Profound architectural and functional readjustments of the secretory pathway in decidualization of endometrial stromal cells.

Certain cell types must expand their exocytic pathway to guarantee efficiency and fidelity of protein secretion. A spectacular case is offered by decidualizing human endometrial stromal cells (EnSCs). In the midluteal phase of the menstrual cycle, progesterone stimulation induces proliferating EnSCs to differentiate into professional secretors releasing proteins essential for efficient blastocyst implantation. Here, we describe the architectural rearrangements of the secretory pathway of a human EnSC line (TERT-immortalized human endometrial stromal cells (T-HESC)). As in primary cells, decidualization entails proliferation arrest and the coordinated expansion of the entire secretory pathway without detectable activation of unfolded protein response (UPR) pathways. Decidualization proceeds also in the absence of ascorbic acid, an essential cofactor for collagen biogenesis, despite also the secretion of some proteins whose folding does not depend on vitamin C is impaired. However, even in these conditions, no overt UPR induction can be detected. Morphometric analyses reveal that the exocytic pathway does not increase relatively to the volume of the cell. Thus, differently from other cell types, abundant production is guaranteed by a coordinated increase of the cell size following arrest of proliferation.

Heparin Protects Human Neural Progenitor Cells from Zika Virus-Induced Cell Death While Preserving Their Differentiation into Mature Neuroglial Cells.

Zika virus (ZIKV) is an arbovirus member of the Flaviviridae family that causes severe congenital brain anomalies in infected fetuses. The key target cells of ZIKV infection, human neural progenitor cells (hNPCs), are highly permissive to infection that causes the inhibition of cell proliferation and induces cell death. We have previously shown that pharmaceutical-grade heparin inhibits virus-induced cell death with negligible effects on in vitro virus replication in ZIKV-infected hNPCs at the "high" multiplicity of infection (MOI) of 1. Here, we show that heparin inhibits formation of ZIKV-induced intracellular vacuoles, a signature of paraptosis, and inhibits necrosis and apoptosis of hNPCs grown as neurospheres (NS). To test whether heparin preserved the differentiation of ZIKV-infected hNPCs into neuroglial cells, hNPCs were infected at the MOI of 0.001. In this experimental condition, heparin inhibited ZIKV replication by ca. 2 log10, mostly interfering with virion attachment, while maintaining its protective effect against ZIKV-induced cytopathicity. Heparin preserved differentiation into neuroglial cells of hNPCs that were obtained from either human-induced pluripotent stem cells (hiPSC) or by fetal tissue. Quite surprisingly, multiple additions of heparin to hNPCs enabled prolonged virus replication while preventing virus-induced cytopathicity. Collectively, these results highlight the potential neuroprotective effect of heparin that could serve as a lead compound to develop novel agents for preventing the damage of ZIKV infection on the developing brain. IMPORTANCE ZIKV is a neurotropic virus that invades neural progenitor cells (NPCs), causing inhibition of their proliferation and maturation into neurons and glial cells. We have shown previously that heparin, an anticoagulant also used widely during pregnancy, prevents ZIKV-induced cell death with negligible inhibition of virus replication. Here, we demonstrate that heparin also exerts antiviral activity against ZIKV replication using a much lower infectious inoculum. Moreover, heparin interferes with different modalities of virus-induced cell death. Finally, heparin-induced prevention of virus-induced NPC death allows their differentiation into neuroglial cells despite the intracellular accumulation of virions. These results highlight the potential use of heparin, or pharmacological agents derived from it, in pregnant women to prevent the devastating effects of ZIKV infection on the developing brain of their fetuses.

Promoting exogenous repair in multiple sclerosis: myelin regeneration.

