Dysfunctional mitochondria elicit bioenergetic decline in the aged heart.

Aging represents a complex biological progression affecting the entire body, marked by a gradual decline in tissue function, rendering organs more susceptible to stress and diseases. The human heart holds significant importance in this context, as its aging process poses life-threatening risks. It entails macroscopic morphological shifts and biochemical changes that collectively contribute to diminished cardiac function. Among the numerous pivotal factors in aging, mitochondria play a critical role, intersecting with various molecular pathways and housing several aging-related agents. In this comprehensive review, we provide an updated overview of the functional role of mitochondria in cardiac aging.

ErbB2-NOTCH1 axis controls autophagy in cardiac cells.

Although the epidermal growth factor receptor 2 (ErbB2) and Notch1 signaling pathways have both significant roles in regulating cardiac biology, their interplay in the heart remains poorly investigated. Here, we present evidence of a crosstalk between ErbB2 and Notch1 in cardiac cells, with effects on autophagy and proliferation. Overexpression of ErbB2 in H9c2 cardiomyoblasts induced Notch1 activation in a post-transcriptional, p38-dependent manner, while ErbB2 inhibition with the specific inhibitor, lapatinib, reduced Notch1 activation. Moreover, incubation of H9c2 cells with lapatinib resulted in stalled autophagic flux and decreased proliferation, consistent with the established cardiotoxicity of this and other ErbB2-targeting drugs. Confirming the findings in H9c2 cells, exposure of primary neonatal mouse cardiomyocytes to exogenous neuregulin-1, which engages ErbB2, stimulated proliferation, and this effect was abrogated by concomitant inhibition of the enzyme responsible for Notch1 activation. Furthermore, the hearts of transgenic mice specifically overexpressing ErbB2 in cardiomyocytes had increased levels of active Notch1 and of Notch-related genes. These data expand the knowledge of ErbB2 and Notch1 functions in the heart and may allow better understanding the mechanisms of the cardiotoxicity of ErbB2-targeting cancer treatments.

Autophagy alterations in obesity, type 2 diabetes, and metabolic dysfunction-associated steatotic liver disease: the evidence from human studies.

Autophagy is an evolutionarily conserved process that plays a pivotal role in the maintenance of cellular homeostasis and its impairment has been implicated in the pathogenesis of various metabolic diseases including obesity, type 2 diabetes (T2D), and metabolic dysfunction-associated steatotic liver disease (MASLD). This review synthesizes the current evidence from human studies on autophagy alterations under these metabolic conditions. In obesity, most data point to autophagy upregulation during the initiation phase of autophagosome formation, potentially in response to proinflammatory conditions in the adipose tissue. Autophagosome formation appears to be enhanced under hyperglycemic or insulin-resistant conditions in patients with T2D, possibly acting as a compensatory mechanism to eliminate damaged organelles and proteins. Other studies have proposed that prolonged hyperglycemia and disrupted insulin signaling hinder autophagic flux, resulting in the accumulation of dysfunctional cellular components that can contribute to ß-cell dysfunction. Evidence from patients with MASLD supports autophagy inhibition in disease progression. Nevertheless, given the available data, it is difficult to ascertain whether autophagy is enhanced or suppressed in these conditions because the levels of autophagy markers depend on the overall metabolism of specific organs, tissues, experimental conditions, or disease duration. Owing to these constraints, determining whether the observed shifts in autophagic activity precede or result from metabolic diseases remains challenging. Additionally, autophagy-modulating strategies are shortly discussed. To conclude, more studies investigating autophagy impairment are required to gain a more comprehensive understanding of its role in the pathogenesis of obesity, T2D, and MASLD and to unveil novel therapeutic strategies for these conditions.

Germline BARD1 variants predispose to mesothelioma by impairing DNA repair and calcium signaling.

