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A large qubit capacity and an individual readout capability are two crucial requirements for large-scale quantum computing and simulation1. As one of the leading physical platforms for quantum information processing, the ion trap has achieved a quantum simulation of tens of ions with site-resolved readout in a one-dimensional Paul trap2-4 and of hundreds of ions with global observables in a two-dimensional (2D) Penning trap5,6. However, integrating these two features into a single system is still very challenging. Here we report the stable trapping of 512 ions in a 2D Wigner crystal and the sideband cooling of their transverse motion. We demonstrate the quantum simulation of long-range quantum Ising models with tunable coupling strengths and patterns, with or without frustration, using 300 ions. Enabled by the site resolution in the single-shot measurement, we observe rich spatial correlation patterns in the quasi-adiabatically prepared ground states, which allows us to verify quantum simulation results by comparing the measured two-spin correlations with the calculated collective phonon modes and with classical simulated annealing. We further probe the quench dynamics of the Ising model in a transverse field to demonstrate quantum sampling tasks. Our work paves the way for simulating classically intractable quantum dynamics and for running noisy intermediate-scale quantum algorithms7,8 using 2D ion trap quantum simulators.
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Generating ion-photon entanglement is a crucial step for scalable trapped-ion quantum networks. To avoid the crosstalk on memory qubits carrying quantum information, it is common to use a different ion species for ion-photon entanglement generation such that the scattered photons are far off-resonant for the memory qubits. However, such a dual-species scheme can be subject to inefficient sympathetic cooling due to the mass mismatch of the ions. Here we demonstrate a trapped-ion quantum network node in the dual-type qubit scheme where two types of qubits are encoded in the S and F hyperfine structure levels of 171Yb+ ions. We generate ion photon entanglement for the S-qubit in a typical timescale of hundreds of milliseconds, and verify its small crosstalk on a nearby F-qubit with coherence time above seconds. Our work demonstrates an enabling function of the dual-type qubit scheme for scalable quantum networks.
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Non-Hermitian systems generically have complex energies, which may host topological structures, such as links or knots. While there has been great progress in experimentally engineering non-Hermitian models in quantum simulators, it remains a significant challenge to experimentally probe complex energies in these systems, thereby making it difficult to directly diagnose complex-energy topology. Here, we experimentally realize a two-band non-Hermitian model with a single trapped ion whose complex eigenenergies exhibit the unlink, unknot, or Hopf link topological structures. Based on non-Hermitian absorption spectroscopy, we couple one system level to an auxiliary level through a laser beam and then experimentally measure the population of the ion on the auxiliary level after a long period of time. Complex eigenenergies are then extracted, illustrating the unlink, unknot, or Hopf link topological structure. Our work demonstrates that complex energies can be experimentally measured in quantum simulators via non-Hermitian absorption spectroscopy, thereby opening the door for exploring various complex-energy properties in non-Hermitian quantum systems, such as trapped ions, cold atoms, superconducting circuits, or solid-state spin systems.
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OBJECTIVE: Chinese herbal medicine (CHM) has been widely used in the treatment of hyperlipidemic acute pancreatitis (HLAP), but the credibility of the evidence for this practice is unclear. We systematically reviewed the efficacy and safety of CHM therapy for HLAP. MATERIALS AND METHODS: In this systematic review and meta-analysis, we searched the Cochrane Central Register of Controlled Trials, Ovid MEDLINE, PubMed, EMBASE, CBM, CNKI, VIP, and Wanfang databases from inception to October 16, 2022, for randomized controlled trials comparing the combination of CHM and Western medicine therapy vs. Western medicine therapy alone in HLAP adults. This study is registered with PROSPERO (No. CRD 42022371052). RESULTS: A total of 50 eligible studies involving 3,635 patients were assessed in this meta-analysis. Compared with Western medicine therapy, the combination of CHM increased the total effective rate by 19% in HLAP patients [relative risk (RR): 1.19, 95% CI: (1.16, 1.23)]. There were significant differences between the two groups in improving clinical symptoms, promoting serum amylase and triglyceride recovery, reducing mortality [RR: 0.28, 95% CI: (0.14, 0.56)] and complication rates [RR:0.40, 95% CI: (0.31, 0.52)], and shortening the length of hospital stay [MD: -3.96, 95% CI: (-4.76, -3.16)]. Adverse reactions were similar between groups. Findings were robust in the sensitivity analysis. CONCLUSIONS: The combined CHM treatment was more effective than Western medicine alone in HLAP patients. However, due to the methodological shortcoming of the eligible studies, caution is needed when interpreting these findings.