PURPOSE OF THE REVIEW: Despite the significant progress in the development of disease-modifying treatments for multiple sclerosis (MS), repair of existing damage is still poorly addressed. Current research focuses on stem cell-based therapies as a suitable alternative or complement to current drug therapies. RECENT FINDINGS: Myelin damage is a hallmark of multiple sclerosis, and novel approaches leading to remyelination represent a promising tool to prevent neurodegeneration of the underlying axon. With increasing evidence of diminishing remyelination capacity of the MS brain with ageing and disease progression, exogenous cell transplantation is a promising therapeutic approach for restoration of oligodendrocyte precursor cell pool reserve and myelin regeneration. SUMMARY: The present review summarizes recent developments of remyelinating therapies in multiple sclerosis, focusing on exogenous cell-based strategies and discussing related scientific, practical, and ethical concerns.

The cannabinoid receptor CB1 contributes to the development of ectopic lesions in a mouse model of endometriosis.

STUDY QUESTION: Does signaling via the cannabinoid (CB1) receptor play a role in the pathogenesis of endometriosis in a mouse model? SUMMARY ANSWER: Mice treated with a CB1 agonist developed larger ectopic lesions, while less severe lesions developed in the absence of functional CB1 expression. WHAT IS KNOWN ALREADY: The expression of components of the endocannabinoid system has been demonstrated in both mouse and human uteri. CB1 receptors are expressed in human epithelial and stromal cell lines derived from eutopic endometrium and deep infiltrating endometriosis nodules. STUDY DESIGN, SIZE, DURATION: This was a randomized study in a mouse model of endometriosis. In a first set of experiments, mice with endometriosis were treated with the CB1 receptor agonist methanandamide (MET) (5 mg/kg, n = 20) on Days 1-5 and 8-12. In a second set of experiments, endometriosis development was evaluated in CB1-/- mice and in their wild-type (WT) littermates. PARTICIPANTS/MATERIALS, SETTING, METHODS: Endometriosis-like lesions were induced in Balb/c and C57/Bl6 mice. Two weeks after disease induction, the lesions were counted, measured and either included for immunohistochemistry analysis or frozen for gene expression profiling by semi-quantitative real-time PCR. To limit the role of chance, the experiments were conducted under standardized laboratory conditions with appropriate controls. MAIN RESULTS AND THE ROLE OF CHANCE: The lesion total volume was significantly higher in MET-treated compared with vehicle-treated mice (P < 0.05). Expression levels of mRNA for survivin, N-cadherin, integrin ß1 and interleukin-6 were increased in the ectopic endometrium of MET-treated versus vehicle-treated mice (P < 0.05). CB1-/- recipients that received endometrial tissue fragments from CB1-/- donors, WT recipients that received endometrial tissue fragments from CB1-/- donors and CB1-/- recipients that received endometrial tissue fragments from WT donors all showed a significant reduction in total lesion volume and lower expression of survivin and N-cadherin compared with WT recipients receiving uterine fragments from WT donors (P < 0.05). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: We provide evidence that endocannabinoid signaling via CB1 receptor plays a role in the development of endometriosis in a mouse model. However, the relative contribution of the CB1-mediated signaling pathways active in inflammatory, uterine and peritoneal cells remains to be ascertained. Since the study was performed in a mouse model, the significance of the findings in the human system warrants further investigation. WIDER IMPLICATIONS OF THE FINDINGS: Clarifying the function and regulation of CB1 and its molecular interactions with endogenous ligands, and how endocannabinoids levels are regulated in women with endometriosis, represent critical areas of research for the potential development of a novel medical treatment of the disease. STUDY FUNDING/COMPETING INTERESTS: A.M.S. was supported by a fellowship from Fondazione Giorgio Pardi. The authors have no conflicts of interest to declare.

Bacterial sensor triggering receptor expressed on myeloid cells-2 regulates the mucosal inflammatory response.

