In vivo labeling of endogenous genomic loci in C. elegans using CRISPR/dCas9

Visualization of genomic loci with open chromatin state has been reported in mammalian tissue culture cells using a CRISPR/Cas9-based system that utilizes an EGFP-tagged endonuclease-deficient Cas9 protein (dCas9::EGFP) (Chen et al. 2013). Here, we adapted this approach for use in Caenorhabditis elegans . We generated a C. elegans strain that expresses the dCas9 protein fused to two nuclear-localized EGFP molecules (dCas9::NLS::2xEGFP::NLS) in an inducible manner. Using this strain, we report the visualization in live C. elegans embryos of two endogenous repetitive loci, rrn-4 and rrn-1 , from which 5S and 18S ribosomal RNAs are constitutively generated

miRNAs cooperate in apoptosis regulation during C. elegans development

Programmed cell death occurs in a highly reproducible manner during Caenorhabditis elegans development. We demonstrate that, during embryogenesis, miR-35 and miR-58 bantam family microRNAs (miRNAs) cooperate to prevent the precocious death of mothers of cells programmed to die by repressing the gene egl-1, which encodes a proapoptotic BH3-only protein. In addition, we present evidence that repression of egl-1 is dependent on binding sites for miR-35 and miR-58 family miRNAs within the egl-1 3\u27 untranslated region (UTR), which affect both mRNA copy number and translation. Furthermore, using single-molecule RNA fluorescent in situ hybridization (smRNA FISH), we show that egl-1 is transcribed in the mother of a cell programmed to die and that miR-35 and miR-58 family miRNAs prevent this mother from dying by keeping the copy number of egl-1 mRNA below a critical threshold. Finally, miR-35 and miR-58 family miRNAs can also dampen the transcriptional boost of egl-1 that occurs specifically in a daughter cell that is programmed to die. We propose that miRNAs compensate for lineage-specific differences in egl-1 transcriptional activation, thus ensuring that EGL-1 activity reaches the threshold necessary to trigger death only in daughter cells that are programmed to die

Genome-wide RNAi screen for regulators of UPRmt in Caenorhabditis elegans mutants with defects in mitochondrial fusion

© The Author(s) 2021. Published by Oxford University Press on behalf of Genetics Society of America.Mitochondrial dynamics plays an important role in mitochondrial quality control and the adaptation of metabolic activity in response to environmental changes. The disruption of mitochondrial dynamics has detrimental consequences for mitochondrial and cellular homeostasis and leads to the activation of the mitochondrial unfolded protein response (UPRmt), a quality control mechanism that adjusts cellular metabolism and restores homeostasis. To identify genes involved in the induction of UPRmt in response to a block in mitochondrial fusion, we performed a genome-wide RNAi screen in Caenorhabditis elegans mutants lacking the gene fzo-1, which encodes the ortholog of mammalian Mitofusin, and identified 299 suppressors and 86 enhancers. Approximately 90% of these 385 genes are conserved in humans, and one-third of the conserved genes have been implicated in human disease. Furthermore, many have roles in developmental processes, which suggests that mitochondrial function and their response to stress are defined during development and maintained throughout life. Our dataset primarily contains mitochondrial enhancers and non-mitochondrial suppressors of UPRmt, indicating that the maintenance of mitochondrial homeostasis has evolved as a critical cellular function, which, when disrupted, can be compensated for by many different cellular processes. Analysis of the subsets “non-mitochondrial enhancers” and “mitochondrial suppressors” suggests that organellar contact sites, especially between the ER and mitochondria, are of importance for mitochondrial homeostasis. In addition, we identified several genes involved in IP3 signaling that modulate UPRmt in fzo-1 mutants and found a potential link between pre-mRNA splicing and UPRmt activation.11Nsciescopu

Outcomes after mechanical versus manual chest compressions in eCPR patients

Background Extracorporeal cardiopulmonary resuscitation (eCPR) is an established treatment option for cardiac arrest. Mechanical reanimation devices are increasingly used but have been associated with complications. This study evaluates typical injury patterns and differences after mechanical versus manual chest compressions among patients undergoing eCPR. Methods From 2016 to 2020, 108 eCPR patients were retrospectively analyzed. Primary endpoints were traumatic, hemorrhagic, or inner organ-related complications, defined as pneumothorax, pulmonary bleeding, major bleeding, gastrointestinal bleeding, gastrointestinal ischemia, cardiac tamponade, aortic dissection, sternal or rib fracture. Results 70 patients were treated with mechanical CPR (mCPR) and 38 with conventional CPR (cCPR). There were more CPR-related injuries in the mCPR group (55% vs. 83%, p = 0.01), CPR duration was longer (cCPR 40 +/- 28 min vs. mCPR 69 +/- 25 min, p = 0.01). There was no significant difference in mortality between the groups. Conclusion Mechanical CPR devices are associated with a higher incidence of traumatic and hemorrhagic injuries in patients undergoing eCPR

