Sex Influence on Autophagy Markers and miRNAs in Basal and Angiotensin II-Treated Human Umbilical Vein Endothelial Cells.

Cardiovascular diseases (CVD) display many sex and gender differences, and endothelial dysfunction, angiotensin II (Ang II), and autophagy represent key factors in the autophagic process Therefore, we studied whether Ang II modulates the mentioned processes in a sex-specific way in HUVECs obtained from healthy male and female newborns. In basal HUVECs, the Parkin gene and protein were higher in FHUVECs than in MHUVECs, while the Beclin-1 protein was more expressed in MHUVECs, and no other significant differences were detected. Ang II significantly increases LAMP-1 and p62 protein expression and decreases the expression of Parkin protein in comparison to basal in MHUVECs. In FHUVECs, Ang II significantly increases the expression of Beclin-1 gene and protein, and Parkin gene. The LC3 II/I ratio and LAMP-1 protein were significantly higher in MHUVECs than in FHUVECs, while Parkin protein was significantly more expressed in Ang II-treated FHUVECs than in male cells. Ang II affects the single miRNA levels: miR-126-3p and miR-133a-3p are downregulated and upregulated in MHUVECs and FHUVECs, respectively. MiR-223 is downregulated in MHUVEC and FHUVECs. Finally, miR-29b-3p and miR-133b are not affected by Ang II. Ang II effects and the relationship between miRNAs and organelles-specific autophagy is sex-dependent in HUVECs. This could lead to a better understanding of the mechanisms underlying sex differences in endothelial dysfunction, providing useful indications for innovative biomarkers and personalized therapeutic approaches.

Stratification of Amniotic Fluid Cells and Amniotic Fluid by Sex Opens Up New Perspectives on Fetal Health.

Amniotic fluid is essential for fetus wellbeing and is used to monitor pregnancy and predict fetal outcomes. Sex affects health and medicine from the beginning of life, but knowledge of its influence on cell-depleted amniotic fluid (AF) and amniotic fluid cells (AFCs) is still neglected. We evaluated sex-related differences in AF and in AFCs to extend personalized medicine to prenatal life. AFCs and AF were obtained from healthy Caucasian pregnant women who underwent amniocentesis at the 16th-18th week of gestation for advanced maternal age. In the AF, inflammation biomarkers (TNFα, IL6, IL8, and IL4), malondialdehyde, nitrites, amino acids, and acylcarnitines were measured. Estrogen receptors and cell fate (autophagy, apoptosis, senescence) were measured in AFCs. TNFα, IL8, and IL4 were higher in female AF, whereas IL6, nitrites, and MDA were similar. Valine was higher in male AF, whereas several acylcarnitines were sexually different, suggesting a mitochondrial involvement in establishing sex differences. Female AFCs displayed higher expression of ERα protein and a higher ERα/ERß ratio. The ratio of LC3II/I, an index of autophagy, was higher in female AFCs, while LC3 gene was similar in both sexes. No significant sex differences were found in the expression of the lysosomal protein LAMP1, while p62 was higher in male AFCs. LAMP1 gene was upregulated in male AFCs, while p62 gene was upregulated in female ones. Finally, caspase 9 activity and senescence linked to telomeres were higher in female AFCs, while caspase 3 and ß-galactosidase activities were similar. This study supports the idea that sex differences start very early in prenatal life and influence specific parameters, suggesting that it may be relevant to appreciate sex differences to cover knowledge gaps. This might lead to improving the diagnosis of risk prediction for pregnancy complications and achieving a more satisfactory monitoring of fetus health, even preventing future diseases in adulthood.

Pleiotropic effects of BAFF on the senescence-associated secretome and growth arrest.

