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Background Methods Results Conclusion Accumulating evidence has suggested that neuropeptides such as orexin, ghrelin, or oxytocin act centrally in the brain to regulate intestinal barrier function through the vagus nerve. It has been reported that the vagal cholinergic anti‐inflammatory pathway was blocked by splenectomy. In the present study, we therefore examined the effect of splenectomy on neuropeptides‐induced improvement of increased intestinal permeability.Colonic permeability was determined in vivo by quantifying the absorbed Evans blue in colonic tissue for 15 min spectrophotometrically in rats.Splenectomy increased colonic permeability. The increased permeability by splenectomy was significantly blocked by vagal activation induced by carbachol or 2‐deoxy‐d‐glucose which was prevented by atropine, suggesting vagal activation could prevent colonic hyperpermeability in splenectomized rats. In the splenectomized rats, intracisternal injection of orexin, ghrelin, oxytocin, or butyrate failed to inhibit increased colonic permeability while intracisternal glucagon‐like peptide‐1 (GLP‐1) analogue, liraglutide, potently blocked the increased colonic permeability in a dose‐dependent manner. The liraglutide‐induced improvement of increased colonic permeability was blocked by atropine in splenectomized rats. Intracisternal injection of GLP‐1 receptor antagonist attenuated 2‐deoxy‐d‐glucose‐induced improvement of colonic hyperpermeability in splenectomized rats.The present results suggested that the spleen is important in the improvement of intestinal barrier function by brain orexin, ghrelin or oxytocin, and butyrate. On the other hand, GLP‐1 acts centrally in the brain to improve colonic hyperpermeability in a spleen‐independent manner. All these results suggest that dual mechanisms (spleen dependent or independent) may exist for the brain‐gut regulation in intestinal barrier function. [ABSTRACT FROM AUTHOR]

• Biomarkers to predict outcomes were assessed from the clinico-statistical approach. • A prospective multicenter observational study was performed. • Six promising biomarkers were successfully identified and validated. • C-reactive protein to platelet ratio was a novel biomarker to predict transplant outcomes. Various biomarkers have been developed and evaluated to predict the prognosis and complications of allogeneic hematopoietic cell transplantation (HCT). Most previous studies conducted on different biomarkers evaluated single effects such as those associated with inflammation, immunology, iron metabolism, and nutrition, and only a few studies have comprehensively analyzed markers. The study aimed to survey comprehensive multiple markers prior to HCT and extract those that significantly predict the outcomes. A prospective multicenter observational study was performed. (UMIN000013506) Patients undergoing HCT for hematologic diseases were consecutively enrolled. Besides the usual clinical biomarkers, serum samples for extra-clinical biomarkers were collected and cryopreserved before starting the conditioning regimen. A total of 32 candidate biomarkers were selected, 23 from hematology, biochemistry, immunology, nutrition, and iron metabolism, and 9 from composite markers. Based on the area under the curve (AUC) values for survival, promising biomarkers was extracted. Internal validation for these markers was applied based on bootstrap methods. Setting the cut-off values for them, log-rank test was applied and outcomes including overall survival (OS), relapse, and non-relapse mortality (NRM) were evaluated using multivariate analyses. Furthermore, detailed analysis including transplant-related complications and external validation were conducted focusing on C-reactive protein (CRP) to platelet (Plt) ratio. A total of 152 patients with hematologic malignancies were enrolled from April 2014 to March 2017. CRP, soluble interleukin-2 receptor (IL2R), CRP to albumin (Alb) ratio, CRP to Plt ratio, Plt to IL2R ratio, and IL2R to Alb ratio were identified as promising markers. Internal validation successfully confirmed their reliability of AUC and multivariate analysis demonstrated the statistical significance between the higher and the lower markers. Above all, a higher CRP to Plt ratio was significantly associated with a lower OS (hazard ratio [HR] 2.77; 95% confidence interval [CI] 1.30−5.91; P = 0.008) and higher non-relapse mortality rates (HR 2.79; 95%CI 1.14−6.80; P = 0.024) at 180 days. Furthermore, univariate analysis showed that a higher CRP to Plt ratio was significantly associated with a higher incidence of sinusoidal obstructive syndrome (P < 0.001) and bloodstream infection (P = 0.027). An external validation test confirmed the significance of the CRP to Plt ratio for these outcomes. The multicenter prospective observational study successfully identified significant biomarkers in patients with hematologic malignancies who received HCT. In particular, CRP to Plt ratio was identified as a novel and useful biomarker for predicting transplant outcomes. Further investigations are needed to validate the novel markers, analysis of the pathophysiology, and application to treatment settings other than HCT. [ABSTRACT FROM AUTHOR]

