Menstrual blood-derived mesenchymal stromal cells: impact of preconditioning on the cargo of extracellular vesicles as potential therapeutics.

BACKGROUND: Mesenchymal stromal cells (MSCs) have been shown to exert their therapeutic effects through the secretion of broad spectrum of paracrine factors, including extracellular vesicles (EVs). Accordingly, EVs are being pursued as a promising alternative to cell-based therapies. Menstrual blood-derived stromal cells (MenSCs) are a type of MSC that, due to their immunomodulatory and regenerative properties, have emerged as an innovative source. Additionally, new strategies of cell priming may potentially alter the concentration and cargo of released EVs, leading to modification of their biological properties. In this study, we aimed to characterize the EVs released by MenSCs and compare their therapeutic potential under three different preconditioning conditions (proinflammatory stimuli, physioxia, and acute hypoxia). METHODS: MenSCs were isolated from five healthy women. Following culturing to 80% confluence, MenSCs were exposed to different priming conditions: basal (21% O2), proinflammatory stimuli (IFNγ and TNFα, 21% O2), physioxia (1-2% O2), and acute hypoxia (< 1% O2) for 48-72 h. Conditioned media from MenSCs was collected after 48 h and EVs were isolated by a combination of ultra-filtration and differential centrifugation. An extensive characterization ranging from nano-flow cytometry (nFC) to quantitative high-throughput shotgun proteomics was performed. Bioinformatics analyses were used to derive hypotheses on their biological properties. RESULTS: No differences in the morphology, size, or number of EVs released were detected between priming conditions. The proteome analysis associated with basal MenSC-EVs prominently revealed their immunomodulatory and regenerative capabilities. Furthermore, quantitative proteomic analysis of differentially produced MenSC-EVs provided sufficient evidence for the utility of the differential preconditioning in purpose-tailoring EVs for their therapeutic application: proinflammatory priming enhanced the anti-inflammatory, regenerative and immunomodulatory capacity in the innate response of EVs, physioxia priming also improves tissue regeneration, angiogenesis and their immunomodulatory capacity targeting on the adaptive response, while acute hypoxia priming, increased hemostasis and apoptotic processes regulation in MenSC-EVs, also by stimulating immunomodulation mainly through the adaptive response. CONCLUSIONS: Priming of MenSCs under proinflammatory and hypoxic conditions affected the cargo proteome of EVs released, resulting in different therapeutic potential, and thus warrants experimental exploration with the aim to generate better-defined MSC-derived bioproducts.

Comparative anatomy of the ovine and human pelvis for laparoscopic sacrocolpopexy: evaluating the effectiveness of the ovine model.

INTRODUCTION AND HYPOTHESIS: Laparoscopic sacrocolpopexy (LSC) is a functional reconstructive surgery used to treat pelvic organ prolapse (POP) in middle-aged women. Although LSC is widely used, its implementation is hindered by perceived technical difficulties and surgical learning curves. Surgeons require adequate experience with LSC prior to performing the procedure on patients to improve their quality of life. This study is aimed at demonstrating the effectiveness of the ovine model (OM) for training and research in LSC, while also comparing anatomical differences between ovine and human models during the procedure. METHODS: The animal model and training were provided by the Jesús Usón Minimally Invasive Surgery Centre. Urologists and gynecologists with experience in LSC participated in a course and their findings were recorded and documented. RESULTS: Differences in patient positioning, trocar placement, and reperitonealization were identified between the ovine and human models. Hysterectomy is always performed in the ovine model, whereas it is not mandatory in humans. There are also differences in the dissection of the levator ani muscle and attachment point of the posterior mesh to the uterus between the two models. Despite differences in some areas, the ovine pelvic structure and vagina are similar in size to those of humans. CONCLUSIONS: The ovine model is a valuable tool for surgeons in their learning curve for LSC, allowing for safe and effective practice prior to performing the procedure on patients. The use of the OM can help to improve the quality of life for women affected by pelvic organ prolapse.