Senescent cells release a variety of cytokines, proteases, and growth factors collectively known as the senescence-associated secretory phenotype (SASP). Sustained SASP contributes to a pattern of chronic inflammation associated with aging and implicated in many age-related diseases. Here, we investigated the expression and function of the immunomodulatory cytokine BAFF (B-cell activating factor; encoded by the TNFSF13B gene), a SASP protein, in multiple senescence models. We first characterized BAFF production across different senescence paradigms, including senescent human diploid fibroblasts (WI-38, IMR-90) and monocytic leukemia cells (THP-1), and tissues of mice induced to undergo senescence. We then identified IRF1 (interferon regulatory factor 1) as a transcription factor required for promoting TNFSF13B mRNA transcription in senescence. We discovered that suppressing BAFF production decreased the senescent phenotype of both fibroblasts and monocyte-like cells, reducing IL6 secretion and SA-ß-Gal staining. Importantly, however, the influence of BAFF on the senescence program was cell type-specific: in monocytes, BAFF promoted the early activation of NF-κB and general SASP secretion, while in fibroblasts, BAFF contributed to the production and function of TP53 (p53). We propose that BAFF is elevated across senescence models and is a potential target for senotherapy.

Origin, Genetic Variation and Molecular Epidemiology of SARS-CoV-2 Strains Circulating in Sardinia (Italy) during the First and Second COVID-19 Epidemic Waves.

Understanding how geography and human mobility shape the patterns and spread of infectious diseases such as COVID-19 is key to control future epidemics. An interesting example is provided by the second wave of the COVID-19 epidemic in Europe, which was facilitated by the intense movement of tourists around the Mediterranean coast in summer 2020. The Italian island of Sardinia is a major tourist destination and is widely believed to be the origin of the second Italian wave. In this study, we characterize the genetic variation among SARS-CoV-2 strains circulating in northern Sardinia during the first and second Italian waves using both Illumina and Oxford Nanopore Technologies Next Generation Sequencing methods. Most viruses were placed into a single clade, implying that despite substantial virus inflow, most outbreaks did not spread widely. The second epidemic wave on the island was actually driven by local transmission of a single B.1.177 subclade. Phylogeographic analyses further suggest that those viral strains circulating on the island were not a relevant source for the second epidemic wave in Italy. This result, however, does not rule out the possibility of intense mixing and transmission of the virus among tourists as a major contributor to the second Italian wave.

RNA-binding proteins in autoimmunity: From genetics to molecular biology.

Autoimmune diseases (ADs) are chronic pathologies generated by the loss of immune tolerance to the body's own cells and tissues. There is growing recognition that RNA-binding proteins (RBPs) critically govern immunity in healthy and pathological conditions by modulating gene expression post-transcriptionally at all levels: nuclear mRNA splicing and modification, export to the cytoplasm, as well as cytoplasmic mRNA transport, storage, editing, stability, and translation. Despite enormous efforts to identify new therapies for ADs, definitive solutions are not yet available in many instances. Recognizing that many ADs have a strong genetic component, we have explored connections between the molecular biology and the genetics of RBPs in ADs. Here, we review the genetics and molecular biology of RBPs in four major ADs, multiple sclerosis (MS), type 1 diabetes mellitus (T1D), systemic lupus erythematosus (SLE), and rheumatoid arthritis (RA). We anticipate that gaining insights into the genetics and biology of ADs can facilitate the discovery of new therapies. This article is categorized under: RNA in Disease and Development > RNA in Disease.

Cross-sectional analysis of the humoral response after SARS-CoV-2 vaccination in Sardinian multiple sclerosis patients, a follow-up study.

Monitoring immune responses to SARS-CoV-2 vaccination and its clinical efficacy over time in Multiple Sclerosis (MS) patients treated with disease-modifying therapies (DMTs) help to establish the optimal strategies to ensure adequate COVID-19 protection without compromising disease control offered by DMTs. Following our previous observations on the humoral response one month after two doses of BNT162b2 vaccine (T1) in MS patients differently treated, here we present a cross-sectional and longitudinal follow-up analysis six months following vaccination (T2, n=662) and one month following the first booster (T3, n=185). Consistent with results at T1, humoral responses were decreased in MS patients treated with fingolimod and anti-CD20 therapies compared with untreated patients also at the time points considered here (T2 and T3). Interestingly, a strong upregulation one month after the booster was observed in patients under every DMTs analyzed, including those treated with fingolimod and anti-CD20 therapies. Although patients taking these latter therapies had a higher rate of COVID-19 infection five months after the first booster, only mild symptoms that did not require hospitalization were reported for all the DMTs analyzed here. Based on these findings we anticipate that additional vaccine booster shots will likely further improve immune responses and COVID-19 protection in MS patients treated with any DMT.

