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Delayed wound healing is a devastating complication of diabetes and supplementation with fish oil, a source of anti-inflammatory omega-3 (ω-3) fatty acids including eicosapentaenoic acid (EPA), seems an appealing treatment strategy. However, some studies have shown that ω-3 fatty acids may have a deleterious effect on skin repair and the effects of oral administration of EPA on wound healing in diabetes are unclear. We used streptozotocin-induced diabetes as a mouse model to investigate the effects of oral administration of an EPA-rich oil on wound closure and quality of new tissue formed. Gas chromatography analysis of serum and skin showed that EPA-rich oil increased the incorporation of ω-3 and decreased ω-6 fatty acids, resulting in reduction of the ω-6/ω-3 ratio. On the tenth day after wounding, EPA increased production of IL-10 by neutrophils in the wound, reduced collagen deposition, and ultimately delayed wound closure and impaired quality of the healed tissue. This effect was PPAR-γ-dependent. EPA and IL-10 reduced collagen production by fibroblasts in vitro. In vivo, topical PPAR-γ-blockade reversed the deleterious effects of EPA on wound closure and on collagen organization in diabetic mice. We also observed a reduction in IL-10 production by neutrophils in diabetic mice treated topically with the PPAR-γ blocker. These results show that oral supplementation with EPA-rich oil impairs skin wound healing in diabetes, acting on inflammatory and non-inflammatory cells.

Introduction: Silver nanoparticles (AgNPs) are of particular interest for their antibacterial properties and are produced by the action of reducing agents on silver ions. Curcumin from Curcuma longa (Zingiberaceae) has been used as a precursor for obtaining biogenic AgNPs, to act as a potential drug. Objectives: This study aimed to evaluate the toxicity of AgNPs synthesized with curcumin (Cur-AgNPs 0.081 mg/mL, ~130 nm) through the Salmonella/microsome (Ames test), one of the first required assays for evaluating toxicity. Methods: The study design was experimental and in vitro. After defining the preliminary toxicity, the mutagenicity was assessed in a concentration range of 0.0010–0.0081 mg/plate Cur-AgNPs using histidine negative (His−) Salmonella Typhimurium strains TA97a, TA98, TA100, and TA102, with (+S9) and without metabolic activation (−S9), in triplicate. Assays were monitored by positive and negative controls. The results were statistically analyzed by Salanal software with p

The detection of Zika virus (ZIKV) in immunoprivileged anatomical sites, potential sites for viral persistence, may guide the confirmation of undefined cases of ZIKV infection and also bring to light unknown pathways of viral transmission. Thus, this study aimed to characterize ZIKV infection in stratified, standardized placental samples in women with exanthematic febrile manifestations during pregnancy and compare findings to the standard investigation protocol of official health agencies. To this end, a case series of placental findings within a prospective cohort study was conducted over a period of 24 months. Serum/urine were obtained at the time of clinical case identification. Placental sampling was performed following standard investigation protocol (samples of 1.0 cm sent to a reference laboratory) and in a systematic way at various regions, such as chorionic plate, chorionic villi, basal plate, amniotic membrane, and umbilical cord, for subsequent ZIKV identification and quantification. Clinical information was obtained and histological preparation with hematoxylin–eosin staining for morphological evaluation was performed. This case series included 17 placentas systematically collected. Of these, 14 were positive by qRT-PCR for ZIKV, 5 in the umbilical cord, 7 in the amniotic membrane, 7 in the chorionic plate, 13 in the chorionic villi, and 7 in the basal plate, whereas none were reported by the reference laboratory. The most common morphological and anatomopathological findings were increased stromal cellularity, villitis, calcification, maternal vascular malperfusion, placental hypoplasia, and maternal–fetal hemorrhage (intervillous thrombi). Seven women presented positive testing for ZIKV in serological and/or molecular tests during gestation in urine. While viral quantification in urine ranged from 101 to 103 FFU eq/ml, that in different placental regions ranged from 103 to 108 FFU eq/g. Thus, ZIKV can infect different regions of the placenta and umbilical cord of pregnant women, showing that the systematic collection and adequate storage of the placenta is fundamental for the detection of ZIKV in this organ. The detection of ZIKV in the placenta after several months of initial symptoms suggests that this tissue may be a site for viral persistence during pregnancy.

