Optimizing cryopreservation conditions for use of fucosylated human mesenchymal stromal cells in anti-inflammatory/immunomodulatory therapeutics.

Mesenchymal stem/stromal cells (MSCs) are being increasingly used in cell-based therapies due to their broad anti-inflammatory and immunomodulatory properties. Intravascularly-administered MSCs do not efficiently migrate to sites of inflammation/immunopathology, but this shortfall has been overcome by cell surface enzymatic fucosylation to engender expression of the potent E-selectin ligand HCELL. In applications of cell-based therapies, cryopreservation enables stability in both storage and transport of the produced cells from the manufacturing facility to the point of care. However, it has been reported that cryopreservation and thawing dampens their immunomodulatory/anti-inflammatory activity even after a reactivation/reconditioning step. To address this issue, we employed a variety of methods to cryopreserve and thaw fucosylated human MSCs derived from either bone marrow or adipose tissue sources. We then evaluated their immunosuppressive properties, cell viability, morphology, proliferation kinetics, immunophenotype, senescence, and osteogenic and adipogenic differentiation. Our studies provide new insights into the immunobiology of cryopreserved and thawed MSCs and offer a readily applicable approach to optimize the use of fucosylated human allogeneic MSCs as immunomodulatory/anti-inflammatory therapeutics.

The TP53-activated E3 ligase RNF144B is a tumour suppressor that prevents genomic instability.

BACKGROUND: TP53, the most frequently mutated gene in human cancers, orchestrates a complex transcriptional program crucial for cancer prevention. While certain TP53-dependent genes have been extensively studied, others, like the recently identified RNF144B, remained poorly understood. This E3 ubiquitin ligase has shown potent tumor suppressor activity in murine Eµ Myc-driven lymphoma, emphasizing its significance in the TP53 network. However, little is known about its targets and its role in cancer development, requiring further exploration. In this work, we investigate RNF144B's impact on tumor suppression beyond the hematopoietic compartment in human cancers. METHODS: Employing TP53 wild-type cells, we generated models lacking RNF144B in both non-transformed and cancerous cells of human and mouse origin. By using proteomics, transcriptomics, and functional analysis, we assessed RNF144B's impact in cellular proliferation and transformation. Through in vitro and in vivo experiments, we explored proliferation, DNA repair, cell cycle control, mitotic progression, and treatment resistance. Findings were contrasted with clinical datasets and bioinformatics analysis. RESULTS: Our research underscores RNF144B's pivotal role as a tumor suppressor, particularly in lung adenocarcinoma. In both human and mouse oncogene-expressing cells, RNF144B deficiency heightened cellular proliferation and transformation. Proteomic and transcriptomic analysis revealed RNF144B's novel function in mediating protein degradation associated with cell cycle progression, DNA damage response and genomic stability. RNF144B deficiency induced chromosomal instability, mitotic defects, and correlated with elevated aneuploidy and worse prognosis in human tumors. Furthermore, RNF144B-deficient lung adenocarcinoma cells exhibited resistance to cell cycle inhibitors that induce chromosomal instability. CONCLUSIONS: Supported by clinical data, our study suggests that RNF144B plays a pivotal role in maintaining genomic stability during tumor suppression.

Retinal response to systemic inflammation differs between sexes and neurons.

Background: Neurological dysfunction and glial activation are common in severe infections such as sepsis. There is a sexual dimorphism in the response to systemic inflammation in both patients and animal models, but there are few comparative studies. Here, we investigate the effect of systemic inflammation induced by intraperitoneal administration of lipopolysaccharide (LPS) on the retina of male and female mice and determine whether antagonism of the NLRP3 inflammasome and the extrinsic pathway of apoptosis have protective effects on the retina. Methods: A single intraperitoneal injection of LPS (5 mg/kg) was administered to two months old C57BL/6J male and female mice. Retinas were examined longitudinally in vivo using electroretinography and spectral domain optical coherence tomography. Retinal ganglion cell (RGC) survival and microglial activation were analysed in flat-mounts. Retinal extracts were used for flow cytometric analysis of CD45 and CD11b positive cells. Matched plasma and retinal levels of proinflammatory cytokines were measured by ELISA. Retinal function and RGC survival were assessed in animals treated with P2X7R and TNFR1 antagonists alone or in combination. Results: In LPS-treated animals of both sexes, there was transient retinal dysfunction, loss of vision-forming but not non-vision forming RGCs, retinal swelling, microglial activation, cell infiltration, and increases in TNF and IL-1ß. Compared to females, males showed higher vision-forming RGC death, slower functional recovery, and overexpression of lymphotoxin alpha in their retinas. P2X7R and TNFR1 antagonism, alone or in combination, rescued vision-forming RGCs. P2X7R antagonism also rescued retinal function. Response to treatment was better in females than in males. Conclusions: Systemic LPS has neuronal and sex-specific adverse effects in the mouse retina, which are counteracted by targeting the NLRP3 inflammasome and the extrinsic pathway of apoptosis. Our results highlight the need to analyse males and females in preclinical studies of inflammatory diseases affecting the central nervous system.