Extracorporeal cardiopulmonary resuscitation for in- and out-of-hospital cardiac arrest: The race against time

Objectives: Extracorporeal cardiopulmonary resuscitation (ECPR) is increasingly used due to its beneficial outcomes and results compared to conventional CPR. Cardiac arrests can be categorized depending on location: in-hospital cardiac arrest (IHCA) and out-of-hospital cardiac arrest (OHCA). Despite this distinction, studies comparing the two are scarce, especially in comparing outcomes after ECPR. This study compared patient characteristics, cardiac arrest characteristics, and outcomes. Methods: Between 2016 and 2022, patients who underwent ECPR for cardiac arrest at our institution were retrospectively analyzed, depending on the arrest location: IHCA and OHCA. We compared periprocedural characteristics and used multinomial regression analysis to indicate parameters contributing to a favorable outcome. Results: A total of n = 157 patients (100%) were analyzed (OHCA = 91; IHCA = 66). Upon admission, OHCA patients were younger (53.2 ± 12.4 vs. 59.2 ± 12.6 years) and predominantly male (91.1% vs. 66.7%, p=<0.001). The low-flow time was significantly shorter in IHCA patients (41.1 ± 27.4 mins) compared to OHCA (63.6 ± 25.1 mins). Despite this significant difference, in-hospital mortality was not significantly different in both groups (IHCA = 72.7% vs. OHCA = 76.9%, p = 0.31). Both groups' survival-to-discharge factors were CPR duration, low flow time, and lactate values upon ECMO initiation. Conclusion: Survival-to-discharge for ECPR in IHCA and OHCA was around 25%, and there was no statistically significant difference between the two cohorts. Factors predicting survival were lower lactate levels before cannulation and lower low-flow time. As such, OHCA patients seem to tolerate longer low-flow times and thus metabolic impairments compared to IHCA patients and may be considered for ECMO cannulation on a broader time span than IHCA

Benign and malignant cardiac masses: long-term outcomes after surgical resection

Introduction Cardiac tumors represent a rare and heterogenous pathologic entity, with a cumulative incidence of up to 0.02%. This study aimed to investigate one of the largest patient cohorts published for clinical presentation and long-term outcomes after surgical resection. Areas covered Between 2009 and 2021, 183 consecutive patients underwent surgery for tumor excision in our center. Preoperative baseline characteristics, intraoperative data, and long-term survival were analyzed. The diagnosis was confirmed postoperatively by histology and Immunohistochemical investigations. Kaplan-Meier curves assessed survival, and the Cox proportional hazards model, was used to identify prognostic factors for overall survival. Results This series included 183 consecutive patients; most (n = 169, 92.3%) were diagnosed with benign cardiac masses. The mean age of patients was 60 +/- 16 years, and 48% (n = 88) were females. The largest group of tumors was myxoma (n = 98; 54%). The most common malignant tumor type was sarcoma (n = 5; 2.7%). The mean hospital stay was 11 +/- 6.5 days, and all-cause mortality after ten years was 14%. Expert Opinion Surgery represents the gold standard in treating primary cardiac tumors; in benign tumors, it is highly effective and curative, whereas, in malignant tumors, it remains associated with more prolonged survival

Surgery for Aortic Prosthetic Valve Endocarditis in the Transcatheter Era

Objectives: As surgical experience with infective endocarditis following transcatheter aortic valve replacement is scarce, this study compared the perioperative and short-term outcomes of patients suffering from endocarditis following surgical aortic valve replacement and transcatheter aortic valve replacement. Methods: Between January 2013 and December 2020, 468 consecutive patients were admitted to our center for surgery for IE. Among them, 98 were operated on for endocarditis following surgical aortic valve replacement and 22 for endocarditis following transcatheter aortic valve replacement. Results: The median EuroSCORE II (52.1 (40.6-62.0) v/s 45.4 (32.6-58.1), p = 0.207) and STS-PROM (1.8 (1.6-2.1) v/s 1.9 (1.4-2.2), p = 0.622) were comparable. Endocarditis following transcatheter aortic valve replacement accounted for 13.7% of the aortic prosthetic valve endocarditis between 2013 and 2015; this increased to 26.9% in the years 2019 and 2020.Concomitant procedures were performed in 35 patients (29.2%). The operative mortality was 26.5% in the endocarditis following surgical aortic valve replacement group and 9.1% in the endocarditis following transcatheter aortic valve replacement group (p = 0.098). Upon follow-up, survival at 6 months was found to be 98% in the group with endocarditis following surgical aortic valve replacement and 89% in the group with endocarditis following transcatheter aortic valve replacement (p = 0.081). Conclusions: Patients suffering from endocarditis following surgical aortic valve replacement and transcatheter aortic valve replacement present with comparable risk profiles and can be surgically treated with comparable results. Surgery as a curative option should not be rejected even in this intermediate-risk cohort