RESUMEN
The tumor suppressor von Hippel-Lindau, pVHL, is a multifaceted protein. One function is to dock to the hypoxia-inducible transcription factor (HIF) and recruit a larger protein complex that destabilizes HIF via ubiquitination, preventing angiogenesis and tumor development. pVHL also binds to the tumor suppressor p53 to activate specific p53 target genes. The oncogene Mdm2 impairs the formation of the p53-pVHL complex and activation of downstream genes by conjugating nedd8 to pVHL. While Mdm2 can impact p53 and pVHL, how pVHL may impact Mdm2 is unclear. Like p53 somatic mutations, point mutations are evident in pVHL that are common in renal clear cell carcinomas (RCC). In patients with RCC, Mdm2 levels are elevated, and we examined whether there was a relationship between Mdm2 and pVHL. TCGA and DepMap analysis revealed that mdm2 gene expression was elevated in RCC with vhl point mutations or copy number loss. In pVHL reconstituted or deleted isogenetically match RCC or MEF cell lines, Mdm2 was decreased in the presence of pVHL. Furthermore, through analysis using genetic and pharmacological approaches, we show that pVHL represses Mdm2 gene expression by blocking the MAPK-Ets signaling pathway and blocks Akt-mediated phosphorylation and stabilization of Mdm2. Mdm2 inhibition results in an increase in the p53-p21 pathway to impede cell growth. This finding shows how pVHL can indirectly impact the function of Mdm2 by regulating signaling pathways to restrict cell growth.
Asunto(s)
Carcinoma de Células Renales , Neoplasias Renales , Proteínas Proto-Oncogénicas c-mdm2 , Transducción de Señal , Proteína p53 Supresora de Tumor , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/genética , Humanos , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/patología , Proteína p53 Supresora de Tumor/metabolismo , Proteína p53 Supresora de Tumor/genética , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión GénicaRESUMEN
The development of reliable methods for producing functional endothelial cells (ECs) is crucial for progress in vascular biology and regenerative medicine. In this study, we present a streamlined and efficient methodology for the differentiation of human induced pluripotent stem cells (iPSCs) into induced ECs (iECs) that maintain the ability to undergo vasculogenesis in vitro and in vivo using a doxycycline-inducible system for the transient expression of the ETV2 transcription factor. This approach mitigates the limitations of direct transfection methods, such as mRNA-mediated differentiation, by simplifying the protocol and enhancing reproducibility across different stem cell lines. We detail the generation of iPSCs engineered for doxycycline-induced ETV2 expression and their subsequent differentiation into iECs, achieving over 90% efficiency within four days. Through both in vitro and in vivo assays, the functionality and phenotypic stability of the derived iECs were rigorously validated. Notably, these cells exhibit key endothelial markers and capabilities, including the formation of vascular networks in a microphysiological platform in vitro and in a subcutaneous mouse model. Furthermore, our results reveal a close transcriptional and proteomic alignment between the iECs generated via our method and primary ECs, confirming the biological relevance of the differentiated cells. The high efficiency and effectiveness of our induction methodology pave the way for broader application and accessibility of iPSC-derived ECs in scientific research, offering a valuable tool for investigating endothelial biology and for the development of EC-based therapies.
RESUMEN
The enzyme 2-methylerythritol 2,4-cyclodiphosphate synthase, IspF, is essential for the biosynthesis of isoprenoids in most bacteria, some eukaryotic parasites, and the plastids of plant cells. The development of inhibitors that target IspF may lead to novel classes of anti-infective agents or herbicides. Enantiomers of tryptophan hydroxamate were synthesized and evaluated for binding to Burkholderia pseudomallei (Bp) IspF. The L-isomer possessed the highest potency, binding BpIspF with a KD of 36 µM and inhibited BpIspF activity 55% at 120 µM. The high-resolution crystal structure of the L-tryptophan hydroxamate (3)/BpIspF complex revealed a non-traditional mode of hydroxamate binding where the ligand interacts with the active site zinc ion through the primary amine. In addition, two hydrogen bonds are formed with active site groups, and the indole group is buried within the hydrophobic pocket composed of side chains from the 60 s/70 s loop. Along with the co-crystal structure, STD NMR studies suggest the methylene group and indole ring are potential positions for optimization to enhance binding potency.