Menstrual blood-derived stromal cells: insights into their secretome in acute hypoxia conditions.

BACKGROUND: Despite constant advances in regenerative medicine, the closure of chronic wounds is still challenging. Therapeutic approaches using locally administered MSCs have been considered a promising option. However, the viability of these cells is seriously threatened by acute hypoxic stress linked to wound healing. In this work, we aimed to study the tolerance of Menstrual blood-derived stromal cells (MenSCs) to acute hypoxia and their therapeutic paracrine effect. METHODS: Isolated MenSCs were phenotypically characterized and evaluated in terms of proliferation, viability, and gene expression, under acute hypoxia (AH) compared with conventional cultured condition or normoxia (N). A step further, the secretome of MenSCs under acute hypoxia was analyzed with respect to their miRNAs content and by in vitro functional assays. For the analysis of differences between the two groups, Student's t-test was performed and one-way ANOVA and Tukey's multiple comparisons test for multiple groups were used. RESULTS: Our results revealed that the viability of MenSCs was not affected under acute hypoxia, although proliferation rate slowed down. Gene analysis revealed 5 up-regulated (BNIP3, ANGPTL4, IL6, IL1B, and PDK1) and 4 down-regulated genes (IDO1, HMOX1, ANGPTL2, and HGF) in AH compared to N. Global gene expression analysis revealed a decrease in the gene ontology functions of migration and wound response with respect to the normoxic condition. In contrast, functions such as angiogenesis were enriched under the AH condition. Regarding the secretome analysis, two miRNAs involved in angiogenic processes (hsa-miR-148a-3p and hsa-miR-378a-3p), were significantly up-expressed when compared to the normoxic condition, being MYC gene, the unique target of both. Functional assays on HUVECs revealed a potential pro-angiogenic capacity of MenSCs cultured in both oxygen conditions (N and AH) based on the wound closure and tube formation results of their released paracrine factors. However, when compared to normoxia, the paracrine factors of MenSCs under acute hypoxia slightly reduced the proliferation, migration, and in vitro wound closure of HUVECs. CONCLUSIONS: MenSC exhibited a good survival capacity under acute hypoxic conditions as well as beneficial properties applicable in the field of tissue regeneration through their secretome, which makes them a potential cell source for wound healing interventions.

Transcriptome Profile Reveals Differences between Remote and Ischemic Myocardium after Acute Myocardial Infarction in a Swine Model.

Acute myocardial infarction (AMI) is the consequence of an acute interruption of myocardial blood flow delimiting an area with ischemic necrosis. The loss of cardiomyocytes initiates cardiac remodeling in the myocardium, leading to molecular changes in an attempt to recover myocardial function. The purpose of this study was to unravel the differences in the molecular profile between ischemic and remote myocardium after AMI in an experimental model. To mimic human myocardial infarction, healthy pigs were subjected to occlusion of the mid-left anterior descending coronary artery, and myocardial tissue was collected from ischemic and remote zones for omics techniques. Comparative transcriptome analysis of both areas was accurately validated by proteomic analysis, resulting in mitochondrion-related biological processes being the most impaired mechanisms in the infarcted area. Moreover, Immune system process-related genes were up-regulated in the remote tissue, mainly due to the increase of neutrophil migration in this area. These results provide valuable information regarding differentially expressed genes and their biological functions between ischemic and remote myocardium after AMI, which could be useful for establishing therapeutic targets for the development of new treatments.

Intracoronary Administration of Microencapsulated HGF in a Reperfused Myocardial Infarction Swine Model.