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Medicamentos Herbarios Chinos , Pancreatitis , Adulto , Humanos , Medicamentos Herbarios Chinos/uso terapéutico , Enfermedad Aguda , Pancreatitis/tratamiento farmacológico , Pancreatitis/inducido químicamente , Ensayos Clínicos Controlados Aleatorios como Asunto , FitoterapiaRESUMEN
OBJECTIVE: The aim of this study was to explore the expression pattern of TRIM56 in Hepatocellular carcinoma (HCC) patients and its influence on the prognosis, and to illustrate the molecular mechanisms of TRIM56 in regulating HCC cell behaviors. PATIENTS AND METHODS: TRIM56 levels in HCC specimens and paracancerous specimens were detected. Then, the influences of TRIM56 on clinical data and prognosis in HCC patients were assessed. Next, the regulatory effects of TRIM56 on proliferative potential in Huh7 and Bel-7402 cells were determined, and the role of TRIM56 on the Wnt signaling was examined. Finally, biological characteristics between TRIM56 and RBM24 in HCC development were illustrated by Luciferase assay and rescue experiments. RESULTS: TRIM56 was lowly expressed in HCC tissues and cell lines. HCC patients expressing a low level of TRIM56 suffered advanced T stage and poor survival. Besides, overexpression of TRIM56 inhibited proliferative potential of Huh7 cells, while knockdown of TRIM56 in Bel-7402 yielded the opposite result. TRIM56 was able to negatively regulate key genes in the Wnt signaling. In addition, RBM24 was proven to be the downstream target of TRIM56, which was involved in TRIM56-influenced HCC development. CONCLUSIONS: Downregulated TRIM56 in HCC samples is closely linked to pathological staging and prognosis. TRIM56 alleviates the malignant development of HCC by inactivating the Wnt signaling and targeting RBM24.
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Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas de Motivos Tripartitos/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Carcinoma Hepatocelular/patología , Proliferación Celular , Células Cultivadas , Femenino , Humanos , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Proteínas de Unión al ARN/genética , Proteínas de Motivos Tripartitos/genética , Ubiquitina-Proteína Ligasas/genética , Vía de Señalización WntRESUMEN
OBJECTIVE: The aim of this study was to explore the effects of long non-coding ribonucleic acid (lncRNA) placenta-specific protein 2 (PLAC2) on the biological behaviors of gastric cancer (GC) cells by regulating the expression of c-Myc gene and its mechanism. PATIENTS AND METHODS: The expression of PLAC2 in GC tissues and different GC cell lines was detected via quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR). The effects of PLAC2 on apoptosis and cycle, migration, and invasion of GC cells were detected using flow cytometry, wound healing assay, and transwell assay, respectively. After interference in PLAC2 expression, the changes in c-Myc expression were determined through qRT-PCR and Western blotting. RESULTS: The expression level of PLAC2 was downregulated in 38 out of 45 cases of GC tissues compared with that in normal gastric tissues, and it also declined in GC cells. The results of flow cytometry showed that after overexpression of PLAC2, the cell cycle was arrested in the G1/G0 phase, and the apoptosis rate was increased. The results of wound healing assay and transwell assay revealed that both migration and invasion of GC cells were inhibited. After overexpression of PLAC2, the mRNA and protein expression levels of c-Myc declined. CONCLUSIONS: LncRNA PLAC2 affects the biological behaviors of GC cells by regulating the expression of c-Myc gene.
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Regulación hacia Abajo , Proteínas Proto-Oncogénicas c-myc/metabolismo , ARN Largo no Codificante/metabolismo , Neoplasias Gástricas/metabolismo , Supervivencia Celular , Células Cultivadas , Humanos , Proteínas Proto-Oncogénicas c-myc/genética , ARN Largo no Codificante/genética , Neoplasias Gástricas/patologíaRESUMEN
Detected in a variety of solid tumors, including lung cancer, the EML4-ALK fusion gene plays an important role in promoting the occurrence and development of cancer. The existing detection methods for EML4-ALK fusion gene are all targeted at surgical or post-sampling tumor tissues, which cannot achieve early detection and real-time monitoring; therefore, a minimally invasive ALK gene fusion detection system is explored and constructed. Vimentin, EpCAM, and EGFR antibodies were grafted, respectively, to prepare multi-site immunoliposome magnetic beads, so as to capture CTC in blood for RT-PCR detection, and then the feasibility of this method was verified by detecting the positive rate of the EML4-ALK fusion gene and clinical information in combination with WB and IHC. The prepared multi-site immunoliposome magnetic beads showed high specificity and stability, and the average proliferation rate and capture rate of cells were 95% and 85%, respectively. In clinical blood samples, the CTC level of the grade I (G1) patients before the operation was lower than grade 2 (G2), and that of grade II (G2) was significantly lower than grade III (G3), but the difference was not significant after the operation. The RT-PCR results of CTC and the RT-PCR, WB, and IHC results of tissues were highly consistent in the fusion gene detection, and the positive rate of ALK gene fusion in 60 lung cancer patients was 31.67% and 28.33% before and after the operation, mostly EML4-ALK (V3) gene fusion. The CTC-ALK gene fusion detection system constructed successfully could avoid the problem of difficult sampling and post-sampling complications, and truly achieve the minimally invasive biopsy, so it was of important clinical significance for the diagnosis and efficacy evaluation of lung.