Asunto(s)
Proteínas Bacterianas/antagonistas & inhibidores , Burkholderia pseudomallei/enzimología , Inhibidores Enzimáticos/farmacología , Triptófano/análogos & derivados , Proteínas Bacterianas/metabolismo , Sitios de Unión/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Modelos Moleculares , Estructura Molecular , Relación Estructura-Actividad , Triptófano/síntesis química , Triptófano/química , Triptófano/farmacologíaRESUMEN
The measurement of trace gases has increasingly become a key technique in healthcare and other medical applications. Quartz-enhanced photoacoustic spectroscopy (QEPAS) is a suitable method that can provide the required characteristics in such applications for a comparatively low cost and small size. The quantitative detection and a low detection limit are also required by applications. In this paper, we present new results on sensing biomedically relevant gases using the on-beam QEPAS technique with some newly developed tunable high-power single-mode laser diodes based on GaSb material. The data processing and detection limit determination are done by a field programmable gate array device, as well as an automatic measurement of the resonance frequency.
RESUMEN
The wavelength, λ, range of 1.8 µm≤λ≤3.5 µm contains strong spectral absorption lines of many gases used in health, industry, safety, and medicine and whose sensitive and quantitative detection is desirable. However, the performance of InP diode lasers markedly deteriorates beyond λâ¼2 µm. In this paper we present new results on developing tunable high power single mode laser diodes based on the GaSb material system with emission in the wavelength range of 1.8 µm≤λ≤2.2 µm.
RESUMEN
Immunotherapy using checkpoint-blocking antibodies against targets such as CTLA-4 and PD-1 can cure melanoma and non-small cell lung cancer in a subset of patients. The presence of CD8 T cells in the tumor correlates with improved survival. We show that immuno-positron emission tomography (immuno-PET) can visualize tumors by detecting infiltrating lymphocytes and, through longitudinal observation of individual animals, distinguish responding tumors from those that do not respond to therapy. We used 89Zr-labeled PEGylated single-domain antibody fragments (VHHs) specific for CD8 to track the presence of intratumoral CD8+ T cells in the immunotherapy-susceptible B16 melanoma model in response to checkpoint blockade. A 89Zr-labeled PEGylated anti-CD8 VHH detected thymus and secondary lymphoid structures as well as intratumoral CD8 T cells. Animals that responded to CTLA-4 therapy showed a homogeneous distribution of the anti-CD8 PET signal throughout the tumor, whereas more heterogeneous infiltration of CD8 T cells correlated with faster tumor growth and worse responses. To support the validity of these observations, we used two different transplantable breast cancer models, yielding results that conformed with predictions based on the antimelanoma response. It may thus be possible to use immuno-PET and monitor antitumor immune responses as a prognostic tool to predict patient responses to checkpoint therapies.
Asunto(s)
Linfocitos T CD8-positivos/fisiología , Antígeno CTLA-4/antagonistas & inhibidores , Neoplasias Mamarias Experimentales/terapia , Animales , Antígeno CTLA-4/fisiología , Femenino , Fragmentos de Inmunoglobulinas/inmunología , Fragmentos de Inmunoglobulinas/uso terapéutico , Inmunoterapia/métodos , Neoplasias Mamarias Experimentales/diagnóstico por imagen , Neoplasias Mamarias Experimentales/inmunología , Ratones , Trasplante de Neoplasias , Tomografía de Emisión de Positrones/métodos , Resultado del TratamientoRESUMEN
We report a unidirectional frequency dissemination scheme for high-fidelity optical carriers deployable over telecommunication networks. For the first time, a 10 Gb/s Binary Phase Shift Keying (BPSK) signal from an ultra-narrow linewidth laser was transmitted through a field-installed optical fibre with round-trip length of 124 km between Cork City and town of Clonakilty, without inline optical amplification. At the receiver, using coherent communication techniques and optical injection-locking the carrier was recovered with noise suppression. The beat signal between the original carrier at the transmitter and recovered carrier at the receiver shows a linewidth of 2.8 kHz. Long term stability measurements revealed fractional instabilities (True Allan deviation) of 3.3 × 10(-14) for 1 s averaging time, prior to phase noise cancellation.