Therapy microencapsulation allows minimally invasive, safe, and effective administration. Hepatocyte growth factor (HGF) has angiogenic, anti-inflammatory, anti-apoptotic, and anti-fibrotic properties. Our objective was to evaluate the cardiac safety and effectiveness of intracoronary (IC) administration of HGF-loaded extended release microspheres in an acute myocardial infarction (AMI) swine model. An IC infusion of 5 × 106 HGF-loaded microspheres (MS+HGF, n = 7), 5 × 106 placebo microspheres (MS, n = 7), or saline (SAL, n = 7) was performed two days after AMI. TIMI flow and Troponin I (TnI) values were assessed pre- and post-treatment. Cardiac function was evaluated with magnetic resonance imaging (cMR) before injection and at 10 weeks. Plasma cytokines were determined to evaluate the inflammatory profile and hearts were subjected to histopathological evaluation. Post-treatment coronary flow was impaired in five animals (MS+HGF and MS group) without significant increases in TnI. One animal (MS group) died during treatment. There were no significant differences between groups in cMR parameters at any time (p > 0.05). No statistically significant changes were found between groups neither in cytokines nor in histological analyses. The IC administration of 5 × 106 HGF-loaded-microspheres 48 h post-AMI did not improve cardiac function, nor did it decrease inflammation or cardiac fibrosis in this experimental setting.

Comparative efficacy and safety of prostatic urethral lift vs prostatic artery embolization for benign prostatic hyperplasia: a systematic review and network meta-analysis.

OBJECTIVE: To assess and compare the clinical efficacy and safety of prostatic urethral lift (PUL) and prostatic artery embolization (PAE) for the treatment of lower urinary tract symptoms (LUTS) in patients with benign prostatic hyperplasia (BPH) by means of a systematic review and network meta-analysis. METHODS: A systematic literature search was performed using PubMed and Web of Science from inception to March 2021 to identify randomized controlled trials (RCTs) that compared PUL or PAE with either transurethral resection of the prostate (TURP) or sham procedures as control interventions. Qualitative and quantitative analyses were performed to pool the data on direct and indirect comparisons between interventions using STATA 14. RESULTS: Eight RCTs with 675 participants were included in our network meta-analysis. Quantitative synthesis revealed that TURP was the most efficacious intervention for clinical (International Prostate Symptoms Score and quality of life) and functional outcomes (maximum urinary flow rate and post-void residual urine volume), and was associated with a lower reintervention rate compared with PAE (risk ratio [RR] 2.08 with 95% confidence interval [CI] 0.96 to 4.51) and PUL (RR 2.33 with 95% CI 0.50 to 10.86), although the difference were not statistically significant. Indirect comparison indicated that PUL and PAE resulted in similar outcomes. PAE was associated with fewer minor adverse events (AEs; RR 0.75 with 95% CI 0.48 to 1.18) and PUL with fewer major AEs (RR 0.72 with 95% CI 0.17 to 3.13) when compared with TURP. Whilst PAE had a better ranking with regard to improvement of most clinical and functional outcomes, PUL was the best ranked procedure regarding erectile function, as measured by the International Index of Erectile Function 5, but no significant difference was observed. CONCLUSION: Current evidence suggests that PUL and PAE have similar clinical efficacy and safety profiles in the management of LUTS associated with BPH. However, the quality of evidence is relatively low because of the paucity of RCTs available, and results should be interpreted with caution.

Popliteal Vascular Lymph Node Resection in the Rabbit Hindlimb for Secondary Lymphedema Induction.

Lymphedema is a common condition often associated with cancer and its treatment, which leads to damage to the lymphatic system, and current treatments are mostly palliative rather than curative. Its high incidence among oncologic patients indicates the need to study both normal lymphatic function and pathologic dysfunction. To reproduce chronic lymphedema, it is necessary to choose a suitable experimental animal. Attempts to establish animal models are limited by the regenerative capacity of the lymphatic system. Among the potential candidates, the rabbit hindlimb is easy to handle and extrapolate to the human clinical scenario, making it advantageous. In addition, the size of this species allows for better selection of lymphatic vessels for vascularized lymph node resection. In this study, we present a procedure of vascular lymph node resection in the rabbit hindlimb for inducing secondary lymphedema. Anesthetized animals were subjected to circumferential measurement, patent blue V infiltration, and indocyanine green lymphography (ICG-L) using real-time near-infrared fluorescence, a technique that allows the identification of single popliteal nodes and lymphatic channels. Access to the identified structures is achieved by excising the popliteal node and ligating the medial and lateral afferent lymphatics. Special care must be taken to ensure that any lymphatic vessel that joins the femoral lymphatic system within the thigh without entering the popliteal node can be identified and ligated. Postoperative evaluation was performed at 3, 6, and 12 months after induction using circumferential measurements of the hindlimb and ICG-L. As demonstrated during follow-up, the animals developed dermal backflow that was maintained until the 12th month, making this experimental animal useful for novel long-term evaluations in the management of lymphedema. In conclusion, the approach described here is feasible and reproducible. Additionally, during the time window presented, it can be representative of human lymphedema, thus providing a useful research tool.