We report that ~1.8% of all mesothelioma patients and 4.9% of those younger than 55, carry rare germline variants of the BRCA1 associated RING domain 1 (BARD1) gene that were predicted to be damaging by computational analyses. We conducted functional assays, essential for accurate interpretation of missense variants, in primary fibroblasts that we established in tissue culture from a patient carrying the heterozygous BARD1V523A mutation. We found that these cells had genomic instability, reduced DNA repair, and impaired apoptosis. Investigating the underlying signaling pathways, we found that BARD1 forms a trimeric protein complex with p53 and SERCA2 that regulates calcium signaling and apoptosis. We validated these findings in BARD1-silenced primary human mesothelial cells exposed to asbestos. Our study elucidated mechanisms of BARD1 activity and revealed that heterozygous germline BARD1 mutations favor the development of mesothelioma and increase the susceptibility to asbestos carcinogenesis. These mesotheliomas are significantly less aggressive compared to mesotheliomas in asbestos workers.

Dysregulation of FLVCR1a-dependent mitochondrial calcium handling in neural progenitors causes congenital hydrocephalus.

Congenital hydrocephalus (CH), occurring in approximately 1/1,000 live births, represents an important clinical challenge due to the limited knowledge of underlying molecular mechanisms. The discovery of novel CH genes is thus essential to shed light on the intricate processes responsible for ventricular dilatation in CH. Here, we identify FLVCR1 (feline leukemia virus subgroup C receptor 1) as a gene responsible for a severe form of CH in humans and mice. Mechanistically, our data reveal that the full-length isoform encoded by the FLVCR1 gene, FLVCR1a, interacts with the IP3R3-VDAC complex located on mitochondria-associated membranes (MAMs) that controls mitochondrial calcium handling. Loss of Flvcr1a in mouse neural progenitor cells (NPCs) affects mitochondrial calcium levels and energy metabolism, leading to defective cortical neurogenesis and brain ventricle enlargement. These data point to defective NPCs calcium handling and metabolic activity as one of the pathogenetic mechanisms driving CH.

Exploring Mitochondrial Interactions with Pulsed Electromagnetic Fields: An Insightful Inquiry into Strategies for Addressing Neuroinflammation and Oxidative Stress in Diabetic Neuropathy.

Pulsed electromagnetic fields (PEMFs) are recognized for their potential in regenerative medicine, offering a non-invasive avenue for tissue rejuvenation. While prior research has mainly focused on their effects on bone and dermo-epidermal tissues, the impact of PEMFs on nervous tissue, particularly in the context of neuropathy associated with the diabetic foot, remains relatively unexplored. Addressing this gap, our preliminary in vitro study investigates the effects of complex magnetic fields (CMFs) on glial-like cells derived from mesenchymal cell differentiation, serving as a model for neuropathy of the diabetic foot. Through assessments of cellular proliferation, hemocompatibility, mutagenicity, and mitochondrial membrane potential, we have established the safety profile of the system. Furthermore, the analysis of microRNAs (miRNAs) suggests that CMFs may exert beneficial effects on cell cycle regulation, as evidenced by the upregulation of the miRNAs within the 121, 127, and 142 families, which are known to be associated with mitochondrial function and cell cycle control. This exploration holds promise for potential applications in mitigating neuropathic complications in diabetic foot conditions.

Thonzonium bromide inhibits progression of malignant pleural mesothelioma through regulation of ERK1/2 and p38 pathways and mitochondrial uncoupling.

BACKGROUND: Malignant Pleural Mesothelioma (MPM) is a rare malignancy with a poor prognosis. Current therapies are unsatisfactory and novel cures are urgently needed. In a previous drug screening, we identified thonzonium bromide (TB) as one of the most active compounds against MPM cells. Since the biological effects of TB are poorly known, in this work we departed from some hints of previous studies and investigated several hypotheses. Moreover, we evaluated the efficacy of TB in an in vivo xenograft rodent model. METHODS: In vitro assessment was made on five MPM (Mero-14, Mero-25, Ren, NCI-H28, MSTO-211H) and one SV40-immortalized mesothelial cell line (MeT-5A). We evaluated TB ability to affect proliferation, apoptosis, mitochondrial functions and metabolism, and the mevalonate pathway. In vivo assay was carried out on MPM-xenograft NOD-SCID mice (4 mg/kg delivered intraperitoneally, twice a week for 4 weeks) and the overall survival was analysed with Kaplan-Meier curves. RESULTS: After TB treatment, we observed the suppression of ERK 1/2 phosphorylation, the increase of BAX expression and p38 phosphorylation. TB affected Ca2+ homeostasis in both mitochondrial and cytosolic compartments, it regulated the mitochondrial functioning, respiration, and ATP production as well as the mevalonate pathway. The in vivo study showed an increased overall survival for TB treated group vs. vehicle control group (P = 0.0076). CONCLUSIONS: Both in vitro and in vivo results confirmed the effect of TB on MPM and unravelled novel targets with translational potential.

