Carboxylesterase 2 (CES2), an endoplasmic reticulum (ER) located phase I enzyme, plays a vital role in the metabolism of various endogenous and exogenous substances, and is regarded as an important target for the design of prodrugs. Unfortunately, superior highly selective ER targeting fluorescent probes for monitoring of CES2 are not currently available. Herein, we report an ER targeting CES2 selective and sensitive ratiometric fluorescent probe ERNB based on the ER localizing group p-toluenesulfonamide. ERNB possessed high specificity, sensitivity, and exhibited excellent subcellular localization when compared to commercial ER tracker, and was used to image CES2 in the ER of living cells. Additionally, using ERNB we evaluated the CES2 regulation under d,l-dithiothreitol and tunicamycin-induced ER stress. Furthermore, we determined the down regulation of CES2 activity and expression in the acetaminophen-induced acute liver injury model. On the basis of these results, we conclude that ERNB is a promising tool for highlighting the role of CES2 in the ER and in exploring the role of CES2 in the development of diseases associated with ER stress.

Uveal melanoma (UVM), the most common primary intraocular malignancy, has a high mortality because of a high propensity to metastasize. Our study analyzed prognostic value and immune-related characteristics of CARD11 in UVM, hoping to provide a potential management and research direction. The RNA-sequence data of 80 UVM patients were downloaded from The Cancer Genome Atlas database and divided them into high- and low-expression groups. We analyzed the differentially expressed genes, enrichment analyses and the infiltration of immune cells using the R package and Gene-Set Enrichment Analysis. A clinical prediction nomogram and protein-protein interaction network were constructed and the first 8 genes were considered as the hub-genes. Finally, we constructed a competing endogenous RNA (ceRNA) network by Cytoscape and analyzed the statistical data via the R software. Here we found that CARD11 expression had notable correlation with UVM clinicopathological features, which was also an independent predictor for overall survival (OS). Intriguingly, CARD11 had a positively correlation to autophagy, cellular senescence and apoptosis. Infiltration of monocytes was significantly higher in low CARD11 expression group, and infiltration of T cells regulatory was lower in the same group. Functional enrichment analyses revealed that CARD11 was positively related to T cell activation pathways and cell adhesion molecules. The expressions of hub-genes were all increased in the high CARD11 expression group and the ceRNA network showed the interaction among mRNA, miRNA and lncRNA. These findings show that high CARD11 expression in UVM is associated with poor OS, indicating that CARD11 may serve as a potential biomarker for the diagnosis and prognosis of the UVM.

Scope: The use of human milk products is increasing for high-risk infants. Human milk contains endogenous enzymes that comprise a dynamic proteolytic system, yet biological properties of these enzymes and their activities in response to variations including pH within infants are unclear. Human milk has a neutral pH around 7, while infant gastric pH varies from 2 to 6 depending on individual conditions. This study is designed to determine the specificity of enzyme-substrate interactions in human milk as a function of pH. Methods and results: Endogenous proteolysis is characterized by incubating freshly expressed human milk at physiologically relevant pH ranging from 2 to 7 without the addition of exogenous enzymes. Results show that the effects of pH on endogenous proteolysis in human milk are protein-specific. Further, specific interactions between cathepsin D and α-lactalbumin are confirmed. The endogenous enzyme cathepsin D in human milk cleaves α-lactalbumin as the milk pH shifts from 7 to 3. Conclusions: This study documents that selective proteolysis activated by pH shift is a mechanism for dynamic interactions between human milk and the infant. Controlled proteolysis can guide the use of human milk products based on individual circumstance.

Objectives The objective of this study was to determine whether a novel fiber formulation improves glucose, insulin, and lipid profiles in overweight men and women consuming a low fiber diet. Methods Twenty individuals were enrolled in this randomized order, placebo-controlled, cross-over study. Participants were young, healthy, overweight (BMI 23.0–32.0) and consumed <15 g/day of fiber. All participants consumed the fiber and placebo supplement for a period of 4 weeks each, with a 4-week washout between intervention arms. Participants recorded their diet for 3 days using dietary records twice during each 4-week segment. They consumed either fiber and/or placebo packet containing a total of 12 g/serving per day. The Fiber and/or Placebo was given out as powder form which include mostly dietary fiber (resistant starch, fructooligosaccharide, sugarcane fiber, and inulin), rice flour, xanthan gum, and fruit powders that was mixed with water for consumption. Questionnaires, anthropometric measurements, blood draws, and stool samples were collected at each study visit. Changes in glucose, insulin, and lipid profile (total cholesterol (TC), triacylglycerols (TG), HDL-C and calculated LDL-C) were assessed using a linear mixed model. Results The mean change in fasted glucose, insulin, and lipid profiles showed a tendency to decrease in response to fiber consumption compared with the placebo but did not meet statistical significance (P = 0.29, 0.42, and 0.61) due to high interindividual variability. This clinical trial was registered at clinicaltrials.gov as NCT03785860. Conclusions Cardiometabolic profiles did not change in response to the fiber supplement. Funding Sources I would like to acknowledge Usana Health Sciences, Inc. for the support in this research.

No abstract