The Oncolytic Adenovirus XVir-N-31, in Combination with the Blockade of the PD-1/PD-L1 Axis,Conveys Abscopal Effects in a Humanized Glioblastoma Mouse Model

Glioblastoma (GBM) is an obligatory lethal brain tumor with a median survival, even with the best standard of care therapy, of less than 20 months. In light of this fact, the evaluation of new GBM treatment approaches such as oncolytic virotherapy (OVT) is urgently needed. Based on our preliminary preclinical data, the YB-1 dependent oncolytic adenovirus (OAV) XVir-N-31 represents a promising therapeutic agent to treat, in particular, therapy resistant GBM. Preclinical studies have shown that XVir-N-31 prolonged the survival of GBM bearing mice. Now using an immunohumanized mouse model, we examined the immunostimulatory effects of XVir-N-31 in comparison to the wildtype adenovirus (Ad-WT). Additionally, we combined OVT with the inhibition of immune checkpoint proteins by using XVir-N-31 in combination with nivolumab, or by using a derivate of XVir-N-31 that expresses a PD-L1 neutralizing antibody. Although in vitro cell killing was higher for Ad-WT, XVir-N-31 induced a much stronger immunogenic cell death that was further elevated by blocking PD-1 or PD-L1. In vivo, an intratumoral injection of XVir-N-31 increased tumor infiltrating lymphocytes (TILs) and NK cells significantly more than Ad-WT not only in the virus-injected tumors, but also in the untreated tumors growing in the contralateral hemisphere. This suggests that for an effective treatment of GBM, immune activating properties by OAVs seem to be of greater importance than their oncolytic capacity. Furthermore, the addition of immune checkpoint inhibition (ICI) to OVT further induced lymphocyte infiltration. Consequently, a significant reduction in contralateral non-virus-injected tumors was only visible if OVT was combined with ICI. This strongly indicates that for an effective eradication of GBM cells that cannot be directly targeted by an intratumoral OV injection, additional ICI therapy is required.     

Coagulation Factor XIIIa and Activated Protein C Activate Platelets via GPVI and PAR1

Platelet and coagulation activation are highly reciprocal processes driven by multi-molecular interactions. Activated platelets secrete several coagulation factors and expose phosphatidylserine, which supports the activation of coagulation factor proteins. On the other hand, the coagulation cascade generates known ligands for platelet receptors, such as thrombin and fibrin. Coagulation factor (F)Xa, (F)XIIIa and activated protein C (APC) can also bind to platelets, but the functional consequences are unclear. Here, we investigated the effects of the activated (anti)coagulation factors on platelets, other than thrombin. Multicolor flow cytometry and aggregation experiments revealed that the ‘supernatant of (hirudin-treated) coagulated plasma’ (SCP) enhanced CRP-XL-induced platelet responses, i.e., integrin α_IIbβ₃ activation, P-selectin exposure and aggregate formation. We demonstrated that FXIIIa in combination with APC enhanced platelet activation in solution, and separately immobilized FXIIIa and APC resulted in platelet spreading. Platelet activation by FXIIIa was inhibited by molecular blockade of glycoprotein VI (GPVI) or Syk kinase. In contrast, platelet spreading on immobilized APC was inhibited by PAR1 blockade. Immobilized, but not soluble, FXIIIa and APC also enhanced in vitro adhesion and aggregation under flow. In conclusion, in coagulation, factors other than thrombin or fibrin can induce platelet activation via GPVI and PAR receptors.     

Unhealthy Diets Induce Distinct and Regional Effects on Intestinal Inflammatory Signalling Pathways and Long-Lasting Metabolic Dysfunction in Rats

The intestinal epithelium is a principal site for environmental agents’ detection. Several inflammation- and stress-related signalling pathways have been identified as key players in these processes. However, it is still unclear how the chronic intake of inadequate nutrients triggers inflammatory signalling pathways in different intestinal regions. We aimed to evaluate the impact of unhealthy dietary patterns, starting at a younger age, and the association with metabolic dysfunction, intestinal inflammatory response, and obesity in adulthood. A rat model was used to evaluate the effects of the consumption of sugary beverages (HSD) and a Western diet (WD), composed of ultra-processed foods. Both diets showed a positive correlation with adiposity index, but a positive correlation was found between the HSD diet and the levels of blood glucose and triglycerides, whereas the WD diet correlated positively with triglyceride levels. Moreover, a distinct inflammatory response was associated with either the WD or HSD diets. The WD induced an increase in TLR2, TLR4, and nuclear factor-kappa B (NF-κB) intestinal gene expression, with higher levels in the colon and overexpression of the inducible nitric oxide synthase. In turn, the HSD diet induced activation of the TLR2-mediated NF-κB signalling pathway in the small intestine. Altogether, these findings support the concept that early intake of unhealthy foods and nutrients are a main exogenous signal for disturbances of intestinal immune mechanisms and in a region-specific manner, ultimately leading to obesity-related disorders in later life.     