A tripartite organelle platform links growth factor receptor signaling to mitochondrial metabolism.

One open question in the biology of growth factor receptors is how a quantitative input (i.e., ligand concentration) is decoded by the cell to produce specific response(s). Here, we show that an EGFR endocytic mechanism, non-clathrin endocytosis (NCE), which is activated only at high ligand concentrations and targets receptor to degradation, requires a tripartite organelle platform involving the plasma membrane (PM), endoplasmic reticulum (ER) and mitochondria. At these contact sites, EGFR-dependent, ER-generated Ca2+ oscillations are sensed by mitochondria, leading to increased metabolism and ATP production. Locally released ATP is required for cortical actin remodeling and EGFR-NCE vesicle fission. The same biochemical circuitry is also needed for an effector function of EGFR, i.e., collective motility. The multiorganelle signaling platform herein described mediates direct communication between EGFR signaling and mitochondrial metabolism, and is predicted to have a broad impact on cell physiology as it is activated by another growth factor receptor, HGFR/MET.

Calcium signaling from sarcoplasmic reticulum and mitochondria contact sites in acute myocardial infarction.

Acute myocardial infarction (AMI) is a serious condition that occurs when part of the heart is subjected to ischemia episodes, following partial or complete occlusion of the epicardial coronary arteries. The resulting damage to heart muscle cells have a significant impact on patient's health and quality of life. About that, recent research focused on the role of the sarcoplasmic reticulum (SR) and mitochondria in the physiopathology of AMI. Moreover, SR and mitochondria get in touch each other through multiple membrane contact sites giving rise to the subcellular region called mitochondria-associated membranes (MAMs). MAMs are essential for, but not limited to, bioenergetics and cell fate. Disruption of the architecture of these regions occurs during AMI although it is still unclear the cause-consequence connection and a complete overview of the pathological changes; for sure this concurs to further damage to heart muscle. The calcium ion (Ca2+) plays a pivotal role in the pathophysiology of AMI and its dynamic signaling between the SR and mitochondria holds significant importance. In this review, we tried to summarize and update the knowledge about the roles of these organelles in AMI from a Ca2+ signaling point of view. Accordingly, we also reported some possible cardioprotective targets which are directly or indirectly related at limiting the dysfunctions caused by the deregulation of the Ca2+ signaling.

ER-mitochondria association negatively affects wound healing by regulating NLRP3 activation.

Methicillin-resistant Staphylococcus aureus (MRSA) is the most common causative agent of acute bacterial skin and skin-structure infections (ABSSSI), one of the major challenges to the health system worldwide. Although the use of antibiotics as the first line of intervention for MRSA-infected wounds is recommended, important side effects could occur, including cytotoxicity or immune dysregulation, thus affecting the repair process. Here, we show that the oxazolidinone antibiotic linezolid (LZD) impairs wound healing by aberrantly increasing interleukin 1 ß (IL-1ß) production in keratinocytes. Mechanistically, LZD triggers a reactive oxygen species (ROS)-independent mitochondrial damage that culminates in increased tethering between the endoplasmic reticulum (ER) and mitochondria, which in turn activates the NLR family pyrin domain-containing 3 (NLRP3) inflammasome complex by promoting its assembly to the mitochondrial surface. Downregulation of ER-mitochondria contact formation is sufficient to inhibit the LZD-driven NLRP3 inflammasome activation and IL-1ß production, restoring wound closure. These results identify the ER-mitochondria association as a key factor for NLRP3 activation and reveal a new mechanism in the regulation of the wound healing process that might be clinically relevant.

