Adipose tissue inflammation drives obesity-related cardiometabolic diseases. Enhancing endogenous resolution mechanisms through administration of lipoxin A4, a specialized pro-resolving lipid mediator, was shown to reduce adipose inflammation and subsequently protects against obesity-induced systemic disease in mice. Here, we demonstrate that lipoxins reduce inflammation in 3D-cultured human adipocytes and adipose tissue explants from obese patients. Approximately 50% of patients responded particularly well to lipoxins by reducing inflammatory cytokines and promoting an anti-inflammatory M2 macrophage phenotype. Responding patients were characterized by elevated systemic levels of C-reactive protein, which causes inflammation in cultured human adipocytes. Responders appeared more prone to producing anti-inflammatory oxylipins and displayed elevated prostaglandin D2 levels, which has been interlinked with transcription of lipoxin-generating enzymes. Using explant cultures, this study provides the first proof-of-concept evidence supporting the therapeutic potential of lipoxins in reducing human adipose tissue inflammation. Our data further indicate that lipoxin treatment may require a tailored personalized-medicine approach.
Dysregulated chronic inflammation plays a crucial role in the pathophysiology of atherosclerosis and may be a result of impaired resolution. Thus, restoring levels of specialized pro-resolving mediators (SPMs) to promote the resolution of inflammation has been proposed as a therapeutic strategy for patients with atherosclerosis, in addition to standard clinical care. Herein, we evaluated the effects of the SPM lipids, lipoxin A4 (LXA4) and lipoxin B4 (LXB4), on neutrophils isolated from patients with atherosclerosis compared with healthy controls. Patients displayed altered endogenous SPM production, and we demonstrated that lipoxin treatment in whole blood from atherosclerosis patients attenuates neutrophil oxidative burst, a key contributor to atherosclerotic development. We found the opposite effect in neutrophils from healthy controls, indicating a potential mechanism whereby lipoxins aid the endogenous neutrophil function in health but reduce its excessive activation in disease. We also demonstrated that lipoxins attenuated upregulation of the high-affinity conformation of the CD11b/CD18 integrin, which plays a central role in clot activation and atherosclerosis. Finally, LXB4 enhanced lymphatic transmigration of human neutrophils isolated from patients with atherosclerosis. This finding is noteworthy, as impaired lymphatic function is now recognized as an important contributor to atherosclerosis. Although both lipoxins modulated neutrophil function, LXB4 displayed more potent effects than LXA4 in humans. This study highlights the therapeutic potential of lipoxins in atherosclerotic disease and demonstrates that the effect of these SPMs may be specifically tailored to the need of the individual.
Communication between mother and offspring in mammals starts at implantation via the maternal–placental–fetal axis, and continues postpartum via milk targeted to the intestinal mucosa. MicroRNAs (miRNAs), short, noncoding single-stranded RNAs, of about 22 nucleotides in length, are actively involved in many developmental and physiological processes. Here we highlight the role of miRNA in the dynamic signaling that guides infant development, starting from implantation of conceptus and persisting through the prenatal and postnatal periods. miRNAs in body fluids, particularly in amniotic fluid, umbilical cord blood, and breast milk may offer new opportunities to investigate physiological and/or pathological molecular mechanisms that portend to open novel research avenues for the identification of noninvasive biomarkers.
Three members of the obscurin protein family that contain tandem kinase domains with important signaling functions for cardiac and striated muscles are the giant protein obscurin, its obscurin-associated kinase splice isoform, and the striated muscle enriched protein kinase (SPEG). While there is increasing evidence for the specific roles that each individual kinase domain plays in cross-striated muscles, their biology and regulation remains enigmatic. Our present study focuses on kinase domain 1 and the adjacent low sequence complexity inter-kinase domain linker in obscurin and SPEG. Using Phos-tag gels, we show that the linker in obscurin contains several phosphorylation sites, while the same region in SPEG remained unphosphorylated. Our homology modeling, mutational analysis and molecular docking demonstrate that kinase 1 in obscurin harbors all key amino acids important for its catalytic function and that actions of this domain result in autophosphorylation of the protein. Our bioinformatics analyses also assign a list of putative substrates for kinase domain 1 in obscurin and SPEG, based on the known and our newly proposed phosphorylation sites in muscle proteins, including obscurin itself.
Abstract Background It remains unclear to what extent the SARS-CoV-2/COVID-19 pandemic disrupted the normal progression of biomedical and medical science graduate programs and if there was a lasting impact on the quality and quantity of supervision of PhD-students. To date, multiple editorials and commentaries indicate the severity of the disruption without providing sufficient evidence with quantifiable data. Methods An online survey was submitted to the administrative offices of biomedical and medical PhD-programs at eight major universities in Sweden to gauge the impact of the pandemic on the students. It consisted of multiple-choice and open-ended questions where students could provide examples of positive and/or negative supervision strategies. Open answered questions were coded as either examples of positive or negative support. Results PhD students were divided into two groups: those with improved or unchanged supervision during the pandemic (group 1, n = 185), versus those whose supervision worsened (group 2, n = 69). Group 1 received more help from supervisors and more frequent supervision via both online and alternative platforms (email/messages and telephone). There was no significant difference in educational-stage, gender or caretaking responsibilities between the groups. Conclusions It is important for the scientific community to learn how to provide the best possible supervision for PhD students during the pandemic. Our data suggests that more frequent supervision, and using a diverse array of meeting platforms is helpful. In addition, it is important for the students to feel that they have their supervisor’s emotional support. Several students also expressed that they would benefit from an extension of their PhD programs due to delays caused by the pandemic.
