OBJECTIVE—We tested the hypothesis that p47phox associates with the actin cytoskeleton, enabling site-directed activation of NAD(P)H oxidase, and assessed whether these actions influence reactive ...oxygen species (ROS) generation and signaling by angiotensin II (Ang II) in vascular smooth muscle cells (VSMCs) from human resistance and coronary arteries.
METHODS AND RESULTS—Electroporation of anti-p47phox antibody into VSMCs abrogated Ang II–mediated generation, establishing the requirement for p47phox in this response. Immunfluorescence confocal microscopy demonstrated a cytosolic distribution of p47phox in basal conditions. After Ang II stimulation, p47phox rearranged in a linear fashion, colocalizing with F-actin. Co-immunoprecipitation studies confirmed an association between p47phox and actin and demonstrated an interaction with the actin-binding protein cortactin. Cytoskeletal disruption with cytochalasin prevented p47phox:actin interaction and attenuated ROS formation and p38MAP kinase and Akt phosphorylation by Ang II. Intracellular ROS generation in response to LY83583 ( generator) or exogenous H2O2 and Ang II–induced ERK1/2 activation were unaltered by cytochalasin.
CONCLUSIONS—The p47phox:actin interaction, through cortactin, plays an important role in Ang II–mediated site-directed assembly of functionally active NAD(P)H oxidase, ROS generation, and activation of redox-sensitive p38MAP kinase and Akt, but not ERK1/2. These findings demonstrate the importance of an intact actin–cytoskeleton in NAD(P)H oxidase regulation and redox signaling by Ang II in human VSMCs.
Activation of cyclic GMP-AMP (cGAMP) synthase (cGAS) plays a critical role in antiviral responses to many DNA viruses. Sensing of cytosolic DNA by cGAS results in synthesis of the endogenous second ...messenger cGAMP that activates stimulator of interferon genes (STING) in infected cells. Critically, cGAMP can also propagate antiviral responses to uninfected cells through intercellular transfer, although the modalities of this transfer between epithelial and immune cells remain poorly defined. We demonstrate here that cGAMP-producing epithelial cells can transactivate STING in cocultured macrophages through direct cGAMP transfer. cGAMP transfer was reliant upon connexin expression by epithelial cells and pharmacological inhibition of connexins blunted
-dependent transactivation of the macrophage compartment. Macrophage transactivation by cGAMP contributed to a positive-feedback loop amplifying antiviral responses, significantly protecting uninfected epithelial cells against viral infection. Collectively, our findings constitute the first direct evidence of a connexin-dependent cGAMP transfer to macrophages by epithelial cells, to amplify antiviral responses.
Recent studies suggest that extracellular cGAMP can be taken up by macrophages to engage STING through several mechanisms. Our work demonstrates that connexin-dependent communication between epithelial cells and macrophages plays a significant role in the amplification of antiviral responses mediated by cGAMP and suggests that pharmacological strategies aimed at modulating connexins may have therapeutic applications to control antiviral responses in humans.
The library described here is a collection of phages with six degenerate codons in gene VIII, specifying amino acids 12, 13, 15-17 and 19 of the major coat protein. The randomized positions are ...surface exposed in the wild-type protein and thus might be expected to tolerate a great diversity of side chains without compromising phage viability. In agreement with this supposition, the new library showed great diversity of amino acids at the randomized positions and diversity did not diminish noticeably during repeated subculture. Despite their diversity, however, the randomized positions should be strongly constrained conformationally because they lie in an extended alpha-helical portion of the protein, stabilized by numerous inter- and intra-subunit contacts--a presupposition corroborated by circular dichroism spectroscopy of many library members. To reflect this conformational homogeneity and the fact that random amino acids subtend a major fraction of the surface `landscape' of the particle, we call the new construct an alpha landscape library. It can be used as a source of alpha-helical ligands and substitute antibodies.
ABSTRACT
We report the discovery of two mini-Neptunes in near 2:1 resonance orbits (P = 7.610303 d for HIP 113103 b and P = 14.245651 d for HIP 113103 c) around the adolescent K-star HIP 113103 (TIC ...121490076). The planet system was first identified from the TESS mission, and was confirmed via additional photometric and spectroscopic observations, including a ∼17.5 h observation for the transits of both planets using ESA CHEOPS. We place ≤4.5 min and ≤2.5 min limits on the absence of transit timing variations over the 3 yr photometric baseline, allowing further constraints on the orbital eccentricities of the system beyond that available from the photometric transit duration alone. With a planetary radius of Rp = $1.829_{-0.067}^{+0.096}$ R⊕, HIP 113103 b resides within the radius gap, and this might provide invaluable information on the formation disparities between super-Earths and mini-Neptunes. Given the larger radius Rp = $2.40_{-0.08}^{+0.10}$ R⊕ for HIP 113103 c, and close proximity of both planets to HIP 113103, it is likely that HIP 113103 b might have lost (or is still losing) its primordial atmosphere. We therefore present simulated atmospheric transmission spectra of both planets using JWST, HST, and Twinkle. It demonstrates a potential metallicity difference (due to differences in their evolution) would be a challenge to detect if the atmospheres are in chemical equilibrium. As one of the brightest multi sub-Neptune planet systems suitable for atmosphere follow up, HIP 113103 b and HIP 113103 c could provide insight on planetary evolution for the sub-Neptune K-star population.
