Applying two approaches to detect unmeasured confounding due to time-varying variables in a self-controlled risk interval design evaluating COVID-19 vaccine safety signals, using myocarditis as a case example.

We test the robustness of the self-controlled risk interval (SCRI) design in a setting where time between doses may introduce time-varying confounding, using both negative control outcomes (NCOs) and quantitative bias analysis (QBA). All vaccinated cases identified from 5 European databases between 1 September 2020 and end of data availability were included. Exposures were doses 1-3 of the Pfizer, Moderna, AstraZeneca, and Janssen COVID-19 vaccines; outcomes were myocarditis and otitis externa (NCO). The SCRI used a 60-day control window and dose-specific 28-day risk windows, stratified by vaccine brand and adjusted for calendar time. The QBA included two scenarios: (i) baseline probability of the confounder was higher in the control window and (ii) vice versa. The NCO was not associated with any of the COVID-19 vaccine types or doses except Moderna dose 1 (IRR = 1.09, 95%CI 1.01-1.09). The QBA suggested even the strongest literature-reported confounder (COVID-19; RRmyocarditis = 18.3) could only explain away part of the observed effect from IRR = 3 to IRR = 1.40. The SCRI seems robust to unmeasured confounding in the COVID-19 setting, although a strong unmeasured confounder could bias the observed effect upward. Replication of our findings for other safety signals would strengthen this conclusion.

Allergic inflammation triggers dyslipidemia via IgG signalling.

BACKGROUND: Allergic diseases begin early in life and are often chronic, thus creating an inflammatory environment that may precede or exacerbate other pathologies. In this regard, allergy has been associated to metabolic disorders and with a higher risk of cardiovascular disease, but the underlying mechanisms remain incompletely understood. METHODS: We used a murine model of allergy and atherosclerosis, different diets and sensitization methods, and cell-depleting strategies to ascertain the contribution of acute and late phase inflammation to dyslipidemia. Untargeted lipidomic analyses were applied to define the lipid fingerprint of allergic inflammation at different phases of allergic pathology. Expression of genes related to lipid metabolism was assessed in liver and adipose tissue at different times post-allergen challenge. Also, changes in serum triglycerides (TGs) were evaluated in a group of 59 patients ≥14 days after the onset of an allergic reaction. RESULTS: We found that allergic inflammation induces a unique lipid signature that is characterized by increased serum TGs and changes in the expression of genes related to lipid metabolism in liver and adipose tissue. Alterations in blood TGs following an allergic reaction are independent of T-cell-driven late phase inflammation. On the contrary, the IgG-mediated alternative pathway of anaphylaxis is sufficient to induce a TG increase and a unique lipid profile. Lastly, we demonstrated an increase in serum TGs in 59 patients after undergoing an allergic reaction. CONCLUSION: Overall, this study reveals that IgG-mediated allergic inflammation regulates lipid metabolism.

Cutting Edge: TCR-ß Selection Is Required at the CD4+CD8+ Stage of Human T Cell Development.

T cell development is predicated on the successful rearrangement of the TCR gene loci, which encode for Ag-specific receptors. Recombination-activating gene (RAG) 2 is required for TCR gene rearrangements, which occur during specific stages of T cell development. In this study, we differentiated human pluripotent stem cells with a CRISPR/Cas9-directed deletion of the RAG2 gene (RAG2-KO) to elucidate the requirement for the TCR ß-chain in mediating ß-selection during human T cell development. In stark contrast to mice, human RAG2-KO T lineage progenitors progressed to the CD4+CD8+ double-positive (DP) stage in the absence of TCRß rearrangements. Nonetheless, RAG2-KO DPs retrovirally transduced to express a rearranged TCR ß-chain showed increased survival and proliferation as compared with control-transduced RAG2-KO DPs. Furthermore, transcriptomic analysis showed that TCRß- and control-transduced RAG2-KO DPs differed in gene pathways related to survival and proliferation. Our results provide important insights as to the distinct requirement for the TCR ß-chain during human T cell development.