Circulating tumor DNA as a biomarker for predicting progression-free survival and overall survival in patients with epithelial ovarian cancer: a systematic review and meta-analysis.

OBJECTIVES: Circulating tumor DNA (ctDNA) is emerging as a potential prognostic biomarker in multiple tumor types. However, despite the many studies available on small series of patients with ovarian cancer, a recent systematic review and meta-analysis is lacking. The objective of this study was to determine the association of ctDNA with progression-free-survival and overall survival in patients with epithelial ovarian cancer. METHODS: An electronic search was conducted using PubMed (MEDLINE), Embase, CENTRAL (Cochrane Library), and CINAHL-Complete from January 2000 to September 15, 2023. To be included in the analysis the studies had to meet the following pre-specified inclusion criteria: (1) evaluable ctDNA; (2) progression-free-survival and overall survival reported as hazard ratio (HR); and (3) the patient population had epithelial ovarian cancer at the time of ctDNA detection. We evaluated the association of ctDNA with progression-free survival and overall survival. Secondary outcomes focused on sub-group analysis of genomic alterations and international Federation of Gynecology and Obstetrics (FIGO) stage. RESULTS: A total of 26 studies reporting on 1696 patients with epithelial ovarian cancer were included. The overall concordance rate between plasma-based and tissue-based analyses was approximately 62%. We found that a high level of ctDNA in epithelial ovarian cancer was associated with worse progression-free survival (HR 5.31, 95% CI 2.14 to 13.17, p<0.001) and overall survival (HR 2.98, 95% CI 1.86 to 4.76, p<0.0001). The sub-group analysis showed a greater than threefold increase in the risk of relapse in patients with positive HOXA9 meth-ctDNA (HR 3.84, 95% CI 1.57 to 9.41, p=0.003). CONCLUSIONS: ctDNA was significantly associated with worse progression-free survival and overall survival in patients with epithelial ovarian cancer. Further prospective studies are needed. PROSPERO REGISTRATION NUMBER: CRD42023469390.

The Wolfram-like variant WFS1E864K destabilizes MAM and compromises autophagy and mitophagy in human and mice.

Dominant variants in WFS1 (wolframin ER transmembrane glycoprotein), the gene coding for a mitochondria-associated endoplasmic reticulum (ER) membrane (MAM) resident protein, have been associated with Wolfram-like syndrome (WLS). In vitro and in vivo, WFS1 loss results in reduced ER to mitochondria calcium (Ca2+) transfer, mitochondrial dysfunction, and enhanced macroautophagy/autophagy and mitophagy. However, in the WLS pathological context, whether the mutant protein triggers the same cellular processes is unknown. Here, we show that in human fibroblasts and murine neuronal cultures the WLS protein WFS1E864K leads to decreases in mitochondria bioenergetics and Ca2+ uptake, deregulation of the mitochondrial quality system mechanisms, and alteration of the autophagic flux. Moreover, in the Wfs1E864K mouse, these alterations are concomitant with a decrease of MAM number. These findings reveal pathophysiological similarities between WS and WLS, highlighting the importance of WFS1 for MAM's integrity and functionality. It may open new treatment perspectives for patients with WLS.Abbreviations: BafA1: bafilomycin A1; ER: endoplasmic reticulum; HSPA9/GRP75: heat shock protein family A (Hsp70) member 9; ITPR/IP3R: inositol 1,4,5-trisphosphate receptor; MAM: mitochondria-associated endoplasmic reticulum membrane; MCU: mitochondrial calcium uniporter; MFN2: mitofusin 2; OCR: oxygen consumption rate; ROS: reactive oxygen species; ROT/AA: rotenone+antimycin A; VDAC1: voltage dependent anion channel 1; WLS: Wolfram-like syndrome; WS: Wolfram syndrome; WT: wild-type.

A mitochondrial NADPH-cholesterol axis regulates extracellular vesicle biogenesis to support hematopoietic stem cell fate.