BACKGROUND AND AIMS: Triggering receptor expressed on myeloid cells (TREM)-2 is a surface receptor detected on macrophages, dendritic cells, and microglia that binds repeated anionic motifs on yeast and Gram-positive and Gram-negative bacteria. Little is known about TREM-2 expression and function in the intestine or its role in inflammatory bowel disease (IBD). We investigated the expression of TREM-2 in the intestinal lamina propria and its role in the development of colonic inflammation. METHODS: We measured levels of TREM-2 in lamina propria mononuclear cells from surgical specimens collected from patients with IBD or cancer (controls). We analyzed the development of colitis in TREM-2 knockout and wild-type mice. Colon samples were isolated from mice and analyzed for cytokine expression, phagocytosis of bacteria, proliferation in colonic crypts, lamina propria mononuclear cell function, and T-cell activation by ovalbumin. RESULTS: TREM-2 was virtually absent from colon samples of control patients, but levels were significantly higher in within the inflamed mucosa of patients with IBD; it was mainly expressed by CD11c(+) cells. Levels of TREM-2 increased as acute or chronic colitis was induced in mice. TREM-2 knockout mice developed less severe colitis than wild-type mice; the knockout mice lost less body weight, had a lower disease activity index, and had smaller mucosal lesions in endoscopic analysis. Colon dendritic cells from TREM-2 knockout mice produced lower levels of inflammatory cytokines and had reduced levels of bacterial killing and T-cell activation than cells from wild-type mice. CONCLUSIONS: TREM-2 contributes to mucosal inflammation during development of colitis in mice. Levels of TREM-2 are increased within the inflamed mucosa of patients with IBD, indicating its potential as a therapeutic target.

Iron availability is increased in individual human ovarian follicles in close proximity to an endometrioma compared with distal ones.

STUDY QUESTION: Does the iron content of an endometrioma represent a potential source of toxicity for the adjacent follicles? SUMMARY ANSWER: The presence of an endometrioma increases iron and H/L ferritin levels, and transferrin receptor (TfR1) mRNA in individual follicles proximal to the endometrioma and is accompanied by reduced oocyte retrieval. WHAT IS KNOWN ALREADY: Levels of free iron in endometriotic ovarian cysts are much higher than those in normal serum or in non-endometriotic ovarian cysts. The presence of an endometrioma exerts a detrimental effect on the surrounding healthy ovarian tissue as reflected by a reduced number of developing follicles and oocytes retrieved in IVF cycles. STUDY DESIGN, SIZE, DURATION: This is a research study with prospective collection and evaluation of individual follicles (follicular fluid and luteinized granulosa cells) from the affected and the healthy ovaries of 13 women with unilateral endometrioma. PARTICIPANTS/MATERIALS, SETTING, METHODS: Individual follicular samples (145) were obtained from 13 women with endometriosis-related infertility undergoing IVF-ICSI procedures from May 2012 to March 2013. All women had unilateral endometrioma not previously treated with surgery; the contralateral ovary was free of endometriomas and previous surgery. The average ± SEM age was 35.36 ± 2.5 years with anti-Mullerian hormone levels of 2.03 ± 0.55 ng/ml. Follicles were classified as: (i) proximal follicles, in physical contact with the endometrioma; (ii) distal follicles, present in the affected ovary but not in close contact with the endometrioma and (iii) contralateral follicles, in the contralateral healthy ovary. Iron content was measured by the FerroZine method. H/L ferritin subunits were evaluated by specific enzyme-linked immunosorbant assays. Expression of H ferritin and TfR1 was examined by semi-quantitative RT-PCR. Oocyte retrieval rates and Day 3 embryo quality were analyzed. MAIN RESULTS AND THE ROLE OF CHANCE: Total iron levels were higher in endometrioma-proximal follicles compared with endometrioma-distal ones (P = 0.009) and to follicles in the healthy ovary (P = 0.02). L ferritin was higher in proximal versus distal follicles (P = 0.044) or follicles from the healthy ovary (P = 0.027). H ferritin was higher in the proximal and distal follicles compared with follicles in the healthy ovary (P = 0.042 and P = 0.0067, respectively). H ferritin transcript levels in granulosa cells were higher in proximal follicles versus follicles from healthy ovary (P = 0.02). TfR1 transcript levels were higher in proximal versus distal follicles (P = 0.03) and versus follicles from the healthy ovary (P = 0.04). The oocyte retrieval rate was lower in proximal and distal follicles than in follicles from the healthy ovary (P = 0.001 and P = 0.04, respectively). LIMITATIONS, REASONS FOR CAUTION: This is a study on a relatively small sample size and confirmation in a larger group of patients may be required. The method used to purify luteinized granulosa cells offers the best combination of purity, viability and total number of cells recovered. However, a minor contamination by CD45(+) cells (<5%) cannot be excluded. WIDER IMPLICATIONS OF THE FINDINGS: This study represents a further in-depth analysis of the toxic influence of the endometrioma content on the surrounding follicles. We demonstrate the presence of iron-related compounds that are potentially toxic to developing ovarian follicles adjacent to the endometrioma during IVF. Our findings provide novel information that suggests that when surgical removal of the endometrioma is not the option, follicle aspiration at sites distant from the endometrioma might increase the probability of retrieving oocytes. STUDY FUNDING/COMPETING INTEREST(S): This project was supported by Fondazione Giorgio Pardi, Milan, Italy. The authors have no competing financial interests in relation to the content of this research paper. TRIAL REGISTRATION NUMBER: NA.