Neuronal Prosurvival Role of Ceramide Synthase 2 by Olidogendrocyte-to-Neuron Extracellular Vesicle Transfer.

Ageing is associated with notorious alterations in neurons, i.e., in gene expression, mitochondrial function, membrane degradation or intercellular communication. However, neurons live for the entire lifespan of the individual. One of the reasons why neurons remain functional in elderly people is survival mechanisms prevail over death mechanisms. While many signals are either pro-survival or pro-death, others can play both roles. Extracellular vesicles (EVs) can signal both pro-toxicity and survival. We used young and old animals, primary neuronal and oligodendrocyte cultures and neuroblastoma and oligodendrocytic lines. We analysed our samples using a combination of proteomics and artificial neural networks, biochemistry and immunofluorescence approaches. We found an age-dependent increase in ceramide synthase 2 (CerS2) in cortical EVs, expressed by oligodendrocytes. In addition, we show that CerS2 is present in neurons via the uptake of oligodendrocyte-derived EVs. Finally, we show that age-associated inflammation and metabolic stress favour CerS2 expression and that oligodendrocyte-derived EVs loaded with CerS2 lead to the expression of the antiapoptotic factor Bcl2 in inflammatory conditions. Our study shows that intercellular communication is altered in the ageing brain, which favours neuronal survival through the transfer of oligodendrocyte-derived EVs containing CerS2.

Predictors of Word and Pseudoword Reading in Languages with Different Orthographic Consistency.

This study analyzes the impact of certain cognitive processes on word and pseudoword reading in languages with different orthographic consistency (Spanish and Arabic) in the first year of Primary Education. The study was conducted with a group of 113 pupils from Algeria and another group of 128 pupils from Ecuador, from a middle-class background and without any special education needs. The participants were assessed in terms of their reading ability of words and pseudowords, knowledge of letters, phonological awareness, rapid automatized naming, and phonological memory. Using a correlational design, descriptive-exploratory, bivariate, and hierarchical multivariate regressions were applied to the different measures of reading in each language. The findings show that knowledge of letters, phonological awareness, rapid automatized naming, and phonological memory contribute differently to the explanation of reading ability in each group at the start of compulsory schooling. These results have important implications for the teaching of reading skills and the prevention of specific learning disabilities, as well as the theory of reading acquisition.

Thromboxane-dependent coronary vasoconstriction in obese mice: Role of peroxynitrite.

Obesity leads to chronic oxidative stress promoting the development of cardiovascular diseases including coronary artery disease and endothelial dysfunction. Increased reactive oxygen species production associated with obesity might lead to endothelial dysfunction through cyclooxygenase (COX) pathway. We evaluated arachidonic acid (AA)-dependent coronary vascular responses and explored COX metabolism in obese C57BL/6 mice. In response to arachidonic acid (AA), isolated hearts from obese mice showed increased vasoconstriction compared with control mice. Released thromboxane (TX) A2 during AA-induced vasoconstriction phase was increased in heart perfusates from obese mice. Indomethacin and 1-benzylimidazole, both reduced vasoconstriction response in control and obese mice. Vasoconstriction response to TXA2 mimetic analog U46619 was 2.7 higher in obese mice. Obesity increased COX-2, TXS and TX receptor protein expression as well as oxidative stress evaluated by nitrotyrosine and peroxynitrite levels, compared with control mice. Obese mice treated with FeTMPyP, a peroxynitrite scavenger, reversed all these parameters to control levels. These data suggest that alterations in COX pathway may be associated with increased generation of free radicals, including peroxynitrite, that result from the oxidative stress observed in obesity.

CRISPR-mediated knock-in in the mouse embryo using long single stranded DNA donors synthesised by biotinylated PCR.

Successful gene knock-in by CRISPR-Cas9 in the mouse zygote requires three components; guideRNA, Cas9 protein and a suitable donor template, which usually comprises homology flanked insert sequence. Recently, long single stranded DNA (lssDNA) donors have emerged as a popular choice of DNA donor, outperforming dsDNA templates in terms of knock-in efficiency for gene tagging and generating conditional alleles. The generation of these donors can be achieved through several methods that may introduce errors in the sequence, result in poor yields, and contain dsDNA contamination. We have developed our own cost-effective lssDNA synthesis methodology that results in high purity, sequence verified, low contamination lssDNA donors. We provide a detailed methodology on the design and generation of such donors for gene tagging experiments and generating conditional alleles.

