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OBJECTIVE: There is concern that the coronavirus disease (COVID-19) vaccine may trigger or worsen autoimmune diseases. The objective of this study was to determine the impacts of COVID-19 vaccination on symptom severity in patients with myasthenia gravis (MG). METHODS: A total of 106 enrolled patients with MG who were vaccinated against COVID-19 were followed up, and a questionnaire was used to document in detail the exacerbation of muscle weakness after vaccination and all other uncomfortable reactions after vaccination. Demographic, clinical characteristics, medication, and vaccination data were collected by follow-up interview. The main observation outcome was whether the MG symptoms of patients were exacerbated. The definition of exacerbation is according to the subjective feeling of the patient or a 2-point increase in daily life myasthenia gravis activity score relative to before vaccination, within 30 days after vaccination. RESULTS: Of 106 enrolled patients [median age (SD) 41.0 years, 38 (35.8%) men, 53 (50.0%) with generalized MG, 74 (69.8%) positive for acetylcholine receptor antibody, and 21 (19.8%) with accompanying thymoma], muscle weakness symptoms were stable in 102 (96.2%) patients before vaccine inoculation. Muscle weakness worsened in 10 (9.4%) people after vaccination, of which 8 patients reported slight symptom worsening that resolved quickly (within a few days). Two (1.9%) of patients showed serious symptom aggravation that required hospitalization. CONCLUSION: Our results suggest that inactivated virus vaccines against COVID-19 may be safe for patients with MG whose condition is stable. Patients with generalized MG may be more likely to develop increased muscle weakness after vaccination.
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Vacinas contra COVID-19 , COVID-19 , Miastenia Gravis , Neoplasias do Timo , Adulto , Feminino , Humanos , Masculino , COVID-19/prevenção & controle , COVID-19/complicações , Vacinas contra COVID-19/efeitos adversos , Debilidade Muscular , Miastenia Gravis/complicações , Neoplasias do Timo/complicações , Vacinação/efeitos adversosRESUMO
Objective: To analyze the clinical characteristics and related genetic variation of juvenile myasthenia gravis (MG) patients. Methods: We collected the clinical data of adolescent MG patients who were treated in the Department of Neurology of the First Affiliated Hospital of Sun Yat-sen University from June 2019 to May 2020. After obtaining the patient's informed consent, the blood samples were collected. The Whole Exome Sequencing (WES) was performed on peripheral blood samples. And use biological information software and SPSS 22.0 for data processing and result analysis. Results: According to the inclusion and exclusion criteria, 54 patients with juvenile MG were included, 28 males and 26 females. And the average age of onset was (3.79±0.89) years. Among the enrolled patients, there were 52 (96.3%) patients with ocular MG, the MG-ADL scores of 54 patients were (3.44±0.44) points, and the titer of AChR antibody was (5.88±2.45) nmol/L. Two patients had thymic hyperplasia, and 5 patients had a family history of MG.A total of 169 variant genes were found in 54 patients, of which TTN gene variants had the largest number, with a total of 17 variants (31.5%). In the TTN gene variant group, 7(41.2%) patients had eye fixation symptoms, and 4 (10.8%) patients in the non-mutation group had eye fixation symptoms. And The difference between the two groups was statistically significant (P=0.016). In addition, the synaptic nucleus envelope protein-1 (SYNE1) and the ryanodine receptor-1 (RYR1) gene variations were also found in 7 cases (13.2%), and no clear relationship between these gene variations and clinical manifestations of MG was found. Conclusions: The incidence of juvenile MG was preschoolers with no gender difference, and ocular MG was more common. The proportion of TTN gene variation in adolescent MG was higher, suggesting that this gene may be a potential therapeutic target for juvenile MG patients.