Differential utilization of surface and arboreal water bodies by birds and mammals in a seasonally dry Neotropical forest in southern Mexico.

Water availability significantly influences bird and mammal ecology in terrestrial ecosystems. However, our understanding of the role of water as a limiting resource for birds and mammals remains partial because most of the studies have focused on surface water bodies in desert and semi-desert ecosystems. This study assessed the use of two types of surface water bodies (waterholes and epikarst rock pools) and one arboreal (water-filled tree holes) by birds and mammals in the seasonally dry tropical forests of the Calakmul Biosphere Reserve in southern Mexico. We deployed camera traps in 23 waterholes, 22 rock pools, and 19 water-filled tree holes in this karstic region to record visits by small, medium, and large-bodied birds and mammals during the dry and rainy seasons. These cameras were set up for recording videos documenting when animals were making use of water for drinking, bathing, or both. We compared the species diversity and composition of bird and mammal assemblages using the different types of water bodies by calculating Hill numbers and conducting nonmetric multidimensional scaling (NMDS), indicator species, and contingency table analyses. There was a greater species richness of birds and mammals using surface water bodies than tree holes during both seasons. There were significant differences in species composition among bird assemblages using the different water bodies, but dominant species and diversity remained the same. Terrestrial and larger mammalian species preferentially used surface water bodies, whereas arboreal and scansorial small and medium mammals were more common in arboreal water bodies. These findings suggest that differences in water body characteristics might favor segregation in mammal activity. The different water bodies may act as alternative water sources for birds and complementary sources for mammals, potentially favoring species coexistence and increasing community resilience to environmental variation (e.g., fluctuations in water availability). Understanding how differences in water bodies favor species coexistence and community resilience is of great relevance from a basic ecological perspective but is also crucial for anticipating the effects that the increased demand for water by humans and climate change can have on wildlife viability.

Molecular mechanisms of postoperative atrial fibrillation in patients with obstructive sleep apnea.

Obstructive sleep apnea (OSA) promotes atrial remodeling and fibrosis, providing a substrate for atrial fibrillation (AF). Herein, we investigate the pathophysiological mechanisms of AF in association with OSA in a cohort of cardiac surgery patients. A prospective study including patients undergoing cardiac surgery. Biomarkers reflective of AF pathophysiology (interleukin [IL-6], C-reactive protein [CRP], von Willebrand factor [vWF], N-terminal pro-brain natriuretic peptide [NT-proBNP], high-sensitivity Troponin T [hs-TnT], and Galectin-3 [Gal-3]) was assessed by functional or immunological assays. miRNAs involved in AF were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Using atrial tissue samples, fibrosis was assessed by Masson's trichrome. Connexin 40 and 43 (Cx40; Cx43) were evaluated by immunolabeling. Fifty-six patients (15 with OSA and 41 non-OSA) were included in this hypothesis-generating pilot study. OSA group had a higher incidence of postoperative AF (POAF) (46.7% vs. 19.5%; p = .042), presented an increased risk of POAF (OR 3.61, 95% CI 1.01-12.92), and had significantly higher baseline levels of NT-proBNP (p = .044), vWF (p = .049), Gal-3 (p = .009), IL-6 (p = .002), and CRP (p = .003). This group presented lower levels of miR-21 and miR-208 (both p < .05). Also, lower Cx40 levels in POAF and/or OSA patients (50.0% vs. 81.8%, p = .033) were found. The presence of interstitial fibrosis (according to myocardial collagen by Masson's trichrome) was raised in OSA patients (86.7% vs. 53.7%, p = .024). Several biomarkers and miRNAs involved in inflammation and fibrosis were dysregulated in OSA patients, which together with a higher degree of interstitial fibrosis, altered miRNA, and Cxs expression predisposes to the development of a substrate that increases the AF risk.