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Quinasa de Linfoma Anaplásico/genética , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Proteínas de Fusión Oncogénica/genética , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Carcinoma de Pulmón de Células no Pequeñas/genética , Fusión Génica , Humanos , Liposomas , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Magnetismo , Proteínas Tirosina Quinasas Receptoras/genéticaRESUMEN
OBJECTIVE: Studies have found that hsa_circ_103809, a newly discovered circRNA in recent years, can serve as an oncogene involved in the progression of hepatocellular carcinoma. However, its role in gastric cancer (GCa) remains elusive. The aim of this study was to reveal the molecular mechanism of hsa_circ_103809 affecting the process of GCa, thus providing new ideas for its treatment. PATIENTS AND METHODS: Hsa_circ_103809 expression in GCa and adjacent tissues specimens were studied by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) analysis, and its effect on the prognosis of GCa patients was analyzed. In GCa cells lines, hsa_circ_103809 was knocked down by small interfering RNA, and GCa cell metastasis ability was detected by cell wound healing test and transwell assay. Finally, the potential target gene of hsa_circ_103809 was predicted through bioinformatics website and verified by Luciferase assay. RESULTS: Hsa_circ_103809 showed an increased expression both in GCa tissues and cell lines, predicting a poor prognosis of GCa patients. Meanwhile, the invasive and migration capacities of GCa cells were remarkably reduced after the knockdown of hsa_circ_103809. Bioinformatics website predicted that there existed binding sites of hsa_circ_103809 on microRNA-101-3p, and Luciferase assay verified that hsa_circ_103809 can adsorb microRNA-101-3p. In GCa tissues, qPCR detected a significantly reduced expression of microRNA-101-3p, which was negatively correlated with that of hsa_circ_103809. In addition, the knockdown of hsa_circ_103809 enhanced microRNA-101-3p expression in GCa cell lines. Subsequent in vitro experiments further detected that the overexpression of hsa_circ_103809 partially reversed the inhibitory effect of microRNA-101-3p overexpression on GCa cell migration ability and invasiveness. CONCLUSIONS: Hsa_circ_103809, highly expressed in GCa, may promote the migration capacity of GCa cells by adsorbing microRNA-101-3p and thus become a new therapeutic target for GCa.
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Movimiento Celular , MicroARNs/metabolismo , Invasividad Neoplásica , ARN Circular/metabolismo , Neoplasias Gástricas/metabolismo , Sitios de Unión , Proliferación Celular , Células Cultivadas , Humanos , MicroARNs/genética , ARN Circular/genética , Neoplasias Gástricas/patologíaRESUMEN
During May of 2009, a new devastating disease was observed on pomegranate (Punica granatum L.) that caused losses estimated at 30% as surveyed by 10 orchards in Panzhihua-Xichang Region of Sichuan Province, Southwest China. Characteristic symptoms were yellow and wilting leaves. Initial symptoms only occurred on shoots, but later, leaves of the whole tree turned yellow and wilted, causing extensive defoliation and dieback and the xylem of the trunk turned brown to black with a star burst-like pattern. Finally, heavy infection resulted in the whole tree dying, causing severe yield losses. A fungus was consistently isolated from basal stems and roots of diseased plants. Single conidia were obtained and cultured on potato dextrose agar (PDA) and incubated at 25 ± 1°C with a 12-h light/dark photoperiod. Mycelium was initially hyaline and then rapidly became dark greenish brown. Two types of endoconidia were produced in 5 days. Barrel-shaped conidia were hyaline, 1-celled, and measured 7.3 to 9.4 × 11.6 to 13.2 µm. Cylindrical conidia were hyaline, 1-celled, and measured 9.2 to 29.6 × 3.1 to 6.8 µm. Aleurioconidia were brownish, thick walled, near globose, and measured 8.7 to 18.1 × 8.2 to 10.7 µm. Perithecia were dark brown to black, globose, measured 90.8 to 149.8 µm in diameter, and had a long thin neck, 254.4 to 533.8 µm long, through which ascospores exuded. Ascospores were small, hyaline, hat shaped, measured 3.7 to 6.5 × 3.1 to 5.7 µm, and accumulated in a sticky matrix at the tip of the ascomal neck. The fungus was identified as Ceratocystis fimbriata (anamorph Chalara sp.) (1). The internal transcribed spacer (ITS) region of rDNA was amplified with universal primers ITS4/ITS5 and sequenced (GenBank Accession No. HQ529711), and comparisons with GenBank showed 99% similarity with C. fimbriata on