An Italian association study and meta-analysis with previous GWAS confirm WNT4, CDKN2BAS and FN1 as the first identified susceptibility loci for endometriosis.

BACKGROUND: Although endometriosis may benefit from primary prevention measures, the epidemiological risk factors identified are equivocal. Two genome-wide association studies (GWAS) have been conducted for endometriosis in two different ethnic populations but results are still to be replicated consistently and across various ethnicities. To confirm the association of GWAS-derived susceptibility loci, we conducted a replication Italian case-control study and a meta-analysis. METHODS: An independent set of 305 laparoscopically-proven endometriosis patients and 2710 controls were recruited. Four SNPs-CDKN2BAS rs1333049, rs7521902 close to WNT4, rs12700667 in an inter-genic region on 7p15.2 and fibronectin 1 rs1250248-were selected for this association study. RESULTS: Rs1333049 risk allele G frequency resulted significantly higher in endometriosis patients compared with controls (OR 1.32, 95% CI 1.11 to 1.57), confirming the role of this locus also in the Caucasian population. The meta-analysis showed that rs7521902 was associated with endometriosis at a genome-wide significance (p(meta)=2.23×10(-9)) while for rs1250248, a genome-wide significant p(meta) value of 3.89×10(-9) was detected only in association with severe forms. An epistatic interaction between rs7521902 and rs1250248 (OR 1.56, p=1.19×10(-2)) was found especially in presence of ovarian disease (OR=2.15, p=3.12×10(-4)). CONCLUSIONS: We confirm WNT4, CDKN2BAS and FN1 as the first identified common loci for endometriosis.

Endogenous neural stem cells characterization using omics approaches: Current knowledge in health and disease.

Neural stem cells (NSCs), an invaluable source of neuronal and glial progeny, have been widely interrogated in the last twenty years, mainly to understand their therapeutic potential. Most of the studies were performed with cells derived from pluripotent stem cells of either rodents or humans, and have mainly focused on their potential in regenerative medicine. High-throughput omics technologies, such as transcriptomics, epigenetics, proteomics, and metabolomics, which exploded in the past decade, represent a powerful tool to investigate the molecular mechanisms characterizing the heterogeneity of endogenous NSCs. The transition from bulk studies to single cell approaches brought significant insights by revealing complex system phenotypes, from the molecular to the organism level. Here, we will discuss the current literature that has been greatly enriched in the "omics era", successfully exploring the nature and function of endogenous NSCs and the process of neurogenesis. Overall, the information obtained from omics studies of endogenous NSCs provides a sharper picture of NSCs function during neurodevelopment in healthy and in perturbed environments.

Heparin Precursors with Reduced Anticoagulant Properties Retain Antiviral and Protective Effects That Potentiate the Efficacy of Sofosbuvir against Zika Virus Infection in Human Neural Progenitor Cells.