Prevalence and Odds of Anxiety Disorders and Anxiety Symptoms in Children and Adults with Psoriasis: Systematic Review and Meta-analysis.

The magnitude of the association between psoriasis and depression has been evaluated, but not that between psoriasis and anxiety. The aim of this systematic review and meta-analysis was to examine the prevalence and odds of anxiety disorders and symptoms in patients with psoriasis. Five medical databases (Cochrane Database, EMBASE, PubMed, PsychINFO, ScienceDirect) were searched for relevant literature. A total of 101 eligible articles were identified. Meta-analysis revealed different prevalence rates depending on the type of anxiety disorder: 15% [95% confidence interval [CI] 9-21] for social anxiety disorder, 11% [9-14] for generalized anxiety disorder, and 9% [95% CI 8-10] for unspecified anxiety disorder. There were insufficient studies assessing other anxiety disorders to be able to draw any conclusions on their true prevalence. Meta-analysis also showed a high prevalence of anxiety symptoms (34% [95% CI 32-37]). Case-control studies showed a positive association between psoriasis and unspecified anxiety disorder (odds ratio 1.48 [1.18; 1.85]) and between psoriasis and anxiety symptoms (odds ratio 2.51 [2.02; 3.12]). All meta-analyses revealed an important heterogeneity, which could be explained in each case by methodological factors. The results of this study raise the necessity of screening for the presence of anxiety disorders, as previously recommended for depressive disorders, in patients with psoriasis and, if necessary, to refer such patients for evaluation by a mental health professional and appropriate treatment.

Toxigenic Microcystis aeruginosa (Cyanobacteria) affects the population growth of two common green microalgae: Evidence of other allelopathic metabolites different to cyanotoxins.

Agriculture runoffs and discharge of wastewaters are the major causes of eutrophication. Although eutrophication could promote the thriving of any phytoplankter, harmful algal blooms (HABs) are dominated frequently by cyanobacteria. Currently, HABs dominated by the toxigenic cyanobacterium Microcystis aeruginosa in lakes and reservoirs are the main environmental concerns worldwide. This study aimed to determine how M. aeruginosa (Ma) modifies the population growth of Pseudokirchneriella subcapitata (Ps) and Ankistrodesmus falcatus (Af). Growth kinetics were determined for each species and in the combinations: Ps-Ma, Af-Ma, Af-Ps, and Ps-Af-Ma. At the end of experiments, photosynthetic pigments, phycobiliproteins, and microcystins were quantified. A logistic equation significantly described the growth trend for all of the tested species, enabling the identification of negative effects on early stages in the population growth of co-cultures with the cyanobacterium; in addition, the interaction effects on the growth rate and in the maximum attainable population density were determined. The biomasses of A. falcatus and P. subcapitata were significantly higher when cultured individually than in all of the combinations with the cyanobacterium. The concentrations of chlorophyll a and b, as well as carotenoids, were lower in combined cultures, but phycobiliprotein content in the cultures with M. aeruginosa was not significantly affected. Microcystis aeruginosa negatively affected the growth of the microalgae, but A. falcatus was significantly more inhibited than P. subcapitata; however, microcystin concentrations were significantly reduced in the co-cultures with microalgae. These results could help to explain the displacements of microalgae when cyanobacteria are present, giving rise to cyanobacterial blooms in eutrophic freshwaters.

Increased APOE ε4 expression is associated with the difference in Alzheimer's disease risk from diverse ancestral backgrounds.