In Vitro and In Vivo Response of Zinc-Containing Mesoporous Bioactive Glasses in a Sheep Animal Model.

Zinc-enriched mesoporous bioactive glasses (MBGs) are bioceramics with potential antibacterial and osteogenic properties. However, few assays have been performed to study these properties in animal models. In this study, MBGs enriched with up to 5% ZnO were synthesized, physicochemically characterized, and evaluated for their osteogenic activity both in vitro and in vivo. The ZnO MBGs showed excellent textural properties despite ZnO incorporation. However, the release of Zn2+ ions inhibited the mineralization process when immersed in simulated body fluid. In vitro assays showed significantly higher values of viability and expression of early markers of cell differentiation and angiogenesis in a ZnO-content-dependent manner. The next step was to study the osteogenic potential in a sheep bone defect model. Despite their excellent textural properties and cellular response in vitro, the ZnO MBGs were not able to integrate into the bone tissue, which can be explained in terms of inhibition of the mineralization process caused by Zn2+ ions. This work highlights the need to develop nanostructured materials for bone regeneration that can mineralize to interact with bone tissue and induce the processes of implant acceptance, cell colonization by osteogenic cells, and regeneration of lost bone tissue.

Cytotoxicity Assessment of a New Design for a Biodegradable Ureteral Mitomycin Drug-Eluting Stent in Urothelial Carcinoma Cell Culture.

Urothelial tumour of the upper urinary tract is a rare neoplasm, but unfortunately, it has a high recurrence rate. The reduction of these tumour recurrences could be achieved by the intracavitary instillation of adjuvant chemotherapy after nephron-sparing treatment in selected patients, but current instillation methods are ineffective. Therefore, the aim of this in vitro study is to evaluate the cytotoxic capacity of a new instillation technology through a biodegradable ureteral stent/scaffold coated with a silk fibroin matrix for the controlled release of mitomycin C as an anti-cancer drug. Through a comparative study, we assessed, in urothelial carcinoma cells in a human cancer T24 cell culture for 3 and 6 h, the cytotoxic capacity of mitomycin C by viability assay using the CCK-8 test (Cell counting Kit-8). Cell viability studies in the urothelial carcinoma cell line confirm that mitomycin C embedded in the polymeric matrix does not alter its cytotoxic properties and causes a significant decrease in cell viability at 6 h versus in the control groups. These findings have a clear biomedical application and could be of great use to decrease the recurrence rate in patients with upper tract urothelial carcinomas by increasing the dwell time of anti-cancer drugs.

Assessment of a Coated Mitomycin-Releasing Biodegradable Ureteral Stent as an Adjuvant Therapy in Upper Urothelial Carcinoma: A Comparative In Vitro Study.

A major limitation of the treatment of low-grade upper tract urothelial carcinoma is the difficulty of intracavitary instillation of adjuvant therapy. Therefore, the aim of this in vitro study was to develop and to assess a new design of biodegradable ureteral stent coated with a silk fibroin matrix for the controlled release of mitomycin C as a chemotherapeutic drug. For this purpose, we assessed the coating of a biodegradable ureteral stent, BraidStent®, with silk fibroin and subsequently loaded the polymeric matrix with two formulations of mitomycin to evaluate its degradation rate, the concentration of mitomycin released, and changes in the pH and the weight of the stent. Our results confirm that the silk fibroin matrix is able to coat the biodegradable stent and release mitomycin for between 6 and 12 h in the urinary environment. There was a significant delay in the degradation rate of silk fibroin and mitomycin-coated stents compared to bare biodegradable stents, from 6-7 weeks to 13-14 weeks. The present study has shown the feasibility of using mitomycin C-loaded silk fibroin for the coating of biodegradable urinary stents. The addition of mitomycin C to the coating of silk fibroin biodegradable stents could be an attractive approach for intracavitary instillation in the upper urinary tract.

