Ovarian Aging: Role of Pituitary-Ovarian Axis Hormones and ncRNAs in Regulating Ovarian Mitochondrial Activity

The number of mitochondria in the oocyte along with their functions (e.g., energy production, scavenger activity) decline with age progression. Such multifaceted functions support several processes during oocyte maturation, ranging from energy supply to synthesis of the steroid hormones. Hence, it is hardly surprising that their impairment has been reported in both physiological and premature ovarian aging, wherein they are crucial players in the apoptotic processes that arise in aged ovaries. In any form, ovarian aging implies the progressive damage of the mitochondrial structure and activities as regards to ovarian germ and somatic cells. The imbalance in the circulating hormones and peptides (e.g., gonadotropins, estrogens, AMH, activins, and inhibins), active along the pituitary-ovarian axis, represents the biochemical sign of ovarian aging. Despite the progress accomplished in determining the key role of the mitochondria in preserving ovarian follicular number and health, their modulation by the hormonal signalling pathways involved in ovarian aging has been poorly and randomly explored. Yet characterizing this mechanism is pivotal to molecularly define the implication of mitochondrial dysfunction in physiological and premature ovarian aging, respectively. However, it is fairly difficult considering that the pathways associated with ovarian aging might affect mitochondria directly or by altering the activity, stability and localization of proteins controlling mitochondrial dynamics and functions, either unbalancing other cellular mediators, released by the mitochondria, such as non-coding RNAs (ncRNAs). We will focus on the mitochondrial ncRNAs (i.e., mitomiRs and mtlncRNAs), that retranslocate from the mitochondria to the nucleus, as active players in aging and describe their role in the nuclear-mitochondrial crosstalk and its modulation by the pituitary-ovarian hormone dependent pathways. In this review, we will illustrate mitochondria as targets of the signaling pathways dependent on hormones and peptides active along the pituitary/ovarian axis and as transducers, with a particular focus on the molecules retrieved in the mitochondria, mainly ncRNAs. Given their regulatory function in cellular activities we propose them as potential diagnostic markers and/or therapeutic targets.

Distinct profiles of immune cell populations underlie in-stent restenosis: a cluster analysis approach

Background In-stent restenosis (ISR) is a major challenge in patients with coronary artery disease due to its association with poor clinical outcomes, quality of life and costs. ISR etiopathogenesis remains unclear, but traditional risk factors cannot fully explain ISR burden. Inflammation-driven loss of endothelial homeostasis and neoatherosclerosis are thought to hallmark ISR. Recently, a number of immune cell subsets have been related to vascular repair failure and endothelial damage, such as angiogenic T-cells (Tang), endothelial progenitor cells (EPC), senescent T-cells (CD4+CD28null), monocyte subsets and low-density granulocytes (LDG). However, these subsets have not been studied in ISR and an integrative analysis is lacking. Purpose 1) to evaluate potential alterations in vascular repair and endothelial damage cellular mediators in ISR and 2) to identify profiles associated with clinical features. Methods Case-control study including 30 patients with ≥1 previous stent implantation (15 bare metal stents (BMS) and 15 drug-eluting stents (DE)) which suffered restenosis and 30 patients with ≥1 BMS without restenosis, both confirmed in a second angiogram performed by clinical symptoms >8 months after index procedure. Cellular mediators of vascular homeostasis were quantified by flow cytometry based on their surface markers in peripheral blood (EPC: CD34+VEGFR2+CD133+; EC: CD34-VEGFR+CD133-; Tang: CD3+CD31+CXCR4+; senescent T-cells: CD4+CD28null) or in peripheral blood mononuclear cells (monocyte subsets, ACE expression; total LDG: CD15+; and LDG subsets: CD15+CD14-CD16- and CD15+CD14lowCD16+). Results Patients with ISR exhibited decreased circulating Tang (p=0.005) and EPC (p<0.001), whereas CD4+CD28null and EC counts were higher (p<0.0001 and p=0.006) compared to ISR-free patients. No differences were observed in the frequency of monocyte subsets (all p>0.050), although ACE expression was found to be increased (non-classical p<0.001; and intermediate p<0.0001) in ISR. Moreover, no differences were noted in the total LDG population (p=0.092), but an increase in the CD14- compartment was observed in ISR (p=0.004). An unsupervised cluster analysis built with these subsets informed the presence of three profiles (Figure 1): group I (hallmarked by a profound impairment in vascular repair and augmented damage, suggestive of central haematopoiesis traits) exhibited an enhanced clinical risk profile compared to group II (hallmarked by a mid-altered vascular repair) and group III (hallmarked by CD16+ shifted LDG and ACE expression) (Figure 2). No differences were observed in stent types or traditional risk factors but hypertension. Conclusions Profound alterations in immune populations related to vascular repair and endothelial damage are found in ISR. Distinct cellular profiles can be distinguished within ISR, suggesting that different alterations may uncover different ISR clinical phenotypes, in terms of severity and extension. FUNDunding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): ISCIIIPrograma Intramural ISPA Figure 1 Figure 2

