of carcinogenesis by HPV, the cells of SHEE31 are in an immortalized state with telomerase activity. The fact that SHEE61 cells remained immortalized and also demonstrated anchorage-independent growth, reveals premalignant character; the cells of SHEE61A exhibited malignant transformation with tumor formation in mice. The results revealed that the telomerase activity, anchorage-independent growth and tumor formation in nude mice are the indicators for immortalization, premalignancy and malignancy, respectively.
Assuntos
Proteínas de Ligação a DNA , Células Epiteliais/enzimologia , Papillomaviridae/genética , Telomerase/metabolismo , Animais , Western Blotting , Adesão Celular , Divisão Celular , Linhagem Celular , Linhagem Celular Transformada , Transformação Celular Neoplásica/genética , Transformação Celular Viral/genética , Células Epiteliais/ultraestrutura , Células Epiteliais/virologia , Esôfago/citologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Camundongos SCID , Microscopia Eletrônica , Transplante de Neoplasias , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/metabolismo , Papillomaviridae/fisiologia , Fenótipo , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
We studied cytogenesis, telomere and telomerase, and c-myc, ras, bcl-2, and p53 genes of cells in the progressive process of immortal epithelial cells from embryonic esophagus induced by human papillomavirus (HPV). The SHEE cell line, established by us, consist of immortalized epithelial cells from the embryonic esophagus induced by genes E6E7 of HPV type 18. It was in initial malignant transformation when cultivated over 60 passages without co-carcinogens. Cells of the 10th, 31st, and 60th passages were represented in the progressive process within the immortal period. In these three stages of the cell line, the modal number of chromosome and karyotypes were analyzed. The telomere length was assayed by Southern blot methods, and the telomerase activity was analyzed by hTR and hTERT assay. C-myc, p53, bcl-2, ras genes were assayed by the multi-PCR method. The morphology of the 10th passage cells exhibited good differentiation, the 60th passage cells were relatively poorly differentiated, and the 31st passage cells differentiated in two distinct ways. The growth characteristics of the 31st and 60th passage cells were weakened at contact-inhibition and anchorage-dependent growth. Karyotypes of three cell passages belonged to hyperdiploid and hypotriploid with abnormal chromosomes +1, +3, +7, +9, +17, +18; del(1)(p32); der(4), t(4;?)(q31;?); der(5),t(5;?)(q31;?); der(13),t(13;13)(p11;q11) and others. Bimodal distribution of chromosomes with more aberrant chromosomes appeared in the 31st and 60th passage cells. Telomere length sharply shortened from normal fetal esophagus to the 10th and 31st passage step by step, but was stable from the 31st to the 60th passage and the telomerase activities measured were expressed at late two passages. p53 mutant was positive in three passages, c-myc was positive in the 31st and the 60th passage K-ras only in the last. The results reveal that changes of chromosomes, telomere length, telomerase activity and certain gene expressions are important events of HPV-immortalized esophageal epithelium cells. All of these changes occurred in dynamic progressive process. This cell line may be useful for the elucidation of the genetic mechanism of cellular immortalization.
Assuntos
Transformação Celular Viral/genética , Células Epiteliais/virologia , Papillomaviridae , Linhagem Celular Transformada , Tamanho Celular , Aberrações Cromossômicas , Bandeamento Cromossômico , Análise Citogenética , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Esôfago/citologia , Esôfago/metabolismo , Esôfago/virologia , Regulação da Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenases/genética , Humanos , Cariotipagem , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-myc/genética , RNA/genética , RNA/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Telomerase/genética , Telômero/genética , Proteína Supressora de Tumor p53/genética , Proteínas ras/genéticaRESUMO
W.-J. Cai, S. Koltai, E. Kocsis, D. Scholz, W. Schaper and J. Schaper. Connexin37, not Cx40 and Cx43, is Induced in Vascular Smooth Muscle Cells During Coronary Arteriogenesis. Journal of Molecular and Cellular Cardiology (2001) 33, 957-967. The hypothesis that an altered expression of gap junction (GJ) proteins, connexin37 (Cx37), Cx40 and Cx43 will contribute to adaptive arteriogenesis was tested in growing coronary collateral vessels (CV) of the dog heart by immunoconfocal microscopy and transmission electron microscopy (TEM). We found that: (1) in the normal coronary system Cx37 and Cx40 were only expressed in endothelial cells (EC) from artery to capillary; (2) during collateral growth Cx37 was significantly induced in smooth muscle cells (SMC) from small-large arteries to precapillary arterioles (Ø=15 microm), while Cx40 was still only present in EC; (3) both homogeneous and heterogeneous distribution of Cx37 was observed in normal vessels (NV) and growing vessels (GV); (4) in mature vessels (MV), Cx37 was downregulated, similar to NV; (5) dual immunostaining revealed an inverse correlation between expression of Cx37 and desmin in GV occurring prior to downregulation of alpha-smooth actin and calponin; (6) Cx43 was undetectable in any vascular cells, both in NV and GV; (7) GJ were not found in SMC by TEM. Our data for the first time show the profile of connexin expression in the coronary system and provide evidence for existence of GJ proteins in capillaries. It is a novel finding that an altered expression of Cx37 is characteristic of adaptive arteriogenesis in the dog heart and may be used as a marker of vascular growth. Induced Cx37 may be an early signal indicating that SMC are responding to haemodynamic changes, i.e. increased shear stress.