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Miastenia Gravis , Hiperplasia do Timo , Adolescente , Anticorpos , Pré-Escolar , Feminino , Variação Genética , Humanos , Masculino , Miastenia Gravis/diagnóstico , Miastenia Gravis/genética , Receptores Colinérgicos/genética , Estudos RetrospectivosRESUMO
Objective: To investigate the clinical manifestations, treatment characteristics and outcomes of myasthenia gravis (MG) dually positive for anti-acetylcholine receptor antibody (AChR-Ab) and anti-muscle-specific tyrosine kinase antibody (MuSK-Ab). Method: MG patients hospitalized in the First Affiliated Hospital of Sun Yat-sen University from August 2017 to November 2020 were retrospectively collected. Thirty-four MuSK-Ab positive MG (MuSK-MG) patients, 11 double-antibodies positive MG (DP-MG) patients, and 80 AChR-Ab positive MG (AChR-MG) patients were included and allocated to three different groups. The clinical data of patients in the three groups were collected, and the differences of demographic characteristics, clinical manifestations and treatment outcomes between DP-MG patients and AChR-MG and MuSK-MG patients were analyzed. Result: The proportion of female and male patients in DP-MG group was 7/11 and 4/11 respectively, and the onset age of DP-MG was (41±27) years.The difference in gender distribution between DP-MG and AChR-MG groups was statistically significant (P<0.05). The proportion of extraocular muscle involvement in the DP-MG and MuSK-MG groups (8/11 and 52.9%) was lower than that in the AChR-MG group (83.8%), and the difference was statistically significant (P<0.05). The incidence of myasthenia crisis in DP-MG and MuSK-MG groups (54.5% and 61.8%) were higher than that in AChR-MG group (20.0%), with astatistically significant difference(P<0.05). The positive rate of neostigmine test in DP-MG and MuSK-MG groups(8/11 and 74.2%) were lower than that of AChR-MG group (96.8%), and the positive rate of low frequency repetitive nerve stimulation (RNS) in DP-MG group (5/10) was lower than that in AChR-MG group (85.1%), with statistically significant differences (all P<0.05). MuSK-Ab titer was positively correlated with the course of disease (r=0.466, P<0.05), and antibody titer decreased after symptom improvement (P<0.05). The response of patients in DP-MG and MuSK-MG groups to cholinesterase inhibitors (2/11 and 9.1%) was worse than that in the AChR-MG group (66.3%), and the incidence of side effects in the two groups (5/11 and 39.4%) was higher than that in the AChR-MG group (15.0%), with statistically significant differences (all P<0.05). There were 4 DP-MG patients underwent thymectomy, and the pathological results detected two cases of thymoma and two cases of thymic hyperplasia. Subsequent follow-up showed that 5 (5/11) DP-MG patients achieved minimal manifestation status or better status. Conclusion: The gender distribution, age of onset, pharmacological characteristics and electrophysiological examination of DP-MG patients were similar to those of MuSK-MG patients, but the severity of DP-MG patients was between that of AChR-MG and MuSK-MG patients.
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Miastenia Gravis , Neoplasias do Timo , Adolescente , Adulto , Idoso , Autoanticorpos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Músculos/patologia , Miastenia Gravis/diagnóstico , Receptores Proteína Tirosina Quinases , Receptores Colinérgicos , Estudos Retrospectivos , Tirosina , Adulto JovemRESUMO
OBJECTIVE: Acquired radioresistance remains the primary obstacle to improving the survival of esophageal cancer (EC). Related hypoxia factors play crucial roles in radioresistance such as hypoxia-inducible factor 1α (HIF-1α), vascular endothelial growth factor (VEGF), and angiogenic factor with G-patch and FHA domains 1 (AGGF1). The aim of this study was to investigate the effect of 2-methoxyestradiol (2ME2) on the radiosensitivity of EC. PATIENTS AND METHODS: The expression of HIF-1α, VEGF, and AGGF1 in 70 EC patients was detected using immunohistochemistry. The relation between the expression levels of the above genes with clinicopathologic characteristics was explored as well. Human ECA-109 cells were subjected to hypoxia and/or radiation in the presence or absence of 2ME2. Subsequently, cell growth, colony formation, and apoptosis were evaluated. Moreover, the mRNA and protein expression levels were determined using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blotting, respectively. RESULTS: HIF-1α, VEGF, and AGGF1 were significantly in EC tissues and were associated with cancer aggressiveness. 2ME2 treatment significantly increased the radiosensitivity of ESCC cells in a dose-dependent manner, which was related to the inhibited expression of HIF-1α. Subsequent immunohistochemical staining results showed that the VEGF expression was positively correlated with the HIF-1α expression in EC. CONCLUSIONS: High levels of HIF-1α, VEGF, and AGGF1 in EC are indicators of poor prognosis in patients treated with radiotherapy, which can offer new ideas and methods for the treatment of ESCC. In addition, 2ME2 inhibits the expression of HIF-1α and confers radiosensitivity in ECA-109 cells. Our findings suggest that 2ME2 has the potential to be applied as an adjuvant treatment with radiotherapy for EC.