Background: As the saturation of transferrin by iron in the serum is approximately 30%, iron loaded to the blood can bind to transferrin not bearing iron. Nevertheless, prolonged iron influx finally results in full transferrin saturation, and iron not bound to transferrin will appear in the serum; this iron is known as non-transferrin-bound iron (NTBI). NTBI damages organs through the production of free radicals. Previously, we established an automated quantification system for NTBI; however, measuring labile plasma iron, which is considered as a highly redox-active component of NTBI, should be a better prognostic factor in iron-overloaded patients. Methods: We designed and developed a novel system for evaluating labile plasma iron utilizing the Trinder reaction. Automated system was utilized because the previously reported methods for labile plasma iron are intricate and the introduction to the clinical stage has been challenging. Validations such as the contribution of serum proteins and metal ions for this system were evaluated using human serum samples. Results: We confirmed that our novel system can evaluate labile plasma iron utilizing Trinder reaction and the oxidative potential of ceruloplasmin in the serum. This system was also confirmed to be clinically practical. Metals other than iron did not influence this system. We observed that samples with high NTBI did not always exhibit high labile plasma iron and vice versa, highlighting the necessity of labile plasma iron quantification in evaluating the toxicity of NTBI. Conclusions: Our novel system should contribute to fundamental and clinical research because it can measure labile plasma iron using the high-throughput automated analyser. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM), a neoplasm of plasma cells (PCs), is a highly heterogeneous disease with multifocal dissemination throughout the body. Minimal residual disease (MRD) detected using PCs in bone marrow (BM) is important for MM management; however, frequent invasive examinations impose a significant burden on patients. Analysis using plasma cell-free DNA (cfDNA) might represent an alternative tool for disease monitoring. In this study, we observed the disease status in a patient with MM by examining the KRAS mutation allele frequency (MAF) in plasma cfDNA using digital PCR. During treatment, the MAF was correlated with serum immunoglobulin A and free light chain-kappa levels. After the second autologous peripheral blood stem cell transplantation, the KRAS MAF became immediately positive after confirming MRD negativity using PCs from BM. Shortly thereafter, the patient experienced clinical relapse primarily involving bone lesions. Mutant KRAS monitoring in cfDNA using serial blood collection might reflect the disease status more accurately than invasive BM examinations, especially in patients with MM whose primary lesions have extra-BM locations. It could also help predict treatment responses and outcomes. [ABSTRACT FROM AUTHOR]

[Display omitted] • Intracisternal injection of GLP-1 analogue improved leaky gut. • Central GLP-1-induced improvement of leaky gut was blocked by atropine or vagotomy. • Orexin 1 receptor antagonist blocked the improvement of leaky gut by central GLP-1. • GLP-1acts centrally to improve leaky gut via brain orexin and the vagal signaling. Leaky gut, an altered intestinal barrier function, has been described in many diseases such as irritable bowel syndrome (IBS). We have recently demonstrated that orexin in the brain blocked leaky gut in rats, suggesting that the brain plays a role in regulation of intestinal barrier function. In the present study, we tried to clarify whether GLP-1 acts centrally in the brain to regulate intestinal barrier function and its mechanism. Colonic permeability was estimated in vivo by quantifying the absorbed Evans blue in colonic tissue in rats. Intracisternal injection of GLP-1 analogue, liraglutide dose-dependently abolished increased colonic permeability in response to lipopolysaccharide. Either atropine or surgical vagotomy blocked the central GLP-1-induced improvement of colonic hyperpermeability. Intracisternal GLP-1 receptor antagonist, exendin (9–39) prevented the central GLP-1-induced blockade of colonic hyperpermeability. In addition, intracisternal injection of orexin receptor antagonist, SB-334867 blocked the GLP-1-induced improvement of intestinal barrier function. On the other hand, subcutaneous liraglutide also improved leaky gut but larger doses of liraglutide were needed to block it. In addition, neither atropine nor vagotomy blocked subcutaneous liraglutide-induced improvement of leaky gut, suggesting that central or peripheral GLP-1 system works separately to improve leaky gut in a vagal-dependent or independent manner, respectively. These results suggest that GLP-1 acts centrally in the brain to reduce colonic hyperpermeability. Brain orexin signaling and the vagal cholinergic pathway play a vital role in the process. We would therefore suggest that activation of central GLP-1 signaling may be useful for leaky gut-related diseases such as IBS. [ABSTRACT FROM AUTHOR]