Colocasia esculenta from Brazil (Accession No. AM712448.1). Pathogenicity tests were conducted. Two-week-old seedlings of pomegranate cv. Qingpiruanzi, germinated in plastic containers in the greenhouse, were wounded with a needle to a depth of 0.5 mm at the base of the stem below the soil level and near the root system, and then inoculated by drenching the wounds with a spore suspension (105 conidia per ml). Control plants were inoculated with sterile water. There were four replicates for each treatment. The treated plants were incubated at 25 ± 1°C with 80 to 95% relative humidity under a 12-h light/dark photoperiod in a greenhouse. All inoculated plants wilted within 25 days after inoculation and C. fimbriata was reisolated. All control plants remained healthily. To our knowledge, this is the first finding of pomegranate wilt caused by C. fimbriata in Sichuan Province. This pathogen may pose a serious threat to pomegranate production in Sichuan where it is a major fruit tree. This pathogen has been previously reported in India (3) and Yunnan Province, China (2), but is not known elsewhere. References: (1) C. J. B. Engelbrecht and T. C. Harrington. Mycologia 97:57, 2005. (2) Q. Huang et al. Plant Dis. 87:1150, 2003. (3) Y. M. Somasekhara. Plant Dis. 83:400, 1999.
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BACKGROUND: Portal vein tumour thrombus (PVTT) is highly associated with the progression and metastasis of hepatocellular carcinoma (HCC). However, there are no appropriate cell models of PVTT with which to study the biological and physiological characteristics of PVTT. METHODS: Primary cell culture was performed by the use of a successive xenograft line called PVTT-#1, which was obtained from a 60-year-old male HCC patient accompanied by PVTT. RESULTS: A successive cell line named CSQT-2 was established. The cell line showed aggressive phenotypes in terms of cell growth, survival, migration, xenograft and metastasis. Moreover, an orthotopic transplantation assay showed that PVTT can be generated in nude mice when CSQT-2 cells were inoculated in the liver and that it shows a typical migratory tendency in the vascular branches of portal vein. Moreover, the established CSQT-2 cells also showed varied expression of tumour-initiating cell (TIC) markers such as CD133, CD90 and EpCAM. CONCLUSION: Establishment of CSQT-2 may provide a suitable model with which to investigate the molecular mechanisms of PVTT-related HCC.
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Carcinoma Hepatocelular/patología , Línea Celular Tumoral/patología , Neoplasias Hepáticas/patología , Vena Porta/patología , Trombosis/patología , Neoplasias Vasculares/secundario , Animales , Carcinoma Hepatocelular/complicaciones , Carcinoma Hepatocelular/genética , Movimiento Celular/fisiología , Análisis Citogenético , Humanos , Neoplasias Hepáticas/complicaciones , Neoplasias Hepáticas/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Persona de Mediana Edad , Metástasis de la Neoplasia , Células Madre Neoplásicas/patología , Trombosis/complicaciones , Trombosis/etiología , Trasplante Heterólogo , Neoplasias Vasculares/complicaciones , Neoplasias Vasculares/genética , Neoplasias Vasculares/patologíaRESUMEN
The retinoid, N-(4-hydroxyphenyl)retinamide (4-HPR), mediates p53-independent cytotoxicity and can increase reactive oxygen species and ceramide in solid tumor cell lines. We determined changes in ceramide and cytotoxicity upon treatment with 4-HPR (3-12 microM) in six human acute lymphoblastic leukemia (ALL) cell lines: T cell (MOLT-3, MOLT-4, CEM), pre-B-cell (NALM-6, SMS-SB), and null cell (NALL-1). Exposure to 4-HPR (12 microM) for 96 h caused 4.7 (MOLT-3), 3.5 (MOLT-4), 3.9 (CEM), 2.9 (NALM-6), 4.7 (SMS-SB), AND 4.5 (NALL-1) logs of cell kill. The average 4-HPR concentration that killed 99% of cells (LC(99)) for all six lines was 4.8 microM (range: 1.5-8.9 microM). Treatment with 4-HPR (9 microM) for 24 h resulted in an 8.9 +/- 1.0-fold (range: 4.9-15.7-fold) increase of ceramide. Ceramide increase was time- and dose-dependent and abrogated by inhibitors of de novo ceramide synthesis. Concurrent inhibition of ceramide glycosylation/acylation by d,l-threo-(1-phenyl-2-hexadecanoylamino-3-morpholino-1-propanol) (PPMP) further increased ceramide levels, and synergistically increased 4-HPR cytotoxicity in four of six ALL cell lines. 4-HPR was minimally cytotoxic to peripheral blood mononuclear cells and a lymphoblastoid cell line, and increased ceramide <2-fold. Thus, 4-HPR was cytotoxic and increased ceramide in ALL cell lines, but not in non-malignant lymphoid cell types.