Differential Plasma Metabolites between High- and Low-Grade Meningioma Cases

Meningiomas (MGMs) are currently classified into grades I, II, and III. High-grade tumors are correlated with decreased survival rates and increased recurrence rates. The current grading classification is based on histological criteria and determined only after surgical tumor sampling. This study aimed to identify plasma metabolic alterations in meningiomas of different grades, which would aid surgeons in predefining the ideal surgical strategy. Plasma samples were collected from 51 patients with meningioma and classified into low-grade (LG) (grade I; n = 43), and high-grade (HG) samples (grade II, n = 5; grade III, n = 3). An untargeted metabolomic approach was used to analyze plasma metabolites. Statistical analyses were performed to select differential biomarkers among HG and LG groups. Metabolites were identified using tandem mass spectrometry along with database verification. Five and four differential biomarkers were identified for HG and LG meningiomas, respectively. To evaluate the potential of HG MGM metabolites to differentiate between HG and LG tumors, a receiving operating characteristic curve was constructed, which revealed an area under the curve of 95.7%. This indicates that the five HG MGM metabolites represent metabolic alterations that can differentiate between LG and HG meningiomas. These metabolites may indicate tumor grade even before the appearance of histological features.     

Targeting Lysosomes in Colorectal Cancer: Exploring the Anticancer Activity of a New Benzo[a]phenoxazine Derivative

Colorectal cancer (CRC) has been ranked as one of the cancer types with a higher incidence and one of the most mortal. There are limited therapies available for CRC, which urges the finding of intracellular targets and the discovery of new drugs for innovative therapeutic approaches. In addition to the limited number of effective anticancer agents approved for use in humans, CRC resistance and secondary effects stemming from classical chemotherapy remain a major clinical problem, reinforcing the need for the development of novel drugs. In the recent years, the phenoxazines derivatives, Nile Blue analogues, have been shown to possess anticancer activity, which has created interest in exploring the potential of these compounds as anticancer drugs. In this context, we have synthetized and evaluated the anticancer activity of different benzo[a]phenoxazine derivatives for CRC therapy. Our results revealed that one particular compound, BaP1, displayed promising anticancer activity against CRC cells. We found that BaP1 is selective for CRC cells and reduces cell proliferation, cell survival, and cell migration. We observed that the compound is associated with reactive oxygen species (ROS) generation, accumulates in the lysosomes, and leads to lysosomal membrane permeabilization, cytosolic acidification, and apoptotic cell death. In vivo results using a chicken embryo choriollantoic membrane (CAM) assay showed that BaP1 inhibits tumor growth, angiogenesis, and tumor proliferation. These observations highlight that BaP1 as a very interesting agent to disturb and counteract the important roles of lysosomes in cancer and suggests BaP1 as a promising candidate to be exploited as new anticancer lysosomal-targeted agent, which uses lysosome membrane permeabilization (LMP) as a therapeutic approach in CRC.     

Relationships between Diabetes and the Intestinal Microbial Population

Diabetes is a metabolic disorder characterized by lower responsiveness of tissues to insulin and consequent large variations in circulating levels of glucose. This fluctuation has harmful effects as both hyperglycemia and hypoglycemia can be very injurious. The causes of diabetes are varied but the consequences are rather uniform. Dietary factors are important especially in adult onset type 2 diabetes (T2D) while type 1 diabetes (T1D) is characterized by having a stronger heritable component and involving autoimmune attach on pancreatic beta cells. This review is focused on the relation of the bacterial components found within the intestine, to the establishment and maintenance of diabetes. The precise composition of the gut microbiome is increasingly recognized as a factor in organismic health and its interaction with a variety of disease states has been described. This is especially marked in the case of diabetes since the nature of the diet is an important factor in establishing both the microbiome and the incidence of diabetes. The bidirectional nature of this relationship is discussed. The effects of disease that lead to altered microbiomal composition together with aberrant metabolic changes are also included. Emphasis is given to the important role of short chain fatty acids (SCFAs) as mediators of the microbiome-diabetes relation.     