Enforced mesenchymal stem cell tissue colonization counteracts immunopathology.

Mesenchymal stem/stromal cells (MSCs) are distributed within all tissues of the body. Though best known for generating connective tissue and bone, these cells also display immunoregulatory properties. A greater understanding of MSC cell biology is urgently needed because culture-expanded MSCs are increasingly being used in treatment of inflammatory conditions, especially life-threatening immune diseases. While studies in vitro provide abundant evidence of their immunomodulatory capacity, it is unknown whether tissue colonization of MSCs is critical to their ability to dampen/counteract evolving immunopathology in vivo. To address this question, we employed a murine model of fulminant immune-mediated inflammation, acute graft-versus-host disease (aGvHD), provoked by donor splenocyte-enriched full MHC-mismatched hematopoietic stem cell transplant. aGvHD induced the expression of E-selectin within lesional endothelial beds, and tissue-specific recruitment of systemically administered host-derived MSCs was achieved by enforced expression of HCELL, a CD44 glycoform that is a potent E-selectin ligand. Compared to mice receiving HCELL- MSCs, recipients of HCELL+ MSCs had increased MSC intercalation within aGvHD-affected site(s), decreased leukocyte infiltrates, lower systemic inflammatory cytokine levels, superior tissue preservation, and markedly improved survival. Mechanistic studies reveal that ligation of HCELL/CD44 on the MSC surface markedly potentiates MSC immunomodulatory activity by inducing MSC secretion of a variety of potent immunoregulatory molecules, including IL-10. These findings indicate that MSCs counteract immunopathology in situ, and highlight a role for CD44 engagement in unleashing MSC immunobiologic properties that maintain/establish tissue immunohomeostasis.

Fatigue and Dyspnoea as Main Persistent Post-COVID-19 Symptoms in Previously Hospitalized Patients: Related Functional Limitations and Disability.

BACKGROUND: Multicentre studies focussing on specific long-term post-COVID-19 symptoms are scarce. OBJECTIVE: The aim of this study was to determine the levels of fatigue and dyspnoea, repercussions on daily life activities, and risk factors associated with fatigue or dyspnoea in COVID-19 survivors at long term after hospital discharge. METHODS: Age, gender, height, weight, symptoms at hospitalization, pre-existing medical comorbidity, intensive care unit admission, and the presence of cardio-respiratory symptoms developed after severe acute respiratory syndrome coronavirus 2 infection were collected from patients who recovered from COVID-19 at 4 hospitals in Madrid (Spain) from March 1 to May 31, 2020 (first COVID-19 wave). The Functional Impairment Checklist was used for evaluating fatigue/dyspnoea levels and functional limitations. RESULTS: A total of 1,142 patients (48% women, age: 61, standard deviation [SD]: 17 years) were assessed 7.0 months (SD 0.6) after hospitalization. Fatigue was present in 61% patients, dyspnoea with activity in 55%, and dyspnoea at rest in 23.5%. Only 355 (31.1%) patients did not exhibit fatigue and/or dyspnoea 7 months after hospitalization. Forty-five per cent reported functional limitations with daily living activities. Risk factors associated with fatigue and dyspnoea included female gender, number of pre-existing comorbidities, and number of symptoms at hospitalization. The number of days at hospital was a risk factor just for dyspnoea. CONCLUSIONS: Fatigue and/or dyspnoea were present in 70% of hospitalized COVID-19 survivors 7 months after discharge. In addition, 45% patients exhibited limitations on daily living activities. Being female, higher number of pre-existing medical comorbidities and number of symptoms at hospitalization were risk factors associated to fatigue/dyspnoea in COVID-19 survivors 7 months after hospitalization.

Obesity is associated with a greater number of long-term post-COVID symptoms and poor sleep quality: A multicentre case-control study.