The blood–brain barrier in systemic infection and inflammation

The vascular blood–brain barrier is a highly regulated interface between the blood and brain. Its primary function is to protect central neurons while signaling the presence of systemic inflammation and infection to the brain to enable a protective sickness behavior response. With increasing degrees and duration of systemic inflammation, the vascular blood–brain barrier becomes more permeable to solutes, undergoes an increase in lymphocyte trafficking, and is infiltrated by innate immune cells; endothelial cell damage may occasionally occur. Perturbation of neuronal function results in the clinical features of encephalopathy. Here, the molecular and cellular anatomy of the vascular blood–brain barrier is reviewed, first in a healthy context and second in a systemic inflammatory context. Distinct from the molecular and cellular mediators of the blood–brain barrier’s response to inflammation, several moderators influence the direction and magnitude at genetic, system, cellular and molecular levels. These include sex, genetic background, age, pre-existing brain pathology, systemic comorbidity, and gut dysbiosis. Further progress is required to define and measure mediators and moderators of the blood–brain barrier’s response to systemic inflammation in order to explain the heterogeneity observed in animal and human studies.

In vivo Pooled Screening: A Scalable Tool to Study the Complexity of Aging and Age-Related Disease

Biological aging, and the diseases of aging, occur in a complex in vivo environment, driven by multiple interacting processes. A convergence of recently developed technologies has enabled in vivo pooled screening: direct administration of a library of different perturbations to a living animal, with a subsequent readout that distinguishes the identity of each perturbation and its effect on individual cells within the animal. Such screens hold promise for efficiently applying functional genomics to aging processes in the full richness of the in vivo setting. In this review, we describe the technologies behind in vivo pooled screening, including a range of options for delivery, perturbation and readout methods, and outline their potential application to aging and age-related disease. We then suggest how in vivo pooled screening, together with emerging innovations in each of its technological underpinnings, could be extended to shed light on key open questions in aging biology, including the mechanisms and limits of epigenetic reprogramming and identifying cellular mediators of systemic signals in aging.

Enhanced NF-κB signaling in type-2 dendritic cells at baseline predicts non-response to adalimumab in psoriasis

Biologic therapies have transformed the management of psoriasis, but clinical outcome is variable leaving an unmet clinical need for predictive biomarkers of response. Here we perform in-depth immunomonitoring of blood immune cells of 67 patients with psoriasis, before and during therapy with the anti-TNF drug adalimumab, to identify immune mediators of clinical response and evaluate their predictive value. Enhanced NF-κBp65 phosphorylation, induced by TNF and LPS in type-2 dendritic cells (DC) before therapy, significantly correlates with lack of clinical response after 12 weeks of treatment. The heightened NF-κB activation is linked to increased DC maturation in vitro and frequency of IL-17+ T cells in the blood of non-responders before therapy. Moreover, lesional skin of non-responders contains higher numbers of dermal DC expressing the maturation marker CD83 and producing IL-23, and increased numbers of IL-17+ T cells. Finally, we identify and clinically validate LPS-induced NF-κBp65 phosphorylation before therapy as a predictive biomarker of non-response to adalimumab, with 100% sensitivity and 90.1% specificity in an independent cohort. Our study uncovers important molecular and cellular mediators underpinning adalimumab mechanisms of action in psoriasis and we propose a blood biomarker for predicting clinical outcome.

