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White matter injury (WMI) is a common neurological issue in premature-born neonates, often causing long-term disabilities. We recently demonstrated a key beneficial role of Wharton’s jelly mesenchymal stromal cell-derived small extracellular vesicles (WJ-MSC-sEVs) microRNAs (miRNAs) in WMI-related processes in vitro. Here, we studied the functions of WJ-MSC-sEV miRNAs in vivo using a preclinical rat model of premature WMI. Premature WMI was induced in rat pups through inflammation and hypoxia-ischemia. Small EVs were purified from the culture supernatant of human WJ-MSCs. The capacity of WJ-MSC-sEV-derived miRNAs to decrease microglia activation and promote oligodendrocyte maturation was evaluated by knocking down (k.d) DROSHA in WJ-MSCs, releasing sEVs containing significantly less mature miRNAs. Wharton’s jelly MSC-sEVs intranasally administrated 24 h upon injury reached the brain within 1 h, remained detectable for at least 24 h, significantly reduced microglial activation, and promoted oligodendrocyte maturation. The DROSHA k.d in WJ-MSCs lowered the therapeutic capabilities of sEVs in experimental premature WMI. Our results strongly indicate the relevance of miRNAs in the therapeutic abilities of WJ-MSC-sEVs in premature WMI in vivo, opening the path to clinical application.

Perinatal derivatives (PnD) are gaining interest as a source for cell-based therapies. Since the eye is easily accessible to local administration, eye diseases may be excellent candidates to evaluate novel therapeutic approaches. With this work, we performed a systematic review of published preclinical and clinical studies addressing PnD in the treatment of ocular diseases. We have set two specific objectives: (i) to investigate the current level of standardization in applied technical procedures in preclinical studies and (ii) to assess clinical efficacy in clinical trials. Hereto, we selected studies that applied amniotic membrane (hAM) and mesenchymal stromal cells derived from amniotic membrane (hAMSC), placenta (hPMSC), umbilical cord (hUC-MSC) and Wharton’s Jelly (hUC-WJ-MSC), excluding those where cells were not transplanted individually, following a systematic PubMed search for preclinical studies and consultation of clinical studies on https://clinicaltrials.gov and https://www.clinicaltrialsregister.eu/. Our bibliographic search retrieved 26 pre-clinical studies and 27 clinical trials. There was a considerable overlap regarding targeted ocular structures. Another common feature is the marked tendency towards (i) locally administered treatments and (ii) the PnD type. In the cornea/ocular surface, hAM was preferred and usually applied directly covering the ocular surface. For neuroretinal disorders, intra-ocular injection of umbilical or placental-derived cells was preferred. In general, basic research reported favourable outcomes. However, due to lack of standardization between different studies, until now there is no clear consensus regarding the fate of administered PnD or their mode of action. This might be accountable for the low index of clinical translation. Regarding clinical trials, only a minority provided results and a considerable proportion is in “unknown status”. Nevertheless, from the limited clinical evidence available, hAM proved beneficial in the symptomatic relief of bullous keratopathy, treating dry eye disease and preventing glaucoma drainage device tube exposure. Regarding neuroretinal diseases, application of Wharton’s Jelly MSC seems to become a promising future approach. In conclusion, PnD-based therapies seem to be beneficial in the treatment of several ocular diseases. However, much is yet to be done both in the pre-clinical and in the clinical setting before they can be included in the daily ophthalmic practice.