Mitochondrial fatty acid oxidation (FAO) is essential for hematopoietic stem cell (HSC) self-renewal; however, the mechanism by which mitochondrial metabolism controls HSC fate remains unknown. Here, we show that within the hematopoietic lineage, HSCs have the largest mitochondrial NADPH pools, which are required for proper HSC cell fate and homeostasis. Bioinformatic analysis of the HSC transcriptome, biochemical assays, and genetic inactivation of FAO all indicate that FAO-generated NADPH fuels cholesterol synthesis in HSCs. Interference with FAO disturbs the segregation of mitochondrial NADPH toward corresponding daughter cells upon single HSC division. Importantly, we have found that the FAO-NADPH-cholesterol axis drives extracellular vesicle (EV) biogenesis and release in HSCs, while inhibition of EV signaling impairs HSC self-renewal. These data reveal the existence of a mitochondrial NADPH-cholesterol axis for EV biogenesis that is required for hematopoietic homeostasis and highlight the non-stochastic nature of HSC fate determination.

All-trans retinoic acid exhibits anti-proliferative and differentiating activity in Merkel cell carcinoma cells via retinoid pathway modulation.

BACKGROUND: The limited therapies available for treating Merkel cell carcinoma (MCC), a highly aggressive skin neoplasm, still pose clinical challenges, and novel treatments are required. Targeting retinoid signalling with retinoids, such as all-trans retinoic acid (ATRA), is a promising and clinically useful antitumor approach. ATRA drives tumour cell differentiation by modulating retinoid signalling, leading to anti-proliferative and pro-apoptotic effects. Although retinoid signalling is dysregulated in MCC, ATRA activity in this tumour is unknown. This study aimed to evaluate the impact of ATRA on the pathological phenotype of MCC cells. METHODS: The effect of ATRA was tested in various Merkel cell polyomavirus-positive and polyomavirus-negative MCC cell lines in terms of cell proliferation, viability, migration and clonogenic abilities. In addition, cell cycle, apoptosis/cell death and the retinoid gene signature were evaluated upon ATRA treatments. RESULTS: ATRA efficiently impaired MCC cell proliferation and viability in MCC cells. A strong effect in reducing cell migration and clonogenicity was determined in ATRA-treated cells. Moreover, ATRA resulted as strongly effective in arresting cell cycle and inducing apoptosis/cell death in all tested MCC cells. Enrichment analyses indicated that ATRA was effective in modulating the retinoid gene signature in MCC cells to promote cell differentiation pathways, which led to anti-proliferative and pro-apoptotic/cell death effects. CONCLUSIONS: These results underline the potential of retinoid-based therapy for MCC management and might open the way to novel experimental approaches with other retinoids and/or combinatorial treatments.

Link between organic nanovescicles from vegetable kingdom and human cell physiology: intracellular calcium signalling.

BACKGROUND: Plant-derived nanovesicles (PDNVs) are a novelty in medical and agrifood environments, with several studies exploring their functions and potential applications. Among fruits, apples (sp. Malus domestica) have great potential as PDNVs source, given their widespread consumption, substantial waste production, and recognized health benefits. Notably, apple-derived nanovesicles (ADNVs) can interact with human cell lines, triggering anti-inflammatory and antioxidant responses. This work is dedicated to the comprehensive biochemical characterization of apple-derived nanovesicles (ADNVs) through proteomic and lipidomic analysis, and small RNAs sequencing. This research also aims to shed light on the underlying mechanism of action (MOA) when ADNVs interface with human cells, through observation of intracellular calcium signalling in human fibroblasts, and to tackles differences in ADNVs content when isolated from fruits derived from integrated and organic production methods cultivars. RESULTS: The ADNVs fraction is mainly composed of exocyst-positive organelles (EXPOs) and MVB-derived exosomes, identified through size and molecular markers (Exo70 and TET-3-like proteins). ADNVs' protein cargo is heterogeneous and exhibits a diverse array of functions, especially in plant's protection (favouring ABA stress-induced signalling, pathogen resistance and Reactive Oxygen Species (ROS) metabolism). Noteworthy plant miRNAs also contribute to phytoprotection. In relation with human cells lines, ADNVs elicit spikes of intracellular Ca2+ levels, utilizing the cation as second messenger, and produce an antioxidant effect. Lastly, organic samples yield a substantial increase in ADNV production and are particularly enriched in bioactive lysophospholipids. CONCLUSIONS: We have conclusively demonstrated that ADNVs confer an antioxidant effect upon human cells, through the initiation of a molecular pathway triggered by Ca2+ signalling. Within ADNVs, a plethora of bioactive proteins, small RNAs, and lipids have been identified, each possessing well-established functions within the realm of plant biology. While ADNVs predominantly function in plants, to safeguard against pathogenic agents and abiotic stressors, it is noteworthy that proteins with antioxidant power might act as antioxidants within human cells.