Genetic Variation among Pharmacogenes in the Sardinian Population.

Pharmacogenetics (PGx) aims to identify the genetic factors that determine inter-individual differences in response to drug treatment maximizing efficacy while decreasing the risk of adverse events. Estimating the prevalence of PGx variants involved in drug response, is a critical preparatory step for large-scale implementation of a personalized medicine program in a target population. Here, we profiled pharmacogenetic variation in fourteen clinically relevant genes in a representative sample set of 1577 unrelated sequenced Sardinians, an ancient island population that accounts for genetic variation in Europe as a whole, and, at the same time is enriched in genetic variants that are very rare elsewhere. To this end, we used PGxPOP, a PGx allele caller based on the guidelines created by the Clinical Pharmacogenetics Implementation Consortium (CPIC), to identify the main phenotypes associated with the PGx alleles most represented in Sardinians. We estimated that 99.43% of Sardinian individuals might potentially respond atypically to at least one drug, that on average each individual is expected to have an abnormal response to about 17 drugs, and that for 27 drugs the fraction of the population at risk of atypical responses to therapy is more than 40%. Finally, we identified 174 pharmacogenetic variants for which the minor allele frequency was at least 10% higher among Sardinians as compared to other European populations, a fact that may contribute to substantial interpopulation variability in drug response phenotypes. This study provides baseline information for further large-scale pharmacogenomic investigations in the Sardinian population and underlines the importance of PGx characterization of diverse European populations, such as Sardinians.

Impact on breast cancer susceptibility and clinicopathological traits of common genetic polymorphisms in TP53, MDM2 and ATM genes in Sardinian women.

Common variants of genes involved in DNA damage correction [tumor protein p53 (TP53), murine double 2 homolog oncoprotein (MDM2) and ataxia-telengiectasia mutated (ATM)] may serve a role in cancer predisposition. The purpose of the present study was to investigate the association of five variants in these genes with breast cancer risk and clinicopathological traits in a cohort of 261 women from northern Sardinia. Polymorphic variants in TP53 (rs17878362, rs1042522 and rs1625895), MDM2 (rs2279744) and ATM (rs1799757) were determined by PCR and TaqMan single nucleotide polymorphism assay in patients with breast cancer (n=136) and healthy controls (n=125). Association with clinicopathological (e.g., age at diagnosis, lymph node involvement, clinical stage) and lifestyle factors (e.g., smoking status, alcohol intake, contraceptive use) was also evaluated. TP53 rs17878362 and rs1625895 polymorphisms were associated with decreased risk of BC diagnosis in patients older than 50 years (codominant and recessive models) and post-menopause (recessive model). Furthermore, there was a significant association between lymph node status (positive vs. negative) and ATM rs1799757-delT in dominant and additive models and between MDM2 rs2279744-allele and use of oral contraceptives. This analysis suggested that TP53 rs17878362 and rs1625895 may affect age of onset of breast cancer and ATM rs1799757 and MDM2 rs2279744 may be associated with lymph node status and prolonged use of oral contraceptives, respectively.

Multiple sclerosis disease-modifying therapies and COVID-19 vaccines: a practical review and meta-analysis.