BACKGROUND: Obesity is a risk factor associated with higher mortality at the acute phase of COVID-19; however, its influence on post-COVID symptoms is not known. OBJECTIVE: Our aim was to investigate if obesity is a risk factor for the presence of long-term post-COVID symptoms in hospitalised COVID-19 survivors. METHODS: A multicentre case-control study including patients hospitalised during the first wave of the pandemic was performed. Patients with obesity were recruited as cases. Two age- and sex-matched patients without obesity per case were considered as controls. Clinical and hospitalisation data were collected from the hospital medical records. Patients were scheduled for a telephonic interview. A list of post-COVID symptoms was systematically evaluated, but participants were free to report any symptom. Anxiety/depressive levels and sleep quality were evaluated with the hospital anxiety and depression scale (HADS) and Pittsburgh sleep quality index (PSQI), respectively. RESULTS: Overall, 88 patients with obesity and 176 without obesity were assessed 7.2 months after the hospital discharge. The most prevalent post-COVID symptoms were fatigue and dyspnea. No significant difference in the prevalence of fatigue, dyspnea, anxiety, depression and limitations of daily living activities was observed between people with and without obesity. Obesity was independently associated with a greater number of post-COVID symptoms (IRR 1.56, 95% CI 1.24-1.95, P < .001) and poor sleep quality (OR 2.10, 95% CI 1.13-3.83, P = .02). CONCLUSIONS: This study found that obesity was associated with a greater number of long-term post-COVID symptoms and poor sleep quality in hospitalised COVID-19 patients.

Gender differences in the renal changes induced by a prolonged high-fat diet in rats with altered renal development.

The mechanisms involved in renal dysfunction induced by high-fat diet (HFD) in subjects with altered renal development (ARDev) are understudied. The objective of this study is to examine whether there are sex-dependent differences in the mechanisms involved in the hypertension and deterioration of renal function in SD rats with prolonged HFD and ARDev. The role of angiotensin II (Ang II) in the arterial pressure (AP) increments, the renal hemodynamic sensitivity to Ang II, glomerular damage and changes in fat abdominal volume, plasma adipokine levels, renal NADPHp67phox expression, and renal infiltration of immune cells were examined. Hypertension and deterioration of renal function were enhanced (P < 0.05) in both sexes of rats with HFD and ARDev. The decrease (P < 0.05) of AP elicited by candesartan in hypertensive rats was similar to that induced by the simultaneous administration of candesartan and apocynin. The greater (P < 0.05) renal vasoconstriction induced by Ang II in both sexes of rats with HFD and ARDev was accompanied by an enhanced (P < 0.05) infiltration of CD-3 cells and macrophages in the renal cortex and renal medulla. The increments (P < 0.05) in the renal expression of NADPHp67phox and glomeruloesclerosis were greater (P < 0.05) in males than in females with HFD and ARDev. Our results suggest that the hypertension and deterioration of renal function induced by HFD in rats with ARDev are Ang II-dependent and mediated by increments in oxidative stress and immune system activation. Sex-dependent increments in oxidative stress and glomerular damage may contribute to the deterioration of renal function in these rats.

Efficiency of Machine Learning Algorithms for the Determination of Macrovesicular Steatosis in Frozen Sections Stained with Sudan to Evaluate the Quality of the Graft in Liver Transplantation.

Liver transplantation is the only curative treatment option in patients diagnosed with end-stage liver disease. The low availability of organs demands an accurate selection procedure based on histological analysis, in order to evaluate the allograft. This assessment, traditionally carried out by a pathologist, is not exempt from subjectivity. In this sense, new tools based on machine learning and artificial vision are continuously being developed for the analysis of medical images of different typologies. Accordingly, in this work, we develop a computer vision-based application for the fast and automatic objective quantification of macrovesicular steatosis in histopathological liver section slides stained with Sudan stain. For this purpose, digital microscopy images were used to obtain thousands of feature vectors based on the RGB and CIE L*a*b* pixel values. These vectors, under a supervised process, were labelled as fat vacuole or non-fat vacuole, and a set of classifiers based on different algorithms were trained, accordingly. The results obtained showed an overall high accuracy for all classifiers (>0.99) with a sensitivity between 0.844 and 1, together with a specificity >0.99. In relation to their speed when classifying images, KNN and Naïve Bayes were substantially faster than other classification algorithms. Sudan stain is a convenient technique for evaluating ME in pre-transplant liver biopsies, providing reliable contrast and facilitating fast and accurate quantification through the machine learning algorithms tested.