Oxidative stress in metabolic dysfunction-associated steatotic liver disease (MASLD): How does the animal model resemble human disease?

Despite decades of research, the pathogenesis of metabolic dysfunction-associated steatotic liver disease (MASLD) is still not completely understood. Based on the evidence from preclinical models, one of the factors proposed as a main driver of disease development is oxidative stress. This study aimed to search for the resemblance between the profiles of oxidative stress and antioxidant defense in the animal model of MASLD and the group of MASLD patients. C57BL/6J mice were fed with the Western diet for up to 24 weeks and served as the animal model of MASLD. The antioxidant profile of mice hepatic tissue was determined by liquid chromatography-MS3 spectrometry (LC-MS/MS). The human cohort consisted of 20 patients, who underwent bariatric surgery, and 6 controls. Based on histological analysis, 4 bariatric patients did not have liver steatosis and as such were also classified as controls. Total antioxidant activity was measured in sera and liver biopsy samples. The hepatic levels of antioxidant enzymes and oxidative damage were determined by Western Blot. The levels of antioxidant enzymes were significantly altered in the hepatic tissue of mice with MASLD. In contrast, there were no significant changes in the antioxidant profile of hepatic tissue of MASLD patients, except for the decreased level of carbonylated proteins. Decreased protein carbonylation together with significant correlations between the thioredoxin system and parameters describing metabolic health suggest alterations in the thiol-redox signaling. Altogether, these data show that even though the phenotype of mice closely resembles human MASLD, the animal-to-human translation of cellular and molecular processes such as oxidative stress may be more challenging.

Molecular mechanisms of naringenin modulation of mitochondrial permeability transition acting on F1FO-ATPase and counteracting saline load-induced injury in SHRSP cerebral endothelial cells.

Naringenin (NRG) was characterized for its ability to counteract mitochondrial dysfunction which is linked to cardiovascular diseases. The F1FO-ATPase can act as a molecular target of NRG. The interaction of NRG with this enzyme can avoid the energy transmission mechanism of ATP hydrolysis, especially in the presence of Ca2+ cation used as cofactor. Indeed, NRG was a selective inhibitor of the hydrophilic F1 domain displaying a binding site overlapped with quercetin in the inside surface of an annulus made by the three α and the three ß subunits arranged alternatively in a hexamer. The kinetic constant of inhibition suggested that NRG preferred the enzyme activated by Ca2+ rather than the F1FO-ATPase activated by the natural cofactor Mg2+. From the inhibition type mechanism of NRG stemmed the possibility to speculate that NRG can prevent the activation of F1FO-ATPase by Ca2+. The event correlated to the protective role in the mitochondrial permeability transition pore opening by NRG as well as to the reduction of ROS production probably linked to the NRG chemical structure with antioxidant action. Moreover, in primary cerebral endothelial cells (ECs) obtained from stroke prone spontaneously hypertensive rats NRG had a protective effect on salt-induced injury by restoring cell viability and endothelial cell tube formation while also rescuing complex I activity.

Spiropiperidine-Based Oligomycin-Analog Ligands To Counteract the Ischemia-Reperfusion Injury in a Renal Cell Model.