Zika virus (ZIKV) infection during pregnancy can result in severe birth defects, such as microcephaly, as well as a range of other related health complications. Heparin, a clinical-grade anticoagulant, is shown to protect neural progenitor cells from death following ZIKV infection. Although heparin can be safely used during pregnancy, it retains off-target anticoagulant effects if directly employed against ZIKV infection. In this study, we investigated the effects of chemically modified heparin derivatives with reduced anticoagulant activities. These derivatives were used as experimental probes to explore the structure-activity relationships. Precursor fractions of porcine heparin, obtained during the manufacture of conventional pharmaceutical heparin with decreased anticoagulant activities, were also explored. Interestingly, these modified heparin derivatives and precursor fractions not only prevented cell death but also inhibited the ZIKV replication of infected neural progenitor cells grown as neurospheres. These effects were observed regardless of the specific sulfation position or overall charge. Furthermore, the combination of heparin with Sofosbuvir, an antiviral licensed for the treatment of hepatitis C (HCV) that also belongs to the same Flaviviridae family as ZIKV, showed a synergistic effect. This suggested that a combination therapy approach involving heparin precursors and Sofosbuvir could be a potential strategy for the prevention or treatment of ZIKV infections.

Potency assays and biomarkers for cell-based advanced therapy medicinal products.

Advanced Therapy Medicinal Products (ATMPs) based on somatic cells expanded in vitro, with or without genetic modification, is a rapidly growing area of drug development, even more so following the marketing approval of several such products. ATMPs are produced according to Good Manufacturing Practice (GMP) in authorized laboratories. Potency assays are a fundamental aspect of the quality control of the end cell products and ideally could become useful biomarkers of efficacy in vivo. Here we summarize the state of the art with regard to potency assays used for the assessment of the quality of the major ATMPs used clinic settings. We also review the data available on biomarkers that may substitute more complex functional potency tests and predict the efficacy in vivo of these cell-based drugs.

Association of Circulating Antiretinal Antibodies With Clinical Outcomes in Retinitis Pigmentosa.

Purpose: To determine if circulating antiretinal antibodies (ARAs) differ between patients affected by retinitis pigmentosa (RP) and control participants and to assess whether ARAs are associated with clinical outcomes in patients with RP. Methods: Cross-sectional study involving a group of patients clinically diagnosed with RP and a control group of healthy participants. Serum autoantibodies against enolase, heat shock protein 70 (HSP70), and carbonic anhydrase II (CAII) were tested in all participants using Jess capillary Western blot. We compared ARA prevalence between the RP and control groups and investigated the association of serum ARA positivity with macular edema and vitreomacular disorders in patients affected by RP. Results: Thirty-six patients affected by RP and a control group of 39 healthy individuals were included. Overall, at least one ARA positivity was detected in 89% and 80% of participants in the RP and control groups, respectively. We observed a similar prevalence of anti-CAII and anti-enolase ARA between patients and controls (P = 0.87 and P = 0.35, respectively). Sera from patients with RP tested positive for anti-HSP70 ARAs more frequently than those from controls (53% vs. 36%), albeit without reaching statistical significance (P = 0.29). Among the 72 eyes with RP, 25% presented with macular edema (most often bilateral) and 33% with epiretinal membrane and/or lamellar macular hole. None of the three ARAs was associated with an increased risk of any macular complications in eyes affected by RP (all P > 0.05). Conclusions: The prevalence of circulating ARAs against enolase, HSP70, and CAII is similar between patients affected by RP and healthy individuals. Our results provide evidence against the association of ARAs with macular edema and vitreomacular interface disorders in RP.

Neural stem cell transplantation in patients with progressive multiple sclerosis: an open-label, phase 1 study.