CD14 release induced by P2X7 receptor restricts inflammation and increases survival during sepsis.

P2X7 receptor activation induces the release of different cellular proteins, such as CD14, a glycosylphosphatidylinositol (GPI)-anchored protein to the plasma membrane important for LPS signaling via TLR4. Circulating CD14 has been found at elevated levels in sepsis, but the exact mechanism of CD14 release in sepsis has not been established. Here, we show for first time that P2X7 receptor induces the release of CD14 in extracellular vesicles, resulting in a net reduction in macrophage plasma membrane CD14 that functionally affects LPS, but not monophosphoryl lipid A, pro-inflammatory cytokine production. Also, we found that during a murine model of sepsis, P2X7 receptor activity is important for maintaining elevated levels of CD14 in biological fluids and a decrease in its activity results in higher bacterial load and exacerbated organ damage, ultimately leading to premature deaths. Our data reveal that P2X7 is a key receptor for helping to clear sepsis because it maintains elevated concentrations of circulating CD14 during infection.

When the immune system detects an infection, it often launches an inflammatory response to fight off the disease. This defense mechanism is activated by a cascade of signaling molecules that can aggravate inflammation, causing it to damage the body's own tissues and organs. This life-threatening reaction is referred to as sepsis, and kills around 11 million people each year. New approaches are therefore needed to help alleviate the damage caused by this condition. The inflammatory response is often triggered by proteins called receptors, which sit on the surface of immune cells. When these receptors are activated, they induce cells to secrete proteins that travel around the body and activate immune cells that can eliminate the infection. In 2016, a group of researchers showed that a receptor called P2X7 stimulates the release of a signaling molecule called CD14. Patients with sepsis often have elevated amounts of CD14 in their bloodstream. Yet, it remained unclear what causes this rise in CD14 and what role this molecule plays in the development of sepsis. Now, Alarcón-Vila et al. ­ including some of the researchers involved in the 2016 study ­ have investigated the role of P2X7 in mice undergoing sepsis. This was done by puncturing the mice's intestines, causing bacteria to leak out and initiate an over-active immune response. Alarcón-Vila et al. found that mice lacking the P2X7 receptor had less CD14 and struggled to eliminate the bacterial infection from their system. This increase in bacteria caused excessive damage to the mice's organs, ultimately leading to premature death. These findings suggest that P2X7 plays an important role in preventing the onset of sepsis by helping maintain high levels of CD14 following infection. This result could help to identify new therapies that reduce the mortality rates of septic infections.

Defibrotide inhibits donor leucocyte-endothelial interactions and protects against acute graft-versus-host disease.

Allogeneic hematopoietic stem cell transplantation (allo-HCT) is an effective therapy for the treatment of high-risk haematological malignant disorders and other life-threatening haematological and genetic diseases. Acute graft-versus-host disease (aGvHD) remains the most frequent cause of non-relapse mortality following allo-HCT and limits its extensive clinical application. Current pharmacologic agents used for prophylaxis and treatment of aGvHD are not uniformly successful and have serious secondary side effects. Therefore, more effective and safe prophylaxis and therapy for aGvHD are an unmet clinical need. Defibrotide is a multi-target drug successfully employed for prophylaxis and treatment of veno-occlusive disease/sinusoidal obstruction syndrome. Recent preliminary clinical data have suggested some efficacy of defibrotide in the prevention of aGvHD after allo-HCT. Using a fully MHC-mismatched murine model of allo-HCT, we report here that defibrotide, either in prophylaxis or treatment, is effective in preventing T cell and neutrophil infiltration and aGvHD-associated tissue injury, thus reducing aGvHD incidence and severity, with significantly improved survival after allo-HCT. Moreover, we performed in vitro mechanistic studies using human cells revealing that defibrotide inhibits leucocyte-endothelial interactions by down-regulating expression of key endothelial adhesion molecules involved in leucocyte trafficking. Together, these findings provide evidence that defibrotide may represent an effective and safe clinical alternative for both prophylaxis and treatment of aGvHD after allo-HCT, paving the way for new therapeutic approaches.