Studies among people with multiple sclerosis (pwMS) receiving disease-modifying therapies (DMTs) have provided adequate evidence for an appraisal of COVID-19 vaccination policies among them. To synthesise the available evidence addressing the effect of MS DMTs on COVID-19 vaccines' immunogenicity and effectiveness, following the Cochrane guidelines, we systematically reviewed all observational studies available in MEDLINE, Scopus, Web of Science, MedRxiv and Google Scholar from January 2021 to January 2022 and extracted their relevant data. Immunogenicity data were then synthesised in a quantitative, and other data in a qualitative manner. Evidence from 28 studies suggests extensively lower B-cell responses in sphingosine-1-phosphate receptor modulator (S1PRM) treated and anti-CD20 (aCD20) treated, and lower T-cell responses in interferon-treated, S1PRM-treated and cladribine-treated pwMS-although most T cell evidence currently comprises of low or very low certainty. With every 10-week increase in aCD20-to-vaccine period, a 1.94-fold (95% CI 1.57 to 2.41, p<0.00001) increase in the odds of seroconversion was observed. Furthermore, the evidence points out that B-cell-depleting therapies may accelerate postvaccination humoral waning, and boosters' immunogenicity is predictable with the same factors affecting the initial vaccination cycle. Four real-world studies further indicate that the comparative incidence/severity of breakthrough COVID-19 has been higher among the pwMS treated with S1PRM and aCD20-unlike the ones treated with other DMTs. S1PRM and aCD20 therapies were the only DMTs reducing the real-world effectiveness of COVID-19 vaccination among pwMS. Hence, it could be concluded that optimisation of humoral immunogenicity and ensuring its durability are the necessities of an effective COVID-19 vaccination policy among pwMS who receive DMTs.

Non-coding RNAs in malaria infection.

Malaria is one of the most severe infectious diseases affecting humans and it is caused by protozoan pathogens of the species Plasmodium (spp.). The malaria parasite Plasmodium is characterized by a complex, multistage life cycle that requires tight gene regulation which allows for host invasion and defense against host immune responses. Unfortunately, the mechanisms regulating gene expression during Plasmodium infection remain largely elusive, though several lines of evidence implicate a major involvement of non-coding RNAs (ncRNAs). The ncRNAs have been found to play a key role in regulating transcriptional and post-transcriptional events in a broad range of organisms including Plasmodium. In Plasmodium ncRNAs have been shown to regulate key events in the multistage life cycle and virulence ability. Here we review recent progress involving ncRNAs (microRNAs, long non-coding RNAs, and circular RNAs) and their role as regulators of gene expression during Plasmodium infection in human hosts with focus on the possibility of using these molecules as biomarkers for monitoring disease status. We also discuss the surprising function of ncRNAs in mediating the complex interplay between parasite and human host and future perspectives of the field. This article is categorized under: RNA in Disease and Development > RNA in Disease.

Effect of Different Disease-Modifying Therapies on Humoral Response to BNT162b2 Vaccine in Sardinian Multiple Sclerosis Patients.

Objectives: Vaccination against COVID-19 is highly recommended to patients affected by multiple sclerosis (MS); however, the impact of MS disease-modifying therapies (DMTs) on the immune response following vaccination has been only partially investigated. Here, we aimed to elucidate the effect of DMTs on the humoral immune response to mRNA-based anti-SARS-CoV-2 vaccines in MS patients. Methods: We obtained sera from 912 Sardinian MS patients and 63 healthy controls 30 days after the second dose of BNT162b2 vaccine and tested them for SARS-CoV-2 response using anti-Spike (S) protein-based serology. Previous SARS-CoV-2 infection was assessed by anti-Nucleocapsid (N) serology. Patients were either untreated or undergoing treatment with a total of 13 different DMTs. Differences between treatment groups comprised of at least 10 patients were assessed by generalized linear mixed-effects model. Demographic and clinical data and smoking status were analyzed as additional factors potentially influencing humoral immunity from COVID-19 vaccine. Results: MS patients treated with natalizumab, teriflunomide, azathioprine, fingolimod, ocrelizumab, and rituximab showed significantly lower humoral responses compared to untreated patients. We did not observe a statistically significant difference in response between patients treated with the other drugs (dimethyl fumarate, interferon, alemtuzumab and glatiramer acetate) and untreated patients. In addition, older age, male sex and active smoking were significantly associated with lower antibody titers against SARS-CoV-2. MS patients previously infected with SARS-CoV-2 had significantly higher humoral responses to vaccine than uninfected patients. Conclusion: Humoral response to BNT162b2 is significantly influenced by the specific DMTs followed by patients, as well as by other factors such as previous SARS-CoV-2 infection, age, sex, and smoking status. These results are important to inform targeted strategies to prevent clinically relevant COVID-19 in MS patients.