Innovative pro-regenerative treatment strategies for progressive multiple sclerosis (PMS), combining neuroprotection and immunomodulation, represent an unmet need. Neural precursor cells (NPCs) transplanted in animal models of multiple sclerosis have shown preclinical efficacy by promoting neuroprotection and remyelination by releasing molecules sustaining trophic support and neural plasticity. Here we present the results of STEMS, a prospective, therapeutic exploratory, non-randomized, open-label, single-dose-finding phase 1 clinical trial ( NCT03269071 , EudraCT 2016-002020-86), performed at San Raffaele Hospital in Milan, Italy, evaluating the feasibility, safety and tolerability of intrathecally transplanted human fetal NPCs (hfNPCs) in 12 patients with PMS (with evidence of disease progression, Expanded Disability Status Scale ≥6.5, age 18-55 years, disease duration 2-20 years, without any alternative approved therapy). The safety primary outcome was reached, with no severe adverse reactions related to hfNPCs at 2-year follow-up, clearly demonstrating that hfNPC therapy in PMS is feasible, safe and tolerable. Exploratory secondary analyses showed a lower rate of brain atrophy in patients receiving the highest dosage of hfNPCs and increased cerebrospinal fluid levels of anti-inflammatory and neuroprotective molecules. Although preliminary, these results support the rationale and value of future clinical studies with the highest dose of hfNPCs in a larger cohort of patients.

The selective vitamin D receptor agonist, elocalcitol, reduces endometriosis development in a mouse model by inhibiting peritoneal inflammation.

BACKGROUND: Endometriosis, which is characterized by the growth of endometrial tissue at ectopic locations as well as vascular development and inflammation, is still an unmet clinical need since an optimal drug that allows for both pain and infertility management does not exist. Since both the eutopic and the ectopic endometrium express the vitamin D receptor (VDR), and VDR agonists are endowed with anti-proliferative and anti-inflammatory properties, we evaluated the effect of elocalcitol, a VDR agonist with low calcaemic liability, in a mouse model of experimentally induced endometriosis. METHODS AND RESULTS: Endometriosis was induced by injection of syngeneic endometrial tissue fragments into adult Balb/c female mice. After having confirmed by immunohistochemistry that endometriotic lesions developing in mice expressed VDR, the mice were administered with elocalcitol (100 µg/kg) or vehicle orally, once a day, for various durations of time. In this model, elocalcitol was able to reduce total lesion weight up to 70% upon treatment for 1 week before and 2 weeks after disease induction. Interestingly, a therapeutic effect was also observed on already established lesions. Elocalcitol was shown to reduce the capacity of mouse endometrial cells to adhere to collagen. In addition in treated mice, a decreased state of peritoneal inflammation was demonstrated by the inhibition of macrophage recruitment and inflammatory cytokine secretion. CONCLUSIONS: The VDR agonist elocalcitol inhibits lesion development in a validated mouse model of endometriosis, and exerts a protective effect on both the implantation and organization of transferred endometrial tissue. These preliminary data in mice provide a sound rationale for further testing in primate models and eventually in humans.

Neurogenesis and Viral Infection.

Neural stem cells (NSCs) are multipotent stem cells that reside in the fetal and adult mammalian brain, which can self-renew and differentiate into neurons and supporting cells. Intrinsic and extrinsic cues, from cells in the local niche and from distant sites, stringently orchestrates the self-renewal and differentiation competence of NSCs. Ample evidence supports the important role of NSCs in neuroplasticity, aging, disease, and repair of the nervous system. Indeed, activation of NSCs or their transplantation into injured areas of the central nervous system can lead to regeneration in animal models. Viral invasion of NSCs can negatively affect neurogenesis and synaptogenesis, with consequent cell death, impairment of cell cycle progression, early differentiation, which cause neural progenitors depletion in the cortical layer of the brain. Herein, we will review the current understanding of Zika virus (ZIKV) infection of the fetal brain and the NSCs, which are the preferential population targeted by ZIKV. Furthermore, the potential neurotropic properties of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which may cause direct neurological damage, will be discussed.