Allogeneic Bone Marrow Mesenchymal Stem Cell Transplantation in Tooth Extractions Sites Ameliorates the Incidence of Osteonecrotic Jaw-Like Lesions in Zoledronic Acid-Treated Rats.

Medication-related osteonecrosis of the jaw (MRONJ) is defined as the exposed necrotic bone involving the maxillofacial structures in bisphosphonate treated patients, and the pathophysiology of this disease remains unclear. The aim of this study was to assess the effects of the allogeneic transplantation of bone marrow-derived mesenchymal stem cells (BM-MSCs) in a model of Wistar mice with induced MRONJ disease. BM-MSCs from five male Wistar rats were characterized and cultured on ß-tricalcium phosphate (ß-TCP) granules. Thirty female Wistar rats were injected intraperitoneally with zoledronic acid and afterwards upper jaw molars were extracted. The animals were randomized to receive: Group 1: 1 × 106 BM-MSCs/ß-TCP construct in the alveolar socket; and Group 2: Saline solution/ß-TCP construct. A clinical and histological analysis was performed. Nested polymerase chain reaction (PCR) was assessed to verify the presence of transplanted male rat cells in the female recipient jaws. Clinical and histological findings evidenced that none of the animals in Group 1 exhibited uncovered sockets or bone exposure associated to MRONJ, whereas we detected 33% of MRONJ cases in Group 2. In addition, male rat cells were detected in the maxillae site four weeks after transplantation in the BM-MSCs-group. Allogeneic BM-MSCs in extractions sites ameliorates MRONJ incidence in zoledronic acid-treated rats compared to non-MSC treatments.

Melatonin as an Agent for Direct Pulp-Capping Treatment.

Melatonin plays an essential role in the regulation of bone growth. The actions that melatonin exerts on odontoblasts may be similar to its action on osteoblasts. This research aimed to evaluate the pulp response to melatonin used for direct pulp capping to evaluate the antioxidant effect of melatonin administered orally and its influence on dental pulp. Direct pulp capping was performed on the upper molars of Sprague Dawley rats using melatonin or Mineral Trioxide Aggregate (MTA). The study groups were: MTA; Melatonin; MTA + Melatonin administered orally; and Melatonin + Melatonin administered orally. In the latter two groups, the animals drank water dosed with melatonin ad libitum (10 mg/100 mL). After 30 days, the animals were sacrificed, and 5 ml of blood, the kidneys, and the liver were extracted in order to evaluate oxidative stress using thiobarbituric acid reactive substances testing (TBARS). Fragments of the maxilla containing the study molars were prepared for histological evaluation. The degree of pulp inflammation and pulp necrosis, the presence of reparative dentin and dentin bridging the pulp chamber, the presence and regularity of the odontoblastic layer, and the presence of pulp fibrosis were evaluated. No significant differences were found between the four study groups for any of the studied histological variables. The oral administration of melatonin did not modify the local effects of MTA or melatonin on dental pulp, or reduce basal-level oxidative stress. The effect of melatonin on pulp is similar to that of MTA and may be used as an agent for direct pulp capping.

Glioblastoma ablates pericytes antitumor immune function through aberrant up-regulation of chaperone-mediated autophagy.

The contractile perivascular cells, pericytes (PC), are hijacked by glioblastoma (GB) to facilitate tumor progression. PC's protumorigenic function requires direct interaction with tumor cells and contributes to the establishment of immunotolerance to tumor growth. Cancer cells up-regulate their own chaperone-mediated autophagy (CMA), a process that delivers selective cytosolic proteins to lysosomes for degradation, with pro-oncogenic effects. However, the possible impact that cancer cells may have on CMA of surrounding host cells has not been explored. We analyzed the contribution of CMA to the GB-induced changes in PC biology. We have found that CMA is markedly up-regulated in PC in response to the oxidative burst that follows PC-GB cell interaction. Genetic manipulations to block the GB-induced up-regulation of CMA in PC allows them to maintain their proinflammatory function and to support the induction of effective antitumor T cell responses required for GB clearance. GB-induced up-regulation of CMA activity in PC is essential for their effective interaction with GB cells that help tumor growth. We show that CMA inhibition in PC promotes GB cell death and the release of high immunogenic levels of granulocyte-macrophage colony stimulating factor (GM-CSF), through deregulation of the expression of cell-to-cell interaction proteins and protein secretion. A GB mouse model grafted in vivo with CMA-defective PC shows reduced GB proliferation and effective immune response compared to mice grafted with control PC. Our findings identify abnormal up-regulation of CMA as a mechanism by which GB cells elicit the immunosuppressive function of PC and stabilize GB-PC interactions necessary for tumor cell survival.