Prostatic artery occlusion versus prostatic artery embolisation for the management of benign prostatic hyperplasia: early results in a canine model.

OBJECTIVE: To compare the technical efficacy and safety between prostatic artery occlusion (PAO) with ethylene vinyl alcohol copolymer (EVOH) and prostatic artery embolizsation (PAE) with microspheres in a canine model. METHODS: 17 adult male beagles underwent PAO (n = 7) with Onyx-18 or PAE (n = 10) with microspheres (300-500 µm). To evaluate the primary outcomes (technical efficacy and safety), MRI evaluations were performed immediately before and 1 week, 2 weeks, and 1 month after procedures to document prostate volume (PV); and all dogs were inspected for procedure-related complications during 1 month follow-up. The secondary outcomes included the prostate ischaemia size detected by MRI and recanalisation of prostatic artery by follow-up angiography. Differences between groups were statistically analysed. RESULTS: Both procedures were bilaterally successful in all animals. Compared with PAE, the mean fluoroscopy time (23.80 vs 36.24 min, p = 0.014) and radiation dose (68.19 vs 125.26 mGy, p = 0.003) were significantly less in PAO procedure. The mean percentage of PV change significantly decreased in both groups at 2 weeks (30.71% vs 37.89%) and 1 month (56.41% %vs 55.56%) after PAO and PAE respectively), without significant differences between groups at either time point. No major complications were observed except one animal after PAO with transient haematuria and acute urinary retention. The mean prostate ischaemia induced by PAO was significant greater compared with PAE at 1 week (43.44% vs 18.91%, p=0.001). PAO with EVOH is technically feasible and with comparable efficacy and safety with PAE. There are possible benefits to PAO over PAE. ADVANCES IN KNOWLEDGE: A new technical modification of the PAE consisting of the use of liquid embolic agent to occlude the prostatic artery trunk and its branches has been developed in pre-clinical study, showing to be an effective and safe procedure which can induce a significant prostate shrinkage for the management of symptomatic benign prostatic hyperplasia in patients. In addition, the findings have showed a similar therapeutic effect comparable with the conventional PAE using microspheres.

Intrapericardial Administration of Secretomes from Menstrual Blood-Derived Mesenchymal Stromal Cells: Effects on Immune-Related Genes in a Porcine Model of Myocardial Infarction.

Acute myocardial infarction (AMI) is a manifestation of ischemic heart disease where the immune system plays an important role in the re-establishment of homeostasis. We hypothesize that the anti-inflammatory activity of secretomes from menstrual blood-derived mesenchymal stromal cells (S-MenSCs) and IFNγ/TNFα-primed MenSCs (S-MenSCs*) may be considered a therapeutic option for the treatment of AMI. To assess this hypothesis, we have evaluated the effect of S-MenSCs and S-MenSCs* on cardiac function parameters and the involvement of immune-related genes using a porcine model of AMI. Twelve pigs were randomly divided into three biogroups: AMI/Placebo, AMI/S-MenSCs, and AMI/S-MenSCs*. AMI models were generated using a closed chest coronary occlusion-reperfusion procedure and, after 72 h, the different treatments were intrapericardially administered. Cardiac function parameters were monitored by magnetic resonance imaging before and 7 days post-therapy. Transcriptomic analyses in the infarcted tissue identified 571 transcripts associated with the Gene Ontology term Immune response, of which 57 were differentially expressed when different biogroups were compared. Moreover, a prediction of the interactions between differentially expressed genes (DEGs) and miRNAs from secretomes revealed that some DEGs in the infarction area, such as STAT3, IGFR1, or BCL6 could be targeted by previously identified miRNAs in secretomes from MenSCs. In conclusion, the intrapericardial administration of secretome early after infarction has a significant impact on the expression of immune-related genes in the infarcted myocardium. This confirms the immunomodulatory potential of intrapericardially delivered secretomes and opens new therapeutic perspectives in myocardial infarction treatment.