MicroRNA-155 (miR-155) is highly expressed in many cancers such as B cell lymphomas and myeloid leukemia and inflammatory disorders such as rheumatoid arthritis, atopic dermatitis, and multiple ...sclerosis. The role of miR-155 as both a promoter of inflammation and an oncogenic agent provides a clear need for miR-155 itself to be stringently regulated. We therefore investigated the transcriptional regulation of miR-155 in response to the respective pro- and anti-inflammatory mediators LPS and IL-10. Bioinformatic analysis revealed Ets binding sites on the miR-155 promoter, and we found that Ets2 is critical for miR-155 induction by LPS. Truncation and mutational analysis of the miR-155 promoter confirmed the role of the Ets2 binding site proximal to the transcription start site for LPS responsiveness. We observed increased binding of Ets2 to the miR-155 promoter and Ets2 deficient mice displayed decreased induction of miR-155 in response to LPS. IL-10 inhibited the induction of Ets2 mRNA and protein by LPS, thereby decreasing Ets2 function on the pri-155 promoter. We have thus identified Ets2 as a key novel regulator in both the positive and negative control of miR-155 in the inflammatory response.
miR-155 is strongly induced by LPS, a response inhibited by IL-10.
The Ets2 transcription factor is required for induction of miR-155 by LPS. IL-10 can subsequently decrease miR-155 via suppression of Ets2.
Ets2 is an important transcription factor for regulation of miR-155.
This study reports a detailed mechanism of induction of miR-155 and provides a new means of inhibition for IL-10 via suppression of Ets2.
The herpes simplex virus type 1 (HSV-1) latency-associated transcript (LAT) promoter 1 (LP1) is an inducible and cell type-specific promoter involved in regulating the production of an 8.3-kb primary ...LAT transcript during acute and latent infection of peripheral sensory neurons and during subsequent virus reactivation. A number of cis-acting regulatory elements have been identified in LP1, including two cyclic-AMP (cAMP) response element (CRE)-like sequences, designated CRE-1 and CRE-2. CRE-1 has previously been shown to confer cAMP responsiveness to LP1 and to regulate reactivation of HSV-1 from latency in vivo. A role for CRE-2 in modulating inducible activity is not yet as clear; however, it has been shown to support basal expression in neuronal cells in vitro. Electrophoretic mobility shift (EMS) analyses demonstrate that the LP1 CRE-like elements interact with distinct subsets of neuronal ATF/CREB and Jun/Fos proteins including CREB-1, CREB-2, ATF-1, and JunD. The factor-binding properties of each LP1 CRE element distinguish them from each other and from a highly related canonical CRE binding site and the TPA response element (TRE). LP1 CRE-1 shares binding characteristics of both a canonical CRE and a TRE. LP1 CRE-2 is more unusual in that it shares more features of a canonical CRE site than a TRE with two notable exceptions: it does not bind CREB-1 very well and it binds CREB-2 better than the canonical CRE. Interestingly, a substantial proportion of the C1300 neuroblastoma factors that bind to CRE-1 and CRE-2 have been shown to be immunologically related to JunD, suggesting that the AP-1 family of transcription factors may be important in regulating CRE-dependent LP1 transcriptional activity. In addition, we have demonstrated the two HSV-1 LP1 CRE sites to be unique with respect to their ability to bind neuronal AP1-related factors that are regulated by cAMP. These studies suggest that both factor binding and activation of bound factors may be involved in cAMP regulation of HSV-1 LP1 through the CRE elements, and indicate the necessity of investigating the expression and posttranslational modification of a variety of ATF/CREB and AP-1 factors during latency and reactivation.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Colorectal cancer (CRC) deaths occur when patients do not receive screening or have inadequate follow-up of abnormal results or when the screening test fails. We have few data on the contribution of ...each to CRC-associated deaths or factors associated with these events.