Perinatal tissues, such as placenta and umbilical cord contain a variety of somatic stem cell types, spanning from the largely used hematopoietic stem and progenitor cells to the most recently described broadly multipotent epithelial and stromal cells. As perinatal derivatives (PnD), several of these cell types and related products provide an interesting regenerative potential for a variety of diseases. Within COST SPRINT Action, we continue our review series, revising and summarizing the modalities of action and proposed medical approaches using PnD products: cells, secretome, extracellular vesicles, and decellularized tissues. Focusing on the brain, bone, skeletal muscle, heart, intestinal, liver, and lung pathologies, we discuss the importance of potency testing in validating PnD therapeutics, and critically evaluate the concept of PnD application in the field of tissue regeneration. Hereby we aim to shed light on the actual therapeutic properties of PnD, with an open eye for future clinical application. This review is part of a quadrinomial series on functional/potency assays for validation of PnD, spanning biological functions, such as immunomodulation, anti-microbial/anti-cancer, anti-inflammation, wound healing, angiogenesis, and regeneration.

The last 18 years have brought an increasing interest in the therapeutic use of perinatal derivatives (PnD). Preclinical studies used to assess the potential of PnD therapy include a broad range of study designs. The COST SPRINT Action (CA17116) aims to provide systematic and comprehensive reviews of preclinical studies for the understanding of the therapeutic potential and mechanisms of PnD in diseases and injuries that benefit from PnD therapy. Here we describe the publication search and data mining, extraction, and synthesis strategies employed to collect and prepare the published data selected for meta-analyses and reviews of the efficacy of PnD therapies for different diseases and injuries. A coordinated effort was made to prepare the data suitable to make statements for the treatment efficacy of the different types of PnD, routes, time points, and frequencies of administration, and the dosage based on clinically relevant effects resulting in clear increase, recovery or amelioration of the specific tissue or organ function. According to recently proposed guidelines, the harmonization of the nomenclature of PnD types will allow for the assessment of the most efficient treatments in various disease models. Experts within the COST SPRINT Action (CA17116), together with external collaborators, are doing the meta-analyses and reviews using the data prepared with the strategies presented here in the relevant disease or research fields. Our final aim is to provide standards to assess the safety and clinical benefit of PnD and to minimize redundancy in the use of animal models following the 3R principles for animal experimentation.

Failed or altered gliogenesis is a major characteristic of diffuse white matter injury in survivors of premature birth. The developmentally regulated long noncoding RNA (lncRNA) H19 inhibits S-adenosylhomocysteine hydrolase (SAHH) and contributes to methylation of diverse cellular components, such as DNA, RNA, proteins, lipids, and neurotransmitters. We showed that the pregnancy-derived synthetic PreImplantation Factor (sPIF) induces expression of the nuclear receptor corepressor 2 (NCOR2) via H19/SAHH-mediated DNA demethylation. In turn, NCOR2 affects oligodendrocyte differentiation markers. Accordingly, after hypoxic-ischemic brain injury in rodents, myelin protection and oligodendrocytes’ fate are in part modulated by sPIF and H19. Our results revealed an unexpected mechanism of the H19/SAHH axis underlying myelin preservation during brain recovery and its use in treating neurodegenerative diseases can be envisioned.

Knowledge of the beneficial effects of perinatal derivatives (PnD) in wound healing goes back to the early 1900s when the human fetal amniotic membrane served as a biological dressing to treat burns and skin ulcerations. Since the twenty-first century, isolated cells from perinatal tissues and their secretomes have gained increasing scientific interest, as they can be obtained non-invasively, have anti-inflammatory, anti-cancer, and anti-fibrotic characteristics, and are immunologically tolerated in vivo. Many studies that apply PnD in pre-clinical cutaneous wound healing models show large variations in the choice of the animal species (e.g., large animals, rodents), the choice of diabetic or non-diabetic animals, the type of injury (full-thickness wounds, burns, radiation-induced wounds, skin flaps), the source and type of PnD (placenta, umbilical cord, fetal membranes, cells, secretomes, tissue extracts), the method of administration (topical application, intradermal/subcutaneous injection, intravenous or intraperitoneal injection, subcutaneous implantation), and the type of delivery systems (e.g., hydrogels, synthetic or natural biomaterials as carriers for transplanted cells, extracts or secretomes). This review provides a comprehensive and integrative overview of the application of PnD in wound healing to assess its efficacy in preclinical animal models. We highlight the advantages and limitations of the most commonly used animal models and evaluate the impact of the type of PnD, the route of administration, and the dose of cells/secretome application in correlation with the wound healing outcome. This review is a collaborative effort from the COST SPRINT Action (CA17116), which broadly aims at approaching consensus for different aspects of PnD research, such as providing inputs for future standards for the preclinical application of PnD in wound healing.