Swine Models of Aneurysmal Diseases for Training and Research.

Large animal models, specifically swine, are widely used to research cardiovascular diseases and therapies, as well as for training purposes. This paper describes two different aneurysmal swine models that may help researchers to study new therapies for aneurysmal diseases. These aneurysmal models are created by surgically adding a pouch of tissue to carotid arteries in swine. When the model is used for research, the pouch must be autologous; for training purposes, a synthetic pouch suffices. First, the right external jugular vein (EJV) and right common carotid artery (CCA) must be surgically exposed. The EJV is ligated and a vein pouch fashioned from a short segment. This pouch is then sutured to an elliptical arteriotomy performed in the CCA. Animals must be kept heparinized during model creation, and local vasodilators may be used to decrease vasospasms. Once the suture is completed, correct blood flow should be inspected, checking for bleeding from the suture line and vessel patency. Finally, the surgical incision is closed by layers and an angiography performed to image the aneurysmal model. A simplification of this aneurysmal carotid model that decreases invasiveness and surgical time is the use of a synthetic, rather than venous, pouch. For this purpose, a pouch is tailored in advance with a segment of a polytetrafluoroethylene (PTFE) prosthesis, one end of which is sutured close using polypropylene vascular suture and sterilized prior to surgery. This "sac" is then attached to an arteriotomy performed in the CCA as described. Although these models do not reproduce many of the physiopathological events related to aneurysm formation, they are hemodynamically similar to the situation found in the clinical setting. Therefore, they can be used for research or training purposes, allowing physicians to learn and practice different endovascular techniques in animal models that are close to the human system.

Prostatic artery occlusion: initial findings on pathophysiological response in a canine prostate model.

Background: Prostatic artery embolization (PAE) is an alternative treatment for symptomatic benign prostatic hyperplasia (BPH) in men. A technical modification of conventional PAE has been developed in a canine prostate model consisting of prostatic artery occlusion (PAO) using Onyx® whose therapeutic effect is prostate shrinkage. However, the underlying mechanisms are not well clarified. The purpose was to evaluate the biological mechanisms responsible for therapeutic effects of PAO in the canine prostate. Methods: Ten adult male beagles (5.0±0.82 years) underwent PAO with Onyx-18 (n=7) and prostatic artery angiography as control (n=3). Blood samples were taken at different time points of follow-up (baseline, 1 week, 2 weeks, 1 month, 3 months and 6 months) to measure the serum canine prostate specific esterase (CPSE). MRI examinations were also performed to document the prostate volume (PV) before and after interventions at different time points of follow-up. Prostates were harvested at 2 weeks (n=2) in the PAO-group, and the remaining ones (n=8) at 6 months for the determinations of intraprostatic testosterone and dihydrotestosterone (DHT) by ELISA, apoptosis by TUNEL assay and histopathological study. Results: The mean serum CPSE concentration started to decrease significantly from 2 weeks to 6 months after PAO along with PV compared with baseline data. In addition, a moderate but significant correlation was observed between CPSE and PV (r=0.655, P=0.000). Regarding intraprostatic androgens, testosterone was significantly higher after PAO than control (19.70 vs. 4.87 ng/mL, P=0.002), whereas DHT was lower but no significant (112.52 vs. 138.35 pg/mL, P=0.144). In histological study, PAO induced a severe hemorrhagic necrosis in the whole prostates along with inflammatory cell infiltration at early 2 weeks, and then diffuse interstitial fibrosis with atrophy of the glandular epithelium and intraprostatic cavity formation at 6 months. Apoptosis was detected in all specimens with higher apoptotic index after PAO at 2 weeks (7.35%) and at 6 months (4.38%) compared with control (2.64%), without statistically significant difference between groups. Conclusions: PAO induces hemorrhagic ischemia predominantly resulting in necrosis rather than apoptosis with prostate shrinkage. CPSE is a potential biomarker to assess the response to PAO in the canine prostate.