Sex-Biased Expression of Pharmacogenes across Human Tissues.

Individual response to drugs is highly variable and largely influenced by genetic variants and gene-expression profiles. In addition, it has been shown that response to drugs is strongly sex-dependent, both in terms of efficacy and toxicity. To expand current knowledge on sex differences in the expression of genes relevant for drug response, we generated a catalogue of differentially expressed human transcripts encoded by 289 genes in 41 human tissues from 838 adult individuals of the Genotype-Tissue Expression project (GTEx, v8 release) and focused our analysis on relevant transcripts implicated in drug response. We detected significant sex-differentiated expression of 99 transcripts encoded by 59 genes in the tissues most relevant for human pharmacology (liver, lung, kidney, small intestine terminal ileum, skin not sun-exposed, and whole blood). Among them, as expected, we confirmed significant differences in the expression of transcripts encoded by the cytochromes in the liver, CYP2B6, CYP3A7, CYP3A5, and CYP1A1. Our systematic investigation on differences between male and female in the expression of drug response-related genes, reinforce the need to overcome the sex bias of clinical trials.

Prolonged B-cell depletion after rituximab in AQP4-IgG-positive neuromyelitis optica spectrum disorder.

Rituximab (a B-cell depleting monoclonal antibody) is increasingly utilized for treatment of different immune-mediated neurologic disorders, including aquaporin-4-IgG-positive neuromyelitis optica spectrum disorder (AQP4-IgG-NMOSD). After an initial treatment course, the drug is generally reinfused when peripheral blood B-cells levels re-increase >1% (usually after 6-12 months), or at fixed pre-planned 6-month intervals. We describe the unusual case of a 40-year-old woman with AQP4-IgG-NMOSD who showed a prolonged B-cell depletion for nearly five years after a single rituximab reinfusion. In similar rare patients with exceptionally long-lasting B-cell depletion, rituximab reinfusions at fixed pre-planned intervals would result in unnecessary treatment-related risks and health-care expenses.

Evolutionarily Selected Overexpression of the Cytokine BAFF Enhances Mucosal Immune Response Against P. falciparum.

We have previously shown that a variant of the TNFSF13B gene that we called BAFF-var increases the production of the cytokine BAFF, upregulating humoral immunity and increasing the risk for certain autoimmune diseases. In addition, genetic population signatures revealed that BAFF-var was evolutionarily advantageous, most likely by increasing resistance to malaria infection, which is a prime candidate for selective pressure. To evaluate whether the increased soluble BAFF (sBAFF) production confers protection, we experimentally assessed the role of BAFF-var in response to malaria antigens. Lysates of erythrocytes infected with Plasmodium falciparum (iRBCs) or left uninfected (uRBCs, control) were used to treat peripheral blood mononuclear cells (PBMCs) with distinct BAFF genotypes. The PBMCs purified from BAFF-var donors and treated with iRBCs showed different levels of specific cells, immunoglobulins, and cytokines as compared with BAFF-WT. In particular, a relevant differential effect on mucosal immunity B subpopulations have been observed. These findings point to specific immune cells and molecules through which the evolutionary selected BAFF-var may have improved fitness during P. falciparum infection.

Overview of the First 6 Months of Clinical Trials for COVID-19 Pharmacotherapy: The Most Studied Drugs.