Asunto(s)
Antiinflamatorios no Esteroideos/administración & dosificación , Inmunoglobulina G/administración & dosificación , Desplazamiento del Disco Intervertebral/tratamiento farmacológico , Disco Intervertebral/efectos de los fármacos , Vértebras Lumbares/efectos de los fármacos , Receptores del Factor de Necrosis Tumoral/administración & dosificación , Femenino , Humanos , MasculinoRESUMEN
STUDY DESIGN: Multicenter, randomized, double-blind, placebo-controlled trial. OBJECTIVE: To examine the safety and efficacy of three different doses of the tumor necrosis factor alpha (TNF-α) inhibitor etanercept versus placebo for the treatment of symptomatic lumbar disc herniation (LDH). SUMMARY OF BACKGROUND DATA: TNF-α is considered to be a major cause of radicular leg pain associated with symptomatic LDH. Systemic administration of TNF-α inhibitors for sciatica has indicated a trend toward efficacy. METHODS: Forty-nine subjects aged between 18 and 70 years, with persistent lumbosacral radicular pain secondary to LDH, and an average leg pain intensity of 5/10 or more were randomized to 1 of 4 groups: 0.5-mg, 2.5-mg, 12.5-mg etanercept, or placebo. Subjects received 2 transforaminal epidural injections, 2 weeks apart, and were assessed for efficacy up to 26 weeks after the second injection. The primary outcome measure was the change in mean daily worst leg pain (WLP). Secondary outcomes included average leg pain, worst back pain, average back pain, in-clinic pain, Oswestry Disability Index, patient global impression of change, and tolerability. RESULTS: Forty-three of the 49 randomized patients completed the study. Patients receiving 0.5-mg etanercept showed a clinically and statistically significant (P< 0.1) reduction in mean daily WLP compared with the placebo cohort from 2 to 26 weeks for both the per protocol population (-5.13 vs. -1.95; P= 0.066) and the intention-to-treat population (-4.40 vs. -1.84; P= 0.058). Fifty percent of these subjects reported a 100% reduction in WLP 4 weeks post-treatment compared with 0% of subjects in the placebo cohort. Improvements in all secondary outcomes were also observed in the 0.5-mg etanercept cohort. The overall incidence of adverse events was similar in placebo and all etanercept cohorts. CONCLUSION: Two transforaminal injections of etanercept provided clinically significant reductions in mean daily WLP and worst back pain compared with placebo for subjects with symptomatic LDH. Epidural etanercept may offer patients with sciatica a safe and effective nonoperative treatment.
Asunto(s)
Antiinflamatorios no Esteroideos/administración & dosificación , Inmunoglobulina G/administración & dosificación , Desplazamiento del Disco Intervertebral/tratamiento farmacológico , Disco Intervertebral/efectos de los fármacos , Vértebras Lumbares/efectos de los fármacos , Receptores del Factor de Necrosis Tumoral/administración & dosificación , Adulto , Antiinflamatorios no Esteroideos/efectos adversos , Australia , Dolor de Espalda/diagnóstico , Dolor de Espalda/tratamiento farmacológico , Evaluación de la Discapacidad , Método Doble Ciego , Esquema de Medicación , Etanercept , Femenino , Humanos , Inmunoglobulina G/efectos adversos , Inyecciones Espinales , Disco Intervertebral/inmunología , Disco Intervertebral/fisiopatología , Desplazamiento del Disco Intervertebral/diagnóstico , Desplazamiento del Disco Intervertebral/inmunología , Desplazamiento del Disco Intervertebral/fisiopatología , Vértebras Lumbares/inmunología , Vértebras Lumbares/fisiopatología , Masculino , Persona de Mediana Edad , Dimensión del Dolor , Factores de Tiempo , Resultado del Tratamiento , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Karpicke and Blunt (Reports, 11 February 2011, p. 772) reported that retrieval practice produces greater gains in learning than elaborative studying with concept mapping and concluded that this strategy is a powerful way to promote meaningful learning of complex concepts commonly found in science education. We question their findings on methodological and epistemological grounds.