A Fibrin Coating Method of Polypropylene Meshes Enables the Adhesion of Menstrual Blood-Derived Mesenchymal Stromal Cells: A New Delivery Strategy for Stem Cell-Based Therapies.

Polypropylene (PP) mesh is well-known as a gold standard of all prosthetic materials of choice for the reinforcement of soft tissues in case of hernia, organ prolapse, and urinary incontinence. The adverse effects that follow surgical mesh implantation remain an unmet medical challenge. Herein, it is outlined a new approach to allow viability and adhesion of human menstrual blood-derived mesenchymal stromal cells (MenSCs) on PP surgical meshes. A multilayered fibrin coating, based on fibrinogen and thrombin from a commercial fibrin sealant, was optimized to guarantee a homogeneous and stratified film on PP mesh. MenSCs were seeded on the optimized fibrin-coated meshes and their adhesion, viability, phenotype, gene expression, and immunomodulatory capacity were fully evaluated. This coating guaranteed MenSC viability, adhesion and did not trigger any change in their stemness and inflammatory profile. Additionally, MenSCs seeded on fibrin-coated meshes significantly decreased CD4+ and CD8+ T cell proliferation, compared to in vitro stimulated lymphocytes (p < 0.0001). Hence, the proposed fibrin coating for PP surgical meshes may allow the local administration of stromal cells and the reduction of the exacerbated inflammatory response following mesh implantation surgery. Reproducible and easy to adapt to other cell types, this method undoubtedly requires a multidisciplinary and translational approach to be improved for future clinical uses.

IFN-Gamma and TNF-Alpha as a Priming Strategy to Enhance the Immunomodulatory Capacity of Secretomes from Menstrual Blood-Derived Stromal Cells.

Mesenchymal stromal cells isolated from menstrual blood (MenSCs) exhibit a potent pro-angiogenic and immunomodulatory capacity. Their therapeutic effect is mediated by paracrine mediators released by their secretomes. In this work, we aimed to evaluate the effect of a specific priming condition on the phenotype and secretome content of MenSCs. Our results revealed that the optimal condition for priming MenSCs was the combination of interferon gamma (IFNγ) and tumor necrosis factor alpha (TNFα) that produced a synergistic and additive effect on IDO1 release and immune-related molecule expression. The analyses of MenSC-derived secretomes after IFNγ and TNFα priming also revealed an increase in EV release and in the differentially expressed miRNAs involved in the immune response and inflammation. Proliferation assays on lymphocyte subsets demonstrated a decrease in CD4+ T cells and CD8+ T cells co-cultured with secretomes, especially in the lymphocytes co-cultured with secretomes from primed cells. Additionally, the expression of immune checkpoints (PD-1 and CTLA-4) was increased in the CD4+ T cells co-cultured with MenSC-derived secretomes. These findings demonstrate that the combination of IFNγ and TNFα represents an excellent priming strategy to enhance the immunomodulatory capacity of MenSCs. Moreover, the secretome derived from primed MenSCs may be postulated as a therapeutic option for the regulation of adverse inflammatory reactions.

Intrapericardial Delivery of APA-Microcapsules as Promising Stem Cell Therapy Carriers in an Experimental Acute Myocardial Infarction Model.