Hyperuricemia is a common feature in pregnancies compromised by pre-eclampsia, a pregnancy disease characterized by hypertension and proteinuria. The role of uric acid in the pathogenesis of pre-eclampsia remains largely unclear. The aim of this study was to investigate the effect of elevated uric acid serum levels during pregnancy on maternal blood pressure and neonatal outcome using two different murine knockout models. Non-pregnant liver-specific GLUT9 knockout (LG9KO) mice showed elevated uric acid serum concentrations but no hypertensive blood pressure levels. During pregnancy, however, blood pressure levels of these animals increased in the second and third trimester, and circadian blood pressure dipping was severely altered when compared to non-pregnant LG9KO mice. The impact of hyperuricemia on fetal development was investigated using a systemic GLUT9 knockout (G9KO) mouse model. Fetal hyperuricemia caused distinctive renal tissue injuries and, subsequently an impaired neonatal growth pattern. These findings provide strong evidence that hyperuricemia plays a major role in the pathogenesis of hypertensive pregnancy disorders such as pre-eclampsia. These novel insights may enable the development of preventive and therapeutic strategies for hyperuricemia-related diseases.

The selection of an appropriate animal model is key to the production of results with optimal relevance to human disease. Particularly in the case of perinatal brain injury, a dearth of affected human neonatal tissue available for research purposes increases the reliance on animal models for insight into disease mechanisms. Improvements in obstetric and neonatal care in the past 20 years have caused the pathologic hallmarks of perinatal white matter injury (WMI) to evolve away from cystic necrotic lesions and toward diffuse regions of reactive gliosis and persistent myelin disruption. Therefore, updated animal models are needed that recapitulate the key features of contemporary disease. Here, we report a murine model of acute diffuse perinatal WMI induced through a two-hit inflammatory–hypoxic injury paradigm. Consistent with diffuse human perinatal white matter injury (dWMI), our model did not show the formation of cystic lesions. Corresponding to cellular outcomes of dWMI, our injury protocol produced reactive microgliosis and astrogliosis, disrupted oligodendrocyte maturation, and disrupted myelination.. Functionally, we observed sensorimotor and cognitive deficits in affected mice. In conclusion, we report a novel murine model of dWMI that induces a pattern of brain injury mirroring multiple key aspects of the contemporary human clinical disease scenario.

Abstract Background Preterm newborns are at high risk of developing neurodevelopmental deficits caused by neuroinflammation leading to perinatal brain injury. Human Wharton’s jelly mesenchymal stem cells (hWJ-MSC) derived from the umbilical cord have been suggested to reduce neuroinflammation, in part through the release of extracellular vesicle-like exosomes. Here, we studied whether exosomes derived from hWJ-MSC have anti-inflammatory effects on microglia-mediated neuroinflammation in perinatal brain injury. Methods Using ultracentrifugation, we isolated exosomes from hWJ-MSC culture supernatants. In an in vitro model of neuroinflammation, we stimulated immortalized BV-2 microglia and primary mixed glial cells with lipopolysaccharide (LPS) in the presence or absence of exosomes. In vivo, we introduced brain damage in 3-day-old rat pups and treated them intranasally with hWJ-MSC-derived exosomes. Results hWJ-MSC-derived exosomes dampened the LPS-induced expression of inflammation-related genes by BV-2 microglia and primary mixed glial cells. The secretion of pro-inflammatory cytokines by LPS-stimulated primary mixed glial was inhibited by exosomes as well. Exosomes interfered within the Toll-like receptor 4 signaling of BV-2 microglia, as they prevented the degradation of the NFκB inhibitor IκBα and the phosphorylation of molecules of the mitogen-activated protein kinase family in response to LPS stimulation. Finally, intranasally administered exosomes reached the brain and reduced microglia-mediated neuroinflammation in rats with perinatal brain injury. Conclusions Our data suggest that the administration of hWJ-MSC-derived exosomes represents a promising therapy to prevent and treat perinatal brain injury.