Clinical Outcome Prediction in COVID-19 Patients by Lymphocyte Subsets Analysis and Monocytes’ iTNF-α Expression

In December 2019, a novel coronavirus, “SARS-CoV-2”, was recognized as the cause of coronavirus disease 2019 (COVID-19). Several studies have explored the changes and the role of inflammatory cells and cytokines in the immunopathogenesis of the disease, but until today, the results have been controversial. Based on these premises, we conducted a retrospective assessment of monocyte intracellular TNF-α expression (iTNF-α) and on the frequencies of lymphocyte sub-populations in twenty-five patients with moderate/severe COVID-19. We found lymphopenia in all COVID-19 infected subjects compared to healthy subjects. On initial observation, in patients with favorable outcomes, we detected a high absolute eosinophil count and a high CD4+/CD8+ T lymphocytes ratio, while in the Exitus Group, we observed high neutrophil and CD8+ T lymphocyte counts. During infection, in patients with favorable outcomes, we observed a rise in the lymphocyte count, in the monocyte and in Treg lymphocyte counts, and in the CD4+ and in CD8+ T lymphocytes count but a reduction in the CD4+/CD8+ T lymphocyte ratio. Instead, in the Exitus Group, we observed a reduction in the Treg lymphocyte counts and a decrease in iTNF-α expression. Our preliminary findings point to a modulation of the different cellular mediators of the immune system, which probably play a key role in the outcomes of COVID-19.

Improved post-stroke spontaneous recovery by astrocytic extracellular vesicles

Spontaneous recovery after a stroke accounts for a major part of the neurological recovery in patients. However limited, the spontaneous recovery is mechanistically driven by axonal restorative processes for which several molecular cues have been previously described. We report the acceleration of spontaneous recovery in a preclinical model of ischemia/reperfusion in rats via a single intracerebroventricular administration of extracellular vesicles released from primary cortical astrocytes. We used MRI, confocal and multiphoton microscopy to correlate the structural remodeling of the corpus callosum and striatocortical circuits with neurological performance over 21 days. We also evaluated the functionality of the corpus callosum by repetitive recordings of compound action potentials to show that the recovery facilitated by astrocytic extracellular vesicles was both anatomical and functional. Our data provide compelling evidence that astrocytes can hasten the basal recovery that naturally occurs post-stroke through the release of cellular mediators contained in extracellular vesicles.

Till Death Do Us Part—The Multifaceted Role of Platelets in Liver Diseases

Platelets are tightly connected with the liver, as both their production and their clearance are mediated by the liver. Platelets, in return, participate in a variety of liver diseases, ranging from non-alcoholic fatty liver diseases, (viral) hepatitis, liver fibrosis and hepatocellular carcinoma to liver regeneration. Due to their versatile functions, which include (1) regulation of hemostasis, (2) fine-tuning of immune responses and (3) release of growth factors and cellular mediators, platelets quickly adapt to environmental changes and modulate disease development, leading to different layers of complexity. Depending on the (patho)physiological context, platelets exert both beneficial and detrimental functions. Understanding the precise mechanisms through which platelet function is regulated at different stages of liver diseases and how platelets interact with various resident and non-resident liver cells helps to draw a clear picture of platelet-related therapeutic interventions. Therefore, this review summarizes the current knowledge on platelets in acute and chronic liver diseases and aims to shed light on how the smallest cells in the circulatory system account for changes in the (patho)physiology of the second largest organ in the human body.