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2-Metoxiestradiol/farmacologia , Neoplasias Esofágicas/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Idoso , Apoptose/efeitos dos fármacos , Relação Dose-Resposta a Droga , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/radioterapia , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/radioterapia , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Lane keeping systems for a keeping a vehicle in the desired lane is key to advanced driving assistance system in autonomous vehicles. This paper presents a cost-effective image sensor with efficient processing algorithm for lane detection and lane control applications to autonomous delivery systems. The algorithm includes (1) lane detection by inverse perspective mapping and random sample consensus parabola fitting and (2) lane control by pure pursuit steering controller and classical proportional integral speed controller based on a nonholonomic kinematic model. The image sensor experiments conducted on a 1/10 scale model car maneuvering in a straightâ»curveâ»straight lane validate the better processing performance before, during, and after the turning section over previous work. The image sensor with the processing algorithm achieves the average lane detection error within 5% and maximum cross-track error within 9% in real-time. The development shall pave the way to cost-effective autonomous delivery systems.
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OBJECTIVE: Disruptions of extracellular matrix (ECM) homeostasis are key events in the pathogenesis of osteoarthritis (OA). MicroRNA-140 (miRNA-140) is expressed specifically in cartilage and regulates ECM-degrading enzymes. Our objective in this study was to determine if intra-articular injection of miRNA-140 can attenuate OA progression in rats. DESIGN: miRNA-140 levels in human normal and OA cartilage derived chondrocytes and synovial fluid were assessed by polymerase chain reaction (PCR). After primary human chondrocytes were transfected with miRNA-140 mimic or inhibitor, PCR and western blotting were performed to quantify Collagen II, MMP-13, and ADAMTS-5 expression. An OA model was induced surgically in rats, and subsequently treated with one single intra-articular injection of miRNA-140 agomir. At 4, 8, and 12 weeks after surgery, OA progression were evaluated macroscopically, histologically, and immunohistochemically in these rats. RESULTS: miRNA-140 levels were significantly reduced in human OA cartilage derived chondrocytes and synovial fluid compared with normal chondrocytes and synovial fluid. Overexpressing miRNA-140 in primary human chondrocytes promoted Collagen II expression and inhibited MMP-13 and ADAMTS-5 expression. miRNA-140 levels in rat cartilage were significantly higher in the miRNA-140 agomir group than in the control group. Moreover, behavioural scores, chondrocyte numbers, cartilage thickness and Collagen II expression levels in cartilage were significantly higher, while pathological scores and MMP-13 and ADAMTS-5 expression levels were significantly lower in the miRNA-140 agomir group than in the control group. CONCLUSION: Intra-articular injection of miRNA-140 can alleviate OA progression by modulating ECM homeostasis in rats, and may have potential as a new therapy for OA.
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Artrite Experimental/tratamento farmacológico , Matriz Extracelular/efeitos dos fármacos , MicroRNAs/administração & dosagem , Osteoartrite/tratamento farmacológico , Proteína ADAMTS5/biossíntese , Proteína ADAMTS5/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Células Cultivadas , Condrócitos/metabolismo , Colágeno Tipo II/biossíntese , Colágeno Tipo II/genética , Progressão da Doença , Avaliação Pré-Clínica de Medicamentos/métodos , Matriz Extracelular/fisiologia , Regulação da Expressão Gênica/fisiologia , Homeostase/efeitos dos fármacos , Humanos , Injeções Intra-Articulares , Masculino , Metaloproteinase 13 da Matriz/biossíntese , Metaloproteinase 13 da Matriz/genética , MicroRNAs/metabolismo , MicroRNAs/farmacologia , MicroRNAs/uso terapêutico , Pessoa de Meia-Idade , Osteoartrite/metabolismo , Osteoartrite/patologia , Ratos Sprague-Dawley , Líquido Sinovial/metabolismo , Adulto JovemRESUMO
A method is proposed to determine the carrier-envelope phase (CEP) of a relativistic few-cycle laser pulse via the frequency of the Thomson backscattering (TBS) light. We theoretically investigate the generation of a flying mirror when a few-cycle drive pulse with relativistic intensity interacts with a target combined with a thin and a thick foil. The frequency of the TBS light generated from the flying mirror shows a sensitive dependence on the CEP of the drive pulse. The obtained results are verified by one-dimensional particle-in-cell simulations and are explained by an analytical model.