Assuntos
Conexina 43/biossíntese , Conexinas/biossíntese , Músculo Liso Vascular/metabolismo , Miocárdio/metabolismo , Neovascularização Fisiológica , Actinas/biossíntese , Animais , Proteínas de Ligação ao Cálcio/biossíntese , Proteínas de Ligação ao Cálcio/metabolismo , Conexinas/metabolismo , Desmina/metabolismo , Cães , Regulação para Baixo , Endotélio Vascular/metabolismo , Imuno-Histoquímica , Lectinas/metabolismo , Proteínas dos Microfilamentos , Microscopia Confocal , Microscopia Eletrônica , Miocárdio/ultraestrutura , Calponinas , Proteína alfa-5 de Junções Comunicantes , Proteína alfa-4 de Junções ComunicantesRESUMO
OBJECTIVE: To identify and clone the differentially expressed genes in brief ischemia and reperfusion myocardium. METHODS: Ischemia and reperfusion were induced by repeated brief ligation of the porcine left anterior descending coronary artery. Total RNA which was isolated from myocardium subjected ischemia and reperfusion was used for mRNA differential display. After cloning and sequencing the cDNA fragments which showed change in expression, their expression were further confirmed by Northern-Blot analysis. RESULTS: Two differentially expressed cDNAs (W12 and W28) were identified and cloned. Their expression were subsequently confirmed to be truly differentially expressed. The expression of both genes in ischemia and reperfusion myocardium was obvious higher than that in nonischemia and reperfusion: W12 expression level was 2-fold (P < 0.05), and W28 expression level 1.9-fold (P < 0.05). In addition, mRNAs of W12 and W28 were existed in all tested organs including heart, liver, lung, kidney, spleen, intestine, brain and skeletal muscle. DNA sequencing analysis showed that there was no homology between W12, W28 and known genes, implying that they would represent novel gene respectively. CONCLUSION: Two novel genes induced by ischemia and reperfusion are identified, cloned and confirmed.
Assuntos
Clonagem Molecular , Traumatismo por Reperfusão Miocárdica/genética , Miocárdio/metabolismo , Animais , DNA Complementar/genética , Expressão Gênica , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Distribuição Aleatória , Porco MiniaturaRESUMO
The formation of collateral arteries as a process adaptive to arterial occlusion is now called 'arteriogenesis' to emphasize the difference to angiogenesis, the formation of capillaries by sprouting from pre-existent ones (W. Schaper, I. Buschmann. Cardiovasc Res 1999; 43: 835-7; I. Buschmann, W. Schaper. J Pathol 2000; 190: 338-42; D. Scholz et al. Virchows Arch 2000; 436: 257-70). The differences are that collaterals develop from pre-existing arterioles and that circulating monocytes adhere to endothelium that had been activated by the high shear stress generated by the large pressure differences between perfusion territories. Monocytes are the major producers of growth factors and of proteolytic enzymes that enable smooth muscle cells to migrate and divide. The nature of the growth factors remains uncertain. Neither FGF-1/2 nor VEGF is expressed on the transcriptional or translational level in collaterals proper and in the tissue surrounding them. Only FGF receptor 1 has a brief window of upregulation shortly after arterial occlusion. While transgenic overexpression of FGF-1 increases number and branching of arterioles, targeted disruption of FGF-1/2 does not negatively influence arteriogenesis. Cytokines that attract monocytes or prolong the life span of monocytes (MCP-1, GM CSF) are strong arteriogenic factors. Collateral vessels exhibit the same morphology whether they had formed in the heart, limbs or brain or in dogs, rabbits or mouse. They are tortuous because they also increase lengthwise in a restricted space. In animals larger than the mouse, they develop an intima, and initially, many arterioles participate in arteriogenesis, but only a few mature into large arterial channels which, when arterial occlusion had proceeded slowly enough, can replace the occluded artery to a significant proportion. Therapy with a single growth factor in animals with occluded femoral arteries significantly increases the speed of arteriogenesis but does not significantly increase the level of adaptation. It appears that the mastergene for arteriogenesis still awaits discovery.