INTRODUCTION: Apolipoprotein E (APOE) ε4 confers less risk for Alzheimer's disease (AD) in carriers with African local genomic ancestry (ALA) than APOE ε4 carriers with European local ancestry (ELA). Cell type specific transcriptional variation between the two local ancestries (LAs) could contribute to this disease risk differences. METHODS: Single-nucleus RNA sequencing was performed on frozen frontal cortex of homozygous APOE ε4/ε4 AD patients: seven with ELA, four with ALA. RESULTS: A total of 60,908 nuclei were sequenced. Within the LA region (chr19:44-46Mb), APOE was the gene most differentially expressed, with ELA carriers having significantly more expression (overall P < 1.8E-317 ) in 24 of 32 cell clusters. The transcriptome of one astrocyte cluster, with high APOE ε4 expression and specific to ELA, is suggestive of A1 reactive astrocytes. DISCUSSION: AD patients with ELA expressed significantly greater levels of APOE than ALA APOE ε4 carriers. These differences in APOE expression could contribute to the reduced risk for AD seen in African APOE ε4 carriers.

Downregulation of MYPT1 increases tumor resistance in ovarian cancer by targeting the Hippo pathway and increasing the stemness.

BACKGROUND: Ovarian cancer is one of the most common and malignant cancers, partly due to its late diagnosis and high recurrence. Chemotherapy resistance has been linked to poor prognosis and is believed to be linked to the cancer stem cell (CSC) pool. Therefore, elucidating the molecular mechanisms mediating therapy resistance is essential to finding new targets for therapy-resistant tumors. METHODS: shRNA depletion of MYPT1 in ovarian cancer cell lines, miRNA overexpression, RT-qPCR analysis, patient tumor samples, cell line- and tumorsphere-derived xenografts, in vitro and in vivo treatments, analysis of data from ovarian tumors in public transcriptomic patient databases and in-house patient cohorts. RESULTS: We show that MYPT1 (PPP1R12A), encoding myosin phosphatase target subunit 1, is downregulated in ovarian tumors, leading to reduced survival and increased tumorigenesis, as well as resistance to platinum-based therapy. Similarly, overexpression of miR-30b targeting MYPT1 results in enhanced CSC-like properties in ovarian tumor cells and is connected to the activation of the Hippo pathway. Inhibition of the Hippo pathway transcriptional co-activator YAP suppresses the resistance to platinum-based therapy induced by either low MYPT1 expression or miR-30b overexpression, both in vitro and in vivo. CONCLUSIONS: Our work provides a functional link between the resistance to chemotherapy in ovarian tumors and the increase in the CSC pool that results from the activation of the Hippo pathway target genes upon MYPT1 downregulation. Combination therapy with cisplatin and YAP inhibitors suppresses MYPT1-induced resistance, demonstrating the possibility of using this treatment in patients with low MYPT1 expression, who are likely to be resistant to platinum-based therapy.

Generation and characterization of murine monoclonal antibodies against immunoreactive trypsinogen for newborn screening of cystic fibrosis.

Cystic fibrosis (CF) is a multisystem disorder that reduces quality of life and survival in affected individuals. In newborns, the release of pancreatic enzymes into the blood raises the levels of immunoreactive trypsinogen (IRT), the main marker for CF screening, which is detected in dried blood samples on filter paper by immunoenzymatic assays. In Cuba, CF has an estimated incidence of 1/9862 live births and should be included in the national basic newborn screening (NBS) panel given its benefits in terms of nutrition, lung function and survival. The Immunoassay Center develops and produces diagnostic kits allowing the establishment of large-scale NBS programs for inherited metabolic disorders in Cuba and other Latin American countries. IRT-specific monoclonal antibodies (MAbs) obtained at the Immunoassay Center are essential for developing an affordable immunoassay for IRT to support CF NBS in our low-income country. An immunization scheme with trypsinogen-1 originated two IgG1-producing murine hybridomas. 4C9C9 and 4C9E11 MAbs recognized different determinants on both trypsin-1 and trypsin-2 molecules. Both antibodies identified conformational epitopes on the molecule of trypsin-1 and of its zymogen. As 4C9E11 MAb cross-reacted with proteins structurally and functionally related to trypsinogen, it was used as revealing antibody in a sandwich-type UMELISA® assay for IRT determination with 4C9C9 MAb for capture. This combination, aside from detecting several commercially available trypsins, adequately quantified IRT from dried blood samples on filter paper of newborns. The evaluation of the assay's accuracy yielded percentage recoveries ranging 93.3-109.2% for commercial controls. The properties of the studied MAbs demonstrate their suitability for being used in a sandwich-type UMELISA® assay for the CF NBS in Cuba.