The increasing cancer incidence has certified oncological management as one of the most critical challenges for the coming decades. New anticancer strategies are still needed, despite the significant advances brought to the forefront in the last decades. The most recent, promising therapeutic approaches have benefitted from the application of human perinatal derivatives (PnD), biological mediators with proven benefits in several fields beyond oncology. To elucidate preclinical results and clinic outcomes achieved in the oncological field, we present a narrative review of the studies resorting to animal models to assess specific outcomes of PnD products. Recent preclinical evidence points to promising anticancer effects offered by PnD mediators isolated from the placenta, amniotic membrane, amniotic fluid, and umbilical cord. Described effects include tumorigenesis prevention, uncontrolled growth or regrowth inhibition, tumor homing ability, and adequate cell-based delivery capacity. Furthermore, PnD treatments have been described as supportive of chemotherapy and radiological therapies, particularly when resistance has been reported. However, opposite effects of PnD products have also been observed, offering support and trophic effect to malignant cells. Such paradoxical and dichotomous roles need to be intensively investigated. Current hypotheses identify as explanatory some critical factors, such as the type of the PnD biological products used or the manufacturing procedure to prepare the tissue/cellular treatment, the experimental design (including human-relevant animal models), and intrinsic pathophysiological characteristics. The effective and safe translation of PnD treatments to clinical practice relies on the collaborative efforts of all researchers working with human-relevant oncological preclinical models. However, it requires proper guidelines and consensus compiled by experts and health workers who accurately describe the methodology of tissue collection, PnD isolation, manufacturing, preservation, and delivery to the final user.

Peripartum cerebral hypoxia and ischemia, and intrauterine infection and inflammation, are detrimental for the precursor cells of the myelin-forming oligodendrocytes in the prematurely newborn, potentially leading to white matter injury (WMI) with long-term neurodevelopmental sequelae. Previous data show that hypomyelination observed in WMI is caused by arrested oligodendroglial maturation rather than oligodendrocyte-specific cell death. In a rat model of premature WMI, we have recently shown that small extracellular vesicles (sEV) derived from Wharton’s jelly mesenchymal stromal cells (WJ-MSC) protect from myelination deficits. Thus, we hypothesized that sEV derived from WJ-MSC directly promote oligodendroglial maturation in oligodendrocyte precursor cells. To test this assumption, sEV were isolated from culture supernatants of human WJ-MSC by ultracentrifugation and co-cultured with the human immortalized oligodendrocyte precursor cell line MO3.13. As many regulatory functions in WMI have been ascribed to microRNA (miR) and as sEV are carriers of functional miR which can be delivered to target cells, we characterized and quantified the miR content of WJ-MSC-derived sEV by next-generation sequencing. We found that WJ-MSC-derived sEV co-localized with MO3.13 cells within 4 h. After 5 days of co-culture, the expression of myelin basic protein (MBP), a marker for mature oligodendrocytes, was significantly increased, while the oligodendrocyte precursor marker platelet-derived growth factor alpha (PDGFRα) was decreased. Notch and MAPK/ERK pathways known to inhibit oligodendrocyte maturation and differentiation were significantly reduced. The pathway enrichment analysis showed that the miR present in WJ-MSC-derived sEV target genes having key roles in the MAPK pathway. Our data strongly suggest that sEV from WJ-MSC directly drive the maturation of oligodendrocyte precursor cells by repressing Notch and MAPK/ERK signaling.