SARS-CoV-2 rapidly spread from China until it was defined a pandemic by WHO in March 2020. Related scientific papers have rapidly extended information regarding the diagnosis, treatment and epidemiology of COVID-19 infection. To date, no vaccine or definitive treatment is available to defeat the virus and therapies are mainly based on existing drugs used to treat other conditions. Existing therapies used in several clinical trials work by affecting the biology of COVID-19 and/or counteracting the harmful host excessive immune response. Here, we have reviewed 526 ongoing clinical trials for COVID-19 to provide a perspective on the first 6 months of global efforts to identify an effective therapy. The drugs most actively tested in various centers include hydroxychloroquine, ritonavir, azithromycin, tocilizumab, lopinavir chloroquine and ivermectin. Our analysis shows that most clinical trials focus on a small number of candidate drugs (namely hydroxychloroquine and chloroquine representing 25% of total clinical trials) while underestimating the potential of other promising drugs. A global coordination in clinical trial management could avoid duplications and increase the effectiveness of the response to the global challenge.

Long Noncoding RNAs and Circular RNAs in Autoimmune Diseases.

Immune responses are essential for the clearance of pathogens and the repair of injured tissues; however, if these responses are not properly controlled, autoimmune diseases can occur. Autoimmune diseases (ADs) are a family of disorders characterized by the body's immune response being directed against its own tissues, with consequent chronic inflammation and tissue damage. Despite enormous efforts to identify new drug targets and develop new therapies to prevent and ameliorate AD symptoms, no definitive solutions are available today. Additionally, while substantial progress has been made in drug development for some ADs, most treatments only ameliorate symptoms and, in general, ADs are still incurable. Hundreds of genetic loci have been identified and associated with ADs by genome-wide association studies. However, the whole list of molecular factors that contribute to AD pathogenesis is still unknown. Noncoding (nc)RNAs, such as microRNAs, circular (circ)RNAs, and long noncoding (lnc)RNAs, regulate gene expression at different levels in various diseases, including ADs, and serve as potential drug targets as well as biomarkers for disease progression and response to therapy. In this review, we will focus on the potential roles and genetic regulation of ncRNA in four autoimmune diseases-systemic lupus erythematosus, rheumatoid arthritis, multiple sclerosis, and type 1 diabetes mellitus.

Direct-to-Consumer Nutrigenetics Testing: An Overview.

At present, specialized companies offering genetic testing services without the involvement of clinicians are growing; this development is a direct consequence of the significant decrease in genotyping and sequencing costs. Online companies offer predictions about the risk of developing complex diseases during one's life course, and they offer suggestions for personal lifestyle. Several companies have been created that provide nutrigenetics services; these companies suggest dietary indications-a central issue in the prevention and etiopathogenesis of specific diseases-based on one's personal genetic background. Dietary patterns are defined on the basis of a limited set of genetic markers. In this article, we analyze the online nutrigenetics services offered by 45 companies worldwide, to obtain an overall picture of the costs, the types of nutritional traits considered and the level of scientific precision of the services proposed. Our analysis clearly highlights the need for specific guidelines, in order to ensure a set of minimum quality standards for the nutrigenetics services offered to the customer.

Circadian timing of injury-induced cell proliferation in zebrafish.

In certain vertebrates such as the zebrafish, most tissues and organs including the heart and central nervous system possess the remarkable ability to regenerate following severe injury. Both spatial and temporal control of cell proliferation and differentiation is essential for the successful repair and re-growth of damaged tissues. Here, using the regenerating adult zebrafish caudal fin as a model, we have demonstrated an involvement of the circadian clock in timing cell proliferation following injury. Using a BrdU incorporation assay with a short labeling period, we reveal high amplitude daily rhythms in S-phase in the epidermal cell layer of the fin under normal conditions. Peak numbers of S-phase cells occur at the end of the light period while lowest levels are observed at the end of the dark period. Remarkably, immediately following amputation the basal level of epidermal cell proliferation increases significantly with kinetics, depending upon the time of day when the amputation is performed. In sharp contrast, we failed to detect circadian rhythms of S-phase in the highly proliferative mesenchymal cells of the blastema. Subsequently, during the entire period of outgrowth of the new fin, elevated, cycling levels of epidermal cell proliferation persist. Thus, our results point to a preferential role for the circadian clock in the timing of epidermal cell proliferation in response to injury.