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By 3D particle-in-cell simulation and analysis, we propose a plasma lens to make high intensity, high contrast laser pulses with a steep front. When an intense, short Gaussian laser pulse of circular polarization propagates in near-critical plasma, it drives strong currents of relativistic electrons which magnetize the plasma. Three pulse shaping effects are synchronously observed when the laser passes through the plasma lens. The laser intensity is increased by more than 1 order of magnitude while the initial Gaussian profile undergoes self-modulation longitudinally and develops a steep front. Meanwhile, a nonrelativistic prepulse can be absorbed by the overcritical plasma lens, which can improve the laser contrast without affecting laser shaping of the main pulse. If the plasma skin length is properly chosen and kept fixed, the plasma lens can be used for varied laser intensity above 10(19) W/cm(2).
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A deuteron radio frequency quadrupoles injector h has been developed at Peking University. A permanent magnetic electron cyclotron resonance (ECR) ion source is used in the injector system. A 50 keV 100 mA proton beam has been extracted from the ECR ion source and the measured normalized rms emittance is 0.11-0.14pi mm mrad. A deuteron beam has also been extracted at 50 kV with 83 mA total current and its emittance is less than 0.18pi mm mrad. The proton beam transmission has been investigated on a low energy beam transport test bench, and up to 93% transmission can be reached. The new injector with two solenoids has been designed and is being constructed. All the development results will be presented in this paper.
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A set of new ion extraction electrodes have been designed for the permanent magnetic electron cyclotron resonance ion source at Peking University to improve beam quality and transmission. PBGUNS has been used to optimize the extraction electrodes and simulate the beam behavior at the extraction region. The experiments showed that with the new system, the beam half divergence angle can be less than 40 mrad and the normalized rms emittance is about 0.13pi mm mrad when the extracted current is 100 mA at 50 keV in pulse mode. The voltage of the suppression electrode has great effect on beam divergence. The effect of the microwave power and gas flow is also studied.
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A major concern in pig-to-human xenotransplantations is the potential risk of transmission of Porcine endogenous retroviruses (PERVs) integrated in the pig genome. Our previous work has shown that PERV provirus genes and gag protein can be detected in human embryonic kidney HEK-293 cells during a long-term infection with PERV (Yu et al., Transplant. Proc. 37, 496-499, 2005). In this study, we continued studying the long-term (>6 months) PERV infection of HEK-293 cells. The results showed no significant differences in morphology, growth, apoptosis, and [(3)H]-thymidine incorporation between PERV-infected and uninfected cells. The PERV LTR sequence showed only an insignifcant mutation after the long-term infection. PERV infection had no effect on the transcription of genes of Human endogenous retrovirus (HERV) naturally occurring in HEK-293 cells. Summing up, this study indicated that a long-term PERV infection of HEK-293 cells in vitro does not result in any significant changes in host cells as well as in PERV LTR sequence.