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Antineoplásicos/farmacología , Ceramidas/biosíntesis , Fenretinida/farmacología , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , Supervivencia Celular/efectos de los fármacos , Ceramidas/metabolismo , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Humanos , Linfocitos/citología , Linfocitos/efectos de los fármacos , Morfolinas/farmacología , Esfingolípidos/farmacología , Factores de Tiempo , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
Endothelial cells express a variety of receptor systems involved in humoral defense, including receptors for the collagen-like and globular domains of the complement component C1q, designated cC1qR and gC1qR, respectively. In the present study a microvascular endothelial cell line was used to test the hypothesis that expression of these C1q-binding proteins may be affected by vascular inflammatory reactions. The results demonstrate that the expression of both cC1qR and gC1qR by bone marrow vascular endothelial cells is up-regulated by inflammatory mediators, interferon-gamma, tumor necrosis factor-alpha, and lipopolysaccharide (Escherichia coli, 055:B5) in a dose- and time-dependent manner, as detected by enzyme-linked immunosorbent assay. cC1qR and gC1qR expression increased significantly (P < .05) within 4 to 7 hours and doubled after 22 hours of stimulation. 3H-thymidine incorporation studies and direct cell counts confirmed that increased C1qR expression was not due to increased cell proliferation. Northern blot analysis revealed that the up-regulation of cC1qR and gC1qR protein expression was preceded by increases in corresponding mRNA levels, suggesting increased gene transcription. Indeed C1qR mRNA up-regulation was prevented by actinomycin D, and C1qR protein synthesis was inhibited by cycloheximide. Bone marrow vascular endothelial cell exposure to C1q, however, did not alter cC1qR or gC1qR expression, but up-regulation of the leukocyte adhesion molecule ICAM-1 was noted in the presence of aggregated C1q. The up-regulation of C1qR by inflammatory mediators and the ability of C1q itself to increase ICAM-1 expression suggest a potential role for these binding sites in vascular inflammation and immune injury.
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Complemento C1q/metabolismo , Citocinas/fisiología , Endotelio Vascular/metabolismo , Receptores de Hialuranos , Inflamación/metabolismo , Glicoproteínas de Membrana , Receptores de Complemento/metabolismo , Northern Blotting , Western Blotting , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/fisiología , Proteínas Portadoras , División Celular , Línea Celular , Citocinas/farmacología , Endotelio Vascular/citología , Escherichia coli/química , Humanos , Interferón gamma/farmacología , Lipopolisacáridos/farmacología , Microcirculación , Proteínas Mitocondriales , ARN Mensajero/metabolismo , Proteínas Recombinantes , Factor de Necrosis Tumoral alfa/farmacología , Regulación hacia ArribaRESUMEN
1. In order to investigate the biological function of the human CLN3 gene that is defective in Batten disease, we created a yeast strain by PCR-targeted disruption of the yeast gene (YHC3), which is a homologue of the human CLN3 gene. 2. The phenotypic characterization revealed that the yhc3 delta mutants are more sensitive to combined heat and alkaline stress than the wild-type strains as determined by inhibition of cell proliferation. 3. This suggests that the yhc3 delta mutant is a good model to investigate the biological function of human CLN3 gene in mammalian cells and to understand the pathophysiology of juvenile Batten disease.