Generation and characterization of monoclonal antibodies against thyroid-stimulating hormone for newborn screening of congenital hypothyroidism.

Congenital hypothyroidism (CH) is one of the most frequent inherited-metabolic diseases in the world, and the main cause of treatable mental retardation in children. Because signs and symptoms of this disease are often scarce and not easily recognizable, newborns are screened for the early CH detection at birth. The Center of Immunoassay (CIE) has developed the UMELISA® TSH Neonatal and UMELISA® TSH to determine neonatal thyroid-stimulating hormone (TSH) levels in dried blood and serum samples. Both reagent kits use the same polystyrene plates coated with anti-ß-TSH monoclonal antibodies (MAbs), but one of these is commercially acquired. Obtaining appropriate anti-TSH MAbs at the CIE would guarantee economic independence and security in the production of these kits. Immunization of mice with TSH led to the generation of 7G11E3, an anti-ß-TSH IgG1-secreting hybridoma. The high affinity of 7G11E3 MAb and its characteristic epitopic recognition explain its better performance when adsorbed to UMELISA® plates for capturing low amounts of TSH in comparison with the studied MAbs. Performance of assays using polystyrene plates coated with 7G11E3 MAb was studied. Recovery percentages (100.0-106.7% for UMELISA® TSH NEONATAL and 97.3-99.0% for UMELISA® TSH) and intra (5.2-7.9% for UMELISA® TSH NEONATAL and 3.2-5.3% for UMELISA® TSH) and inter (6.6-7.7% for UMELISA® TSH NEONATAL and 5.2-8.0% for UMELISA® TSH) coefficients of variation were similar to the ones described for the commercial kits. Limits of detection and quantification were 1.0 and 3.8 mIU/L for UMELISA® TSH NEONATAL, and 0.3 and 0.6 mIU/L for UMELISA® TSH, respectively. The results also showed high overall concordance between assays (n = 2 019, ρc = 0.90 for UMELISA® TSH NEONATAL and n = 200, ρc = 0.94 for UMELISA® TSH). The 7G11E3 MAb meets the requirements for its use in the plates of UMELISA® TSH kits for CH newborn screening in Cuba. Abbreviations: CECMED, Center for the State Control of Medicaments and Medical Equipment and Devices; CH, congenital hypothyroidism; CIE, Center of Immunoassay; CLSI, Clinical and Laboratory Standards Institute; CV coefficient of variation; DBS, dried blood spots; LOB, limit of blank; LOD, limit of detection; LOQ, limit of quantitation; SD, standard deviation; Sr, repeatability standard deviation; SUMA, Ultra Micro Analytic System; UMELISA, ultramicro enzyme-linked immunosorbent assay.

Molecular Identification and Susceptibility Testing of Molds Isolated in a Prospective Surveillance of Triazole Resistance in Spain (FILPOP2 Study).

Antifungal resistance is increasing by the emergence of intrinsically resistant species and by the development of secondary resistance in susceptible species. A previous study performed in Spain revealed levels of azole resistance in molds of between 10 and 12.7%, but secondary resistance in Aspergillus fumigatus was not detected. We used itraconazole (ITZ)-supplemented medium to select resistant strains. A total of 500 plates supplemented with 2 mg/liter of ITZ were sent to 10 Spanish tertiary hospitals, and molecular identification and antifungal susceptibility testing were performed. In addition, the cyp51A gene in those A. fumigatus strains showing azole resistance was sequenced. A total of 493 isolates were included in the study. Sixteen strains were isolated from patients with an infection classified as proven, 104 were isolated from patients with an infection classified as probable, and 373 were isolated from patients with an infection classified as colonization. Aspergillus was the most frequent genus isolated, at 80.3%, followed by Scedosporium-Lomentospora (7.9%), Penicillium-Talaromyces (4.5%), Fusarium (2.6%), and the order Mucorales (1%). Antifungal resistance was detected in Scedosporium-Lomentospora species, Fusarium, Talaromyces, and Mucorales Three strains of A. fumigatus sensu stricto were resistant to azoles; two of them harbored the TR34+L98H mechanism of resistance, and the other one had no mutations in cyp51A The level of azole resistance in A. fumigatus remains low, but cryptic species represent over 10% of the isolates and have a broader but overall higher range of antifungal resistance.

