Differential expression of microRNAs in mouse pain models

<p>Abstract</p> <p>Background</p> <p>MicroRNAs (miRNAs) are short non-coding RNAs that inhibit translation of target genes by binding to their mRNAs. The expression of numerous brain-specific miRNAs with a high degree of temporal and spatial specificity suggests that miRNAs play an important role in gene regulation in health and disease. Here we investigate the time course gene expression profile of miR-1, -16, and -206 in mouse dorsal root ganglion (DRG), and spinal cord dorsal horn under inflammatory and neuropathic pain conditions as well as following acute noxious stimulation.</p> <p>Results</p> <p>Quantitative real-time polymerase chain reaction analyses showed that the mature form of miR-1, -16 and -206, is expressed in DRG and the dorsal horn of the spinal cord. Moreover, CFA-induced inflammation significantly reduced miRs-1 and -16 expression in DRG whereas miR-206 was downregulated in a time dependent manner. Conversely, in the spinal dorsal horn all three miRNAs monitored were upregulated. After sciatic nerve partial ligation, miR-1 and -206 were downregulated in DRG with no change in the spinal dorsal horn. On the other hand, axotomy increases the relative expression of miR-1, -16, and 206 in a time-dependent fashion while in the dorsal horn there was a significant downregulation of miR-1. Acute noxious stimulation with capsaicin also increased the expression of miR-1 and -16 in DRG cells but, on the other hand, in the spinal dorsal horn only a high dose of capsaicin was able to downregulate miR-206 expression.</p> <p>Conclusions</p> <p>Our results indicate that miRNAs may participate in the regulatory mechanisms of genes associated with the pathophysiology of chronic pain as well as the nociceptive processing following acute noxious stimulation. We found substantial evidence that miRNAs are differentially regulated in DRG and the dorsal horn of the spinal cord under different pain states. Therefore, miRNA expression in the nociceptive system shows not only temporal and spatial specificity but is also stimulus-dependent.</p

Expression and cellular localization of microRNA-29b and RAX, an activator of the RNA-dependent protein kinase (PKR), in the retina of streptozotocin-induced diabetic rats

Purpose: The apoptosis of retinal neurons plays a critical role in the pathogenesis of diabetic retinopathy (DR), but the molecular mechanisms underlying this phenomenon remain unclear. The purpose of this study was to investigate the cellular localization and the expression of microRNA-29b (miR-29b) and its potential target PKR associated protein X (RAX), an activator of the pro-apoptotic RNA-dependent protein kinase (PKR) signaling pathway, in the retina of normal and diabetic rats. Methods: Retinas were obtained from normal and diabetic rats within 35 days after streptozotocin (STZ) injection. In silico analysis indicated that RAX is a potential target of miR-29b. The cellular localization of miR-29b and RAX was assessed by in situ hybridization and immunofluorescence, respectively. The expression levels of miR-29b and RAX mRNA were evaluated by quantitative reverse transcription PCR (qRT-PCR), and the expression of RAX protein was evaluated by western blot. A luciferase reporter assay and inhibition of endogenous RAX were performed to confirm whether RAX is a direct target of miR-29b as predicted by the in silico analysis. Results: We found that miR-29b and RAX are localized in the retinal ganglion cells (RGCs) and the cells of the inner nuclear layer (INL) of the retinas from normal and diabetic rats. Thus, the expression of miR-29b and RAX, as assessed in the retina by quantitative RT-PCR, reflects their expression in the RGCs and the cells of the INL. We also revealed that RAX protein is upregulated (more than twofold) at 3, 6, 16, and 22 days and downregulated (70%) at 35 days, whereas miR-29b is upregulated (more than threefold) at 28 and 35 days after STZ injection. We did not confirm the computational prediction that RAX is a direct target of miR-29b. Conclusions: Our results suggest that RAX expression may be indirectly regulated by miR-29b, and the upregulation of this miRNA at the early stage of STZ-induced diabetes may have a protective effect against the apoptosis of RGCs and cells of the INL by the pro-apoptotic RNA-dependent protein kinase (PKR) signaling pathway.FAPESP[08/58325-4]FAPESP[08/50294-2]Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Detection of <it>P. aeruginosa </it> harboring <it>bla </it>_CTX-M-2, <it>bla </it>_GES-1 and <it>bla </it>_GES-5,<it>bla </it>_IMP-1 and <it>bla </it>_SPM-1 causing infections in Brazilian tertiary-care hospital

<p>Abstract</p> <p>Background</p> <p>Nosocomial infections caused by <it>Pseudomonas aeruginosa</it> presenting resistance to beta-lactam drugs are one of the most challenging targets for antimicrobial therapy, leading to substantial increase in mortality rates in hospitals worldwide. In this context, <it>P. aeruginosa</it> harboring acquired mechanisms of resistance, such as production of metallo-beta-lactamase (MBLs) and extended-spectrum beta-lactamases (ESBLs) have the highest clinical impact. Hence, this study was designed to investigate the presence of genes codifying for MBLs and ESBLs among carbapenem resistant <it>P. aeruginosa</it> isolated in a Brazilian 720-bed teaching tertiary care hospital.</p> <p>Methods</p> <p>Fifty-six carbapenem-resistant <it>P. aeruginosa</it> strains were evaluated for the presence of MBL and ESBL genes. Strains presenting MBL and/or ESBL genes were submitted to pulsed-field gel electrophoresis for genetic similarity evaluation.</p> <p>Results</p> <p>Despite the carbapenem resistance, genes for MBLs (<it>bla</it>_SPM-1 or <it>bla</it>_IMP-1) were detected in only 26.7% of isolates. Genes encoding ESBLs were detected in 23.2% of isolates. The <it>bla</it>_CTX-M-2 was the most prevalent ESBL gene (19.6%), followed by <it>bla</it>_GES-1 and <it>bla</it>_GES-5 detected in one isolate each. In all isolates presenting MBL phenotype by double-disc synergy test (DDST), the <it>bla</it>_SPM-1 or <it>bla</it>_IMP-1 genes were detected. In addition, <it>bla</it>_IMP-1 was also detected in three isolates which did not display any MBL phenotype. These isolates also presented the <it>bla</it>_CTX-M-2 gene. The co-existence of <it>bla</it>_CTX-M-2 with <it>bla</it>_IMP-1 is presently reported for the first time, as like as co-existence of <it>bla</it>_GES-1 with <it>bla</it>_IMP-1.</p> <p>Conclusions</p> <p>In this study MBLs production was not the major mechanism of resistance to carbapenems, suggesting the occurrence of multidrug efflux pumps, reduction in porin channels and production of other beta-lactamases. The detection of <it>bla</it>_CTX-M-2,<it>bla</it>_GES-1 and <it>bla</it>_GES-5 reflects the recent emergence of ESBLs among antimicrobial resistant <it>P. aeruginosa</it> and the extraordinary ability presented by this pathogen to acquire multiple resistance mechanisms. These findings raise the concern about the future of antimicrobial therapy and the capability of clinical laboratories to detect resistant strains, since simultaneous production of MBLs and ESBLs is known to promote further complexity in phenotypic detection. Occurrence of intra-hospital clonal dissemination enhances the necessity of better observance of infection control practices.</p