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Retrovirus Endógenos/genética , Suínos/virologia , Animais , Apoptose , Linhagem Celular , Retrovirus Endógenos/crescimento & desenvolvimento , Retrovirus Endógenos/ultraestrutura , Produtos do Gene gag/análise , Humanos , Mutação , Sequências Repetidas Terminais , Transplante HeterólogoRESUMO
OBJECTIVES: The diabetic rhesus monkey seems to be a useful model for preclinical investigations of islet transplantation and new drug treatments for type 1 diabetes mellitus (T1DM). Information is limited regarding a standard technique to induce and assess diabetes in rhesus monkeys as well as the strategy to apply insulin administration. Herein, we have established and characterized a model of diabetic rhesus macaques. METHODS: Four monkeys were divided into 2 groups of 2 each: group 1, total pancreatectomy; and group 2, partial pancreatectomy (75%) with low-dose streptozotocin (STZ) administration. Pancreatic function was measured using intravenous glucose tolerance tests before the operation. Spiral computed tomography (CT) scans of the pancreas were obtained before and after pancreatectomy. Fasting blood glucose and postprandial blood glucose levels were monitored twice daily using blood samples from the fingers or toes. Various types and doses of insulin were administered twice daily. We performed regular assessments of hematological and serum biochemical parameters, insulin, and C-peptide. RESULTS: Both total pancreatectomy and partial pancreatectomy (75%) with STZ administration induced T1DM in rhesus monkeys; there was interindividual variation in the STZ dose. Excluding C-peptide and insulin, the hematological and serum biochemical parameters did not differ significantly from normal values postoperatively. The various insulin treatment strategies are achieved stable blood glucose (BG) levels. CONCLUSIONS: STZ injection after partial pancreatectomy may be a safe, reproducible method to induce T1DM. Porcine insulin administration was a safe, economical method to control BG levels in a diabetic rhesus monkey before islet transplantation.
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Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/veterinária , Insulina/uso terapêutico , Doenças dos Macacos/tratamento farmacológico , Animais , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Tipo 1/sangue , Modelos Animais de Doenças , Esquema de Medicação , Teste de Tolerância a Glucose , Humanos , Insulina/administração & dosagem , Macaca mulatta , Pancreatectomia/métodos , Pancreatectomia/veterinária , Tomografia Computadorizada EspiralRESUMO
A new ion acceleration method, namely, phase-stable acceleration, using circularly-polarized laser pulses is proposed. When the initial target density n(0) and thickness D satisfy a(L) approximately (n(0)/n(c))D/lambda(L) and D>l(s) with a(L), lambda(L), l(s), and n(c) the normalized laser amplitude, the laser wavelength in vacuum, the plasma skin depth, and the critical density of the incident laser pulse, respectively, a quasiequilibrium for the electrons is established by the light pressure and the space charge electrostatic field at the interacting front of the laser pulse. The ions within the skin depth of the laser pulse are synchronously accelerated and bunched by the electrostatic field, and thereby a high-intensity monoenergetic proton beam can be generated. The proton dynamics is investigated analytically and the results are verified by one- and two-dimensional particle-in-cell simulations.
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INTRODUCTION: To provide standard implemental cells for biomedical research, an immortalized mesenchymal stem cell (MSC) lineage was established from the bone marrow of Chinese Guizhou minipigs. We investigated the characteristics of the MSC transfected with hTERT. METHODS: MSCs isolated from bone marrow (BM) were steadily transfected using a plasmid containing human telomerase reverse transcriptase gene (pCI-neo-hTERT) at population doubling (PD) time of 3. The expression of hTERT was analyzed by reverse transcriptase polymerase chain reaction; the morphology of the cells, by phase contrast microscope; cell growth curve, by MTS assay, and phenotype, flow cytometry and immunofluorescence. Karyotype and osteogenic differentiation of transfected cells were compared with normal MSCs. RESULTS: After transfection with the hTERT gene, the MSCs showed vigorous proliferation activity undergoing more than 60 PDs. Expression of the hTERT gene was detected in these cells. Transfected MSCs showed a prolonged life span, maintaining similar morphology and karyotype compared with normal MSCs. The apoptotic rate of transfected MSCs at PD60 was far below that of the normal MSCs at PD15. More than 99% of transfected MSCs were positive for stem cell markers, including SH-2, SH-3, SB-21, and CD29, and negative for CD34, CD45, and SLA-II. Transfected MSCs possessed the ability to differentiate into osteogenic as same as that of normal MSCs. CONCLUSION: The hTERT gene-induced immortalization prolonged the life span of MSCs, maintaining the overall properties identical to the original cells, findings that will be useful for basic research.