Finding a therapy for ischemia-reperfusion injury, which consists of cell death following restoration of blood flowing into the artery affected by ischemia, is a strong medical need. Nowadays, only the use of broad-spectrum molecular therapies has demonstrated a partial efficacy in protecting the organs following reperfusion, while randomized clinical trials focused on more specific drug targets have failed. In order to overcome this problem, we applied a combination of molecular modeling and chemical synthesis to identify novel spiropiperidine-based structures active in mitochondrial permeability transition pore opening inhibition as a key process to enhance cell survival after blood flow restoration. Our results were confirmed by biological assay on an in vitro cell model on HeLa and human renal proximal tubular epithelial cells and pave the way to further investigation on an in vivo model system.

Tofacitinib restores psoriatic arthritis fibroblast-like synoviocytes function via autophagy and mitochondrial quality control modulation.

OBJECTIVES: To evaluate the in vitro effect of tofacitinib on autophagy activity of psoriatic arthritis (PsA) fibroblast-like synoviocytes (FLS), and to confirm its activity on inflammatory and invasive properties of FLS and synovial cells, deepening the impact on mitochondrial function. METHODS: FLS, peripheral blood mononuclear cells (PBMCs), and synovial cells from active PsA patients were cultured with tofacitinib 1 µM or vehicle control for 24 h. Autophagy was measured by Western blot and by fluorescence microscopy. Chemokines/cytokines released into culture supernatants were quantified by ELISA, while invasive properties of FLS by migration assays. Specific mitochondrial probes were adopted to measure intracellular reactive oxygen species (ROS), mitochondrial potential, morphology, turnover and mitophagy. Oxygen consumption rate (OCR), reflecting oxidative phosphorylation, was quantified using the Seahorse technology. Differences were determined by adopting the non-parametric Wilcoxon signed rank test. RESULTS: 18 patients with moderately-to-severely active PsA were enrolled. Tofacitinib significantly increased the levels of the autophagy markers LC3-II and ATG7 in PsA FLS compared to vehicle control, suggesting an increase in spontaneous autophagy activity; no effect was highlighted in PBMCs and synovial cells cultures. Tofacitinib reduced migration properties of PsA FLS, and reduced MCP-1 and IL-6 release into FLS and synovial cells cultures supernatants. Furthermore, tofacitinib decreased intracellular ROS production, increased basal OCR, ATP production and maximal respiratory capacity, and enhanced mitophagy and mitochondrial turnover. CONCLUSIONS: The JAK inhibitor tofacitinib reduces the pro-invasive and pro-inflammatory properties of PsA FLS. Autophagy induction and mitochondrial quality control modulation by tofacitinib might contribute to FLS function restoration.

Hsa-microRNA-1249-3p/Homeobox A13 axis modulates the expression of ß-catenin gene in human epithelial cells.

Intercellular adhesion is a key function for epithelial cells. The fundamental mechanisms relying on epithelial cell adhesion have been partially uncovered. Hsa-microRNA-1249-3p (hsa-miR-1249-3p) plays a role in the epithelial mesenchymal transition in carcinoma cells, but its physiological function in epithelial cells is unknown. We aimed to investigate the role and molecular mechanisms of hsa-miR-1249-3p on epithelial cell functions. Hsa-miR-1249-3p was overexpressed in human epithelial cells and uterine cervical tissues, compared to cervical carcinoma cells and precancerous tissues, respectively. Hsa-miR-1249-3p was analyzed to verify its regulatory function on Homeobox A13 (HOXA13) target gene and its downstream cell adhesion gene ß-catenin. Functional experiments indicated that hsa-miR-1249-3p inhibition prompted the mRNA and protein overexpression of HOXA13 which, in turn, led to the ß-catenin protein expression. Moreover, hsa-miR-1249-3p inhibition induced a strong colony forming ability in epithelial cells, suggesting the miR involvement in cell adhesion machinery. These data indicate that hsa-miR-1249-3p regulates the expression of HOXA13 and its downstream cell adhesion gene ß-catenin, possible resulting in cell adhesion modification in epithelial cells. This study will allow the set-up of further investigations aimed at exploring the relationship between the hsa-miR-1249-3p/HOXA13 axis and downstream cell adhesion genes.