Proteomics can map extracellular vesicles (EVs), including exosomes, across disease states between organisms and cell types. Due to the diverse origin and cargo of EVs, tailoring methodological and analytical techniques can support the reproducibility of results. Proteomics scans are sensitive to in-sample contaminants, which can be retained during EV isolation procedures. Contaminants can also arise from the biological origin of exosomes, such as the lipid-rich environment in human milk. Human milk (HM) EVs and exosomes are emerging as a research interest in health and disease, though the experimental characterization and functional assays remain varied. Past studies of HM EV proteomes have used data-dependent acquisition methods for protein detection, however, improvements in data independent acquisition could allow for previously undetected EV proteins to be identified by mass spectrometry. Depending on the research question, only a specific population of proteins can be compared and measured using isotope and other labelling techniques. In this review, we summarize published HM EV proteomics protocols and suggest a methodological workflow with the end-goal of effective and reproducible analysis of human milk EV proteomes.
Breast milk is an intricate source of key nutritional complexes required to nourish an infant as well as facilitate the development of the infant's immune system. During the postnatal period, the gastrointestinal tract (GIT) matures following enteral feeds and colonization with commensal bacteria. The development of the GIT could be delayed or altered in a premature infant, thus predisposing the infant to inflammatory diseases such as necrotizing enterocolitis (NEC). NEC is the leading cause of death among premature infants. The pathophysiology is currently unknown, however, prematurity, formula feeding and an abnormal intestinal microbiota are thought to cause an inflammatory response in the intestine which can lead to NEC and sepsis. Breast milk is the gold standard of infant nutrition and is largely thought to help prevent NEC. The components of breast milk responsible for the development of the GIT have yet to be identified, although, novel research has discovered the presence of microRNA (miRNA) in breast milk exosomes. Due to stable structure, exosomes are protected against degradation, suggesting breast milk exosomal miRNAs continue to be biologically active following ingestion by the infant. miRNAs have previously been described as functioning in cellular communication, implying a method of “listening in” to the cross‐talk between mother and infant via breastfeeding. To characterize miRNAs and identify the role they play in the development of the GIT, exosomes from breast milk must be isolated in a reproducible method to extract miRNAs. Isolating and storing procedures of unprocessed breast milk are underexposed. In recent studies, it has been revealed that storing unprocessed breast milk at −80°C or −4°C lead to cell death within the milk, causing contamination of the native breast milk exosomes. The profiling of miRNA fluctuates depending on the milk fraction analyzed, preparation of milk and exosome fraction as well as the miRNA extraction protocol. Extra care should be taken when preparing samples and following extraction procedures in this area of research. During this early stage of exploring the ways in which the mother communicates with her newborn, it is necessary to collaborate with other research groups to standardize procedures and to synchronize approaches.
After birth, upon exposure to the ex utero environment, the gastrointestinal tract of a newborn transitions from near sterile conditions to being exposed to a plethora of bacteria. In normal, term births the gastrointestinal tract matures through enteral feeds and commensal bacteria colonization. However, premature birth perturbs in utero organ maturation, thus depriving preterm infants of immune tolerance and increasing their susceptibility to gastrointestinal disorders. Human milk is the gold standard of infant nutrition, but in some cases, it is unavailable or insufficient. It is necessary to understand the components of human milk which facilitate development and promote immune tolerance to optimize feeding methods, ensuring best possible outcomes. Human milk exosomes have been shown to have immunomodulatory properties. Previous reports have demonstrated that milk exosomes survive in vivo digestion in animals, in vitro digestion in humans and can be taken up by human intestinal epithelial cells. We hypothesized that exosomes may promote immune tolerance in human intestinal epithelial cells. Exosomes were isolated from human milk through a series of centrifugations and ultracentrifugations. Post‐confluent Caco‐2/15 cells were incubated with either culture medium containing milk exosomes or PBS (control) for 24 hours. Caco‐2/15 cells, once maintained at post‐confluency, recapitulate mid‐gestation small intestine absorptive cells morphologically, functionally and at the gene level. An inflammatory response was induced by incubating the Caco‐2/15 cells with heat‐killed bacteria ( E. coli + S. typhimurium ). Expression of pro‐inflammatory markers, CXCL8, IL‐6 and CXCL10, was significantly decreased in cells pretreated with human milk exosomes compared to the control. Milk exosomes and their cargo caused a dampened immune response in human intestinal epithelial cells, suggesting that they may be promoting immune tolerance in the gastrointestinal tract of newborns. Infants born prematurely would benefit from such effects as their intestine is underdeveloped and more permeable, therefore, more susceptible to react to bacteria, causing an inflammatory cascade that could lead to sepsis. This is a relevant finding in identifying future directions to optimize feeding strategies for preterm infants. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
Dietary lipids can affect metabolic health through gut microbiota-mediated mechanisms, but the influence of lipid-microbiota interaction on liver steatosis is largely unknown. We investigate the impact of dietary lipids on human gut microbiota composition and the effects of microbiota-lipid interactions on steatosis in male mice. In humans, low intake of saturated fatty acids (SFA) is associated with increased microbial diversity independent of fiber intake. In mice, poorly absorbed dietary long-chain SFA, particularly stearic acid, induce a shift in bile acid profile and improved metabolism and steatosis. These benefits are dependent on the gut microbiota, as they are transmitted by microbial transfer. Diets enriched in polyunsaturated fatty acids are protective against steatosis but have minor influence on the microbiota. In summary, we find that diets enriched in poorly absorbed long-chain SFA modulate gut microbiota profiles independent of fiber intake, and this interaction is relevant to improve metabolism and decrease liver steatosis.