Screen for multi-SUMO-binding proteins reveals a multi-SIM-binding mechanism for recruitment of the transcriptional regulator ZMYM2 to chromatin.

Protein SUMOylation has emerged as an important regulatory event, particularly in nuclear processes such as transcriptional control and DNA repair. In this context, small ubiquitin-like modifier (SUMO) often provides a binding platform for the recruitment of proteins via their SUMO-interacting motifs (SIMs). Recent discoveries point to an important role for multivalent SUMO binding through multiple SIMs in the binding partner as exemplified by poly-SUMOylation acting as a binding platform for ubiquitin E3 ligases such as ring finger protein 4. Here, we have investigated whether other types of protein are recruited through multivalent SUMO interactions. We have identified dozens of proteins that bind to multi-SUMO platforms, thereby uncovering a complex potential regulatory network. Multi-SUMO binding is mediated through multi-SIM modules, and the functional importance of these interactions is demonstrated for the transcriptional corepressor ZMYM2/ZNF198 where its multi-SUMO-binding activity is required for its recruitment to chromatin.

An enzyme immunoassay for determining epidermal growth factor (EGF) in human serum samples using an ultramicroanalytical system.

Human epidermal growth factor is a small peptide consisting of 53 amino acid residues, which stimulates cell proliferation and is associated with several human carcinomas. A simple sandwich-type ultramicroELISA assay (UMELISA), based on the advantages of high affinity reaction between streptavidin and biotin has been developed for the measurement of EGF in human serum samples. Strips coated with a high affinity monoclonal antibody directed against EGF are used as solid phase, to ensure the specificity of the assay. The EGF assay was completed in 18 hr, with a measuring range of 39-2500 pg/mL. The intra- and inter-assay coefficients of variation were 4.4-7.3% and 0-5.1%, respectively, depending on the EGF concentrations evaluated. Percentage recovery ranged from 96-104%. Regression analysis showed a good correlation with the commercially available Human EGF Immunoassay Quantikine® ELISA kit (n = 130, r = 0.92, P < 0.01). The analytical performance characteristics of our UMELISA EGF endorse its use for the quantification of EGF in human serum samples.

[Implementation and evaluation of a blended learning course on gastroesophageal reflux disease for physicians in Latin America]. / Implementación y evaluación de un curso semipresencial sobre enfermedad por reflujo gastroesofágico para médicos de América Latina.

Integrating evidence-based clinical practice guidelines on gastroesophageal reflux disease into medical practice is of prime importance in Latin America, given its high prevalence in this region. The aim of this project was to implement and assess an educational intervention on gastroesophageal reflux disease, aimed at primary care physicians in Latin America, with contents based on current clinical guidelines. The course included initial activities, whether face-to-face or through distance learning, and a 2-month period of Internet study and interaction. A pilot test was carried out in Uruguay, which was then repeated in 5 countries (Mexico, Colombia, Venezuela, Argentina and again in Uruguay). A global template was designed, which was then adapted to each of the countries: this was done with the participation of local institutions and leaders. Local credits were given for recertification. Participation was free. Of 3,110 physicians invited to participate, 1,143 (36.8%) started the course. Of these, 587 (51.4%) accessed at least half the contents of the course and 785 (68.7%) took part in the clinical discussions. A total of 338 (29.6%) completed all the requirements of the course and received a certificate. Among physicians who took both the pre- and post-intervention knowledge tests, scores improved from 60 to 80% (P<.001). Ninety-two percent of planned changes in clinical practice were related to the pedagogic aims of the course. In conclusion, a multifaceted, 2-phase continuing education course was successfully imparted in Latin America, with an overall design that was adapted to each country. Determination of specific needs and the participation of national experts were fundamental to the success of the course.