Assuntos
Adaptação Fisiológica , Arteriopatias Oclusivas/fisiopatologia , Artérias/crescimento & desenvolvimento , AnimaisRESUMO
OBJECTIVE: To identify and clone genes induced by ischemia and reperfusion. METHODS: cDNA fragments responding to ischemia and reperfusion in porcine myocardium were isolated by mRNA differential display and then the cDNA library from porcine heart was screened. After cloning, sequencing, and hybridization, the basic characteristics of the gene were analyzed. RESULTS: One cDNA fragment from a novel gene up-regulated by ischemia and reperfusion was identified and cloned subsequently; a novel full-length cDNA containing an open reading frame encoding 608 amino acid was obtained by screening the library. Sequencing results revealed that both DNA and amino acid sequences had no substantial homology to previously described DNA or protein sequences. Northern blot analysis confirmed that this gene expression in ischemia and reperfusion myocardium was significantly increased, and the expression was also able to be detected in normal porcine heart, liver, lung, kidney, spleen, intestine, brain, and skeletal muscle. Southern blot analysis clearly showed that this novel gene had high evolutional conservation in human beings, pigs, rabbits, and mice. CONCLUSION: The cloned gene is a novel one induced by ischemia and reperfusion, and it may play a very important role in the response to ischemia and reperfusion.
Assuntos
DNA Complementar/biossíntese , Isquemia Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/genética , Miocárdio/metabolismo , Sequência de Aminoácidos , Animais , Clonagem Molecular , DNA Complementar/genética , Dados de Sequência Molecular , Isquemia Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Fases de Leitura Aberta/genética , Distribuição Aleatória , Análise de Sequência , Porco MiniaturaRESUMO
It is accepted that inorganic arsenic trioxide is an inducer of apoptosis for many types of cancer. Our previous studies have demonstrated that arsenic trioxide induces apoptosis of esophageal carcinoma cells. Administration of arsenic trioxide results in the inhibition of growth and survival of tumor cells. Esophageal carcinoma cells treated with arsenic trioxide for 3 days demonstrated a typical morphological appearance of apoptosis. To further examine molecular mechanism of arsenic trioxide induced apoptosis of esophageal carcinoma cells, we have investigated the early changes of the apoptotic cell induced by arsenic trioxide. Our results indicated that arsenic trioxide induced apoptosis of esophageal carcinoma cells occurs as early as 2 h after treatment. Annexin-v staining has further proved that the phosphatidylserine is exposed at 2 h. The early morphological change of arsenic trioxide treated cells was in the mitochondria. Arsenic trioxide treated cells displayed aggregated mitochondria. It induces accumulation of high electron-density amorphous substances, swollen and disruption of mitochondria in oesophageal carcinoma cells after 2 h treatment. The alteration of mitochondria induced by arsenic trioxide seems to occur before the condensation of chromatin. Thus, our data demonstrated that the primary target of arsenic trioxide induced apoptosis of esophageal carcinoma cells may be the mitochondria. It is possible that arsenic trioxide is a mitochondriotoxic agent.
Assuntos
Antineoplásicos/farmacologia , Apoptose , Arsenicais/farmacologia , Mitocôndrias/efeitos dos fármacos , Óxidos/farmacologia , Trióxido de Arsênio , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/ultraestrutura , Citometria de Fluxo , Humanos , Microscopia Eletrônica , Mitocôndrias/patologia , Células Tumorais CultivadasRESUMO
Administration of the neuroactive steroid hormone estrogen has been shown to effect cholinergic basal forebrain neuronal function. Antibodies directed against the estrogen receptor alpha (ERalpha) revealed dark (type 1) and light (type 2) nuclear positive neurons within the islands of Calleja, endopiriform nucleus, lateral septum, subfields of the cholinergic basal forebrain, bed nucleus of the stria terminalis, striohypothalamic region, medial preoptic region, periventricular, ventromedial, arcuate and tuberal mammillary nuclei of the hypothalamus, reuniens and anterior medial thalamic nuclei, amygdaloid complex, piriform cortex and subfornical organ. In contrast, only a few scattered ERalpha labeled neurons were found in cortex and hippocampus. ERalpha stained cell bodies were not seen in the striatum. Counts of ERalpha labeled neurons in intact female rats revealed significantly more type 2 neurons within the basal forebrain subfields. Quantitation of ERalpha immunoreactive neurons revealed a significant decrease in the relative number of type 1 neurons within the medial septum (MS), horizontal limb of the diagonal band (HDB) and substantia innominata/nucleus basalis (SI/NB) following ovariectomy. Quantitation following choline acetyltransferease (ChAT) immunohistochemistry revealed a significant decrease in the number of ChAT positive neurons within the MS, HDB and SI/NB, but not VDB following ovariectomy. Following ovx, the percentage of double labeled cholinergic basal forebrain neurons also declined significantly within the MS, VDB, HDB and SI/NB. These observations suggest that estrogen effects a subpopulation of cholinergic basal forebrain neurons and may provide insight into the biologic actions of this steroid in Alzheimer's disease.