P2X7 receptor induces mitochondrial failure in monocytes and compromises NLRP3 inflammasome activation during sepsis.

Sepsis is characterized by a systemic inflammatory response followed by immunosuppression of the host. Metabolic defects and mitochondrial failure are common in immunocompromised patients with sepsis. The NLRP3 inflammasome is important for establishing an inflammatory response after activation by the purinergic P2X7 receptor. Here, we study a cohort of individuals with intra-abdominal origin sepsis and show that patient monocytes have impaired NLRP3 activation by the P2X7 receptor. Furthermore, most sepsis-related deaths are among patients whose NLRP3 activation is profoundly altered. In monocytes from sepsis patients, the P2X7 receptor is associated with mitochondrial dysfunction. Furthermore, activation of the P2X7 receptor results in mitochondrial damage, which in turn inhibits NLRP3 activation by HIF-1α. We show that mortality increases in a mouse model of sepsis when the P2X7 receptor is activated in vivo. These data reveal a molecular mechanism initiated by the P2X7 receptor that contributes to NLRP3 impairment during infection.

Evaluation of rat liver with ARFI elastography: In vivo and ex vivo study.

OBJECTIVE: The aim of this study was to compare in vivo vs ex vivo liver stiffness in rats with acoustic radiation force impulse (ARFI) elastography using the histological findings as the gold standard. METHODS: Eighteen male Wistar rats aged 16-18 months were divided into a control group (n = 6) and obese group (n = 12). Liver stiffness was measured with shear wave velocity (SWV) using the ARFI technique both in vivo and ex vivo. The degree of fibrosis, steatosis and liver inflammation was evaluated in the histological findings. Pearson's correlation coefficient was applied to relate the SWV values to the histological parameters. RESULTS: The SWV values acquired in the ex vivo study were significantly lower than those obtained in vivo (P < 0.004). A significantly higher correlation value between the degree of liver fibrosis and the ARFI elastography assessment was observed in the ex vivo study (r = 0.706, P < 0.002), than the in vivo study (r = 0.623, P < 0.05). CONCLUSION: Assessment of liver stiffness using ARFI elastography yielded a significant correlation between SWV and liver fibrosis in both the in vivo and ex vivo experiments. We consider that by minimising the influence of possible sources of artefact we could improve the accuracy of the measurements acquired with ARFI.

Silk fibroin scaffolds seeded with Wharton's jelly mesenchymal stem cells enhance re-epithelialization and reduce formation of scar tissue after cutaneous wound healing.

BACKGROUND: The treatment of extensive and/or chronic skin wounds is a widespread and costly public health problem. Mesenchymal stem cells (MSCs) have been proposed as a potential cell therapy for inducing wound healing in different clinical settings, alone or in combination with biosynthetic scaffolds. Among them, silk fibroin (SF) seeded with MSCs has been shown to have increased efficacy in skin wound healing experimental models. METHODS: In this report, we investigated the wound healing effects of electrospun SF scaffolds cellularized with human Wharton's jelly MSCs (Wj-MSCs-SF) using a murine excisional wound splinting model. RESULTS: Immunohistopathological examination after transplant confirmed the presence of infiltrated human fibroblast-like CD90-positive cells in the dermis of the Wj-MSCs-SF-treated group, yielding neoangiogenesis, decreased inflammatory infiltrate and myofibroblast proliferation, less collagen matrix production, and complete epidermal regeneration. CONCLUSIONS: These findings indicate that Wj-MSCs transplanted in the wound bed on a silk fibroin scaffold contribute to the generation of a well-organized and vascularized granulation tissue, enhance reepithelization of the wound, and reduce the formation of fibrotic scar tissue, highlighting the potential therapeutic effects of Wj-MSC-based tissue engineering approaches to non-healing wound treatment.