Purpose: This study aimed to quantify the combined effects of heat exposure and exercise of increasing intensity on pulmonary blood flow using lung diffusing capacity for carbon monoxide (DLCO) as an indirect measure. We hypothesized that, during exercise in the heat, the well-documented increase in skin blood flow for thermoregulation would lead to alterations in pulmonary blood flow and a subsequent fall in DLCO versus a thermoneutral condition. Methods: Nine healthy subjects (4 F/5 M, 20–45 years, VO2max 46.7 ± 5.8 mL/kg/min) completed three 15-min stages including rest and during cycling at 20 and 40% of maximum workload (Wmax) in either thermoneutral (TN; 22.2 ± 0.6 °C) or heat (HT; 39.4 ± 0.4 °C) conditions. DLCO, minute ventilation (VE), oxygen consumption ( VO 2 ), heart rate (HR), and core (TC) and skin temperature (Tsk) were measured. Results: DLCO showed a significant interaction between exercise intensity and heat (P = 0.019); post hoc testing revealed that DLCO was higher at 40% of Wmax in HT vs. TN (53.2 ± 10.6 vs 50.0 ± 10.3 mL/min/mmHg, P = 0.003) only. VE and VO 2 showed no difference in HT vs. TN. HR was higher in HT vs. TN (P < 0.001). TC and Tsk showed a significant interaction between temperature and intensity (P < 0.05). Conclusion: The unexpected increase in DLCO during exercise in HT vs. TN conditions suggests a larger lung surface area for gas exchange, perhaps due to increased pulmonary capillary recruitment and/or distension secondary to a higher cardiac output (Q) in the heat. This study furthers our understanding of how heat exposure might impact pulmonary blood flow, specifically as assessed via DLCO. [ABSTRACT FROM AUTHOR]

Hypoxic-ischemic (HI) insult in the perinatal phase harbors a high risk of encephalopathy in the neonate. Brain cells undergo apoptosis, initiating neurodegeneration. So far, therapeutic approaches such as cooling remain limited. Transplantation of mesenchymal stem cells (MSCs) exhibits therapeutic success despite the short-time survival in the host brain, providing strong evidence that their beneficial effects are largely based on secreted factors, including extracellular vesicles (EVs). The aim of this study was to investigate the effects of human Wharton’s jelly MSC (hWJ-MSC)-derived EVs on neuroprotection and neuroregeneration, using an in vitro model of oxygen–glucose deprivation/reoxygenation (OGD/R) mimicking HI injury in the mouse neuroblastoma cell line neuro2a (N2a). hWJ-MSC-derived EVs were isolated from cell culture supernatants by multistep centrifugation and identified by endosomal marker expression and electron microscopy. OGD/R significantly increased DNA fragmentation and caspase 3 ( Casp3 ) transcription in N2a cells relative to undamaged cells. OGD/R-mediated DNA fragmentation and Casp3 expression could be prevented as well as resolved by the addition of hWJ-MSC-derived EV before and after OGD, respectively. hWJ-MSC-derived EV also tended to increase the phosphorylation of the B cell lymphoma 2 (Bcl2) family member Bcl-2-antagonist of cell death (BAD) in N2a cells, when added prior or post OGD, thereby inactivating the proapoptotic function of BAD. Fluorescence confocal microscopy revealed the close localization of hWJ-MSC-derived EVs to the nuclei of N2a cells. Furthermore, EVs released their RNA content into the cells. The expression levels of the microRNAs (miRs) let-7a and let-7e, known regulators of Casp3 , were inversely correlated to Casp3 . Our data suggest that hWJ-MSC-derived EVs have the potential to prevent and resolve HI-induced apoptosis in neuronal cells in the immature neonatal brain. Their antiapoptotic effect seems to be mediated by the transfer of EV-derived let-7-5p miR.