The teratogenic mechanisms triggered by ZIKV are still obscure due to the lack of a suitable animal model. Here we present a mouse model of developmental disruption induced by ZIKV hematogenic infection. The model utilizes immunocompetent animals from wild-type FVB/NJ and C57BL/6J strains, providing a better analogy to the human condition than approaches involving immunodeficient, genetically modified animals, or direct ZIKV injection into the brain. When injected via the jugular vein into the blood of pregnant females harboring conceptuses from early gastrulation to organogenesis stages, akin to the human second and fifth week of pregnancy, ZIKV infects maternal tissues, placentas and embryos/fetuses. Early exposure to ZIKV at developmental day 5 (second week in humans) produced complex manifestations of anterior and posterior dysraphia and hydrocephalus, as well as severe malformations and delayed development in 10.5 days post-coitum (dpc) embryos. Exposure to the virus at 7.5-9.5 dpc induces intra-amniotic hemorrhage, widespread edema, and vascular rarefaction, often prominent in the cephalic region. At these stages, most affected embryos/fetuses displayed gross malformations and/or intrauterine growth restriction (IUGR), rather than isolated microcephaly. Disrupted conceptuses failed to achieve normal developmental landmarks and died in utero. Importantly, this is the only model so far to display dysraphia and hydrocephalus, the harbinger of microcephaly in humans, as well as arthrogryposis, a set of abnormal joint postures observed in the human setting. Late exposure to ZIKV at 12.5 dpc failed to produce noticeable malformations. We have thus characterized a developmental window of opportunity for ZIKV-induced teratogenesis encompassing early gastrulation, neurulation and early organogenesis stages. This should not, however, be interpreted as evidence for any safe developmental windows for ZIKV exposure. Late developmental abnormalities correlated with damage to the placenta, particularly to the labyrinthine layer, suggesting that circulatory changes are integral to the altered phenotypes.

Royal Jelly (RJ) is widely consumed in diets throughout the world due to its beneficial effects: antioxidant, antitumor and anti-inflammatory. We have investigated the role of RJ in the development of TNBS colitis in mice. Colitis was induced by a rectal instillation of TNBS at 0.1 mL per mouse. Intestine samples of the animals orally treated with RJ (100, 150, and 200 mg/kg) were collected for antioxidant assays (GSH and GSH-Px), proinflammatory protein quantification (COX-2 and NF-κB), and histological analyses. RJ 100 mg/kg maintained GSH levels and increased the activity of GSH-Px, downregulated key inflammatory mediators (COX-2 and NF-κB), and decreased the lesions caused by TNBS as shown by the histological analyses. In conclusion, RJ showed anti-inflammatory and antioxidant properties in experimental colitis, resulting in the amelioration of the macroscopic and histological analyses. These results corroborate with the RJ supplementation in diets.

Clinical and experimental data indicate that severe acute respiratory syndrome coronavirus (SARS-CoV)-2 infection is associated with significant changes in the composition and function of intestinal microbiota. However, the relevance of these effects for SARS-CoV-2 pathophysiology is unknown. In this study, we analyzed the impact of microbiota depletion after antibiotic treatment on the clinical and immunological responses of K18-hACE2 mice to SARS-CoV-2 infection. Mice were treated with a combination of antibiotics (kanamycin, gentamicin, metronidazole, vancomycin, and colistin, Abx) for 3 days, and 24 h later, they were infected with SARS-CoV-2 B lineage. Here, we show that more than 80% of mice succumbed to infection by day 11 post-infection. Treatment with Abx had no impact on mortality. However, Abx-treated mice presented better clinical symptoms, with similar weight loss between infected–treated and non-treated groups. We observed no differences in lung and colon histopathological scores or lung, colon, heart, brain and kidney viral load between groups on day 5 of infection. Despite some minor differences in the expression of antiviral and inflammatory markers in the lungs and colon, no robust change was observed in Abx-treated mice. Together, these findings indicate that microbiota depletion has no impact on SARS-CoV-2 infection in mice.

Colorectal cancer (CRC) is in the top 10 cancers most prevalent worldwide, affecting equally men and women. Current research on tumor-derived extracellular vesicles (EVs) suggests that these small extracellular vesicles (sEVs) play an important role in mediating cell-to-cell communication and thus potentially affecting cancer progression via multiple pathways. In the present study, we hypothesized that sEVs derived from different CRC cell lines differ in their ability to reprogram normal human fibroblasts through a process called tumor education. The sEVs derived from CRC cell lines (HT29 and HCT116) were isolated by a combination of ultrafiltration and polymeric precipitation, followed by characterization based on morphology, size, and the presence or absence of EV and non-EV markers. It was observed that the HT29 cells displayed a higher concentration of sEVs compared with HCT116 cells. For the first time, we demonstrated that HT29-derived sEVs were positive for low-molecular-weight protein tyrosine phosphatase (Lmwptp). CRC cell-derived sEVs were uptake by human fibroblasts, stimulating migratory ability via Rho-Fak signaling in co-incubated human fibroblasts. Another important finding showed that HT29 cell-derived sEVs are much more efficient in activating human fibroblasts to cancer-associated fibroblasts (CAFs). Indeed, the sEVs produced by the HT29 cells that are less responsive to a cytotoxic agent display higher efficiency in educating normal human fibroblasts by providing them advantages such as activation and migratory ability. In other words, these sEVs have an influence on the CRC microenvironment, in part, due to fibroblasts reprogramming.