CREB3L1 and CREB3L2 control Golgi remodelling during decidualization of endometrial stromal cells.

Upon progesterone stimulation, Endometrial Stromal Cells (EnSCs) undergo a differentiation program into secretory cells (decidualization) to release in abundance factors crucial for embryo implantation. We previously demonstrated that decidualization requires massive reshaping of the secretory pathway and, in particular, of the Golgi complex. To decipher the underlying mechanisms, we performed a time-course transcriptomic analysis of in vitro decidualizing EnSC. Pathway analysis shows that Gene Ontology terms associated with vesicular trafficking and early secretory pathway compartments are the most represented among those enriched for upregulated genes. Among these, we identified a cluster of co-regulated genes that share CREB3L1 and CREB3L2 binding elements in their promoter regions. Indeed, both CREB3L1 and CREB3L2 transcription factors are up-regulated during decidualization. Simultaneous downregulation of CREB3L1 and CREB3L2 impairs Golgi enlargement, and causes dramatic changes in decidualizing EnSC, including Golgi fragmentation, collagen accumulation in dilated Endoplasmic Reticulum cisternae, and overall decreased protein secretion. Thus, both CREB3L1 and CREB3L2 are required for Golgi reshaping and efficient protein secretion, and, as such, for successful decidualization.

Neural precursor cells tune striatal connectivity through the release of IGFBPL1.

The adult brain retains over life endogenous neural stem/precursor cells (eNPCs) within the subventricular zone (SVZ). Whether or not these cells exert physiological functions is still unclear. In the present work, we provide evidence that SVZ-eNPCs tune structural, electrophysiological, and behavioural aspects of striatal function via secretion of insulin-like growth factor binding protein-like 1 (IGFBPL1). In mice, selective ablation of SVZ-eNPCs or selective abrogation of IGFBPL1 determined an impairment of striatal medium spiny neuron morphology, a higher failure rate in GABAergic transmission mediated by fast-spiking interneurons, and striatum-related behavioural dysfunctions. We also found IGFBPL1 expression in the human SVZ, foetal and induced-pluripotent stem cell-derived NPCs. Finally, we found a significant correlation between SVZ damage, reduction of striatum volume, and impairment of information processing speed in neurological patients. Our results highlight the physiological role of adult SVZ-eNPCs in supporting cognitive functions by regulating striatal neuronal activity.

Chromatin Velocity reveals epigenetic dynamics by single-cell profiling of heterochromatin and euchromatin.

Recent efforts have succeeded in surveying open chromatin at the single-cell level, but high-throughput, single-cell assessment of heterochromatin and its underlying genomic determinants remains challenging. We engineered a hybrid transposase including the chromodomain (CD) of the heterochromatin protein-1α (HP-1α), which is involved in heterochromatin assembly and maintenance through its binding to trimethylation of the lysine 9 on histone 3 (H3K9me3), and developed a single-cell method, single-cell genome and epigenome by transposases sequencing (scGET-seq), that, unlike single-cell assay for transposase-accessible chromatin with sequencing (scATAC-seq), comprehensively probes both open and closed chromatin and concomitantly records the underlying genomic sequences. We tested scGET-seq in cancer-derived organoids and human-derived xenograft (PDX) models and identified genetic events and plasticity-driven mechanisms contributing to cancer drug resistance. Next, building upon the differential enrichment of closed and open chromatin, we devised a method, Chromatin Velocity, that identifies the trajectories of epigenetic modifications at the single-cell level. Chromatin Velocity uncovered paths of epigenetic reorganization during stem cell reprogramming and identified key transcription factors driving these developmental processes. scGET-seq reveals the dynamics of genomic and epigenetic landscapes underlying any cellular processes.

Macrophages are alternatively activated in patients with endometriosis and required for growth and vascularization of lesions in a mouse model of disease.