The administration of cardiosphere-derived cells (CDCs) after acute myocardial infarction (AMI) is very promising. CDC encapsulation in alginate-poly-l-lysine-alginate (APA) could increase cell survival and adherence. The intrapericardial (IP) approach potentially achieves high concentrations of the therapeutic agent in the infarcted area. We aimed to evaluate IP therapy using a saline vehicle as a control (CON), a dose of 30 × 106 CDCs (CDCs) or APA microcapsules containing 30 × 106 CDCs (APA-CDCs) at 72 h in a porcine AMI model. Magnetic resonance imaging (MRI) was used to determine the left ventricular ejection fraction (LVEF), infarct size (IS), and indexed end diastolic and systolic volumes (EDVi; ESVi) pre- and 10 weeks post-injection. Programmed electrical stimulation (PES) was performed to test arrhythmia inducibility before euthanasia. Histopathological analysis was carried out afterwards. The IP infusion was successful in all animals. At 10 weeks, MRI revealed significantly higher LVEF in the APA-CDC group compared with CON. No significant differences were observed among groups in IS, EDVi, ESVi, PES and histopathological analyses. In conclusion, the IP injection of CDCs (microencapsulated or not) was feasible and safe 72 h post-AMI in the porcine model. Moreover, CDCs APA encapsulation could have a beneficial effect on cardiac function, reflected by a higher LVEF at 10 weeks.

Laparoscopic Diagnosis and Treatment of Gossypiboma Postconventional Ovariohysterectomy in a Bitch.

Introduction. Gossypiboma is a rare surgical complication in small animals. The authors reported the laparoscopic diagnosis and treatment of an abdominal gossypiboma and chronic draining fistula postopen ovariohysterectomy (OVH) unresponsive to medical treatment in a bitch. Case Presentation. The patient had undergone OVH and exploratory laparotomy in other veterinary practice 3 years previously. The animal, presenting a chronic fistula, was then referred to UFSM Veterinary Hospital. Ultrasonographic examination revealed a structure compatible with a granuloma. For the laparoscopic procedure, a 3-port (two at 11 mm; one at 6 mm) access was used. Adhesiolysis and mass removal were performed by blunt dissection and bipolar electrocoagulation. The fistula was treated by mobilising the omentum through it. There were no intra- or postoperative complications. The chronic wound showed first intention healing. The mass was composed of fibrous tissues surrounding one surgical gauze. Discussion and Conclusion. The removal of a retained surgical gauze in the abdomen by laparoscopy has already been described in medicine. However, a laparoscopic approach for treating a fistulous draining tract due to a gossypiboma has not been previously reported in dogs. Laparoscopic exploration of the fistula allowed the use of a pedicled omental flap through infected sites to control chronic infection. Laparoscopic surgery can be used to identify and treat abdominal gossypiboma in dogs, including those with chronic abdominal sinus.

Preclinical Evaluation of the Safety and Immunological Action of Allogeneic ADSC-Collagen Scaffolds in the Treatment of Chronic Ischemic Cardiomyopathy.

The use of allogeneic adipose-derived mesenchymal stromal cells (alloADSCs) represents an attractive approach for treating myocardial infarction (MI). Furthermore, adding a natural support improves alloADSCs engraftment and survival in heart tissues, leading to a greater therapeutic effect. We aimed to examine the safety and immunological reaction induced by epicardial implantation of a clinical-grade collagen scaffold (CS) seeded with alloADSCs for its future application in humans. Thus, cellularized scaffolds were myocardially or subcutaneously implanted in immunosuppressed rodent models. The toxicological parameters were not significantly altered, and tumor formation was not found over the short or long term. Furthermore, biodistribution analyses in the infarcted immunocompetent rats displayed cell engraftment in the myocardium but no migration to other organs. The immunogenicity of alloADSC-CS was also evaluated in a preclinical porcine model of chronic MI; no significant humoral or cellular alloreactive responses were found. Moreover, CS cellularized with human ADSCs cocultured with human allogeneic immune cells produced no alloreactive response. Interestingly, alloADSC-CS significantly inhibited lymphocyte responses, confirming its immunomodulatory action. Thus, alloADSC-CS is likely safe and does not elicit any alloreactive immunological response in the host. Moreover, it exerts an immunomodulatory action, which supports its translation to a clinical setting.