Abstract The active transport of glycolytic pyruvate across the inner mitochondrial membrane is thought to involve two mitochondrial pyruvate carrier subunits, MPC1 and MPC2, assembled as a 150 kDa heterotypic oligomer. Here, the recombinant production of human MPC through a co-expression strategy is first described; however, substantial complex formation was not observed, and predominantly individual subunits were purified. In contrast to MPC1, which co-purifies with a host chaperone, we demonstrated that MPC2 homo-oligomers promote efficient pyruvate transport into proteoliposomes. The derived functional requirements and kinetic features of MPC2 resemble those previously demonstrated for MPC in the literature. Distinctly, chemical inhibition of transport is observed only for a thiazolidinedione derivative. The autonomous transport role for MPC2 is validated in cells when the ectopic expression of human MPC2 in yeast lacking endogenous MPC stimulated growth and increased oxygen consumption. Multiple oligomeric species of MPC2 across mitochondrial isolates, purified protein and artificial lipid bilayers suggest functional high-order complexes. Significant changes in the secondary structure content of MPC2, as probed by synchrotron radiation circular dichroism, further supports the interaction between the protein and ligands. Our results provide the initial framework for the independent role of MPC2 in homeostasis and diseases related to dysregulated pyruvate metabolism.

Skeletal muscle atrophy occurs in several pathological conditions, such as cancer, especially during cancer-induced cachexia. This condition is associated with increased morbidity and poor treatment response, decreased quality of life, and increased mortality in cancer patients. A leucine-rich diet could be used as a coadjutant therapy to prevent muscle atrophy in patients suffering from cancer cachexia. Besides muscle atrophy, muscle function loss is even more important to patient quality of life. Therefore, this study aimed to investigate the potential beneficial effects of leucine supplementation on whole-body functional/movement properties, as well as some markers of muscle breakdown and inflammatory status. Adult Wistar rats were randomly distributed into four experimental groups. Two groups were fed with a control diet (18% protein): Control (C) and Walker 256 tumour-bearing (W), and two other groups were fed with a leucine-rich diet (18% protein + 3% leucine): Leucine Control (L) and Leucine Walker 256 tumour-bearing (LW). A functional analysis (walking, behaviour, and strength tests) was performed before and after tumour inoculation. Cachexia parameters such as body weight loss, muscle and fat mass, pro-inflammatory cytokine profile, and molecular and morphological aspects of skeletal muscle were also determined. As expected, Walker 256 tumour growth led to muscle function decline, cachexia manifestation symptoms, muscle fibre cross-section area reduction, and classical muscle protein degradation pathway activation, with upregulation of FoxO1, MuRF-1, and 20S proteins. On the other hand, despite having no effect on the walking test, inflammation status or muscle oxidative capacity, the leucine-rich diet improved muscle strength and behaviour performance, maintained body weight, fat and muscle mass and decreased some protein degradation markers in Walker 256 tumour-bearing rats. Indeed, a leucine-rich diet alone could not completely revert cachexia but could potentially diminish muscle protein degradation, leading to better muscle functional performance in cancer cachexia.

Summary: Antibiotic-induced dysbiosis is a key factor predisposing intestinal infection by Clostridium difficile. Here, we show that interventions that restore butyrate intestinal levels mitigate clinical and pathological features of C. difficile-induced colitis. Butyrate has no effect on C. difficile colonization or toxin production. However, it attenuates intestinal inflammation and improves intestinal barrier function in infected mice, as shown by reduced intestinal epithelial permeability and bacterial translocation, effects associated with the increased expression of components of intestinal epithelial cell tight junctions. Activation of the transcription factor HIF-1 in intestinal epithelial cells exerts a protective effect in C. difficile-induced colitis, and it is required for butyrate effects. We conclude that butyrate protects intestinal epithelial cells from damage caused by C. difficile toxins via the stabilization of HIF-1, mitigating local inflammatory response and systemic consequences of the infection. : Fachi et al. demonstrate that butyrate is able to protect the intestinal epithelium from the damage caused by Clostridium difficile toxins by stabilizing HIF-1 and increasing tight junctions, which reduces intestinal epithelial permeability, thus inhibiting intestinal inflammation and bacterial translocation. Keywords: short chain fatty acids, microbiota, hypoxia, colitis, intestinal epithelial cells, infection

Animal models commonly serve as a bridge between in vitro experiments and clinical applications; however, few physiological processes in adult animals are sufficient to serve as proof-of-concept models for cartilage regeneration. Intriguingly, some rodents, such as young adult mice, undergo physiological connective tissue modifications to birth canal elements such as the pubic symphysis during pregnancy; therefore, we investigated whether the differential expression of cartilage differentiation markers is associated with cartilaginous tissue morphological modifications during these changes. Our results showed that osteochondral progenitor cells expressing Runx2, Sox9, Col2a1 and Dcx at the non-pregnant pubic symphysis proliferated and differentiated throughout pregnancy, giving rise to a complex osteoligamentous junction that attached the interpubic ligament to the pubic bones until labour occurred. After delivery, the recovery of pubic symphysis cartilaginous tissues was improved by the time-dependent expression of these chondrocytic lineage markers at the osteoligamentous junction. This process potentially recapitulates embryologic chondrocytic differentiation to successfully recover hyaline cartilaginous pads at 10 days postpartum. Therefore, we propose that this physiological phenomenon represents a proof-of-concept model for investigating the mechanisms involved in cartilage restoration in adult animals.