We performed a retrospective cohort study of patients in the Kaiser Permanente Northern and Southern California systems (55–90 years old) who died of CRC from 2006 through 2012 and had ≥5 years of enrollment before diagnosis. We compared data from patients with those from a matched cohort of cancer-free patients in the same system. Receipt, results, indications, and follow-up of CRC tests in the 10-year period before diagnosis were obtained from electronic databases and chart audits.
Of 1750 CRC deaths, 75.9% (n = 1328) occurred in patients who were not up to date in screening and 24.1% (n = 422) occurred in patients who were up to date. Failure to screen was associated with fewer visits to primary care physicians. Of 3486 cancer-free patients, 44.6% were up to date in their screening. Patients who were up to date in their screening had a lower risk of CRC death (odds ratio, 0.38; 95% confidence interval, 0.33–0.44). Failure to screen, or failure to screen at appropriate intervals, occurred in a 67.8% of patients who died of CRC vs 53.2% of cancer-free patients; failure to follow-up on abnormal results occurred in 8.1% of patients who died of CRC vs 2.2% of cancer-free patients. CRC death was associated with higher odds of failure to screen or failure to screen at appropriate intervals (odds ratio, 2.40; 95% confidence interval, 2.07–2.77) and failure to follow-up on abnormal results (odds ratio, 7.26; 95% confidence interval, 5.26–10.03).
Being up to date on screening substantially decreases the risk of CRC death. In 2 health care systems with high rates of screening, most people who died of CRC had failures in the screening process that could be rectified, such as failure to follow-up on abnormal findings; these significantly increased the risk for CRC death.
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Positron emission tomography and single photon emission computed tomography scanning have 87-94% sensitivity and 80-100% specificity to differentiate patients with Parkinson's disease (PD) from ...control subjects and patients with essential (ET) or atypical tremor. More than 10% of patients diagnosed as early PD can have scans without evidence of dopaminergic deficiency (SWEDDs). This study investigated whether smell tests can help identify possible cases with SWEDDs.
The 40 item University of Pennsylvania Smell Test (UPSIT) was used to evaluate the sense of smell in 21 SWEDDs patients. Twenty-six ET patients, 16 patients with a diagnosis of idiopathic adult onset dystonia (D), 191 non-demented PD patients and 136 control subjects were also tested. Multiple regression analyses were used to compare the mean UPSIT score in the SWEDDs group with the other four groups (ET, D, PD and controls) after adjusting for the effects of relevant covariates.
The mean UPSIT score for the SWEDDs group was greater than in the PD group (p<0.001) and not different from the mean UPSIT in the control (p = 0.7), ET (p = 0.4) or D (p = 0.9) groups. Smell tests indicated a high probability of PD in only 23.8% of SWEDDs as opposed to 85.3% of PD patients.
In a patient with suspected PD, a high PD probability on smell testing favours the diagnosis of PD, and a low PD probability strengthens the indication for dopamine transporter imaging.
The aims of this retrospective cohort study were to 1) identify new alignment risk factors for proximal junctional kyphosis (PJK) in adult spinal deformity (ASD) patients with lower thoracic upper ...instrumented vertebra (UIV) and 2) determine the effect of junctional tethers on PJK and UIV alignment.
We analyzed consecutive ASD patients who underwent posterior instrumented fusion with lower thoracic UIV (T9-T11). Posteriorly anchored junctional tethers were used more recently for ligamentous augmentation to prevent PJK. In addition to regional and global parameters, upper segmental lumbar lordosis (ULL) versus lower segmental lumbar lordosis and UIV angle (measured from UIV inferior endplate to horizontal) were assessed. Primary outcome of PJK was defined as proximal junctional angle >10° and >10° greater than the corresponding preoperative measurement. Univariable and multivariable analyses were performed.
The study cohort comprised 120 ASD patients (mean age, 67 years) with minimum 1-year follow-up. Preoperative ULL (P = 0.034) and UIV angle (P = 0.026) were associated with PJK. No independent preoperative alignment risk factors of PJK were identified in multivariable analysis. Tether use was protective against PJK (odds ratio, 0.063 0.016–0.247; P < 0.001). PJK in tethered patients was more common with greater postoperative ULL (P = 0.047) and UIV angle (P = 0.026).
Junctional tethers significantly reduced PJK in ASD patients with lower thoracic UIV. In tethered patients, PJK was more common with greater postoperative lordosis of the upper lumbar spine and greater UIV angle. This finding suggests potential benefit of tethers to mitigate effects of segmental lumbar and focal UIV malalignment that may occur after deformity surgery.
•No preoperative alignment risk factors of PJK were identified in multivariable analysis.•Lower thoracic junctional tethers have a protective effect against PJK.•PJK in tethered patients was more common with greater postoperative ULL and UIV angle.•Tethers may mitigate effects of segmental and focal malalignment or overcorrection.
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