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Glicoproteínas de Membrana , Chaperonas Moleculares , Lipofuscinosis Ceroideas Neuronales/genética , Proteínas/genética , Saccharomyces cerevisiae/genética , Alcalosis/fisiopatología , Antineoplásicos/farmacología , Ciclinas/genética , Inhibidores Enzimáticos/farmacología , Humanos , Concentración de Iones de Hidrógeno , Lipofuscinosis Ceroideas Neuronales/fisiopatología , Oligomicinas/farmacología , Proteínas/fisiología , Saccharomyces cerevisiae/efectos de los fármacos , Proteínas de Saccharomyces cerevisiae/genéticaRESUMEN
Juvenile neuronal ceroid lipofuscinosis or Batten disease (JNCL) is a neurodegenerative disorder characterized by blindness, seizures, cognitive decline and early death. Brain atrophy and retinitis pigmentosa ensue because of neuronal and photoreceptor apoptosis. The CLN3 gene defective in JNCL encodes a novel 438 amino acid protein. Most affected genes harbor a deletion resulting in a truncated protein. CLN3 overexpression in NT2 cells enhances growth, reverses growth inhibition induced by serum starvation and protects from apoptosis induced by vincristine, staurosporine, and etoposide but not from death caused by ceramide. CLN3 modulates endogenous and vincristine-activated ceramide, and therefore suppresses apoptosis by impacting generation of ceramide.
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Apoptosis , Ceramidas/metabolismo , Ciclinas , Glicoproteínas de Membrana/metabolismo , Chaperonas Moleculares/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Proteínas de Saccharomyces cerevisiae , Esfingosina/análogos & derivados , Western Blotting , División Celular , Línea Celular , Supervivencia Celular , Células Cultivadas , Relación Dosis-Respuesta a Droga , Etopósido/farmacología , Humanos , Modelos Biológicos , Poli(ADP-Ribosa) Polimerasas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Esfingosina/metabolismo , Estaurosporina/farmacología , Factores de Tiempo , Transfección , Vincristina/farmacologíaRESUMEN
Mutations in presenilin 1 (PS-1) and presenilin 2 (PS-2) have been linked to early onset, autosomal dominant Alzheimer's disease. Neither the normal function(s) of the presenilins nor their role(s) in mediating the devastating neurological and pathological changes associated with Alzheimer's Disease, however, are well understood. The results of the experiments described here demonstrate that expression of wild-type PS-1 or PS-2 increases outward K+ current densities in HEK-293 cells relative to untransfected or mock-transfected cells. Western blot analysis reveals that there is a marked increase in full-length, rather than processed, presenilins in transiently transfected HEK-293 cells, suggesting that full-length PS-1 (or PS-2) underlies the observed increases in outward K+ current densities. Consistent with this hypothesis, EXpression of an N-terminal proteolytic fragment of PS-1 is without effects on the membrane properties of HEK-293 cells. Mean outward K+ current densities are also shown to be increased in HEK-293 cells expressing the exon 9 splice site PS-1 mutation (deltaex9/PS-1), a mutant that does not undergo proteolytic processing. In HEK- 293 cells transiently transfected with a missense (G209V) PS-1 mutant, however, mean K+ current densities were not significantly different from controls. Expression of wild-type PS-1 in neonatal rat ventricular myocytes also results in increased outward K+ currents, whereas no detectable effects on membrane currents were seen in PS-1-transfected COS-7 cells. These results suggest that the presenilins do not actually form K+ channels, but rather that these proteins upregulate functional K+ channel expression either directly by associating with K+ channel pore-forming subunits or indirectly by increasing the synthesis, assembly, and/or transport of these subunits. The observation that PS-1 and PS-2 are highly expressed in neurons, localized to the endoplasmic reticulum, suggests that the presenilins could regulate neuronal K+ channel expression; mutations in PS-1/PS-2 would then be expected to result in profound changes in neuronal excitability and contribute to the cognitive decline commonly associated with Alzheimer's Disease.