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Células-Tronco Mesenquimais/citologia , Telomerase/genética , Animais , Apoptose , Células da Medula Óssea/citologia , Divisão Celular , Sobrevivência Celular , Humanos , Células-Tronco Mesenquimais/enzimologia , Células-Tronco Mesenquimais/fisiologia , Suínos , Porco Miniatura , TransfecçãoRESUMO
OBJECTIVES: A pig-to-monkey transplant model was initiated to investigate the outcome of pig-to-human xenotransplantation. Though monkey is close to human in biology and physiology, the genetic differences between the two species remains unclear. This study sought to compare the gene expressions of three tissues from humans and rhesus monkey. METHODS: RNA samples extracted from liver, spleen, and peripheral blood cells were hybridized onto Illumina gene expression microarray. Genes with detected signals greater than 1000 and diff-scores higher than 100 were selected as significant results. The data were analyzed with Illumina software. mRNA expression levels were confirmed by semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR) analysis. RESULTS: Of the 47,293 transcripts tested on every gene chip, more than 6000 genes were expressed in three tissues. Total numbers of genes detected and the similarity ratios followed the same rule as liver < PBC < spleen. The 136 IRI-related genes, 192 immunological-related genes, and 131 cell cycle-related genes selected and analyzed showed gene expression concordance rates of 82.35%, 72.92%, and 77.10%, respectively. RT-PCR tests indicated similar mRNA expression levels of RTN4, interleukin (IL)-1beta, NF-kappaB1, IL-8, and G0S2 to the results on chips. CONCLUSIONS: The detected mRNA expressions in human and monkey tissues showed an average consistency in 85.78%, indicating that a human microarray might provide a part of the information for monkey sample testing. Therefore, in pig-to-monkey transplant models, monkey microarray may be used to determine recipient gene expressions. The genetic difference between human and monkey must be taken into account in interpreting the experimental results.
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Perfilação da Expressão Gênica , Macaca mulatta/genética , Análise de Sequência com Séries de Oligonucleotídeos , Transplante Heterólogo , Animais , Amplificação de Genes , Humanos , Masculino , RNA/genética , RNA/isolamento & purificação , Valores de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Suínos , Transcrição GênicaRESUMO
OBJECTIVE: Hemorrhagic diseases have been considered to be one of the main causes of xenotransplantation failure. To explore the role of the rhesus monkey (Macaca mulatta) coagulation system in a pig-to-human xenotransplantation model, we primarily studied the full-length cDNA and the three-dimensional (3-D) structures of the important coagulation factor-prothrombin of the rhesus. MATERIALS AND METHODS: The full-length cDNA of rhesus prothrombin was obtained by rhesus monkey liver cDNA library screening and a 5'RACE technique. The 3-D protein structure was modeled using the SWISS-MODEL program. The important macromolecular interaction sites were compared with human and porcine prothrombin. RESULTS: At first, the full-length rhesus prothrombin cDNA was cloned; the sequence was submitted to the Genebank (accession number: EF057490). The full-length cDNA is 2029 bp, with a complete open reading frame of 1884 bp, coding 626 amino acids. The deduced protein sequence contains a signal peptide, propeptide, Gla domain, two Kringle domains, and a Trypsin domain. The nucleotide similarities of rhesus-human and rhesus-porcine genes are 95.64% and 86.14%, and those of the amino acids, 94.51% and 82.82%, respectively. Some important functional sites, such as the catalytic triad DHS, RGD, Na+ -binding, and the carboxylase-binding site, are highly conserved. However, among the three species some variations are observed in potential N-glycosylation, O-glycosylation, phosphorylation, cleavage site, FXa-binding sites. Especially, the autolysis loop shows great differences in both amino acid sequence and 3-D model. CONCLUSIONS: The great similarity of prothrombin among rhesus, human, and porcine confirmed the great value of the pig-to-rhesus xenotransplantation model. The variation especially in some important recognition sites between rhesus and pig would vary the binding affinity of enzymes to xenosubstrates and their reaction velocities in postoperative coagulation processes, which would be one possible reason for the disordered regulation of clotting seen during the xenorejection in animal models.