The mechanisms that sustain endometrial tissues at ectopic sites in patients with endometriosis are poorly understood. Various leukocytes, including macrophages, infiltrate endometriotic lesions. In this study, we depleted mouse macrophages by means of either clodronate liposomes or monoclonal antibodies before the injection of syngeneic endometrial tissue. In the absence of macrophages, tissue fragments adhered and implanted into the peritoneal wall, but endometriotic lesions failed to organize and develop. When we depleted macrophages after the establishment of endometriotic lesions, blood vessels failed to reach the inner layers of the lesions, which stopped growing. Macrophages from patients with endometriosis and experimental mice, but not nonendometriotic patients who underwent surgery for uterine leiomyomas or control mice, expressed markers of alternative activation. These markers included high levels of scavenger receptors, CD163 and CD206, which are involved in both the scavenging of hemoglobin with iron transfer into macrophages and the silent clearance of inflammatory molecules. Macrophages in both inflammatory liquid and ectopic lesions were equally polarized, suggesting a critical role of environmental cues in the peritoneal cavity. Adoptively transferred, alternatively activated macrophages dramatically enhanced endometriotic lesion growth in mice. Inflammatory macrophages effectively protected mice from endometriosis. Therefore, endogenous macrophages involved in tissue remodeling appear as players in the natural history of endometriosis, required for effective vascularization and ectopic lesion growth.

The Emerging Role of Stem Cells in Regenerative Dentistry.

Progress of modern dentistry is accelerating at a spectacular speed in the scientific, technological and clinical areas. Practical examples are the advancement in the digital field, which has guaranteed an average level of prosthetic practices for all patients, as well as other scientific developments, including research on stem cell biology. Given their plasticity, defined as the ability to differentiate into specific cell lineages with a capacity of almost unlimited self-renewal and release of trophic/immunomodulatory factors, stem cells have gained significant scientific and commercial interest in the last 15 years. Stem cells that can be isolated from various tissues of the oral cavity have emerged as attractive sources for bone and dental regeneration, mainly due to their ease of accessibility. This review will present the current understanding of emerging conceptual and technological issues of the use of stem cells to treat bone and dental loss defects. In particular, we will focus on the clinical application of stem cells, either directly isolated from oral sources or in vitro reprogrammed from somatic cells (induced pluripotent stem cells). Research aimed at further unraveling stem cell plasticity will allow to identify optimal stem cell sources and characteristics, to develop novel regenerative tools in dentistry.

The surface-exposed chaperone, Hsp60, is an agonist of the microglial TREM2 receptor.

Triggering receptor expressed in myeloid (TREM) cells 2, a receptor expressed by myeloid cells, osteoclasts and microglia, is known to play a protective role in bones and brain. Mutations of the receptor (or of its coupling protein, DAP12) sustain in fact a genetic disease affecting the two organs, the polycystic lipomembraneous osteodysplasia with sclerosing leukoencephalopathy (PLOSL or Nasu-Hakola disease). So far, specific agonist(s) of TREM2 have not been identified and its (their) transduction mechanisms are largely unknown. Heat shock protein 60 (Hsp60) is a mitochondrial chaperone that can also be harboured at the cell surface. By using constructs including the extracellular domain of TREM2 and the Fc domain of IgGs we have identified Hsp60 as the only TREM2-binding protein exposed at the surface of neuroblastoma N2A cells and astrocytes, and lacking in U373 astrocytoma. Treatment with Hsp60 was found to stimulate the best known TREM2-dependent process, phagocytosis, however, only in the microglial N9 cells rich in the receptor. Upon TREM2 down-regulation, the Hsp60-induced stimulation of N9 phagocytosis was greatly attenuated. Hsp60 is also released by many cell types, segregated within exosomes or shedding vesicles which might then undergo dissolution. However, the affinity of its binding (K(d) = 3.8 microM) might be too low for the soluble chaperone released from the vesicles to the extracellular space to induce a significant activation of TREM2. It might in contrast be appropriate for the binding of TREM2 to Hsp60 exposed at the surface of cells closely interacting with microglia. The ensuing stimulation of phagocytosis could play protective effects on the brain.