Risk of herpes zoster in adults with SARS-CoV-2 infection in Spain: A population-based, retrospective cohort study.

OBJECTIVES: We aimed to compare the risk of herpes zoster (HZ) in adults with and without laboratory-confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. METHODS: This retrospective dynamic cohort study analyzed data from a public healthcare database in Spain between November 2020 and October 2021. The main outcome was incident cases of HZ in individuals ≥18-year-old. Relative risk (RR) of HZ in SARS-CoV-2-confirmed versus SARS-CoV-2-free individuals was estimated by a multivariable negative binomial regression adjusted by age, sex, and comorbidities. RESULTS: Data from 4,085,590 adults were analyzed. The overall HZ incidence rate in adults was 5.76 (95% confidence interval [CI], 5.66-5.85) cases per 1000 person-years. Individuals ≥18-year-old with SARS-CoV-2-confirmed infection had a 19% higher risk of developing HZ versus SARS-CoV-2-free ≥18-year-olds (adjusted RR = 1.19; 95% CI, 1.09-1.29); this percentage was 16% (adjusted RR = 1.16; 95% CI, 1.05-1.29) in ≥50-year-olds. Severe (hospitalized) cases of SARS-CoV-2 infection had a 64% (if ≥18 years old) or 44% (if ≥50 years old) higher risk of HZ versus nonhospitalized cases. CONCLUSION: These results support an association between SARS-CoV-2 infection and HZ, with a greater HZ risk in severe cases of SARS-CoV-2 infection.

Protein homeostasis maintained by HOOK1 levels promotes the tumorigenic and stemness properties of ovarian cancer cells through reticulum stress and autophagy.

BACKGROUND: Ovarian cancer has a high mortality rate mainly due to its resistance to currently used therapies. This resistance has been associated with the presence of cancer stem cells (CSCs), interactions with the microenvironment, and intratumoral heterogeneity. Therefore, the search for new therapeutic targets, particularly those targeting CSCs, is important for improving patient prognosis. HOOK1 has been found to be transcriptionally altered in a substantial percentage of ovarian tumors, but its role in tumor initiation and development is still not fully understood. METHODS: The downregulation of HOOK1 was performed in ovarian cancer cell lines using CRISPR/Cas9 technology, followed by growth in vitro and in vivo assays. Subsequently, migration (Boyden chamber), cell death (Western-Blot and flow cytometry) and stemness properties (clonal heterogeneity analysis, tumorspheres assay and flow cytometry) of the downregulated cell lines were analysed. To gain insights into the specific mechanisms of action of HOOK1 in ovarian cancer, a proteomic analysis was performed, followed by Western-blot and cytotoxicity assays to confirm the results found within the mass spectrometry. Immunofluorescence staining, Western-blotting and flow cytometry were also employed to finish uncovering the role of HOOK1 in ovarian cancer. RESULTS: In this study, we observed that reducing the levels of HOOK1 in ovarian cancer cells reduced in vitro growth and migration and prevented tumor formation in vivo. Furthermore, HOOK1 reduction led to a decrease in stem-like capabilities in these cells, which, however, did not seem related to the expression of genes traditionally associated with this phenotype. A proteome study, along with other analysis, showed that the downregulation of HOOK1 also induced an increase in endoplasmic reticulum stress levels in these cells. Finally, the decrease in stem-like properties observed in cells with downregulated HOOK1 could be explained by an increase in cell death in the CSC population within the culture due to endoplasmic reticulum stress by the unfolded protein response. CONCLUSION: HOOK1 contributes to maintaining the tumorigenic and stemness properties of ovarian cancer cells by preserving protein homeostasis and could be considered an alternative therapeutic target, especially in combination with inducers of endoplasmic reticulum or proteotoxic stress such as proteasome inhibitors.