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Clonagem Molecular , DNA Complementar/genética , Macaca mulatta/genética , Protrombina/genética , Transplante Heterólogo/normas , Sequência de Aminoácidos , Animais , Sequência de Bases , Biologia Computacional , Feminino , Humanos , Masculino , Modelos Moleculares , Dados de Sequência Molecular , Protrombina/química , Segurança , Suínos , Porco Miniatura/genéticaRESUMO
INTRODUCTION: Our previous experiments indicated that bone marrow mesenchymal stem cells of rhesus monkey (RhBMSCs) have a low proliferative ability with a finite life span, which will hamper their application in biomedical research. Establishing an immortalized RhBMSC lineage might solve the problem. METHODS: RhBMSCs isolated from the bone marrow of rhesus monkeys using density gradient centrifugation were purified using adherence separation. Then, the cells were steadily transfected by plasmid containing human telomerase reverse transcriptase gene (pCI-neo-hTERT). We analyzed expression of hTERT, proliferation, phenotype (SH-2, SH-3, SB-10, CD29, CD34, CD45, and HLA-DR), differentiation toward osteogenic lineage, karyotype, and tumorigenesis of transfected cells. RESULTS: After transfection, the RhBMSCs proliferated vigorously, undergoing more than 50 population doublings (PDs). Apoptotic rate of transfected RhBMSCs at PD40 was only 4.5%, versus untransfected RhBMSCs at PD15, which was more than 33.5%. Compared with normal RhBMSC, the life span of transfected RhBMSCs was prolonged, retaining similar morphology, karyotype, and potential to differentiate into an osteogenic lineage. More than 99% of transfected RhBMSCs were positive for stem cell markers, including SH-2, SH-3, SB-10, and CD29, and negative for CD34, CD45, and HLA-DR. Furthermore, the transfected cell line was benign in nude mice tumor formation. CONCLUSION: Our results demonstrated that hTERT gene had been transfected into RhBMSCs. The transfected RhBMSCs proliferated vigorously. Phenotype, differentiation, and karyotype of transfected RhBMSC showed no significant difference from untransfected cells. The transfected RhBMSCs are a potential cell source for transplantation as well as tissue engineering.
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Células da Medula Óssea/citologia , Células-Tronco Mesenquimais/citologia , Telomerase/genética , Animais , Diferenciação Celular , Divisão Celular , Separação Celular/métodos , Primers do DNA , Humanos , Cinética , Macaca mulatta , Osteogênese , Plasmídeos , Transcrição Gênica , TransfecçãoRESUMO
In an electron cyclotron resonance (ECR) ion source, the magnetic field along the axis of the plasma chamber and extraction system is a key parameter. At Peking University, a new 2.45 GHz ECR ion source (PMECR III), dedicated to proton production, has been developed to investigate the influence of the magnetic field on the gas discharge and beam characteristics. The magnetic configuration is provided by two permanent magnet rings independently tunable along the source axis. Moreover, the beam extraction position changes by moving the whole magnetic system along the source axis and by using different lengths of plasma electrode. A brief description of the source is reported and the magnetic field influence results are presented.
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Xenotransplantation is associated with the risk of Porcine endogenous retrovirus (PERV) transmission, since it has been shown that PERV can infect human cells in vitro (Specke et al., Virology 285, 177-180, 2001). We evaluated the possibility of PERV infection of human cells in nude mice model. Porcine kidney cells PK15 carrying PERV and human liver cancer cells SMMC-7721 were injected separately into the right and left axilla of nude mice, respectively. Two months later, pig cytochrome oxidase II (COII) gene, PERV DNA, PERV mRNA, and PERV-Gag protein were detected in the mass formed in both axillas and in several organs of nude mice. The pig COII genes were detected in the right and left axilla, but not in other organs of nude mice implicating that the microchimerism of pig cells occurred in human SMMC-7721 cells and induced the formation of the mass. PERV gene and gag protein were detected in all mouse tissues except liver. These data indicated that (i) PERV may be transmitted from porcine to mouse cells, (ii) PERV genes and proteins were detectable in the mass formed by injection of human cells and consequently (iii) there was a possibility of PERV transmission to human cells after xenotransplantation.