Hepcidin is a main regulator of iron metabolism, of which abnormal expression affects intestinal absorption and reticuloendothelial sequestration of iron by interacting with ferroportin. It is also noted that abnormal iron accumulation is one of the key factors to facilitate promotion and progression of cancer including hepatoma. By RT-PCR/agarose gel electrophoresis of hepcidin mRNA in a hepatocellular carcinoma cell line HLF, a smaller mRNA band was shown in addition to the wild-type hepcidin mRNA. From sequencing analysis, this additional band was a selective splicing variant of hepcidin mRNA lacking exon 2 of HAMP gene, producing the transcript that encodes truncated peptide lacking 20 amino acids at the middle of preprohepcidin. In the present study, we used the digital PCR, because such a small amount of variant mRNA was difficult to quantitate by the conventional RT-PCR amplification. Among seven hepatoma-derived cell lines, six cell lines have significant copy numbers of this variant mRNA, but not in one cell line. In the transient transfection analysis of variant-type hepcidin cDNA, truncated preprohepcidin has a different character comparing with native preprohepcidin: its product is insensitive to digestion, and secreted into the medium as a whole preprohepcidin form without maturation. Loss or reduction of function of HAMP gene by aberrantly splicing may be a suitable phenomenon to obtain the proliferating advantage of hepatoma cells. [ABSTRACT FROM AUTHOR]

Background Iron is an essential metal in the body, but its excessive accumulation causes damage in various organs through free radical production. Iron homeostasis is therefore tightly regulated. However, when iron balance collapses, such as in prolonged transfusion, transferrin (Tf) is fully saturated and non-Tf-bound iron (NTBI) appears in the serum. Monitoring serum NTBI levels is therefore crucial in the assessment of the clinical status of patients with iron overload, since NTBI is associated with cellular and organ damage. Several methods for NTBI determination have been reported, but these are extremely complicated and very few laboratories can quantify NTBI at present. Methods We established a novel assay system utilizing automated analyzers that are widely used in clinical laboratories for diagnostic testing. In this assay, NTBI is chelated by nitrilotriacetic acid (NTA), after which the iron is reduced and transferred to nitroso-PSAP, a chromogen. Results The assay shows excellent linearity, reproducibility, and compatibility with HPLC, one of the most reliable conventional methods for NTBI quantification. Conclusions Our novel method for NTBI measurement is high-throughput and may be a useful and powerful tool in the study of the physiological and clinical importance of NTBI. [ABSTRACT FROM AUTHOR]

Orexins are neuropeptides implicated in several physiological functions. Accumulating findings suggest a relationship between orexin and sepsis. A recent study demonstrated that orexin acts centrally to improve conditions in sepsis. The present study aims to clarify the precise mechanisms by which central orexin could induce a protective action against septic conditions. We established a new septic model by treating rats with lipopolysaccharide (LPS) and colchicine and used this to examine the effect of brain orexin on survival. Observation of survival was stopped three days after the chemicals injection or at death. We established a lethal model (rats died within 24 h) by injecting subcutaneously a combination of 1 mg/kg LPS and 1 mg/kg colchicine. A Toll-like receptor 4 (TLR4) inhibitor completely blocked lethality, suggesting a vital role of LPS-TLR4 signaling in the process. Intracisternal orexin-A dose-dependently reduced lethality in the sepsis model while neither intracisternal orexin-B nor intraperitoneal orexin-A changed the mortality rate. Vagal stimulation with carbachol or 2-deoxy-D-glucose improved survival and atropine potently blocked the protection by carbachol or 2-deoxy-D-glucose. The orexin-A-induced reduction of lethality was significantly blocked by atropine or surgical vagotomy. Intracisternal injection of an OX 1 receptor antagonist blocked the improvement of survival by intracisternal injection of orexin-A, carbachol, or 2-deoxy-D-glucose. These results suggest that orexin acts centrally to reduce the lethality in our septic model treated (LPS and colchicine). Activation of the vagal cholinergic pathway may mediate the action of orexin, and the OX 1 receptor in the brain might play a role in the process. Since the efferent vagus nerve mediates anti-inflammatory mechanisms, we speculate that the vagal cholinergic anti-inflammatory pathway is implicated in the mechanisms of septic lethality reduction by brain orexin. [ABSTRACT FROM AUTHOR]