Relevance of the Exocyst in Arabidopsis exo70e2 Mutant for Cellular Homeostasis under Stress

Plants must adapt to cope with adverse environmental conditions that affect their growth and development. To overcome these constraints, they can alter their developmental patterns by modulating cellular processes and activating stress-responsive signals. Alongside the activation of the antioxidant (AOX) system, a high number of genes are expressed, and proteins must be distributed to the correct locations within the cell. The endomembrane system and associated vesicles thus play an important role. Several pathways have been associated with adverse environmental conditions, which is the case for the exocyst-positive organelle—EXPO. The present work, using Arabidopsis mutants with T-DNA insertions in the gene EXO70, essential for EXPO vesicles formation, was designed to characterise the anatomical (morphology and root length), biochemical (quantification of stress markers and antioxidant system components), and molecular responses (gene expression) to abiotic stresses (saline, drought, oxidative, and metal-induced toxicity). The results obtained showed that mutant plants behave differently from the wild type (WT) plants. Therefore, in the exo70 mutant, morphological changes were more noticeable in plants under stress, and the non-enzymatic component of the antioxidant system was activated, with no alterations to the enzymatic component. Furthermore, other defence strategies, such as autophagy, did not show important changes. These results confirmed the EXPO as an important structure for tolerance/adaptation to stress.     

Relative Contribution of Blood Pressure and Renal Sympathetic Nerve Activity to Proximal Tubular Sodium Reabsorption via NHE3 Activity

We examined the effects of an acute increase in blood pressure (BP) and renal sympathetic nerve activity (rSNA) induced by bicuculline (Bic) injection in the paraventricular nucleus of hypothalamus (PVN) or the effects of a selective increase in rSNA induced by renal nerve stimulation (RNS) on the renal excretion of sodium and water and its effect on sodium-hydrogen exchanger 3 (NHE3) activity. Uninephrectomized anesthetized male Wistar rats were divided into three groups: (1) Sham; (2) Bic PVN: (3) RNS + Bic injection into the PVN. BP and rSNA were recorded, and urine was collected prior and after the interventions in all groups. RNS decreased sodium (58%) and water excretion (53%) independently of BP changes (p < 0.05). However, after Bic injection in the PVN during RNS stimulation, the BP and rSNA increased by 30% and 60% (p < 0.05), respectively, diuresis (5-fold) and natriuresis (2.3-fold) were increased (p < 0.05), and NHE3 activity was significantly reduced, independently of glomerular filtration rate changes. Thus, an acute increase in the BP overcomes RNS, leading to diuresis, natriuresis, and NHE3 activity inhibition.     

COVID-19 Mimics Pulmonary Dysfunction in Muscular Dystrophy as a Post-Acute Syndrome in Patients

Although progressive wasting and weakness of respiratory muscles are the prominent hallmarks of Duchenne muscular dystrophy (DMD) and long-COVID (also referred as the post-acute sequelae of COVID-19 syndrome); however, the underlying mechanism(s) leading to respiratory failure in both conditions remain unclear. We put together the latest relevant literature to further understand the plausible mechanism(s) behind diaphragm malfunctioning in COVID-19 and DMD conditions. Previously, we have shown the role of matrix metalloproteinase-9 (MMP9) in skeletal muscle fibrosis via a substantial increase in the levels of tumor necrosis factor-α (TNF-α) employing a DMD mouse model that was crossed-bred with MMP9-knockout (MMP9-KO or MMP9^-/-) strain. Interestingly, recent observations from clinical studies show a robust increase in neopterin (NPT) levels during COVID-19 which is often observed in patients having DMD. What seems to be common in both (DMD and COVID-19) is the involvement of neopterin (NPT). We know that NPT is generated by activated white blood cells (WBCs) especially the M1 macrophages in response to inducible nitric oxide synthase (iNOS), tetrahydrobiopterin (BH4), and tetrahydrofolate (FH4) pathways, i.e., folate one-carbon metabolism (FOCM) in conjunction with epigenetics underpinning as an immune surveillance protection. Studies from our laboratory, and others researching DMD and the genetically engineered humanized (hACE2) mice that were administered with the spike protein (SP) of SARS-CoV-2 revealed an increase in the levels of NPT, TNF-α, HDAC, IL-1β, CD147, and MMP9 in the lung tissue of the animals that were subsequently accompanied by fibrosis of the diaphragm depicting a decreased oscillation phenotype. Therefore, it is of interest to understand how regulatory processes such as epigenetics involvement affect DNMT, HDAC, MTHFS, and iNOS that help generate NPT in the long-COVID patients.