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Proteínas de la Membrana/genética , Canales de Potasio/fisiología , Regulación hacia Arriba/fisiología , Enfermedad de Alzheimer/metabolismo , Células Cultivadas , Reacciones Cruzadas , ADN Complementario , Humanos , Riñón/citología , Potenciales de la Membrana/fisiología , Proteínas de la Membrana/análisis , Proteínas de la Membrana/inmunología , Mutagénesis/fisiología , Neuronas/química , Neuronas/fisiología , Técnicas de Placa-Clamp , Presenilina-1 , Presenilina-2 , TransfecciónRESUMEN
Kaposi's sarcoma, a sexually dimorphic disease inflicting high mortality in AIDS, remains at present without effective treatment. A recent report (Nature 375:64, 1995) showed that the placental glycoprotein hormone, human chorionic gonadotropin (HCG), and surprisingly its beta subunit, inhibit tumorigenicity and metastasis of Kaposi's sarcoma cells in mice xenografts. The anti-KS efficacy of a commercial HCG was subsequently demonstrated in clinical trials. Experimental data presented herein confirm that commercial HCG preparations (known to be about 25% pure) display significant inhibitory action in a dose-dependent manner. However, pure and biologically active HCG has no effect on Kaposi's sarcoma growth in culture. In fact, incubation of Kaposi's sarcoma cells with either one of four different well characterized preparations of pure HCG dimer or any of its two subunits did not alter cellular proliferation suggesting that a contaminant (or degradation product) may be the active agent. Commercial HCG preparations were subfractionated based on molecular size and each fraction was tested with respect to inhibition of KS cell growth, HCG radioreceptor binding and steroidogenic bioactivity. Results demonstrate that the anti-KS activity resides among low molecular weight components, and not in bona fide (macromolecular) HCG. Our study indicates that HCG activity and anti-KS action are separable. Interestingly, the active components in the crude HCG markedly down-regulate AP-1, a complex of transcription factors of the immediate-early response genes associated with cell growth. We conclude that, as yet unidentified molecules, present in the commercial HCG preparations, are responsible for the growth inhibitory effects presumably via the AP-1 signalling pathway.
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División Celular/efectos de los fármacos , Gonadotropina Coriónica/aislamiento & purificación , Gonadotropina Coriónica/farmacología , Sarcoma de Kaposi/metabolismo , Sarcoma de Kaposi/patología , Factor de Transcripción AP-1/metabolismo , Síndrome de Inmunodeficiencia Adquirida/complicaciones , Fraccionamiento Químico , Cromatografía en Gel , Dimerización , Humanos , Cinética , Peso Molecular , Células Tumorales CultivadasRESUMEN
X-linked immune deficient (Xid) mice fail to produce anti-phosphocholine (PC) antibodies even after immunization with Streptococcus pneumoniae. Consequently, Xid mice are extremely susceptible to infection with S. pneumoniae, PC-specific B cells appear to undergo clonal deletion in Xid mice; however, a new thymus-dependent form of PC, 6-(O-phosphocholine)hydroxyhexanoate (EPC), can rescue PC-specific B cells from the bone marrow presumably by providing T cell help before clonal deletion. Analysis of PC-specific IgG hybridomas from Xid mice revealed utilization of several V-D junctional variants of the VH1 gene segment rearranged to different D and JH gene segments. The majority of Xid anti-PC antibodies exhibit an Asp-->Gly95H replacement at the V-D junction. These Gly95H VH1 variants associate with kappa 1C L chains to produce anti-PC antibodies that: (1) have low relative affinity for PC, (ii) are heteroclitic for nitrophenylphosphocholine and (iii) fall to bind to or provide protection against S. pneumoniae. Single prototypic V-D variants of the T15 idiotype (Asp95H), M603 idiotype (Asn95H) and M167 idiotype (Asp95H-Ala96H) were also induced in Xid mice. The M603-like and M167-like antibodies bound to and protected against S. pneumoniae even though they exhibited Kas for PC which were lower than T15 idiotype+ antibodies. These data demonstrate that small changes in the V-D junctional sequence of the T15 (VH1) heavy chain alter L chain usage and the structure of the PC binding site so that the PC expressed on S. pneumoniae is no longer recognized.
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Anticuerpos Antifosfolípidos/metabolismo , Sitios de Unión de Anticuerpos , Reordenamiento Génico de Cadena Pesada de Linfocito B , Cadenas Pesadas de Inmunoglobulina/metabolismo , Región de Unión de la Inmunoglobulina/metabolismo , Región Variable de Inmunoglobulina/metabolismo , Fosforilcolina/inmunología , Infecciones Neumocócicas/prevención & control , Secuencia de Aminoácidos , Animales , Anticuerpos Antifosfolípidos/genética , Afinidad de Anticuerpos , Adhesión Bacteriana/inmunología , Secuencia de Bases , Caproatos/inmunología , Haptenos/inmunología , Hibridomas/química , Inmunización Pasiva , Alotipos de Inmunoglobulinas/análisis , Cadenas Pesadas de Inmunoglobulina/análisis , Cadenas Pesadas de Inmunoglobulina/genética , Isotipos de Inmunoglobulinas/análisis , Región de Unión de la Inmunoglobulina/análisis , Región de Unión de la Inmunoglobulina/genética , Región Variable de Inmunoglobulina/análisis , Región Variable de Inmunoglobulina/genética , Ratones , Ratones Endogámicos CBA , Ratones Mutantes , Datos de Secuencia Molecular , Fosforilcolina/análogos & derivados , Infecciones Neumocócicas/microbiología , Streptococcus pneumoniae/inmunología , Streptococcus pneumoniae/patogenicidad , VirulenciaRESUMEN
We have cloned and sequenced mouse utrophin cDNA, and successfully expressed full length utrophin (400 kDa) in both muscle and non-muscle cells. The expression of recombinant utrophin is compared with that of its homologue, dystrophin (427 kDa). We demonstrate that recombinant utrophin is targeted into agrin-induced acetylcholine receptor (AChR) clusters, while recombinant dystrophin is evenly distributed along cell membranes in cultured Sol 8 muscle cells. This observation suggests that utrophin and dystrophin may interact with different cytoskeletal proteins. The C-terminal domains are found to be responsible for the association of utrophin with AChR clusters.