Asunto(s)
Disciplinas de las Ciencias Biológicas/educación , Aprendizaje , Memoria , HumanosRESUMEN
We present a phase locking scheme that enables the demonstration of a practical dual pump degenerate phase sensitive amplifier for 10 Gbit/s non-return to zero amplitude shift keying signals. The scheme makes use of cascaded Mach Zehnder modulators for creating the pump frequencies as well as of injection locking for extracting the signal carrier and synchronizing the local lasers. An in depth optimization study has been performed, based on measured error rate performance, and the main degradation factors have been identified.
Asunto(s)
Amplificadores Electrónicos , Óptica y Fotónica/instrumentación , Óptica y Fotónica/métodos , Procesamiento de Señales Asistido por Computador/instrumentación , Artefactos , Diseño de Equipo , Rayos LáserRESUMEN
Cost effective lasers meeting the linewidth requirements for coherent communication systems are a key element in reducing the overall cost of future coherent systems. We report on monolithic devices with linewidths as low as 138 kHz which operate in a narrow linewidth, single wavelength mode with high sidemode suppression ratio over a wide temperature tuning range of -10 °C < T < 110 °C. A linewidth variation of only 23 kHz was measured at a constant emitted power of 4 mW as the device temperature is varied in the range 0 °C < T < 85 °C.
RESUMEN
A "black-box" phase sensitive amplifier is presented achieving simultaneous suppression of deterministic phase distortion on two independent 42.66 Gbit/s DPSK modulated signal wavelengths.
RESUMEN
Return-to-Zero (RZ) and Non-Return-to-Zero (NRZ) Differential Phase Shift Keyed (DPSK) systems require cheap and optimal transmitters for widespread implementation. The authors report on a gain switched Discrete Mode (DM) laser that can be employed as a cost efficient transmitter in a 10.7 Gb/s RZ DPSK system and compare its performance to that of a gain switched Distributed Feed-Back (DFB) laser. Experimental results show that the gain switched DM laser readily provides error free performance and a receiver sensitivity of -33.1 dBm in the 10.7 Gbit/s RZ DPSK system. The standard DFB laser on the other hand displays an error floor at 10(-1) in the same RZ DPSK system. The difference in performance, between the two types of gain switched transmitters, is analysed by investigating their linewidths. We also demonstrate, for the first time, the generation of a highly coherent gain switched pulse train which displays a spectral comb of approximately 13 sidebands spaced by the 10.7 GHz modulation frequency. The filtered side-bands are then employed as narrow linewidth Continuous Wave (CW) sources in a 10.7 Gb/s NRZ DPSK system.
RESUMEN
A novel single frequency stabilized Fabry-Perot (SFP) laser diode with an emission wavelength of lambda = 1590 nm for H2S gas sensing is reported. Sculpting of the multi-mode spectral distribution of a FP laser to achieve single frequency emission is carried out using post growth photolitographic processing of the device. The resulting longitudinal-mode controlled FP laser has a stabilized single frequency emission with a side mode suppression ratio (SMSR) of 40 dB. The application of this device to spectroscopic based H2S sensing is demonstrated by targeting absorption lines in the wavelength range 1588 < or = lambda < or = 1591 nm. Using wavelength modulation spectroscopy (WMS), a low detection limit of 120 ppm x m x Hz(-1/2) was estimated while targeting the absorption line at 1590.08 nm. These initial results demonstrate the potential of the stabilized FP laser diode at this wavelength as a tunable, single frequency source for spectroscopic based gas sensing.