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Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Distrofina/metabolismo , Proteínas de la Membrana , Receptores Colinérgicos/metabolismo , Agrina/farmacología , Secuencia de Aminoácidos , Animales , Células CHO , Línea Celular , Membrana Celular/metabolismo , Clonación Molecular , Cricetinae , Proteínas del Citoesqueleto/química , Expresión Génica , Humanos , Ratones , Datos de Secuencia Molecular , Músculos/citología , Músculos/metabolismo , Proteínas Recombinantes/metabolismo , Eliminación de Secuencia , Transfección , UtrofinaRESUMEN
A phenol sulfotransferase from rat liver (EC 2.8.2.9), expressed in Escherichia coli from a single cDNA, was purified as two separable but catalytically active proteins. The proteins appeared to be identical to each other and to the natural liver sulfotransferase by comparison of their amino acid constitution, amino-terminal end group, and interaction with a polyclonal antibody raised against the liver enzyme. Each of the recombinant forms, alpha and beta, catalyzed the sulfuryl group transfer from 4-nitrophenylsulfate to an acceptor phenol, a reaction in which 3'-phospho-adenosine 5'-phosphate (PAP) is a necessary intermediate. Only form beta, however, catalyzes the physiological transfer of a sulfuryl group from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to the free phenol. Evidence is presented that sulfotransferase alpha, but not beta, has 1 mol of PAP tightly bound per enzyme dimer. The ability to utilize PAPS as a sulfate donor could be altered: form alpha could be treated and purified as form beta to acquire the ability to use PAPS, whereas form beta was treated by extended incubation with PAP, lost its ability to use PAPS, and was purified as form alpha.
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Arilsulfotransferasa/metabolismo , Hígado/enzimología , Animales , Dicroismo Circular , ADN Complementario/análisis , Escherichia coli/metabolismo , Cinética , Nitrofenoles/metabolismo , Fosfoadenosina Fosfosulfato/metabolismo , RatasRESUMEN
Development of Kaposi's sarcoma (KS) after glucocorticoid therapy has been observed in a variety of clinical states including human immunodeficiency virus-1 infection and recent in vitro studies provided evidence for a direct stimulation effect of glucocorticoid hormones on KS cell proliferation. The importance of glucocorticoids in KS pathogenesis is further highlighted by the finding that glucocorticoids synergize with cytokines to promote acquired immune deficiency syndrome (AIDS)-associated KS (AIDS-KS) growth. Furthermore, cytokine effects were abrogated by the glucocorticoid antagonist RU-486. As glucocorticoid action is mediated through activation of their intracellular cognate receptors, we hypothesized that enhanced responsiveness of AIDS-KS cells to glucocorticoids may be due to elevated glucocorticoid receptor (GR) content. Indeed, high expression of GRs in AIDS-KS tumor biopsies was detected both at the level of mRNA and protein. Quantitative measurements of GRs in these specimens by a sensitive immunoassay showed that GR content was significantly elevated in the tumor tissue (4663 fmol/mg protein) compared with the uninvolved skin of the same patients (2777 fmol/mg protein), both of which were markedly above the normal skin of healthy donors (893 fmol/mg protein). Immunocytochemical analysis confirmed the presence of GRs in the cytoplasm and the nucleus of KS cells. Interestingly, four major KS cytokines, namely interleukin-1beta, interleukin-6, tumor necrosis factor-alpha, and oncostatin M, all of which are known autocrine growth factors for AIDS-KS cells, significantly increased the expression of functional GRs in cultured AIDS-KS cells. The latter result may explain, at least in part, the synergistic effect of glucocorticoid and oncostatin M on AIDS-KS cell proliferation. Thus, the high levels of GR expression in AIDS-KS and the up-regulation of GRs by KS-growth-promoting factors may confer enhanced and sustained sensitivity to the stimulatory effects of glucocorticoids. The data presented also provide molecular bases for therapeutic interventions targeting GRs in this disease.